Methods for treatment of splenomegaly in cell based therapeutics

Abstract

Embodiments of the disclosure include methods and compositions for the treatment of splenomegaly. In specific embodiments, fibroblasts and / or fibroblast-derived materials, are utilized in at least some cases to affect the immune response of the individual in order to reduce the pathogenic levels of immune cells or cytokines and / or chemokines excreted by those immune cells.

Description

METHODS FOR TREATMENT OF SPLENOMEGALY IN CELL BASED THERAPEUTICS

[0001] This application claims priority to U.S. Provisional Patent Application Serial No.63 / 709,795, filed October 21, 2024, which is incorporated by reference herein in its entirety.BACKGROUNDI. Technical Field

[0002] This disclosure relates at least to the fields of cell biology, molecular biology, immunology, and medicine.II. Background

[0003] Fibroblasts, traditionally known for their role in tissue repair and structural support, have emerged as key players in immune modulation. They influence both innate and adaptive immune responses through interactions with immune cells, such as T cells, macrophages, and dendritic cells (1-4). This unique immunomodulatory capacity makes fibroblasts a promising therapeutic option for addressing immune dysregulation.

[0004] Splenomegaly, or spleen enlargement, frequently arises from systemic conditions, particularly those involving immune dysregulation and is associated with a wide range of diseases not limited to inflammatory disease. The spleen plays a crucial role in blood filtration, immune surveillance, and red blood cell management, but in immune-related disorders, it can become overactive. Autoimmune diseases (e.g., psoriasis and systemic lupus erythematosus), chronic infections (e.g., HIV and tuberculosis), and hematological malignancies (e.g., lymphoma and leukemia) are common triggers of immune-related splenomegaly. In these cases, the spleen enlarges in response to the increased workload of filtering abnormal or excess immune cells and inflammatory by-products, resulting in discomfort, anemia, and increased vulnerability to infections due to compromised immune regulation (5).SUMMARY

[0005] Embodiments of the present disclosure include methods and compositions related to the treatment or prevention of immune-related splenomegaly. Individuals that are treated with methods and compositions of the disclosure include those that have one or more symptoms of splenomegaly. Individuals that are prevented from having splenomegaly, or in which theonset is delayed and / or the severity is reduced, include those at risk for splenomegaly. Factors that predispose an individual to be at risk include extrinsic and intrinsic risk factors. Extrinsic risk factors include at least mechanical stress, air pollution, drugs, vaccination, infection, smoking, alcohol, or a combination thereof. Intrinsic risk factors include metabolic syndrome, obesity, diabetes mellitus, dyslipidemia, hypertension, mental stress, or a combination thereof. An individual may be genetically predisposed to splenomegaly if they have genetic disorders that affect blood cell production, metabolism, or immune function, such as hemoglobinopathies, storage diseases, infections, blood disorders, liver disease, and / or certain cancers or autoimmune diseases.

[0006] Embodiments of the disclosure encompass compositions, methods, and systems for utilizing fibroblasts (including single cell fibroblasts), fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast culture, or any other fibroblast-derived material.

[0007] In some embodiments, the fibroblasts and / or fibroblast-derived material(s) may be given to an individual that has splenomegaly, or that might have splenomegaly, or that is at risk for splenomegaly for any reason, by any suitable route of delivery to the individual. In specific embodiments, the fibroblasts and / or fibroblast-derived material(s) may be delivered to the individual topically, subcutaneously, intrasplenic injection, and / or intravenously, as examples only.

[0008] In some embodiments, the fibroblasts are modified ex vivo prior to delivery to the individual. In some embodiments, following delivery of the fibroblasts and / or fibroblast-derived material(s) to the individual, the existing fibroblast cells within the skin become modified for regeneration of their function (9). In certain embodiments, the fibroblasts are activated by one or more activating agents prior to use. In certain embodiments, the fibroblasts from which the fibroblast-derived materials are derived are activated by one or more activating agents prior to use of the fibroblast-derived materials.

[0009] In particular embodiments, one or more endogenous genes in the fibroblasts may be modified and / or the fibroblasts may harbor one or more exogenously provided genes. Any gene modification system may be utilized, such as CRISPR / CAS systems. In specific cases, the gene modification system may be used to add, delete, or modify one or more fibroblast cell surface markers, e.g., to target the cells to a specific organ or tissue of interest (10).

[0010] In particular embodiments the disclosure concerns interaction of fibroblasts with a second type of cells and / or certain agent(s) and includes modification(s) to the fibroblasts and / or second type of cells as a result of the interaction. In specific embodiments, thedisclosure includes compositions, methods, and systems in which fibroblasts are modified upon exposure to certain cells and / or one or a combination of certain agent(s) including nucleic acids, cytokines, chemokines, or growth factors. In certain embodiments, delivery of these modified or unmodified fibroblasts to the individual can regenerate, repair, and / or reprogram the necessary functional splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells, and / or resident immune cells. In specific embodiments, this results in the initiation of the foundation for new tissue growth.

[0011] In particular embodiments, the regeneration of these splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells, and / or resident immune cells in the individual can be either through one or more possible steps, such as:

[0012] (1) The differentiation of the modified or unmodified fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or any other fibroblast-derived materials. The delivery to the individual may be intravenously, subcutaneously, or topically, including by injection.

[0013] (2) Epigenetic reprogramming of senescent or pathogenic fibroblasts and red blood cells in the individual through contact with the delivered modified or unmodified fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or any other fibroblast-derived materials (11).

[0014] (3) Reprogramming or activating pathogenic splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells, and / or resident immune cells by one or more types of products excreted from the modified or unmodified fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or any other fibroblast-derived materials.

[0015] (4) Recruitment, activation, and differentiation of local stem cells into active splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells, through the direct / indirect action of the injected modified / unmodified single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or fibroblast-derived materials from the modified / unmodified fibroblasts.

[0016] (5) Modulation of the immune system through single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblasts, and / or fibroblast-derived materials to control the expansion and localization of pathogenic immune cells, and their expression of pathogenic level of any one or more cytokines, one or more chemokines and / or one or more growth factors associated withsplenomegaly. These cells include T cells, dendritic cells macrophages, B cells, and neutrophils, expressing pathogenic levels of one or more of IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF, INF- y, TGF- pi, as examples.

[0017] In some embodiments, the regeneration of these splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells, and / or resident immune cells may occur by one or more other mechanisms.

[0018] In specific embodiments, methods, and compositions herein allow for the reduction of the pathogenic levels of immune cells and their impact on splenomegaly. Reduction of inflammation, and levels of pathogenic cytokines, pathogenic chemokines, and / or pathogenic growth factors will lead to reprogramming of the immune regulation in splenomegaly. In specific embodiments, the methods and compositions allow for normalizing of the immune response in order to reduce the pathogenic levels of immune cells or cytokines, growth factors, and / or chemokines excreted by those cells.

[0019] In specific embodiments, the disclosure concerns compositions, methods, and systems in which certain cells are modified upon exposure to fibroblasts and / or certain agent(s) excreted from fibroblasts. In particular embodiments, the interaction of fibroblasts with one or more other types of cells (and optionally that interaction also includes one or more certain agent(s)) results in modification of the fibroblasts and / or the other type of cells. In specific embodiments, the other types of cells include at least immune cells.

[0020] In specific embodiments, methods of the disclosure occur ex vivo, such as in a culture and can also occur in vivo. In particular cases, the methods occur by the hand of man and do not encompass ordinary or random occurrences in a body. The methods of the disclosure are non-natural, in particular aspects. In specific embodiments, the concentrations of cells used in a method of exposing one type of cells to another type of cells does not occur in nature and does not happen randomly in nature. In specific embodiments, the concentration of one or more agents used in a method of exposing the one or more agents to one or more types of cells does not occur in nature and does not happen randomly in nature. The modification of any types of cells encompassed by the disclosure that occurs ex vivo or in vitro does not occur in vivo naturally in the same manner. In such embodiment, tissue biopsy from the donor is used to isolate, characterize, if necessary activate, expand, and reintroduce back into the donor one or a combination of the cells for the purpose of modulating the immune system, reprogramming splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells.

[0021] In some embodiments, therapeutic uses of cells may include dendritic cells, macrophages, B lymphocytes, myoid cells, endothelial cells, fibroblasts, immune cells, and mixtures thereof. In some embodiments, the single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials have been modified prior to their exposure to the immune cells and other cells found in the blood and skin tissue, such as chemically, physically, and / or epigenetically activated, and / or exposed to conditions that are not normally found in the body, and in other cases immune cells, or their derivatives, have been modified, such as activated, prior to their exposure to the fibroblasts and / or fibroblast-derived materials.

[0022] Embodiments of the disclosure provide means of utilizing fibroblasts as allogeneic, autologous, xenogeneic, and / or syngeneic therapeutic cells through modification of culture conditions. In one embodiment of the disclosure, fibroblasts are extracted from sources with lower immunogenicity (e.g., placental fibroblasts, omental tissue derived fibroblasts, cord blood-derived fibroblasts, etc. .

[0023] In one embodiment of the disclosure, single cell fibroblasts and fibroblast spheroids are cultured in vitro for preserving viability and proliferative ability of fibroblasts. The disclosure provides for the modification of known culture techniques to decrease recognition of fibroblasts by the recipient immune system. In one embodiment, fibroblasts and / or fibroblast spheroids (for example) may be cultured in conditions that lack xenogeneic components, such as xenogeneic-free medium; in some cases, for example, the media may be free of fetal calf serum. In specific embodiments, the disclosure encompasses the substitution of fetal calf serum with one or more other agents, such as those that facilitate reduction of immunogenicity of fibroblasts, for example, human platelet rich plasma, platelet lysate, umbilical cord blood serum, autologous serum, and / or one or more defined cytokines, such as one or a combination of fibroblast growth factor, epidermal growth factor, leukemia inhibitory factor, insulin like growth factor, angiopoietin, and vascular endothelial growth factor.

[0024] In one embodiment of the disclosure, effective amounts of single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials as prepared in methods encompassed by the disclosure are administered to an individual for a therapy or prevention of one or more medical conditions. In specific embodiments, the fibroblasts are administered to improve fibroblast, endothelial cells, and / or resident immune cell responsiveness, modulation of immune cell activity, and cell differentiation.

[0025] Embodiments of the disclosure provide methods for co-administration of universal donor single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials with one or more agents that modulate the immune cell activity in the site of inflammation. In a specificembodiment of the disclosure, methods are provided for co-administration of universal donor single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials with one or a combination of growth factors, chemokines, and / or cytokines. In one embodiment of the disclosure, universal donor fibroblasts derived from fibroblasts that have been treated under conditions to reduce immunogenicity are utilized to stimulate vascular endothelial growth factor (VEGF) production by the introduced modified / unmodified fibroblasts or from endogenous cells under the regulatory control of nerve growth factor (NGF) of the individual thereby leading to restored spleen function.

[0026] Embodiments of the disclosure provide methods of reducing immunogenicity of particular types of fibroblasts. Single cell fibroblasts, fibroblast spheroids, exosomes, lysates, apoptotic cells or fibroblast-derived materials may be derived from various tissues or organs, including, but not limited to skin, heart, blood vessels, bone marrow, skeletal muscle, liver, pancreas, brain, foreskin, which can be obtained by biopsy (where appropriate) or upon autopsy. In some aspects, the cells comprise fibroblasts, which can be from a fetal, neonatal, adult origin, or a combination thereof.

[0027] In some embodiments, single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials or products for use in any methods of the disclosure may be exposed to certain medium component s), in specific embodiments.

[0028] In some embodiments, one may monitor the levels of particular immune cells and / or one or more agents expressed by the cells. This may occur prior to treatment, during treatment, and / or following treatment. Levels of T cells, dendritic cells, B cells, macrophages, and / or neutrophils may be monitored by Complete Blood Count, and White Blood Count, and tissue levels of T cells, dendritic cells, macrophages, and IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF, INF- y, TGF- pi may be monitored using ELISA and / or bead-based immunoassays, for example.

[0029] Embodiments of the disclosure encompass an in vivo method of treating splenomegaly using a topical, intravenous, or subcutaneous injection of single cell fibroblasts, fibroblast spheroids , fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or fibroblast-derived materials, such as to modulate the immune system and to reduce the production / infiltration of splenomegaly-associated pathogenic T cells, pathogenic dendritic cells, pathogenic macrophages, and / or pathogenic neutrophils that express pathogenic levels of IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF,INF- y, TGF- pi. The fibroblast may be autologous, allogeneic, xenogeneic, or syngeneic with respect to the individual, and in some cases a combination may be utilized.

[0030] Particular embodiments include introduction into an individual of single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or fibroblast-derived materials, or these products wherein the fibroblasts may be activated with one or more agents. In some cases, the single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials plus one or more type of immune cells or derivative agents thereof are delivered into the blood stream or skin tissue to modulate the proliferation of pathogenic immune cells therein.

[0031] Some embodiments include the introduction of single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or fibroblast-derived materials, wherein these may be activated with one or more agents or conditions. In some cases, the single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials plus one or more type of immune cells and / or derivative agents are provided into the bloodstream of the individual to modulate the expression of pathogenic levels of IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF, INF- y, and / or TGF- pi.

[0032] Particular embodiments include introduction of single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials, single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials or products activated with one or more agents or conditions. In some cases, the single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from the fibroblast, and / or fibroblast-derived materials plus one or more type of immune cells and / or one or more derivative agents are introduced into the bloodstream of the individual to enable the epigenetic reprogramming of localized splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells of the individual.

[0033] In some embodiments, introduction of single cell fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, conditioned media from fibroblast, and / or fibroblast-derived materials or products, may be activated with one or more agents. In some cases, the single cell fibroblasts, fibroblast spheroids, or fibroblast-derived materials plus one or more types of immune cells and / or one or more derivative agents thereof are introduced into the bloodstream of the individual to enable the activation and migration of localized stem cell niches to replace the pathogenetic splenic fibroblasts, splenic reticular cells, splenic endothelial cells, splenic epithelial cells.

[0034] In some embodiments, there is use of one or more adjuvants in combination with single cell fibroblasts, fibroblast spheroids, and / or fibroblast-derived materials to activate the immune system. These adjuvants could be chemical, viral, bacterial product based or a mixture thereof.

[0035] The foregoing has outlined rather broadly the features and technical advantages of the present invention in order that the detailed description of the invention that follows may be better understood. Additional features and advantages of the invention will be described hereinafter which form the subject of the claims of the invention. It should be appreciated by those skilled in the art that the conception and specific embodiment disclosed may be readily utilized as a basis for modifying or designing other structures for carrying out the same purposes of the present invention. It should also be realized by those skilled in the art that such equivalent constructions do not depart from the spirit and scope of the invention as set forth in the appended claims. The novel features which are believed to be characteristic of the invention, both as to its organization and method of operation, together with further objects and advantages will be better understood from the following description when considered in connection with the accompanying figures. It is to be expressly understood, however, that each of the figures is provided for the purpose of illustration and description only and is not intended as a definition of the limits of the present invention.BRIEF DESCRIPTION OF THE DRAWINGS

[0036] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein.

[0037] FIGS. 1A, IB, 1C, ID, and IE. An imiquimod (IMQ)-induced psoriasis mouse model. (1A) Representative images of mouse back skin after a single administration of fibroblasts via the tail vein. (1B-1E) Clinical scores for thickness, erythema, scaling, and their average scores (a scale from 0 to 4, 0-none; 1-slight; 2-moderate; 3-marked; 4-very marked). A single intravenous administration of fibroblasts on Day 4 significantly alleviated IMQ-induced psoriasis from Day 6 to Day 8.

[0038] FIGS. 2A, 2B, and 2C. Fibroblasts significantly attenuated the IMQ-induced increase in (2A-2C) spleen weight and length.

[0039] FIGS. 3A, 3B, 3C, 3D, and 3E. Comparative analysis of therapeutic efficacy between fibroblasts and anti-IL-23 treatment in psoriasis. (3 A) Representative images of mouse back skin after treatments. (3B-3E) Clinical scores for thickness, erythema, scaling, and their average scores (a scale from 0 to 4, 0-none; 1-slight; 2-moderate; 3-marked; 4-very marked). Both fibroblasts and anti-IL-23 treatments effectively reduced IMQ-induced psoriasis over the 7-day monitoring period.

[0040] FIGS. 4A, 4B, and 4C. Psoriasis mice treated with fibroblasts showed no splenomegaly and exhibited improved platelet counts following the treatment (4A-4C). In contrast, mice treated with anti-IL-23 did not demonstrate these improvements, as splenomegaly was still present, and platelet counts were significantly lower than those of the healthy controls.

[0041] FIGS. 5A, 5B, 5C. Flow cytometry analysis of isolated mouse splenocytes (5 A) revealed a marked reduction in the population of macrophages (CDllb+F4 / 80+ cells) in fibroblasts-treated mice (5B and 5C). Anti-IL-23 treatment did not exhibit a similar reduction in splenic macrophages.

[0042] FIGS. 6A, 6B, 6C, and 6D. Fibroblasts exerted inhibitory effects on splenic T cell subsets in a psoriatic mouse model. Flow cytometry analysis showed a marked reduction in CD3+(6A), CD4+(6B), CD8+(6C), and CD25+(6D) T cell populations following fibroblast treatment, demonstrating that fibroblasts broadly modulate adaptive immune responses in the spleen.DETAILED DESCRIPTIONI. Examples of Definitions

[0043] In keeping with long-standing patent law convention, the words “a” and “an” when used in the present specification in concert with the word comprising, including the claims, denote “one or more.” Some embodiments of the disclosure may consist of or consist essentially of one or more elements, method steps, and / or methods of the disclosure. It is contemplated that any method or composition described herein can be implemented with respect to any other method or composition described herein.

[0044] The phrase “and / or” means “and” or “or”. To illustrate, A, B, and / or C includes: A alone, B alone, C alone, a combination of A and B, a combination of A and C, a combination of B and C, or a combination of A, B, and C. In other words, “and / or” operates as an inclusive or.

[0045] As used herein, the term “about” or “approximately” refers to a quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length that varies by as much as 30, 25, 20, 25, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 % to a reference quantity, level, value, number, frequency, percentage, dimension, size, amount, weight or length. In particular embodiments, the terms “about” or “approximately” when preceding a numerical value indicates the value plus or minus a range of 15%, 10%, 5%, or 1%. With respect to biological systems or processes, the term can mean within an order of magnitude, preferably within 5-fold, and more preferably within 2-fold, of a value. Unless otherwise stated, the term 'about' means within an acceptable error range for the particular value.

[0046] Throughout this specification, unless the context requires otherwise, the words “comprise”, “comprises” and “comprising” will be understood to imply the inclusion of a stated step or element or group of steps or elements but not the exclusion of any other step or element or group of steps or elements. By “consisting of’ is meant including, and limited to, whatever follows the phrase “consisting of.” Thus, the phrase “consisting of’ indicates that the listed elements are required or mandatory, and that no other elements may be present. By “consisting essentially of’ is meant including any elements listed after the phrase, and limited to other elements that do not interfere with or contribute to the activity or action specified in the disclosure for the listed elements. Thus, the phrase “consisting essentially of’ indicates that the listed elements are required or mandatory, but that no other elements are optional and may or may not be present depending upon whether or not they affect the activity or action of the listed elements.

[0047] Reference throughout this specification to “one embodiment,” “an embodiment,” “a particular embodiment,” “a related embodiment,” “a certain embodiment,” “an additional embodiment,” or “a further embodiment” or combinations thereof means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment of the present disclosure. Thus, the appearances of the foregoing phrases in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

[0048] As used herein, the term “activated immune cells” refers to immune cells treated with one or more stimuli capable of inducing one or more alterations in the cell: metabolic, immunological, epigenetic, growth factor secreting, surface marker expression, and production and excretion of microvesicles.

[0049] The term "administered" or "administering", as used herein, refers to any method of providing a composition to an individual such that the composition has its intended effect on the individual. For example, one method of administering is by a direct mechanism such as, local tissue administration, intravenous administration, oral ingestion, transdermal patch, topical, inhalation, suppository, etc. Another example includes an indirect mechanism using a medical device such as, but not limited to a catheter, applicator gun, syringe, etc.

[0050] As used herein, “allogeneic” refers to tissues or cells from another body that in a natural setting are immunologically incompatible or capable of being immunologically incompatible, although from one or more individuals of the same species.

[0051] As used herein, “autologous” refers to tissues or cells that are derived or transferred from the same individual's body (i.e., autologous blood donation; an autologous bone marrow transplant).

[0052] As used herein, “agent” refers to nucleic acids, cytokines, chemokines, transcription factors, epigenetics factors, growth factors, or hormones.

[0053] As used herein, “xenogeneic” refers to tissues or cells from a species different from the patient.

[0054] Cell culture" may be an artificial in vitro system containing viable cells, whether quiescent, senescent or (actively) dividing. In a cell culture, cells are grown and maintained at an appropriate temperature, typically a temperature of 37°C and under an atmosphere typically containing oxygen and CO2. Culture conditions may vary widely for each cell type though, and variation of conditions for a particular cell type can result in different phenotypes being expressed. The most commonly varied factor in culture systems is the growth medium. Growth media can vary in concentration of nutrients, growth factors, and the presence of other components. The growth factors used to supplement media are often derived from animal blood, such as calf serum.

[0055] The term “fibroblast-derived material” as used herein refers to at least fibroblast cell fragments, fibroblast conditioned media, exosomes secreted from fibroblasts, and / or fibroblast lysate and includes fibroblast-like cells; spheroids that comprise fibroblasts; and / or extracellular vesicles including exosomes, microvesicles, apoptotic bodies or any other fragments or biologic components of fibroblast cells.

[0056] The term "individual", as used herein, refers to a human or animal that may or may not be housed in a medical facility and may be treated as an outpatient of a medical facility. The individual may be receiving one or more medical compositions via the internet. An individual may comprise any age of a human or non-human animal and therefore includes bothadults and juveniles ( / .< ., children) and infants. It is not intended that the term "individual" connotes a need for medical treatment, therefore, an individual may voluntarily or involuntarily be part of experimentation whether clinical or in support of basic science studies. The term “subject” or “individual” may be used interchangeably and refers to any organism or animal subject that is an object of a method or material, including mammals, e.g., humans, laboratory animals (e.g., primates, rats, mice, rabbits), livestock (e.g., cows, sheep, goats, pigs, turkeys, and chickens), household pets (e.g., dogs, cats, and rodents), horses, and transgenic non-human animals.

[0057] The terms "reduce," "inhibit," "diminish," "suppress," "decrease," "prevent" and grammatical equivalents (including "lower," "smaller," etc.) when in reference to the expression of any symptom in an untreated subject relative to a treated subject, mean that the quantity and / or magnitude of the symptoms in the treated subject is lower than in the untreated subject by any amount that is recognized as clinically relevant by any medically trained personnel. In one embodiment, the quantity and / or magnitude of the symptoms in the treated subject is at least 10% lower than, at least 25% lower than, at least 50% lower than, at least 75% lower than, and / or at least 90% lower than the quantity and / or magnitude of the symptoms in the untreated subject.

[0058] As used herein, the term “transplantation” refers to the process of taking living tissue or cells and implanting it in another part of the body or into another body.“Treatment,” “treat,” or “treating” means a method of reducing the effects of a disease or condition. Treatment can also refer to a method of reducing the disease or condition itself rather than just the symptoms. The treatment can be any reduction from pre-treatment levels and can be but is not limited to the complete ablation of the disease, condition, or the symptoms of the disease or condition. Therefore, in the disclosed methods, treatment” can refer to a 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% reduction in the severity of an established disease or the disease progression, including reduction in the severity of at least one symptom of the disease. For example, a disclosed method for reducing the immunogenicity of cells is considered to be a treatment if there is a detectable reduction in the immunogenicity of cells when compared to pre-treatment levels in the same subject or control subjects. Thus, the reduction can be a 10, 20, 30, 40, 50, 60, 70, 80, 90, 100%, or any amount of reduction in between as compared to native or control levels. It is understood and herein contemplated that “treatment” does not necessarily refer to a cure of the disease or condition, but an improvement in the outlook of a disease or condition. In specific embodiments, treatment refers to thelessening in severity or extent of at least one symptom and may alternatively or in addition refer to a delay in the onset of at least one symptom.

[0059] Although the present disclosure and its advantages have been described in detail, it should be understood that various changes, substitutions and alterations can be made herein without departing from the spirit and scope of the invention as defined by the appended claims. Moreover, the scope of the present application is not intended to be limited to the particular embodiments of the process, machine, manufacture, composition of matter, means, methods and steps described in the specification. As one of ordinary skill in the art will readily appreciate from the disclosure of the present disclosure, processes, machines, manufacture, compositions of matter, means, methods, or steps, presently existing or later to be developed that perform substantially the same function or achieve substantially the same result as the corresponding embodiments described herein may be utilized according to the present disclosure. Accordingly, the appended claims are intended to include within their scope such processes, machines, manufacture, compositions of matter, means, methods, or steps.

[0060] It is contemplated that any embodiment discussed in this specification can be implemented with respect to any method or composition of the invention, and vice versa. Furthermore, compositions of the invention can be used to achieve methods of the invention.

[0061] Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the specific examples, while indicating specific embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.II. Administration of Therapeutic Compositions

[0062] Particular embodiments of the disclosure concern therapeutic compositions for the treatment of splenomegaly. The compositions may cure splenomegaly of the individual or may reduce the severity of one symptom of splenomegaly and / or delay the onset of splenomegaly.

[0063] The splenomegaly may be of any type, including those associated with hematologic conditions such as hemolytic anemia or leukemia, infectious diseases like mononucleosis or malaria, metabolic disorders such as Gaucher disease or Niemann-Pick disease, autoimmuneconditions like systemic lupus erythematosus (SLE) or rheumatoid arthritis, and chronic liver diseases such as cirrhosis or portal hypertension.

[0064] In some embodiments, the methods and compositions of the disclosure are utilized in addition to one or more other splenomegaly treatments, such as corticosteroids, immunosuppressive drugs (e.g., methotrexate, cyclosporine), and / or biologies (e.g., rituximab). In some embodiments, the individual in need of the methods and compositions of the disclosure has employed one or more of these other splenomegaly treatments, such as to little or no effect.

[0065] The therapeutic agents of the disclosure may be administered by any suitable administration route. In embodiments wherein more than one splenomegaly therapy is utilized, the therapeutic agent(s) of the disclosure may be administered by the same route of administration or by different routes of administration than the additional therapy. In some embodiments, the therapy is administered intravenously, topically, intramuscularly, subcutaneously, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. In some embodiments, the antibiotic is administered intravenously, intramuscularly, subcutaneously, topically, orally, transdermally, intraperitoneally, intraorbitally, by implantation, by inhalation, intrathecally, intraventricularly, or intranasally. The appropriate dosage may be determined based on the type of disease to be treated, severity and course of the disease, the clinical condition of the individual, the individual's clinical history and response to the treatment, and the discretion of the attending physician.

[0066] The treatments may include various “unit doses.” Unit dose is defined as containing a predetermined-quantity of the therapeutic composition. The quantity to be administered, and the particular route and formulation, is within the skill of determination of those in the clinical arts. A unit dose need not be administered as a single injection but may comprise continuous infusion over a set period of time. In some embodiments, a unit dose comprises a single administrable dose.

[0067] In some embodiments, a single dose of the therapy is administered. In some embodiments, multiple doses of the therapy are administered. In some embodiments, the therapy is administered at a dose of between 1 mg / kg and 5000 mg / kg. In some embodiments, the therapy is administered at a dose of at least, at most, or about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86,87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151, 152, 153, 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, 170, 171, 172, 173, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, 217, 218, 219, 220, 221, 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 290, 291, 292, 293, 294, 295, 296, 297, 298, 299, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335, 336, 337, 338, 339, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358, 359, 360, 361, 362, 363, 364, 365, 366, 367, 368, 369, 370, 371, 372, 373, 374, 375, 376, 377, 378, 379, 380, 381, 382, 383, 384, 385, 386, 387, 388, 389, 390, 391, 392, 393, 394, 395, 396, 397, 398, 399, 400, 401, 402, 403, 404, 405, 406, 407, 408, 409, 410, 411, 412, 413, 414, 415, 416, 417, 418, 419, 420, 421, 422, 423, 424, 425, 426, 427, 428, 429, 430, 431, 432, 433, 434, 435, 436, 437, 438, 439, 440, 441, 442, 443, 444, 445, 446, 447, 448, 449, 450, 451, 452, 453, 454, 455, 456, 457, 458, 459, 460, 461, 462, 463, 464, 465, 466, 467, 468, 469, 470, 471, 472, 473, 474, 475, 476, 477, 478, 479, 480, 481, 482, 483, 484, 485, 486, 487, 488, 489, 490, 491, 492, 493, 494, 495, 496, 497, 498, 499, 500, 501, 502, 503, 504, 505, 506, 507, 508, 509, 510, 511, 512, 513, 514, 515, 516, 517, 518, 519, 520, 521, 522, 523, 524, 525, 526, 527, 528, 529, 530, 531, 532, 533, 534, 535, 536, 537, 538, 539, 540, 541, 542, 543, 544, 545, 546, 547, 548, 549, 550, 551, 552, 553, 554, 555, 556, 557, 558, 559, 560, 561, 562, 563, 564, 565, 566, 567, 568, 569, 570, 571, 572, 600, 700, 800, 900, 1000, 1100, 1200, 1300, 1400, 1500, 1600, 1700, 1800, 1900, 2000, 2100, 2200, 2300, 2400, 2500, 2600, 2700, 2800, 2900, 3000, 3100, 3200, 3300, 3400, 3500, 3600, 3700, 3800, 3900, 4000, 4100, 4200, 4300, 4400, 4500, 4600, 4700, 4800, 4900, or 5000 mg / kg.

[0068] The quantity to be administered, both according to number of treatments and unit dose, depends on the treatment effect desired. An effective dose is understood to refer to an amount necessary to achieve a particular effect. In the practice in certain embodiments, it is contemplated that doses in the range from 10 mg / kg to 200 mg / kg can affect the protective capability of these agents. Thus, it is contemplated that doses include doses of about 0.1, 0.5,1, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, and 200, 300, 400, 500, 1000 pg / kg, mg / kg, pg / day, or mg / day or any range derivable therein. Furthermore, such doses can be administered at multiple times during a day, and / or on multiple days, weeks, or months.

[0069] In certain embodiments, the effective dose of the pharmaceutical composition is one which can provide a blood level of about 1 pM to 150 pM. In another embodiment, the effective dose provides a blood level of about 4 pM to 100 pM.; or about 1 pM to 100 pM; or about 1 pM to 50 pM; or about 1 pM to 40 pM; or about 1 pM to 30 pM; or about 1 pM to 20 pM; or about 1 pM to 10 pM; or about 10 pM to 150 pM; or about 10 pM to 100 pM; or about 10 pM to 50 pM; or about 25 pM to 150 pM; or about 25 pM to 100 pM; or about 25 pM to 50 pM; or about 50 pM to 150 pM; or about 50 pM to 100 pM (or any range derivable therein). In other embodiments, the dose can provide the following blood level of the agent that results from a therapeutic agent being administered to a subject: about, at least about, or at most about 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100 pM or any range derivable therein. In certain embodiments, the therapeutic agent that is administered to a subject is metabolized in the body to a metabolized therapeutic agent, in which case the blood levels may refer to the amount of that agent. Alternatively, to the extent the therapeutic agent is not metabolized by a subject, the blood levels discussed herein may refer to the unmetabolized therapeutic agent.

[0070] Precise amounts of the therapeutic composition also depend on the judgment of the practitioner and may be tailored to each individual. Factors affecting dose include physical and clinical state of the patient, the route of administration, the intended goal of treatment (alleviation of symptoms versus cure) and the potency, stability and toxicity of the particular therapeutic substance or other therapies a subject may be undergoing.

[0071] It will be understood by those skilled in the art and made aware that dosage units of pg / kg or mg / kg of body weight can be converted and expressed in comparable concentration units of pg / ml or mM (blood levels), such as 4 pM to 100 pM. It is also understood that uptake is species and organ / tissue dependent. The applicable conversion factors and physiological assumptions to be made concerning uptake and concentration measurement are well-known and would permit those of skill in the art to convert one concentration measurement to anotherand make reasonable comparisons and conclusions regarding the doses, efficacies and results described herein.

[0072] In certain instances, it will be desirable to have multiple administrations of the composition, e.g., 2, 3, 4, 5, 6 or more administrations. The administrations can be at 1, 2, 3, 4, 5, 6, 7, 8, to 5, 6, 7, 8, 9, 10, 11, or 12 week intervals, including all ranges there between.

[0073] The phrases “pharmaceutically acceptable” or “pharmacologically acceptable” refer to molecular entities and compositions that do not produce an adverse, allergic, or other untoward reaction when administered to an animal or human. As used herein, “pharmaceutically acceptable carrier” includes any and all solvents, dispersion media, coatings, anti-bacterial and anti-fungal agents, isotonic and absorption delaying agents, and the like. The use of such media and agents for pharmaceutical active substances is well known in the art. Except insofar as any conventional media or agent is incompatible with the active ingredients, its use in immunogenic and therapeutic compositions is contemplated. Supplementary active ingredients, such as other anti-infective agents and vaccines, can also be incorporated into the compositions.

[0074] The active compounds can be formulated for parenteral administration, e.g., formulated for injection via the intravenous, intramuscular, subcutaneous, or intraperitoneal routes. Typically, such compositions can be prepared as either liquid solutions or suspensions; solid forms suitable for use to prepare solutions or suspensions upon the addition of a liquid prior to injection can also be prepared; and, the preparations can also be emulsified.

[0075] The pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions; formulations including, for example, aqueous propylene glycol; and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases the form must be sterile and must be fluid to the extent that it may be easily injected. It also should be stable under the conditions of manufacture and storage and must be preserved against the contaminating action of microorganisms, such as bacteria and fungi.

[0076] The proteinaceous compositions may be formulated into a neutral or salt form. Pharmaceutically acceptable salts, include the acid addition salts (formed with the free amino groups of the protein) and which are formed with inorganic acids such as, for example, hydrochloric or phosphoric acids, or such organic acids as acetic, oxalic, tartaric, mandelic, and the like. Salts formed with the free carboxyl groups can also be derived from inorganic bases such as, for example, sodium, potassium, ammonium, calcium, or ferric hydroxides, and such organic bases as isopropylamine, trimethylamine, histidine, procaine and the like.

[0077] A pharmaceutical composition can include a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene glycol, and the like), suitable mixtures thereof, and vegetable oils. The proper fluidity can be maintained, for example, by the use of a coating, such as lecithin, by the maintenance of the required particle size in the case of dispersion, and by the use of surfactants. The prevention of the action of microorganisms can be brought about by various anti-bacterial and anti-fungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars or sodium chloride. Prolonged absorption of the injectable compositions can be brought about by the use in the compositions of agents delaying absorption, for example, aluminum monostearate and gelatin.

[0078] Sterile injectable solutions are prepared by incorporating the active compounds in the required amount in the appropriate solvent with various other ingredients enumerated above, as required, followed by filtered sterilization or an equivalent procedure. Generally, dispersions are prepared by incorporating the various sterilized active ingredients into a sterile vehicle which contains the basic dispersion medium and the required other ingredients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred methods of preparation are vacuum-drying and freeze-drying techniques, which yield a powder of the active ingredient, plus any additional desired ingredient from a previously sterile-filtered solution thereof.

[0079] Administration of the compositions may be via any route. This includes, but is not limited to topical and / or intravenous administration. In some embodiments both topical and intravenous are utilized. For example, a first dose may be a bolus that is intravenous, followed by topical administration thereafter. Alternatively, administration may be by orthotopic, intradermal, subcutaneous, intramuscular, intraperitoneal, or intranasal administration. Such compositions would normally be administered as pharmaceutically acceptable compositions that include physiologically acceptable carriers, buffers or other excipients.

[0080] Upon formulation, solutions are administered in a manner compatible with the dosage formulation and in such amount as is therapeutically or prophylactically effective. The formulations are easily administered in a variety of dosage forms, such as the type of injectable solutions described above.III. Cellular TherapiesA. Cells and Cell Culture

[0081] In certain embodiments, the fibroblasts may be derived from tissues comprising skin, heart, blood vessels, bone marrow, skeletal muscle, liver, pancreas, brain, adipose tissue, foreskin, placental, umbilical cord, or a mixture thereof. In specific embodiments, the fibroblasts are placental, fetal, neonatal or adult or mixtures thereof.

[0082] In some embodiments, fibroblast cells may be cultured for at least between about 10 days and about 40 days, for at least between about 15 days and about 35 days, for at least between about 15 days and 21 days, such as for at least about 15, 16, 17, 18, 19 or 21 days. In some embodiments, the cells of the disclosure may be cultured for no longer than 60 days, or no longer than 50 days, or no longer than 45 days. The cells may be cultured for 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, or 40 days. The cells may be cultured in the presence of a liquid culture medium. Typically, the medium may comprise a basal medium formulation as known in the art. Many basal media formulations can be used to culture cells herein, including but not limited to Eagle's Minimum Essential Medium (MEM), Dulbecco's Modified Eagle's Medium (DMEM), alpha modified Minimum Essential Medium (alpha-MEM), Basal Medium Essential (BME), Iscove's Modified Dulbecco's Medium (IMDM), BGJb medium, F-12 Nutrient Mixture (Ham), Liebovitz L-15, DMEM / F-12, Essential Modified Eagle's Medium (EMEM), RPMI-1640, and modifications and / or combinations thereof. Compositions of the above basal media are generally known in the art, and it is within the skill of one in the art to modify or modulate concentrations of media and / or media supplements as necessary for the cells cultured. In some embodiments, a culture medium formulation may be explants medium (CEM) which is composed of IMDM supplemented with 10% fetal bovine serum (FBS), 100 U / ml penicillin G, 100 pg / ml streptomycin and 2 mmol / L L-glutamine. Other embodiments may employ further basal media formulations, such as chosen from the ones above.

[0083] Any medium capable of supporting cells in vitro may be used to culture the cells. Media formulations that can support the growth of cells include, but are not limited to, Dulbecco's Modified Eagle's Medium (DMEM), alpha modified Minimal Essential Medium (aMEM), and Roswell Park Memorial Institute Media 1640 (RPMI Media 1640) and the like. Typically, up to 20% fetal bovine serum (FBS) or 1-20% horse serum is added to the above medium in order to support the growth of cells. A defined medium, however, also can be used if the growth factors, cytokines, and hormones necessary for culturing cells are provided at appropriate concentrations in the medium. Media useful in the methods of the disclosure maycomprise one or more compounds of interest, including, but not limited to, antibiotics, mitogenic compounds, or differentiation compounds useful for the culturing of cells. The cells may be grown at temperatures between 27° C to 40° C, such as 31° C to 37° C, and may be in a humidified incubator. The carbon dioxide content may be maintained between 2% to 10% and the oxygen content may be maintained between 1% and 22%. The disclosure, however, should in no way be construed to be limited to any one method of isolating and culturing cells. Rather, any method of isolating and culturing cells should be construed to be included in the present disclosure.

[0084] For use in the cell culture, media can be supplied with one or more further components. For example, additional supplements can be used to supply the cells with the necessary trace elements and substances for optimal growth and expansion. Such supplements include insulin, transferrin, selenium salts, and combinations thereof. These components can be included in a salt solution such as, but not limited to, Hanks' Balanced Salt Solution (HBSS), Earle's Salt Solution. Further antioxidant supplements may be added, e.g., P-mercaptoethanol. While many media already contain amino acids, some amino acids may be supplemented later, e.g., L-glutamine, which is known to be less stable when in solution. A medium may be further supplied with antibiotic and / or antimycotic compounds, such as, typically, mixtures of penicillin and streptomycin, and / or other compounds, exemplified but not limited to, amphotericin, ampicillin, gentamicin, bleomycin, hygromycin, kanamycin, mitomycin, mycophenolic acid, nalidixic acid, neomycin, nystatin, paromomycin, polymyxin, puromycin, rifampicin, spectinomycin, tetracycline, tylosin, and zeocin. Also contemplated is supplementation of cell culture medium with mammalian plasma or sera. Plasma or sera often contain cellular factors and components that are necessary for viability and expansion. The use of suitable serum replacements is also contemplated.

[0085] Reference to particular buffers, media, reagents, cells, culture conditions and the like, or to some subclass of same, is not intended to be limiting, but should be read to include all such related materials that one of ordinary skill in the art would recognize as being of interest or value in the particular context in which that discussion is presented. For example, it is often possible to substitute one buffer system or culture medium for another, such that a different but known way is used to achieve the same goals as those to which the use of a suggested method, material or composition is directed. In particular embodiments, cells are cultured in a cell culture system comprising a cell culture medium, preferably in a culture vessel, in particular a cell culture medium supplemented with a substance suitable and determined for protecting the cells from in vitro aging and / or inducing in an unspecific or specific reprogramming.B. Cell Generation

[0086] Certain methods of the disclosure concern culturing the cells obtained from human tissue samples. In particular embodiments of the present disclosure, cells are plated onto a substrate that allows for adherence of cells thereto. This may be carried out, for example, by plating the cells in a culture plate that displays one or more substrate surfaces compatible with cell adhesion. When the one or more substrate surfaces contact the suspension of cells (e.g., suspension in a medium) introduced into the culture system, cell adhesion between the cells and the substrate surfaces may ensue. Accordingly, in certain embodiments cells are introduced into a culture system that features at least one substrate surface that is generally compatible with adherence of cells thereto, such that the plated cells can contact the said substrate surface, such embodiments encompass plating onto a substrate, which allows adherence of cells thereto.

[0087] Cells of the present disclosure may be identified and characterized by their expression of specific marker proteins, such as cell-surface markers. Detection and isolation of these cells can be achieved, for example, through flow cytometry, ELISA, and / or magnetic beads. Reverse-transcription polymerase chain reaction (RT-PCR) may be used to quantify cell-specific genes and / or to monitor changes in gene expression in response to differentiation. In certain embodiments, the marker proteins used to identify and characterize the cells are selected from the list consisting of c-Kit, Nanog, Sox2, Heyl, SMA, Vimentin, Cyclin D2, Snail, E-cadherin, Nkx2.5, GATA4, CD105, CD90, CD29, CD73, Wtl, CD34, CD45, and a combination thereof.C. Pharmaceutical Compositions

[0088] In certain aspects, the compositions or agents for use in the methods, such as fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast microvesicles, fibroblast lysate, fibroblast apoptotic bodies, fibroblast conditioned media, or a combination thereof, are suitably contained in a pharmaceutically acceptable carrier. The carrier is non-toxic, biocompatible and is selected so as not to detrimentally affect the biological activity of the agent. The agents in some aspects of the disclosure may be formulated into preparations for local delivery (i.e., to a specific location of the body, such as on or in the skin or other tissue) or systemic delivery, in solid, semi-solid, gel, liquid or gaseous forms such as tablets, capsules, powders, granules, ointments, solutions, depositories, inhalants and injections allowing for oral, parenteral or surgical administration. Certain aspects of the disclosure also contemplate local administration of the compositions by coating medical devices and the like.

[0089] Suitable carriers for parenteral delivery via injectable, infusion or irrigation and topical delivery include distilled water, physiological phosphate-buffered saline, normal or lactated Ringer's solutions, dextrose solution, Hank's solution, or propanediol. In addition, sterile, fixed oils may be employed as a solvent or suspending medium. For this purpose, any biocompatible oil may be employed including synthetic mono- or di-glycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables. The carrier and agent may be compounded as a liquid, suspension, polymerizable or non-polymerizable gel, paste or salve.

[0090] The carrier may also comprise a delivery vehicle to sustain (i.e., extend, delay or regulate) the delivery of the agent(s) or to enhance the delivery, uptake, stability or pharmacokinetics of the therapeutic agent(s). Such a delivery vehicle may include, by way of non-limiting examples, microparticles, microspheres, nanospheres or nanoparticles composed of proteins, liposomes, carbohydrates, synthetic organic compounds, inorganic compounds, polymeric or copolymeric hydrogels and polymeric micelles.

[0091] In certain aspects, the actual dosage amount of a composition administered to a patient or subject can be determined by physical and physiological factors such as body weight, severity of condition, the type of disease being treated, previous or concurrent therapeutic interventions, idiopathy of the patient and on the route of administration. The practitioner responsible for administration will, in any event, determine the concentration of active ingredient(s) in a composition and appropriate dose(s) for the individual subject.

[0092] Solutions of pharmaceutical compositions can be prepared in water suitably mixed with a surfactant, such as hydroxypropylcellulose. Dispersions also can be prepared in glycerol, liquid polyethylene glycols, mixtures thereof and in oils. Under ordinary conditions of storage and use, these preparations contain a preservative to prevent the growth of microorganisms.

[0093] In certain aspects, the pharmaceutical compositions are advantageously administered in the form of injectable compositions either as liquid solutions or suspensions; solid forms suitable or solution in, or suspension in, liquid prior to injection may also be prepared. These preparations also may be emulsified. A typical composition for such purpose comprises a pharmaceutically acceptable carrier. For instance, the composition may contain 10 mg or less, 25 mg, 50 mg or up to about 100 mg of human serum albumin per milliliter of phosphate buffered saline. Other pharmaceutically acceptable carriers include aqueous solutions, non-toxic excipients, including salts, preservatives, buffers and the like.

[0094] Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, vegetable oil and injectable organic esters such as ethyloleate. Aqueous carriers include water,alcoholic / aqueous solutions, saline solutions, parenteral vehicles such as sodium chloride, Ringer's dextrose, etc. Intravenous vehicles include fluid and nutrient replenishers. Preservatives include antimicrobial agents, antifungal agents, anti-oxidants, chelating agents and inert gases. The pH and exact concentration of the various components the pharmaceutical composition are adjusted according to well-known parameters.

[0095] Additional formulations may be suitable for oral administration. Oral formulations include such typical excipients as, for example, pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate and the like. The compositions take the form of solutions, suspensions, tablets, pills, capsules, sustained release formulations or powders.

[0096] In further aspects, the pharmaceutical compositions may include classic pharmaceutical preparations. Administration of pharmaceutical compositions according to certain aspects may be via any common route so long as the target tissue is available via that route. This may include oral, nasal, buccal, rectal, vaginal or topical. Alternatively, administration may be by orthotopic, intradermal, subcutaneous, intramuscular, intraperitoneal or intravenous injection. Such compositions would normally be administered as pharmaceutically acceptable compositions that include physiologically acceptable carriers, buffers or other excipients. For treatment of conditions of the lungs, aerosol delivery can be used. Volume of the aerosol may be between about 0.01 ml and 0.5 ml, for example.

[0097] An effective amount of the pharmaceutical composition is determined based on the intended goal. The term “unit dose” or “dosage” refers to physically discrete units suitable for use in a subject, each unit containing a predetermined-quantity of the pharmaceutical composition calculated to produce the desired responses discussed above in association with its administration, i.e., the appropriate route and treatment regimen. The quantity to be administered, both according to number of treatments and unit dose, depends on the protection or effect desired.

[0098] Precise amounts of the pharmaceutical composition also depend on the judgment of the practitioner and are peculiar to each individual. Factors affecting the dose include the physical and clinical state of the patient, the route of administration, the intended goal of treatment (e.g., alleviation of symptoms versus cure) and the potency, stability and toxicity of the particular therapeutic substance.II. Formulations and Culture of the Cells

[0099] In particular embodiments, the cells of the disclosure may be specifically formulated and / or they may be cultured in a particular medium. The cells may be formulated in such a manner as to be suitable for delivery to a recipient without deleterious effects.

[0100] The medium in certain aspects can be prepared using a medium used for culturing animal cells as their basal medium, such as any of AIM V, X-VIVO-15, NeuroBasal, EGM2, TeSR, BME, BGJb, CMRL 1066, Glasgow MEM, Improved MEM Zinc Option, IMDM, Medium 199, Eagle MEM, aMEM, DMEM, Ham, RPMI-1640, and Fischer's media, as well as any combinations thereof, but the medium may not be particularly limited thereto as far as it can be used for culturing animal cells. Particularly, the medium may be xeno-free or chemically defined.

[0101] The medium can be a serum-containing or serum-free medium, or xeno-free medium. From the aspect of preventing contamination with heterogeneous animal-derived components, serum can be derived from the same animal as that of the stem cell(s). The serum-free medium refers to medium with no unprocessed or unpurified serum and accordingly, can include medium with purified blood-derived components or animal tissue-derived components (such as growth factors).

[0102] The medium may contain or may not contain any alternatives to serum. The alternatives to serum can include materials which appropriately contain albumin (such as lipid-rich albumin, bovine albumin, albumin substitutes such as recombinant albumin or a humanized albumin, plant starch, dextrans and protein hydrolysates), transferrin (or other iron transporters), fatty acids, insulin, collagen precursors, trace elements, 2-mercaptoethanol, 3'-thiolgiycerol, or equivalents thereto. The alternatives to serum can be prepared by the method disclosed in International Publication No. 98 / 30679, for example (incorporated herein in its entirety). Alternatively, any commercially available materials can be used for more convenience. The commercially available materials include knockout Serum Replacement (KSR), Chemically-defined Lipid concentrated (Gibco), and Glutamax (Gibco).

[0103] In certain embodiments, the medium may comprise 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20 or more of the following: Vitamins such as biotin; DL Alpha Tocopherol Acetate; DL Alpha-Tocopherol; Vitamin A (acetate); proteins such as BSA (bovine serum albumin) or human albumin, fatty acid free Fraction V; Catalase; Human Recombinant Insulin; Human Transferrin; Superoxide Dismutase; Other Components such as Corticosterone; D-Galactose; Ethanolamine HC1; Glutathione (reduced); L-Carnitine HC1;Linoleic Acid; Linolenic Acid; Progesterone; Putrescine 2HC1; Sodium Selenite; and / or T3 (triiodo-I-thyronine). . In specific embodiments, one or more of these may be explicitly excluded.

[0104] In some embodiments, the medium further comprises vitamins. In some embodiments, the medium comprises 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13 of the following (and any range derivable therein): biotin, DL alpha tocopherol acetate, DL alpha-tocopherol, vitamin A, choline chloride, calcium pantothenate, pantothenic acid, folic acid nicotinamide, pyridoxine, riboflavin, thiamine, inositol, vitamin B12, or the medium includes combinations thereof or salts thereof. In some embodiments, the medium comprises or consists essentially of biotin, DL alpha tocopherol acetate, DL alpha-tocopherol, vitamin A, choline chloride, calcium pantothenate, pantothenic acid, folic acid nicotinamide, pyridoxine, riboflavin, thiamine, inositol, and vitamin B 12. In some embodiments, the vitamins include or consist essentially of biotin, DL alpha tocopherol acetate, DL alpha-tocopherol, vitamin A, or combinations or salts thereof. In some embodiments, the medium further comprises proteins. In some embodiments, the proteins comprise albumin or bovine serum albumin, a fraction of BSA, catalase, insulin, transferrin, superoxide dismutase, or combinations thereof. In some embodiments, the medium further comprises one or more of the following: corticosterone, D-Galactose, ethanolamine, glutathione, L-camitine, linoleic acid, linolenic acid, progesterone, putrescine, sodium selenite, or triiodo-I-thyronine, or combinations thereof. In some embodiments, the medium comprises one or more of the following: a B-27® supplement, xeno-free B-27® supplement, GS21TM supplement, or combinations thereof. In some embodiments, the medium comprises or further comprises amino acids, monosaccharides, inorganic ions. In some embodiments, the amino acids comprise arginine, cystine, isoleucine, leucine, lysine, methionine, glutamine, phenylalanine, threonine, tryptophan, histidine, tyrosine, or valine, or combinations thereof. In some embodiments, the inorganic ions comprise sodium, potassium, calcium, magnesium, nitrogen, or phosphorus, or combinations or salts thereof. In some embodiments, the medium further comprises one or more of the following: molybdenum, vanadium, iron, zinc, selenium, copper, or manganese, or combinations thereof. In certain embodiments, the medium comprises or consists essentially of one or more vitamins discussed herein and / or one or more proteins discussed herein, and / or one or more of the following: corticosterone, D-Galactose, ethanolamine, glutathione, L-camitine, linoleic acid, linolenic acid, progesterone, putrescine, sodium selenite, or triiodo-I-thyronine, a B-27® supplement, xeno-free B-27® supplement, GS21TM supplement, an amino acid (such as arginine, cystine, isoleucine, leucine, lysine, methionine, glutamine, phenylalanine, threonine, tryptophan, histidine, tyrosine, or valine),monosaccharide, inorganic ion (such as sodium, potassium, calcium, magnesium, nitrogen, and / or phosphorus) or salts thereof, and / or molybdenum, vanadium, iron, zinc, selenium, copper, or manganese. In specific embodiments, one or more of these may be explicitly excluded.

[0105] The medium can also contain one or more externally added fatty acids or lipids, amino acids (such as non-essential amino acids), vitamin(s), growth factors, cytokines, antioxidant substances, 2-mercaptoethanol, pyruvic acid, buffering agents, and / or inorganic salts. In specific embodiments, one or more of these may be explicitly excluded.

[0106] One or more of the medium components may be added at a concentration of at least, at most, or about 0.1, 0.5, 1, 2, 3, 4, 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 150, 180, 200, 250 ng / L, ng / ml, pg / ml, mg / ml, or any range derivable therein.

[0107] In specific embodiments, the cells of the disclosure are specifically formulated. They may or may not be formulated as a cell suspension. In specific cases they are formulated in a single dose form. They may be formulated for systemic or local administration. In some cases the cells are formulated for storage prior to use, and the cell formulation may comprise one or more cryopreservation agents, such as DMSO (for example, in 5% DMSO). The cell formulation may comprise albumin, including human albumin, with a specific formulation comprising 2.5% human albumin. The cells may be formulated specifically for intravenous administration; for example, they are formulated for intravenous administration over less than one hour. In particular embodiments the cells are in a formulated cell suspension that is stable at room temperature for 1, 2, 3, or 4 hours or more from time of thawing.IV. Kits of the Disclosure

[0108] Any of the cellular and / or non-cellular compositions described herein or similar thereto may be comprised in a kit. In a non-limiting example, one or more reagents for use in methods for preparing cellular therapy may be comprised in a kit. Such reagents may include cells, IFN-gamma, platelet rich plasma, platelet lysate, one or more angiogenic factors, one or more growth factors, vector(s) one or more costimulatory factors, media, enzymes, buffers, nucleotides, salts, primers, and so forth. The kit may comprise any protein listed in the disclosure. The kit components are provided in suitable container means.

[0109] Some components of the kits may be packaged either in aqueous media or in lyophilized form. The container means of the kits will generally include at least one vial, test tube, flask, bottle, syringe or other container means, into which a component may be placed,and preferably, suitably aliquoted. Where there are more than one component in the kit, the kit also will generally contain a second, third or other additional container into which the additional components may be separately placed. However, various combinations of components may be comprised in a vial. The kits of the present disclosure also will typically include a means for containing the components in close confinement for commercial sale. Such containers may include injection or blow molded plastic containers into which the desired vials are retained.

[0110] When the components of the kit are provided in one and / or more liquid solutions, the liquid solution is an aqueous solution, with a sterile aqueous solution being particularly useful. In some cases, the container means may itself be a syringe, pipette, and / or other such like apparatus, or may be a substrate with multiple compartments for a desired reaction.[OHl] Some components of the kit may be provided as dried powder(s). When reagents and / or components are provided as a dry powder, the powder can be reconstituted by the addition of a suitable solvent. It is envisioned that the solvent may also be provided in another container means. The kits may also comprise a second container means for containing a sterile acceptable buffer and / or other diluent.

[0112] In specific embodiments, reagents and materials include primers for amplifying desired sequences, nucleotides, suitable buffers or buffer reagents, salt, and so forth, and in some cases the reagents include apparatus or reagents for isolation of a particular desired cell(s).

[0113] In particular embodiments, there are one or more apparatuses in the kit suitable for extracting one or more samples from an individual. The apparatus may be a syringe, fine needles, scalpel, and so forth.V. Examples

[0114] The following examples are included to demonstrate preferred embodiments of the invention. It should be appreciated by those of skill in the art that the techniques disclosed in the examples which follow represent techniques discovered by the inventor to function well in the practice of the invention, and thus can be considered to constitute preferred modes for its practice. However, those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments which are disclosed and still obtain a like or similar result without departing from the spirit and scope of the disclosure.EXAMPLE 1TREATMENT OF IMMUNE-RELATED SPLENOMEGALY

[0115] In a mouse model of imiquimod-induced psoriasis, human dermal fibroblast demonstrated significant therapeutic effects, not only by reducing the severity of psoriasis but also by attenuating imiquimod-induced splenomegaly. Indeed, a single intravenous dose of fibroblasts on Day 4 notably reduced skin inflammation, reflected in improved clinical scores for thickness, erythema, and scaling, from Day 6 to Day 8 (FIGS. 1 A-1E). Further, mice treated with fibroblasts showed a marked reduction in spleen weight and length (as seen at least in FIG. 2A-2C), as the fibroblasts effectively modulated systemic inflammation beyond skin pathology. Flow cytometry analysis of splenocytes revealed a significant reduction in the population of macrophages (CDllb+F4 / 80+ cells) in the spleens of fibroblasts-treated mice. Such a finding shows that the treatment suppresses macrophage infiltration and / or activation in the spleen. Additionally, comparative analysis showed that both fibroblast and anti-IL-23 treatments effectively reduced IMQ-induced psoriasis over a 7-day period, as evidenced by improvements in skin appearance and clinical scores for thickness, erythema, and scaling (FIGS. 3A-3E). Importantly, no significant changes were observed in other immune cell populations, such as dendritic cells, B cells, T cells, and NK cells. This selective reduction in splenic macrophages, coupled with the absence of splenomegaly, highlights fibroblasts' ability to specifically target macrophage-driven inflammation (FIG. 5B-5C). Additionally, mice treated with fibroblast showed normalized platelet counts. That is, flow cytometry revealed marked reductions in CD3+, CD4+, CD8+, and CD25+T cell populations (FIG. 6A-6D). Such results indicate that fibroblasts effectively modulate adaptive immune responses within the spleen.

[0116] As a comparator, an anti-IL-23 monoclonal antibody was evaluated alongside fibroblasts. Over the monitoring period, the anti-IL-23 treatment did not exhibit the same systemic benefits as fibroblasts. Mice treated with anti-IL-23 showed no significant reduction in splenic macrophage populations and continued to exhibit splenomegaly, with enlarged spleens and notably lower platelet counts compared to healthy controls (FIG 4A-4C). These findings demonstrate that anti-IL-23 treatment does not address macrophage-mediated systemic inflammation in the spleen. In contrast, fibroblasts offer a unique therapeutic capability by mitigating splenomegaly and restoring hematological balance, making them a useful avenue for treatment of inflammation-related splenomegaly in conditions like psoriasis.

[0117] Animals and psoriasis-related splenomegaly model. For psoriasis induction on the back, mice were anesthetized using an isoflurane respirator. A rectangular area of 3 cm x 4 cm on the back skin was shaved and treated with Nair™ hair-removal cream on Day 0. Psoriasis was induced via topical application of 31.25 mg of IMQ cream (5% IMQ, Taro Pharmaceutical, Hawthorne, NY, USA) to the lower dorsal skin for 7 consecutive days. Mouse body weight was recorded daily. All experimental procedures followed the NIH Guide for the Care and Use of Laboratory Animals and were approved by the Institutional Animal Care and Use Committee at the K2Bio Lab (Houston, TX, USA) and StillMeadow, Inc. (Sugar Land, TX, USA).

[0118] Clinical assessment of psoriasis and spleen mass. The clinical features of IMQ-induced psoriasis were evaluated daily using the Psoriasis Area and Severity Index (PASI) as described previously (12). Briefly, double-folded skin thickness was measured using a digital micrometer (Rexbeti, Auburn, Washington, USA). Psoriatic skin lesions, including thickness, erythema, and scaling, were assessed by two observers daily on a scale of 0 to 4 (0-no incidence; 1-slight; 2-moderate; 3-marked; 4-severe), respectively. No significant deviation was noted between observers. The scores from two observers were then averaged to determine the mean PASI for each mouse. Tail vein intravenous injection of fibroblast spheroids occurred at mild psoriasis (average PASI score 2). To compare the therapeutic efficacy of fibroblast spheroids with a pre-clinical monoclonal antibody targeting mouse IL-23 (anti-IL-23, BioxCell, Lebanon, NH, USA), anti-IL-23 was administered intraperitoneally on Days 1, 3, and 5. Euthanasia occurred on Day 8. Mouse spleens were harvested and measured for length and weight.

[0119] Cell culture and treatment. Human dermal fibroblasts (HDFs) purchased from Gibco (Thermo Fisher Scientific, Waltham, MA, USA) were cultured in Cellartis MSC Xeno-Free Culture Medium (Takara Bio, Japan) at 37°C with 5% CO2 until they reached 70-80% confluence. For 3D spheroid culture, Elplasia® ultra-low attachment microcavity plates and flasks (Corning, New York, USA) were used according to the manufacturer's instructions. HDF spheroids cultured in 6-well Elplasia® plates (seeded at 5xl06cells / well) or 12K flasks (seeded at 20xl06cells / flask) were washed twice with PBS, resuspended in plasma-Lyte A (PLA), and 577 spheroids were administered via the tail vein at a dosage of IxlO6cells / 100 mL / mouse.

[0120] Flow cytometry. Spleens were placed in cold RPMI-1640 medium supplemented with 10% FBS and 1% penicillin-streptomycin. Spleens were then digested in the medium containing 150 mg / mL Liberase (Millipore Sigma, St Louis, MO, USA) for 1 hour at 37°C and homogenized by gently pressing through a 70 pm cell strainer (Greiner Bio-One,Kremsmunster, Austria) using the plunger of a syringe. The homogenate was collected into a 15 mL conical tube, and the cell suspension was centrifuged at 300 x g for 7 minutes at 4°C. The supernatant was discarded, and the cell pellet was resuspended in 1 mL of RBC lysis buffer (Thermo Fisher Scientific, Waltham, MA, USA) and incubated for 3-5 minutes at RT. Cells were washed twice with PBS by centrifugation at 300 x g for 5 minutes at 4°C and resuspended in 1 mL of PBS containing 2% FBS. Cells were blocked with anti-mouse CD16 / CD32 (TruStain FcX™, BioLegend, San Diego, CA, USA) and stained with fluorophore-conjugated antibodies, including CDllb (BV421 -conjugated, BD Biosciences, Franklin Lakes, New Jersey, USA) and F4 / 80 (Alexa Fluor 488-conjugated, Thermo Fisher Scientific, Waltham, MA, USA). Following 10 min incubation at 4°C in the dark, cells were washed twice with PBS and resuspended in 100 mL PBS containing 2% PF A. Stained cells were analyzed using a CytoFLEX flow cytometer (Beckman Coulter, Brea, CA, USA). The percentage of macrophages within the splenocyte population was determined based on F4 / 80 and CDllb, and data were analyzed using FlowJo software (Tree Star, Inc.). The absolute number of macrophages was calculated by multiplying the total number of splenocytes, determined by cell counting using a NucleoCounter NC-3000 instrument, by the percentage of macrophages identified through flow cytometry (13).

[0121] Statistical analysis. Statistical analysis was performed using GraphPad Prism version 10 (GraphPad Software, La Jolla, CA, USA). Results are expressed as the Mean ± Standard Error of the Mean (SEM). Normality tests were performed, and statistical differences between groups were analyzed using one-way ANOVA followed by Holm-Sidak's multiple comparison test for parametric data or Kruskal-Wallis test followed by Dunn's multiple comparison test for non-parametric data. A p-value of less than 0.05 was considered statistically significant.REFERENCES

[0122] The following references, to the extent that they provide exemplary procedural or other details supplementary to those set forth herein, are specifically incorporated herein by reference.1. Jalili RB, Zhang Y, Hosseini-Tabatabaei A, Kilani RT, Khosravi Maharlooei M, Li Y, et al. Fibroblast Cell-Based Therapy for Experimental Autoimmune Diabetes. PLoS One. 2016;l l(l):e0146970.2. Jalili RB, Kilani RT, Li Y, Khosravi-Maharlooie M, Nabai L, Wang EHC, et al.Fibroblast cell-based therapy prevents induction of alopecia areata in an experimental model. Cell Transplant. 2018;27(6):994-1004.3. Bouffi C, Bony C, Jorgensen C, Noel D. Skin fibroblasts are potent suppressors of inflammation in experimental arthritis. Ann Rheum Dis. 2011;70(9):1671-6.4. Ichim TE, O’Heeron P, Perez J, Liu P, Min W-P, Kesari S. Fibroblasts as an Alternative to Mesenchymal Stem Cells with Successful Treatment and Immune Modulation in EAE Model of Multiple Sclerosis. bioRxiv. 2020:2020.06. 04.133249.5. Pozo AL, Godfrey EM, Bowles KM. Splenomegaly: investigation, diagnosis and management. Blood reviews. 2009;23(3):105-ll.6. Lowes MA, Suarez-Farinas M, Krueger JG. Immunology of psoriasis. Annu Rev Immunol. 2014;32:227-55.7. Rendon A, Schakel K. Psoriasis Pathogenesis and Treatment. Int J Mol Sci.2019;20(6).8. Samotij D, Nedoszytko B, Bartosinska J, Batycka-Baran A, Czajkowski R, Dobrucki IT, et al. Pathogenesis of psoriasis in the "omic" era. Part I. Epidemiology, clinical manifestation, immunological and neuroendocrine disturbances. Postepy Dermatol Alergol.2020;37(2): 135-53.9. Thangapazham RL, Darling TN, Meyerle J. Alteration of skin properties with autologous dermal fibroblasts. Int J Mol Sci. 2014;15(5):8407-27.10. Kim S, Hupperetz C, Lim S, Kim CH. Genome editing of immune cells using CRISPR / Cas9. BMB Rep. 2021;54(l):59-69.11. Basu A, Tiwari VK. Epigenetic reprogramming of cell identity: lessons from development for regenerative medicine. Clin Epigenetics. 2021 ; 13(1): 144.12. Neu SD, Strzepa A, Martin D, Sorci-Thomas MG, Pritchard KA, Jr., Dittel BN.Myeloperoxidase Inhibition Ameliorates Plaque Psoriasis in Mice. Antioxidants (Basel). 2021; 10(9).13. Fujiyama S, Nakahashi-Oda C, Abe F, Wang Y, Sato K, Shibuya A. Identification and isolation of splenic tissue-resident macrophage sub-populations by flow cytometry. Int Immunol. 2019;31(1):51-6.* * *

[0123] All of the methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methodsof this invention have been described in terms of preferred embodiments, it will be apparent to those of skill in the art that variations may be applied to the methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the invention. More specifically, it will be apparent that certain agents which are both chemically and physiologically related may be substituted for the agents described herein while the same or similar results would be achieved. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the invention as defined by the appended claims.ADDITIONAL RECITED EMBODIMENTS:

[0124] Embodiment 1 : A method of treating splenomegaly in an individual, comprising the step of administering to the individual a therapeutically effective amount of fibroblasts and / or fibroblast-derived products.

[0125] Embodiment 2: The method of embodiment 1, wherein the fibroblast-derived products comprise fibroblast spheroids, fibroblast exosomes, fibroblast microvesicles, fibroblast lysate, fibroblast apoptotic bodies, fibroblast conditioned media, or a combination thereof.

[0126] Embodiment 3: The method of embodiment 1 or 2, wherein following the administering, there is a reduction of the spleen mass and the product! on / infiltrati on of immune cells in the spleen.

[0127] Embodiment 4: The method of embodiment 3, wherein the splenic immune cells express pathogenic levels of IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF, IFN-y, TGF-pl, or a combination thereof.

[0128] Embodiment 5: The method of embodiment 3 or 4, wherein the splenic immune cells are pathogenic B cells, T cells, dendritic cells, monocytes, macrophages, NK cells, or a combination thereof.

[0129] Embodiment 6: The method of any one of embodiments 1-3, wherein the administering is topically, intravenously, or by subcutaneous injection.

[0130] Embodiment 7: The method of any one of claims 1-6, wherein the fibroblasts or fibroblast-derived products are autologous with respect to the individual.

[0131] Embodiment 8: The method of any one of embodiments 1-6, wherein the fibroblasts or fibroblast-derived products are allogeneic with respect to the individual.

[0132] Embodiment 9: The method of any one of embodiments 1-6, wherein the fibroblasts or fibroblast-derived products are xenogeneic with respect to the individual.

[0133] Embodiment 10: The method of any one of embodiments 1-6, wherein the fibroblasts or fibroblast-derived products are syngeneic with respect to the individual.

[0134] Embodiment 11: A method of treating splenomegaly in an individual, comprising the step of providing to the individual an effective amount of: (1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; and (2) one or more types of immune cells and / or one or more derivative agents of the immune cells, optionally wherein the derivative agents are cytokines and / or chemokines.

[0135] Embodiment 12: The method of embodiment 11, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual, optionally via intravenous injection or subcutaneous injection.

[0136] Embodiment 13: The method of embodiment 11 or 12, wherein following the providing step, there is decrease of the production / infiltration of one or more types of pathogenic immune cells.

[0137] Embodiment 14: The method of any one of embodiments 11-13, wherein the providing of (1) and (2) occurs generally simultaneously.

[0138] Embodiment 15: The method of any one of embodiments 11-13, wherein the providing of (1) and (2) occurs at different times.

[0139] Embodiment 16: The method of embodiment 15, wherein (1) occurs before (2).

[0140] Embodiment 17: The method of embodiment 15, wherein (1) occurs after (2).

[0141] Embodiment 18: The method of embodiment 11, wherein the fibroblasts are activated and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts.

[0142] Embodiment 19: The method of embodiment 12, wherein the fibroblasts are activated ex vivo with one or more nucleic acids, one or more transcription factor activators, one or more growth factors, one or more cytokines, one or more chemokines, and / or with coculture with non-fibroblast cells, optionally wherein the fibroblasts are activated ex vivo with IL-1, IL-6, IL-8, MCP-1, CCL-2, prostaglandins, miRNA-21, miRNA-23b, miRNA-31, miR-215, miR-1, miR-133, miR-208, and miR-499, transcription factors Mcm3, Dicerl, Cdc25A, Ick, Tripl3, McmlO, OCT4, SOX2, KLF4, and MYC, Pax3, or a combination thereof.

[0143] Embodiment 20: The method of embodiment 19, wherein the non-fibroblast cells are one or more types of immune cells.

[0144] Embodiment 21: The method of embodiment 20, wherein the immune cells are T cells, dendritic cells, natural killer cells, natural killer T cells, macrophages, microglia, B cells, neutrophils, or a combination thereof.

[0145] Embodiment 22: A method of treating splenomegaly in an individual, comprising the step of providing to the individual an effective amount of: (1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and (2) one or more types of immune cells and / or one or more derivative agents thereof.

[0146] Embodiment 23: The method of embodiment 22, wherein the fibroblasts are activated ex vivo with one or more nucleic acids, one or more transcription factor activators, one or more growth factors, one or more cytokines, one or more chemokines, and / or with coculture with non-fibroblast cells.

[0147] Embodiment 24: The method of embodiment 22, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

[0148] Embodiment 25: The method of embodiment 22 or 23, wherein following the providing step, there is a decrease of the expression in immune cells of the individual of pathogenic levels from the cells of one or more of IL-23, IL-1, IL-10, IL-22, IL-17, IL-12, TNF, INF-y, TGF-pl.

[0149] Embodiment 26: A method of epigenetic reprogramming of localized fibroblasts and endothelial cells in an individual with splenomegaly, comprising the step of providing to the individual an effective amount of: (1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and (2) one or more types of immune cells or derivative agent.

[0150] Embodiment 27: The method of embodiment 26, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

[0151] Embodiment 28: A method of enabling the activation and migration of localized stem cell niches to replace the pathogenetic fibroblasts and / or endothelial cells in the spleen ofan individual, comprising the step of providing to the individual an effective amount of: (1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and (2) one or more types of immune cells or derivative agent.

[0152] Embodiment 29: The method of embodiment 28, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

[0153] Embodiment 30: A method of eliciting an immune response in an individual with splenomegaly, comprising the step of providing to the individual an effective amount of: (1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; and (2) one or more adjuvants.

[0154] Embodiment 31 : The method of embodiment 30, wherein the adjuvant is chemical product-based, viral product-based, bacterial product-based, or a mixture thereof.

[0155] Embodiment 32: The method of any one of the embodiments 1-31, wherein the method of culturing the composition comprises a three-dimensional culture.

[0156] Embodiment 33: The method of any one of embodiments 1-32, whether the individual has an enlarged spleen.

[0157] Embodiment 34: The method of any one of embodiments 1-33, that the individual has an enlarged spleen.

[0158] Embodiment 35: The method of any one of embodiments 1-34, wherein the individual is without psoriasis.

[0159] Embodiment 36: The method of any one of embodiments 1-35, wherein the individual is with hematologic conditions comprising hemolytic anemia or leukemia, infectious diseases like mononucleosis or malaria, metabolic disorders such as Gaucher disease or Niemann-Pick disease.

[0160] Embodiment 37: The method of any one of embodiments 1-36, wherein the individual is with autoimmune conditions like systemic lupus erythematosus (SLE) or rheumatoid arthritis.

[0161] Embodiment 38: The method of any one of embodiments 1-37, wherein the individual is with chronic liver diseases such as cirrhosis or portal hypertension.

[0162] Embodiment 39: The method of embodiments 1-38, wherein following the administering, there is a normalization of platelet count within the individual.

Claims

CLAIMSWHAT IS CLAIMED IS:

1. A method of treating splenomegaly in an individual, comprising the step of administering to the individual a therapeutically effective amount of fibroblasts and / or fibroblast-derived products.

2. The method of claim 1, wherein the fibroblast-derived products comprise fibroblast spheroids, fibroblast exosomes, fibroblast microvesicles, fibroblast lysate, fibroblast apoptotic bodies, fibroblast conditioned media, or a combination thereof.

3. The method of claim 1 or 2, wherein following the administering, there is a reduction of the spleen mass and the production / infiltration of immune cells in the spleen.

4. The method of claim 3, wherein the splenic immune cells express pathogenic levels of IL- 23, IL-1, IL- 10, IL-22, IL- 17, IL- 12, TNF, IFN-y, TGF-pl, or a combination thereof.

5. The method of claim 3 or 4, wherein the splenic immune cells are pathogenic B cells, T cells, dendritic cells, monocytes, macrophages, NK cells, or a combination thereof.

6. The method of any one of claims 1-3, wherein the administering is topically, intravenously, or by subcutaneous injection.

7. The method of any one of claims 1-6, wherein the fibroblasts or fibroblast-derived products are autologous with respect to the individual.

8. The method of any one of claims 1-6, wherein the fibroblasts or fibroblast-derived products are allogeneic with respect to the individual.

9. The method of any one of claims 1-6, wherein the fibroblasts or fibroblast-derived products are xenogeneic with respect to the individual.

10. The method of any one of claims 1-6, wherein the fibroblasts or fibroblast-derived products are syngeneic with respect to the individual.

11. A method of treating splenomegaly in an individual, comprising the step of providing to the individual an effective amount of:(1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; and(2) one or more types of immune cells and / or one or more derivative agents of the immune cells, optionally wherein the derivative agents are cytokines and / or chemokines.

12. The method of claim 11, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual, optionally via intravenous injection or subcutaneous injection.

13. The method of claim 11 or 12, wherein following the providing step, there is decrease of the production / infiltration of one or more types of pathogenic immune cells.

14. The method of any one of claims 11-13, wherein the providing of (1) and (2) occurs generally simultaneously.

15. The method of any one of claims 11-13, wherein the providing of (1) and (2) occurs at different times.

16. The method of claim 15, wherein (1) occurs before (2).

17. The method of claim 15, wherein (1) occurs after (2).

18. The method of claim 11, wherein the fibroblasts are activated and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts.

19. The method of claim 12, wherein the fibroblasts are activated ex vivo with one or more nucleic acids, one or more transcription factor activators, one or more growth factors, one or more cytokines, one or more chemokines, and / or with co-culture with non- fibroblast cells, optionally wherein the fibroblasts are activated ex vivo with IL-1, IL- 6, IL-8, MCP-1, CCL-2, prostaglandins, miRNA-21, miRNA-23b, miRNA-31, miR- 215, miR-1, miR-133, miR-208, and miR-499, transcription factors Mcm3, Dicerl, Cdc25A, Ick, Tripl3, McmlO, OCT4, SOX2, KLF4, and MYC, Pax3, or a combination thereof.

20. The method of claim 19, wherein the non-fibroblast cells are one or more types of immune cells.

21. The method of claim 20, wherein the immune cells are T cells, dendritic cells, natural killer cells, natural killer T cells, macrophages, microglia, B cells, neutrophils, or a combination thereof.

22. A method of treating splenomegaly in an individual, comprising the step of providing to the individual an effective amount of:(1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and(2) one or more types of immune cells and / or one or more derivative agents thereof.

23. The method of claim 22, wherein the fibroblasts are activated ex vivo with one or more nucleic acids, one or more transcription factor activators, one or more growth factors, one or more cytokines, one or more chemokines, and / or with co-culture with non- fibroblast cells.

24. The method of claim 22, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

25. The method of claim 22 or 23, wherein following the providing step, there is a decrease of the expression in immune cells of the individual of pathogenic levels from the cells of one or more of IL-23, IL-1, IL- 10, IL-22, IL- 17, IL- 12, TNF, INF-y, TGF-pl.

26. A method of epigenetic reprogramming of localized fibroblasts and endothelial cells in an individual with splenomegaly, comprising the step of providing to the individual an effective amount of:(1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and(2) one or more types of immune cells or derivative agent.

27. The method of claim 26, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

28. A method of enabling the activation and migration of localized stem cell niches to replace the pathogenetic fibroblasts and / or endothelial cells in the spleen of an individual, comprising the step of providing to the individual an effective amount of(1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; wherein the fibroblasts are activated with one or more agents and / or the fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, or fibroblast-derived materials come from activated fibroblasts are activated with one or more agents; and(2) one or more types of immune cells or derivative agent.

29. The method of claim 28, wherein the providing is (a) into the bloodstream of the individual and / or (b) into and / or onto the skin tissue of the individual.

30. A method of eliciting an immune response in an individual with splenomegaly, comprising the step of providing to the individual an effective amount of(1) fibroblasts, fibroblast spheroids, fibroblast exosomes, fibroblast lysates, fibroblast apoptotic bodies, and / or fibroblast-derived materials; and(2) one or more adjuvants.

31. The method of claim 30, wherein the adjuvant is chemical product-based, viral productbased, bacterial product-based, or a mixture thereof.

32. The method of any one of the claims 1-31, wherein the method of culturing the composition comprises a three-dimensional culture.

33. The method of any one of claims 1-32, whether the individual has an enlarged spleen.

34. The method of any one of claims 1-33, that the individual has an enlarged spleen.

35. The method of any one of claims 1-34, wherein the individual is without psoriasis.

36. The method of any one of claims 1-35, wherein the individual is with hematologic conditions comprising hemolytic anemia or leukemia, infectious diseases like mononucleosis or malaria, metabolic disorders such as Gaucher disease or Niemann- Pick disease.

37. The method of any one of claims 1-36, wherein the individual is with autoimmune conditions like systemic lupus erythematosus (SLE) or rheumatoid arthritis.

38. The method of any one of claims 1-37, wherein the individual is with chronic liver diseases such as cirrhosis or portal hypertension.

39. The method of claims 1-38, wherein following the administering, there is a normalization of platelet count within the individual.

Images