Magnetic microgels for composite musculoskeletal tissue regeneration

Abstract

Various implementations described herein relate to the production and use of magnetic microgels. According to some implementations, the magnetic microgels include a therapeutic agent. Magnetic microgels can facilitate controlled spatiotemporal delivery of the therapeutic agent. Various implementations provide scalable methods for production of magnetic microgels. Various implementations described herein can be used for research, diagnostic, or therapeutic uses.

Description

MAGNETIC MICROGELS FOR COMPOSITE MUSCULOSKELETAL TISSUEREGENERATIONCROSS-REFERENCE TO RELATED APPLICATION(S)

[0001] This application claims the priority of U.S. Provisional Patent Application No. 63 / 662,901 , filed on June 21 , 2024, which is incorporated by reference herein in its entirety.REFERENCE TO SEQUENCE LISTING

[0002] The Sequence Listing associated with this application is provided in XML format in lieu of a paper copy and is hereby incorporated by reference into the specification. The name of the file containing the Sequence Listing is W149- 6005PCT.xml. The file is 4,338 bytes, was created June 17, 2025, and is being submitted electronically via Patent CenterTECHNICAL FIELD

[0003] This application relates to techniques for delivering therapeutic agents to specific areas within the body of a subject using magnetic microgels. The temporal release of the therapeutic agents from the magnetic microgels can be controlled using various implementations described herein.BACKGROUND

[0004] Humans have poor regenerative capacity of amputated appendages, including the limbs and digits. Controlled delivery of growth factors that stimulate pro-regenerative signaling pathways could promote limb regrowth, thus improving the prognosis for amputees. To that end, the mouse digit tip is used to investigate the mechanisms of composite musculoskeletal tissue regeneration following amputation, where stem cells (e.g., progenitor cells) consistently regrow the bone and soft tissues after distal digit amputation. However, amputations that occur more proximally lead to fibrotic scarring, and resected synovial joints typically do not naturally regenerate in humans. Previous studies demonstrated that the consecutive delivery of growth factors to the amputated bone stump, such as bone morphogenetic protein-2 (BMP-2) followed by BMP-9, can induce skeletal elongation and subsequent joint regeneration. However, these studies relied on multiple surgeries to deliver proteins in a time- and location-dependent manner, which can be clinically challenging.BRIEF DESCRIPTION OF THE DRAWINGS

[0005] FIG. 1A illustrates an example environment for the treatment of a subject suffering from a musculoskeletal condition. FIG. 1 B illustrates an example environment for the production of microgels.

[0006] FIG. 2 illustrates a schematic of microgel synthesis.

[0007] FIG. 3A illustrates a schematic of desired growth factor release profiles. FIG. 3B illustrates the expected outcome of magnetic bioactive microgel delivery.

[0008] FIGs. 4A-4D illustrate example characterization of the disclosed magnetic microgels. FIG. 4A illustrates a PEG- NB microgel schematic. FIG. 4B illustrates microgels with varying levels of Fe3O4 incorporation. Scale: 0.1 mm. FIG. 4C illustrates quantification of microgel size, shape, and opacity (n=73-120 / group, mean ± SD). *: p<0.05 between groups.FIG. 4D illustrates velocity of low FegC microgels in response to a magnetic field (n=5-9 / group, mean ± SD) *: p<0.05 between groups.

[0009] FIGs. 5A and 5B illustrate schematics of microgel synthesis using (FIG. 5A) mineral oil and (FIG. 5B) synthetic oil.

[0010] FIG. 6 shows images of control, low Fe3O4, and high FesC microgels made with mineral and synthetic oil. Dashed line: microgel border. Scale: 50 pm.

[0011] FIG. 7A illustrates microgel diameter, circularity, and opacity (n=70- 103 / group, mean ± SD). $: p<0.05 vs Low. *: p<0.05 vs. High. #: p<0.05 vs. Mineral. FIG. 7B illustrates a schematic of the experimental setup to evaluate microgel translocation. FIGs. 7C and 7D illustrate velocity of microgels fabricated with (FIG. 7C) mineral and (FIG. 7D) synthetic oil with or without magnetic stimulation (n=6- 8 / group, mean ± SD). $: p<0.05 vs Low. *: p<0.05 vs. High. #: p<0.05 vs. No Magnet.

[0012] FIG. 8 illustrates the effect of vortex mixing speed and time on microgel size. $: p<0.05 vs. Medium speed. *: p<0.05 vs. High speed. #: p<0.05 vs. 5 seconds. %: p<0.05 vs. 7 seconds. n=70— 10O / group, mean ± SD.

[0013] FIGs. 9A-9C illustrate (FIG. 9A) a schematic of the experimental setup for evaluating translocation in a collagen hydrogel and (FIG. 9B) images of magnetic microgels translocating in the collagen hydrogel. FIG. 9C shows a quantification of the microgel displacement illustrated in FIG. 9B.

[0014] FIG. 10 illustrates the attachment of cells to the microgels using the cell-adhesive peptide arginylglycylaspartic acid (RGD).DETAILED DESCRIPTION

[0015] In 2017, an estimated 57.7 million people worldwide were living with limb amputations resulting from traumatic injuries, diabetes mellitus, peripheral vascular disease, or neuropathy (McDonald, et al. Prosthet Orthot Int.2021; 45(2): 105-14). Unfortunately, humans have poor regenerative capacity of amputated appendages, including the limbs and digits (Yokoyama, Dev Growth Differ. 2007;50(1 ): 13-22 ). Controlled delivery of growth factors that stimulate pro-regenerative signaling pathways could promote limb regrowth, thus improving the prognosis for amputees. To that end, the mouse digit tip is used to investigate the mechanisms of composite musculoskeletal tissue regeneration following amputation, where stem / progenitor cells consistently regrow the bone and soft tissues after distal digit amputation (Storer and Miller, Open Biol. 2020; 10(9):200194; Lehoczky, et al. PNAS. 2011 ; 108(51):20609-14). However, amputations that occur more proximally lead to fibrotic scarring, and resected synovial joints do not naturally regenerate (Qu, et al. FASEB J 2020;34:9740-54; Simkin, et al. Methods Mol Biol. 2013;1037:419-35). Previous studies demonstrated that the consecutive delivery of growth factors to the amputated bone stump, such as bone morphogenetic protein-2 (BMP-2) followed by BMP-9, can induce skeletal elongation and subsequent joint regeneration (Yu, et al. Nat Commun. 2019;10:424). However, these studies relied on multiple surgeries to deliver proteins in a time- and locationdependent manner, which would be clinically challenging. Developing methods to regenerate composite musculoskeletal tissues through the controlled delivery of growth factors can enhance the quality of life for amputees.

[0016] Various implementations described herein relate to the production and use of magnetic microgels. In various implementations, magnetic microgels can enable spatiotemporal control of the delivery of therapeutic agents. The present disclosure describes use of magnetic microgels for simultaneous delivery of multiple therapeutic agents with distinct delivery parameters (e.g., release rate, release duration, spatial distribution of release, etc.). Accordingly, the disclosed magnetic microgels can facilitate complex tissue repair, such as the regrowth of multiple tissue types including epithelial tissue, connective tissue, muscle tissue, and nervous tissue. The present disclosure enables bone and joint formation without the need for repeated invasive procedures to administer growth factors and other agents into a subject.

[0017] Conventionally, microgels are often generated using microfluidic technology, which suffers from low throughput. Various implementations of the present disclosure provide scalable methods to generate magnetic microgels with a controlled size (e.g., diameter) and size distribution. Accordingly, various implementations can enhance control of drug delivery using both the magnetic microgels and non-magnetic microgels.

[0018] Implementations of the present disclosure will now be described with reference to the accompanying figures.

[0019] FIG. 1A illustrates an example environment 100 for the treatment of a subject 102 suffering from a musculoskeletal condition. In various implementations of the present disclosure, musculoskeletal conditions include conditions in which tissue is damaged or lost. For instance, examples of musculoskeletal conditions include loss of a body part (e.g., after an amputation), wounds (e.g., open wounds, closed wounds, burns, etc.), injuries (e.g., fractures, sprains, dislocations, etc.), bone disorders (e.g., bone cancer, osteoporosis, osteopenia, etc.), muscular disorders (e.g., muscular dystrophy), surgical tissue resections, or the like. In some examples, the subject 102 may use a prosthetic device to restore function of a lost body part. The subject 102 may be a human, a non-human primate, a mammal, or another animal.

[0020] In various examples, the subject 102 presents to a healthcare facility for treatment of the musculoskeletal condition. One or more care providers 104 are responsible for treating the subject 102. The care provider(s) 104 may include a clinician, a laboratory technician, or the like. In some examples, the care provider(s) may image an affected area 105 of the body of the subject 102. For instance, the affected area 105 may be visualized using ultrasound, x-ray, computed tomography (CT), magnetic resonance imaging (MRI), positron emission tomography (PET), single photon emission CT (SPECT), or any combination thereof. In some examples, the subject 102 has an amputated limb, and the care provider(s) 104 may image the residual limb as well as, in some cases, the contralateral limb of the subject 102. Visualizing the contralateral area of the subject 102 can, in various examples, provide information for the care provider(s) 104 to guide the treatment of the residual limb.

[0021] In various implementations, the care provider(s) 104 may determine that the subject 102 may benefit from treatment with one or more therapeutic agents in the affected area 105 Examples of therapeutic agents include growth factors, pain management medications, anti-inflammatory medications, therapeutic cells (e.g., stem cells), peptides, immune signaling molecules (e.g., cytokines, chemokines, etc.), angiogenic compounds, gene therapy reagents, exosomes, extracellular vesicles, and the like. Examples of growth factors include bone morphogenetic protein (BMP; e.g., BMP-2, BMP-4, BMP-7, BMP-9), fibroblast growth factor (FGF; e.g., FGF-2, FGF-8, FGF-18), transforming growthfactor (TGF; e.g., TGF- 1, TGF-p3), vascular endothelial growth factor (VEGF; e.g., VEGF-A), nerve growth factor, brain- derived neurotrophic factor, insulin-like growth factors (e.g., IGF-1), or the like.

[0022] In various examples, treatment of the musculoskeletal condition of the subject 102 involves the repair of more than one tissue type. For instance, following a traumatic injury, the subject 102 may benefit from regrowth of blood vessels, muscle, and epithelial tissue. In some examples, the subject 102 may benefit from bone and joint formation following an amputation. In some examples, the subject 102 may benefit from regrowth of muscle and epithelial tissue to improve usability (e.g., comfort) of a prosthetic device. However, the complex repair of multiple tissue types conventionally involves repeated procedures to apply various therapeutic agents to particular locations in the affected area 105 in order to facilitate the growth of each tissue type. In various cases, the care provider(s) 104 may generate a treatment plan that includes repeated procedures to replenish and / or change the therapeutic agents administered to the subject 102. Repeated procedures increase the risk of harm to the health of the subject 102 (e.g., due to infections, complications, physical pain, stress, etc.). Furthermore, the subject 102 may have to repeatedly visit the healthcare facility, which can cause a financial burden (e.g., lost wages, transportation costs, treatment-related costs, etc.) on the subject 102.

[0023] These issues can be addressed, in various examples, by using therapeutic agent(s) formulated as a controlled- release system that can be applied to the affected area 105 with a single procedure. In various implementations of the present disclosure, a therapeutic agent can be translocated within the body of the subject 102, thereby improving control over the spatial delivery of individual therapeutic agents to targeted regions within the body. Accordingly, implementations of the present disclosure can simplify the delivery of multiple therapeutic agents for complex tissue repair in the treatment of musculoskeletal conditions.

[0024] In various examples, the care provider(s) 104 identify a first therapeutic agent 106 and a second therapeutic agent 108 to treat the musculoskeletal condition of the subject 102. In some examples, the care provider(s) 104 analyze an image (e.g., a CT scan) of the affected area 105 and / or the contralateral area. In some examples, the care provider(s) 104 perform a physical exam of the subject 102. Based on analyzing the image and / or the physical exam of the subject 102, the care provider(s) identify distinct physical locations for the delivery of the first therapeutic agent 106 and of the second therapeutic agent 108. For instance, the care provider(s) 104 may prescribe the first therapeutic agent 106 to facilitate muscle growth and the second therapeutic agent 108 to facilitate bone growth in the subject 102. Accordingly, the subject 102 may benefit from the first therapeutic agent 106 being therapeutically active at a first physical location and the second therapeutic agent 108 being therapeutically active at a second physical location. In particular examples, the first therapeutic agent 106 includes BMP-9, and the second therapeutic agent 108 includes BMP-2.

[0025] In some examples, the first therapeutic agent 106 may be formulated within magnetic microgels 110 that further include metal nanoparticles 112. The second therapeutic agent 108 may be formulated as non-magnetic microgels 114. In various examples, the magnetic microgels 110 and the non-magnetic microgels 114 include one or more polymers. For instance, the magnetic microgels 110 and the non-magnetic microgels 114 may include one or more hydrogels made of the polymer(s). In various instances, the polymer(s) may include polyethylene glycol) (PEG), poly(vinyl alcohol) (PVA),poly(lactic-co-glycolic acid) (PLGA), alginate, collagen, fibrin, chitosan, cellulose, or a combination thereof. In some cases, the magnetic microgels 110 and the non-magnetic microgels 114 are biocompatible and / or biodegradable. In some examples, the magnetic microgels 110 and the non-magnetic microgels 114 include a crosslinker configured to form a three-dimensional network of the polymer. Examples of crosslinking chemistries include thiol-based reactions, amine-based reactions, photopolymerization, enzymatic crosslinking, or the like. In various cases, the magnetic microgels 110 and the non-magnetic microgels 114 include a crosslinker, such as PEG-dithiol or a protease-sensitive crosslinker (e.g., GCRDVPMSMRGGDRCG (VPM; SEQ ID NO: 2), GPQGIWGQK (GPQ-W; SEQ ID NO: 3), GCRDGPQGIAGQDRCG (GPQ-A; SEQ ID NO: 4), or the like). In various cases, the magnetic microgels 110 and the non-magnetic microgels 114 include a photoinitiator, such as lithium phenyl(2,4,6-trimethylbenzoyl)phosphinate (LAP), ruthenium and sodium persulfate (SPS), or the like. For instance, upon receiving a photon with a particular energy level (e.g., a photon in visible or ultraviolet (UV) spectra), the photoinitiator may break down into a reactive species (e.g., including a free radical) that reacts with nearby cross-linkers, monomers, oligomers, polymers, or a combination thereof. In various examples, the photoinitiator is configured to activate in response to photons (e.g., light) having a particular wavelength. The photoinitiator may be configured to mediate the crosslinking of the polymer (e.g., cause the crosslinking in response to being activated). In some cases, the precursors of the magnetic microgels 110 and the non-magnetic microgels 114 include an enzyme that is configured to mediate the crosslinking of the polymer.

[0026] In some examples, the magnetic microgels 110 and / or the non-magnetic microgels 114 include a degradable component. For instance, the magnetic microgels 110 may include a degradable crosslinker. The degradable component may be configured to dissolve in the presence of an enzyme (e.g., a protease), a reducing agent, an oxidizing agent, or water. Enzyme-sensitive degradable crosslinkers degrade in the presence of, for instance, matrix metalloproteinase (MMP), collagenase, elastase, or trypsin. In some examples, the degradable component is configured to break down in response to a change in pH (e.g., when the pH reaches a particular threshold). In particular examples, the degradable component include a peptide that includes the sequence GCRDVPMSMRGGDRCG (SEQ ID NO: 1). In various examples, the magnetic microgels 110 and / or the non-magnetic microgels 114 may include different types of and / or different concentrations of the degradable component. Accordingly, the release profile of the first therapeutic agent 106 from the magnetic microgels 110 may be different that the release profile of the second therapeutic agent 108 from the non-magnetic microgels 114.

[0027] The metal nanoparticles 112 may include a ferromagnetic material. For instance, the metal nanoparticles 112 may include iron nanoparticles, iron oxide nanoparticles, cobalt nanoparticles, nickel nanoparticles, or alloy nanoparticles. In some examples, the metal nanoparticles 112 include magnetic beads. The metal nanoparticles 112, in some cases, are spherical, rod-shaped, cubic, triangular, star-shaped, dendritic (e.g., branched), or amorphous. In some cases, the metal nanoparticles 112 are solid, porous, hollow, or have a core-shell structure. The metal nanoparticles 112, in various examples, have a diameter in a range of 1 nanometer (nm) to 100 nm. The metal nanoparticles 112, in various examples, have a diameter in a range of 10 nm to 80 nm. In some instances, the metal nanoparticles 112 are disposed within the magnetic microgels 110. In some instances, the metal nanoparticles 112 are disposed on a surface of themagnetic microgels 110. For instance, the metal nanoparticles 112 may be conjugated to the surface of the magnetic microgels 110 by streptavidin-biotin bonds, avidin-biotin bonds, or the like.

[0028] The magnetic microgels 110, in various cases, have a diameter in a range of 100 nm to 500 microns (pm). In various cases, the diameter of the magnetic microgels 110 is in a range of 30 pm to 150 pm. In various cases, the diameter of the magnetic microgels 110 is in a range of 35 pm to 65 pm. In some examples, the diameter of the nonmagnetic microgels 114 is in a range of 100 nm to 500 pm. In some examples, the diameter of the non-magnetic microgels 114 is in a range of 30 pm to 150 pm. In some examples, the diameter of the non-magnetic microgels 114 is in a range of 35 pm to 65 pm. In various examples, the circularity of the magnetic microgels 110 is in a range of 0.6 to 1. In various examples, the circularity of the magnetic microgels 110 is in a range of 0.8 to 1. In various examples, the circularity of the non-magnetic microgels 114 is in a range of 0.6 to 1. In various examples, the circularity of the nonmagnetic microgels 114 is in a range of 0.8 to 1. In various examples, the opacity to visible light of the magnetic microgels 110 is in a range of 150 arbitrary units (AU) to 255 AU. In various examples, the opacity to visible light of the magnetic microgels 110 is in a range of 200 AU to 250 AU. In various examples, the opacity to visible light of the nonmagnetic microgels 114 is in a range of 0 AU to 5 AU. According to some implementations, the opacity to visible light may correspond to the 8-bit grayscale intensity scale. For instance, an image of the microgels may be obtained and converted to the 8-bit grayscale intensity scale, in which each pixel of the image is assigned a numerical value from 0 AU to 255 AU, in order to determine the opacity to visible light of the microgels.

[0029] The care provider(s) 104 may administer a therapeutic formulation that includes the magnetic microgels 110 and the non-magnetic microgels 114 to the subject 102. In some examples, the care provider(s) 104 may administer, to the subject 102, a first therapeutic formulation that includes the magnetic microgels 110 and a second therapeutic formulation that includes the non-magnetic microgels 114. The first therapeutic formulation and the second therapeutic formulation may be administered to the subject 102 simultaneously or at different time points. The first therapeutic formulation and the second therapeutic formulation may be administered at the same physical location on or within the body of the subject 102. Administration to the same physical location, as used herein, refers to administering two or more compositions to a particular physical position or within a threshold distance of the particular physical position (e.g., within 2 millimeter (mm), within 5 mm, within 10 mm, within 50 mm, or the like). In some cases, the first therapeutic formulation and the second therapeutic formulation may be delivered to distinct physical locations on or within the body of the subject 102. In various cases, the therapeutic formulation(s) are administered by injection. Routes of administration can include subcutaneous and / or intradermal administration. In some cases, routes of administration include intramuscular, intralesional, intranodal, intravesicular, intrathecal, intraperitoneal, or intranasal administration.

[0030] After administering the magnetic microgels 110 and the non-magnetic microgels 114 to the subject 102, the care provider(s) 104 may apply a magnet 116 to the affected area 105 of the subject 102. The magnet 116 may include a permanent magnet and / or an electromagnet. In various cases, the care provider(s) 104 use the magnet 116 to cause the magnetic microgels 110 to move within the body of the subject 102 in response to the magnetic field generated by the magnet 116. In some examples, the magnet 116 generates a magnetic field that exerts a magnetic force of the metalnanoparticles 112, thereby causing the rotation and / or translocation of the metal nanoparticles 112 relative to the magnet 116. For instance, the magnet 116 may generate a magnetic field between a north pole and a south pole of the magnet 116, such that the metal nanoparticles 112 are pulled toward one pole of the magnet 116. In various cases, applying the north pole of the magnet 116 on or near the skin of the subject 102 may cause iron nanoparticles 112 in the magnetic microgels 110 to move towards the magnet 116. In other cases, applying the south pole of the magnet 116 on or near the skin of the subject 102 may cause iron nanoparticles 112 in the magnetic microgels 110 to move away from the magnet 116. Accordingly, the magnet 116 can be used to disperse the magnetic microgels 110 away from the administration site (e.g., where the magnetic microgels 110 were injected into the subject 102). In some examples, the care provider(s) 104 may use an imaging technique or an image (e.g., an x-ray) of the subject 102 in order to visualize and improve the translocation of magnetic microgels 110, using the magnet 116, towards a particular location within the affected area 105. For instance, the care provider(s) 104 may use the magnet 116 to move the magnetic microgels 110 toward a distal end of an amputated bone of the subject 102.

[0031] In some examples, the subject 102 may apply the magnet 116 to the affected area 105. For instance, the magnet 116 may be integrated into a wearable device, such as a glove, a cast, a bandage, a brace, a sleeve, a belt, an article of clothing, a helmet, or the like. In some examples, the subject 102 may place a device that includes the magnet 116 on or around the affected area 105. In various cases, the subject 102 may apply the magnet 116 to the affected area 105 for a particular duration and / or at a particular interval. For instance, the subject 102 may apply the magnet 116 to the affected area 105 for 10 minutes, 20 minutes, 30 minutes, 60 minutes, 90 minutes, 2 hours, 3 hours, 6 hours, 9 hours, 12 hours, or more than 12 hours. In various cases, the subject 102 applies the magnet 116 to the affected area 105 every hour, every 3 hours, every 6 hours, every 12 hours, every day, every 2 days, every 3 days, every 4 days, every 5 days, every 6 days, every week, every 2 weeks, every month, or the like.

[0032] In various examples, the molecular weight of the polymer precursor! 22 and the binding affinity of the crosslinker, among other factors, can affect the density of the magnetic microgels 110 and the non-magnetic microgels 114 and the temporal release profile of the first and second therapeutic agents 106 and 108. For instance, the magnetic microgels 110 may be formulated such that the magnetic microgels 110 deform (e.g., compress) when they are translocated within the body of the subject 102 using the magnet 116. In various examples, the shear stress applied to the magnetic microgels 110 as they contact tissues within the body of the subject 102 may cause degradation of the magnetic microgels 110 and release of the first therapeutic agent 106 from the magnetic microgels 110.

[0033] While FIG. 1A illustrates the administration of the magnetic microgels 110 and the non-magnetic microgels 114 to the subject 102, implementations of the present disclosure are not so limited. In various examples, the care provider(s) 104 may administer the magnetic microgels 110 to the subject 102 and use the magnet 116 to guide the movement of the magnetic microgels 110 over time. For instance, the magnetic microgels 110 may be translocated within the body of the subject 102 in order to facilitate tissue growth in a particular direction. In some examples, the care provider(s) 104 may administer a first subset of magnetic microgels 110 that have a first concentration of metal nanoparticles 112 and a second subset of the magnetic microgels 110 that have a second concentration of the metal nanoparticles 112. Inresponse to the application of the magnet 116, the first subset of the magnetic microgels 110 may travel further towards the magnet 116 than the second subset of the magnetic microgels 110 due to the first concentration of the metal nanoparticles 112 being greater than than the second concentration of the metal nanoparticles 112.

[0034] In some cases, the care provider(s) 104 may administer a protease to the subject 102 in order to cause the degradation of a protease-sensitive degradable component in the magnetic microgels 110 and / or the non-magnetic microgels 114. For instance, the care provider(s) 104 may administer MMP1 and / or MMP2 to the subject 102 in order to facilitate release of, for instance, the first therapeutic agent 106 from the magnetic microgels 110.

[0035] In various examples, the care provider(s) 104 may perform an imaging procedure to visualize (e.g., using x-ray imaging, CT imaging, or the like) the magnetic microgels 110 within the body of the subject 102. In some cases, the care provider(s) 104 may periodically image the subject 102 in order to ensure that the magnetic microgels 110 are appropriately distributed within the body of the subject 102. Accordingly, implementations of the present disclosure can help the care provider(s) 104 confirm delivery of the first therapeutic agent 106 to the affected area 105.

[0036] While FIG. 1A illustrates the use of the magnetic microgels 110 for therapeutic purposes, implementations of the present disclosure are not so limited. In various examples, the magnetic microgels 110 can be used for diagnosis and / or research purposes. In various examples, the magnetic microgels 110 may be administered to the subject 102 in order to visualize the distribution of the magnetic microgels 110 within the subject 102. For instance, the magnetic microgels 110 may be conjugated to a targeting molecule, and by visualizing the magnetic microgels 110, it can be determined whether the targeting molecule specifically binds to a particular cell type that expresses a ligand for the targeting molecule (e.g., a tumor cell). In various cases, the magnetic microgels 110 and / or the non-magnetic microgels 114 include a fluorescent particle and / or a fluorescent tag. In some examples, the magnetic microgels 110 and / or the non-magnetic microgels 114 include or a cell-adhesive peptide. Accordingly, the magnetic microgels 110 can be bound to stem cells, immune cells, modified cells, or other cells used for the treatment of the musculoskeletal condition of the subject 102.

[0037] In some implementations, the magnetic microgels 110 are administered with a pharmaceutically acceptable carrier. Exemplary pharmaceutically acceptable carriers and formulations are disclosed in Remington's Pharmaceutical Sciences, 18th Ed. Mack Printing Company, 1990. Moreover, formulations can be prepared to meet sterility, pyrogenicity, general safety and purity standards as required by United States FDA Office of Biological Standards and / or other relevant foreign regulatory agencies.

[0038] Exemplary generally used pharmaceutically acceptable carriers include any and all bulking agents or fillers, solvents or co-solvents, dispersion media, coatings, surfactants, antioxidants (e.g., ascorbic acid, methionine, vitamin E), preservatives, isotonic agents, absorption delaying agents, salts, stabilizers, buffering agents, chelating agents (e.g., EDTA), gels, binders, disintegration agents, and / or lubricants.

[0039] Exemplary buffering agents include citrate buffers, succinate buffers, tartrate buffers, fumarate buffers, gluconate buffers, oxalate buffers, lactate buffers, acetate buffers, phosphate buffers, histidine buffers and / or trimethylamine salts.

[0040] Exemplary preservatives include phenol, benzyl alcohol, meta-cresol, methyl paraben, propyl paraben, octadecyldimethylbenzyl ammonium chloride, benzalkonium halides, hexamethonium chloride, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol and 3-pentanol.

[0041] Exemplary isotonic agents include polyhydric sugar alcohols including trihydric or higher sugar alcohols, such as glycerin, erythritol, arabitol, xylitol, sorbitol or mannitol.

[0042] Exemplary stabilizers include organic sugars, polyhydric sugar alcohols, polyethylene glycol, sulfur-containing reducing agents, amino acids, low molecular weight polypeptides, proteins, immunoglobulins, hydrophilic polymers or polysaccharides.

[0043] In particular implementations, treatments described herein (e.g., the magnetic microgels 110, the non-magnetic microgels 114) can be formulated into a carrier in a therapeutically effective amount. As described herein, exemplary carriers include saline, buffered saline, physiological saline, water, Hanks' solution, Ringer's solution, Normosol-R (Abbott Labs), PLASMA-LYTE A® (Baxter Laboratories, Inc., Morton Grove, IL), and combinations thereof.

[0044] Compositions including the magnetic microgels 110 and / or other components (e.g., the non-magnetic microgels 114) described herein may be administered in a formulation that includes one or more carriers, stabilizers, anesthetics, preservatives, or any combinations thereof.

[0045] In particular implementations, carriers can be supplemented with human serum albumin (HSA) or other human serum components or fetal bovine serum. In particular implementations, a carrier for infusion includes buffered saline with 5% HSA or dextrose. Additional isotonic agents include polyhydric sugar alcohols including trihydric or higher sugar alcohols, such as glycerin, erythritol, arabitol, xylitol, sorbitol, or mannitol.

[0046] Carriers can include buffering agents, such as citrate buffers, succinate buffers, tartrate buffers, fumarate buffers, gluconate buffers, oxalate buffers, lactate buffers, acetate buffers, phosphate buffers, histidine buffers, and / or trimethylamine salts.

[0047] Stabilizers refer to a broad category of excipients which can range in function from a bulking agent to an additive which helps to prevent cell adherence to container walls. Typical stabilizers can include polyhydric sugar alcohols; amino acids, such as arginine, lysine, glycine, glutamine, asparagine, histidine, alanine, ornithine, L-leucine, 2-phenylalanine, glutamic acid, and threonine; organic sugars or sugar alcohols, such as lactose, trehalose, stachyose, mannitol, sorbitol, xylitol, ribitol, myoinisitol, galactitol, glycerol, and cyclitols, such as inositol; PEG; amino acid polymers; sulfur-containing reducing agents, such as urea, glutathione, thioctic acid, sodium thioglycolate, thioglycerol, alpha-monothioglycerol, and sodium thiosulfate; low molecular weight polypeptides (e.g., <10 residues); proteins such as HSA, bovine serum albumin, gelatin or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; monosaccharides such as xylose, mannose, fructose and glucose; disaccharides such as lactose, maltose and sucrose; trisaccharides such as raffinose, and polysaccharides such as dextran.

[0048] Where beneficial, formulations can include a local anesthetic such as lidocaine to ease pain at a site of injection.

[0049] Exemplary preservatives include phenol, benzyl alcohol, meta-cresol, methyl paraben, propyl paraben, octadecyldimethylbenzyl ammonium chloride, benzalkonium halides, hexamethonium chloride, alkyl parabens such as methyl or propyl paraben, catechol, resorcinol, cyclohexanol, and 3-pentanol.

[0050] FIG. 1 B illustrates an example environment 118 for the production of microgels 120. In various examples, magnetic and non-magnetic microgels 120 can be produced in the environment 118. In various implementations, the processes described with reference to FIG. 1 B are performed by one or more entities, such as a computing device, laboratory equipment (e.g., a vortexer), or a user (e.g., a trained user, a laboratory technician, a researcher, or the like). “Microgels," "microparticles,” and their equivalents, as used herein, refer to particles with dimensions (e.g., diameter, height, length, width, etc.) between 1 m and 999 pm. In various implementations, the microparticles described herein have a spherical shape, an ovoid shape, or an amorphous shape. In various examples, the microparticles described herein are solid or porous.

[0051] In various implementations, a polymer precursor 122, a crosslinker 124, and optionally, metal nanoparticles 126 are mixed together to form a microgel precursor 128. The polymer precursor 122, in various cases, includes a monomer, macromer, oligomer, or the like. In various instances, the polymer precursor 122 is biocompatible and / or biodegradable. The polymer precursor 122 may include ethylene glycol, vinyl alcohol, p-D-mannuronic acid, a-L-guluronic acid, an amino acid (e.g., glycine, proline, hydroxyproline), fibrinogen, glucosamine, D-glucose, or a combination thereof. In various cases, the polymer precursor 122 includes a functionalized monomer, a functionalized macromer, or the like. In some examples, the polymer precursor 122 includes ethylene glycol. The polymer precursor 122 may include monomers and / or macromers of ethylene glycol. In some cases, an ethylene glycol monomer may be modified by one or more functional groups. For instance, the polymer precursor 122 may include a ethylene glycol monomer functionalized with norbornene.

[0052] In some examples, the crosslinker 124 includes PEG-dithiol. The number average molecular weight (Mn) of the crosslinker 124, in various cases, is in a range of 2400 Da to 4400 Da. In some cases, the Mnof the crosslinker 124 is in a range of 2900 Da to 3900 Da. In some examples in which the microgels 120 are non-magnetic, the microgel precursor 128 does not include the metal nanoparticles 126. In the examples in which the microgels 120 are magnetic, the microgel precursor 128 includes the metal nanoparticles 126.

[0053] In some examples, the microgel precursor 128 includes a photoinitiator. For instance, the microgel precursor 128 may include LAP. In various examples, the molecular weight of the photoinitiator is in a range of 200 Da to 400 Da. In various examples, the molecular weight of the photoinitiator is in a range of 250 Da to 350 Da. In some examples, the microgel precursor 128 includes a surfactant. For instance, the surfactant may include a fluorinated oil. In some examples, the surfactant has a viscosity in a range of 600-1200 centistoke (cSt) at 40°C. In various cases, the microgel precursor 128 includes a buffer solution, such as phosphate-buffered saline (PBS).

[0054] In particular implementations, a batch of microgels includes 0.2% weight per weight of the batch (w / w) to 10% w / w of the metal nanoparticles 126. In particular implementations, a batch of microgels includes 0.5% w / w to 5% w / w of the metal nanoparticles 126. In particular implementations, a batch of microgels includes 4% w / w to 7% w / w of the polymer precursor 122. In particular implementations, a batch of microgels includes 5% w / w to 6% w / w of the polymerprecursor 122. In particular implementations, a batch of microgels includes 1 % w / w to 3% w / w of the crosslinker 124. In particular implementations, a batch of microgels includes 1.5% w / w to 2.5% w / w of the crosslinker 124. In particular implementations, a batch of microgels includes 0.6% w / w to 1.0% w / w of the photoinitiator. In particular implementations, a batch of microgels includes 0.7% w / w to 0.9% w / w of the photoinitiator. According to various examples, a batch of microgels may include a 2% to 10% crosslinked hydrogel. According to various examples, a batch of microgels may include a 4% to 6% crosslinked hydrogel. According to various examples, a batch of microgels may include a 5% crosslinked hydrogel. In some instances, the amount of the metal nanoparticles 126, the polymer precursor 122, the crosslinker 124, and the photoinitiator depend on the level of crosslinking in the hydrogel.

[0055] The microgel precursor 128, in various examples, is mixed with a volume of an oil 130. The oil 130 may include mineral oil, a synthetic oil, or a combination thereof. The oil-microgel precursor mixture is vortexed in order to form a water-in-oil emulsion. According to some implementations, the oil-microgel precursor mixture is vortexed for 1 to 20 seconds. In some examples, the oil-microgel precursor mixture is vortexed for 2 to 6 seconds. In various cases, the time duration of the vortexing depends on the volume of the oil-microgel precursor mixture and / or the ratio of the volume of the microgel precursor 128 to volume of the oil 130.

[0056] In various cases, the polymer precursor 122 is crosslinked by receiving photons (e.g., light) to the oil-microgel precursor, causing the activation of the photoinitiator to form the microgels 120. In various cases, the photons have a predetermined wavelength that corresponds to the photoinitiator in the microgel precursor 128. For instance, the photons may have a wavelength between 10 nm to 400 nm. In various cases, the photons may be delivered to the polymer precursor 122 for 20 seconds to 10 minutes. In various cases, the photons may be delivered to the polymer precursor 122 for 1 minute to 5 minutes. In various examples, the time duration of the emission of the photons depends on the volume of the oil-microgel precursor mixture and / or the ratio of the volume of the microgel precursor 128 to volume of the oil 130.

[0057] In some examples, the polymer precursor 122 forms a polymer in response to receiving the photons. In some examples, the polymer includes PEG. In some examples, the polymer includes PEG-norbornene. The molecular weight of the polymer, in various instances, is in a range of 20 kilodaltons (kDa) to 60 kDa. In various instances, the molecular weight of the polymer is in a range of 30 kDa to 50 kDa.

[0058] In response to the vortexing and the application of the photons, the microgel precursor 128 may form the microgels 120 that include the polymer. The microgels 120 may be isolated by separating the oil 130 from the microgels 120. For instance, the oil 130 may be removed from a mixture that includes the microgels 120. After the formation of the microgels 120, the microgels 120 may be washed using, for instance, a washing oil that includes a volume of the oil 130. In some cases in which the microgel precursor 128 includes a surfactant, the washing oil may include a surfactant remover. For instance, the washing oil may include 1 H, 1 H,2H,2H-perfluoro-1 -octanol or another suitable surfactant remover. In various examples, the microgels 120 may be washed repeatedly until the surfactant is removed from the microgels 120.

[0059] In various implementations of the present disclosure, the microgels 120 can be formed using microfluidic technology. For instance, an apparatus may include a first inlet configured to receive the microgel precursor 128 and a second inlet configured to receive the oil 130. The apparatus may include a nozzle or a pump configured to output droplets of the microgel precursor 128 into the oil 130. For instance, the pump may output a volume of the microgel precursor 128 followed by a volume of the oil 130 into a channel of the apparatus. In various examples, the channel has a non-linear orientation (e.g., the channel does not extend along a straight line). For instance, the channel may have a serpentine orientation. In some examples, the channel has a linear orientation. In various cases, a light source may be configured to output photons onto the channel of the apparatus in order to crosslink (e.g., polymerize) the polymer precursor 122 to form the microgels 120. The apparatus may include an outlet configured to output the microgels 120 and the oil 130.EXPERIMENTAL EXAMPLE 1

[0060] Regenerative capacity of human musculoskeletal tissues is limited. Instead of limb regrowth, a fibrotic scar forms after amputation (Quijano, et al. Tissue Eng Part B Rev. 2016;22(3):251 -62). This Example describes the development of magnetic microgels that can be translocated using a user-controlled magnetic field to instruct composite tissue regeneration (Mora-Boza, et al. Mater Sci Eng C Mater Biol Appl. 2021 ;120:111716). Previous studies show that the combined delivery of bone morphogenetic protein-2 (BMP-2) and BMP-9 to proximally amputated murine digits induces skeletal elongation and synovial joint regeneration, respectively (Yu, supra] Dawson, et al. Regeneration. 2017;4:140-50). To interrogate the potential for mammalian limb regeneration, the murine digit tip, which regenerates after distal, but not proximal, amputation, is often used (Lehoczky, supra Simkin, supra] Qu, supra] Yu, supra] Dawson, supra). Accordingly, this Example describes the use of stimuli-responsive, bioactive hydrogels to direct the sequential and spatial formation of bone followed by articular cartilage in the murine digit.

[0061] Methods.

[0062] 8-arm 40 kDa Poly (ethylene glycol)-norbonene (PEG-NB, 5% w / v), PEG-dithiol (3 8 mM), Biotin-PEG-thiol (2 mM), and photoinitiator (Lithium phenyl-2,4,6-trimethylbenzoylphosphinate, (LAP) 2.5 mM) were dissolved in PBS (Control) (Lin, et al. J Appl Polym Sci. 2015; 132:41563). Microgel droplets were generated by agitating this mixture in a water-in-oil emulsion and photopolymerized by applying a field of light (A: 365 nm) (FIG. 2). To generate magnetically responsive microgels, either a low (23.3 millimolar (mM)) or high (221.4 mM) concentration of iron oxide (FeaOi) nanoparticles was incorporated in place of Biotin-PEG-thiol (Franco, et al. Acta Biomater. 2011;7(9):3267-76). Microgels in PBS were imaged with a bright field microscope and analyzed using Imaged, where the average diameter, circularity (4TT(area / perimeter2)), and opacity (0: transparent, 255: opaque) of individual microgels were quantified from binarized images. To test if microgels could respond to a magnetic field, Low FesO4 microgels were injected into a linear polydimethylsiloxane (PDMS) channel with a cylindrical neodymium magnet (5 / 8" thick, 9 / 16” diameter, 19.5 lb pull force, D9A; K&J Magnetics, Pipersville, PA) placed approximately 1 cm away. Velocity was determined by tracking the microgel centroid location over a 10-second time course. To test whether magnetic particle incorporation could be localized to the microgel surface, superparamagnetic beads coupled with streptavidin (Dynabead; Invitrogen, Waltham, MA; 1 :10) wereincubated with Control microgels and assessed by bright field microscopy. Significance was assessed by 2-tailed Student's t-test or 1-way ANOVA with Tukey’s post-hoc test (p<0.05).

[0063] Results.

[0064] The desired growth factor release profile and expected outcome of the magnetic bioactive microgel delivery are illustrated in FIGs. 3A and 3B. Magnetically responsive PEG-NB microgels were successfully fabricated by encapsulating Fe3O4 nanoparticles (FIGs. 4A and 4B). Microgel size, shape, and opacity depended on the Fe3O4 concentration (FIG. 4C). The average diameter and circularity of both low and high Fe3O4 microgels were significantly less than the control values. As expected, microgel opacity increased with FesCU concentration. High FesC microgels exhibited the greatest morphometric heterogeneity, indicating that higher nanoparticle concentrations may disrupt microgel droplet formation. In the presence of a magnetic field, low FesO4 microgels moved in a linear direction through PBS at approximately 0.2 millimeters per second (mm / s) (FIG. 4D). In contrast, microgel movement was negligible in the absence of a magnet. Similarly, control microgels with surface-conjugated Dynabeads were magnetically responsive (Wang, et al. Mol Ther Oncolytics. 2016;3: 16015).

[0065] Magnetic PEG-NB microgels represent a versatile biomaterial platform that may be adapted for spatially controlled drug delivery. This Example describes the development of a clinically translatable strategy to promote composite musculoskeletal tissue regeneration after digit amputation.EXPERIMENTAL EXAMPLE 2

[0066] This Example describes the generation of injectable micron-sized hydrogels (‘microgels') that can be translocated using an external magnetic field to spatially direct growth factor release.

[0067] Methods.

[0068] Synthesis: 8-arm 40 kDa Polyethylene glycolj-norbornene (PEG-NB, 5% w / v), PEG-dithiol (3.8 mM), and photoinitiator (Lithium phenyl-2,4,6-trimethylbenzoylphosphinate, 2.5 mM) were dissolved in PBS to create the polymer precursor. To generate magnetically responsive microgels, the precursor was mixed with either 0 (Control), 25 (Low), or 256 (High) pg / pL of iron oxide (Fe3O4) particles (Sigma-Aldrich, St. Louis, MO). Afterwards, the precursor was agitated in a water-in-oil emulsion to generate microgel droplets. To tune microgel size, two types of oil were tested: (1) mineral oil (Mineral) and (2) HFE-7500 3MTM NovecTM engineered fluid (Oakwood Chemical, Estill, SC) with 20% w / v KrytoxTM 157 FSH oil (Synthetic) (Garcia-Herreros, et al. BioRxiv. 2023) (FIGs. 5A, 5B). Photopolymerization was achieved by applying a field of light (A: 365 nm) for two minutes after agitation. Afterwards, microgels were washed and collected via centrifugation. Microgels in PBS were imaged with a bright field microscope and analyzed using Imaged The diameter, circularity (4rr(area / peri meter2)), and opacity (0: transparent, 255: opaque) of individual microgels were quantified from binarized images (n=70— 103 / group). To assess microgel translocation in response to an external magnetic field, microgels were injected into a PBS-filled polydimethylsiloxane (PDMS) channel bonded to a glass slide. Either a cylindrical neodymium magnet (19.5 lb pull force, K&J Magnetics) was placed approximately 5 cm from the injection site or no magnet was applied. Velocity was determined by tracking the microgel’s centroid location over time with video recordings (n=6— 8 / group). Significance was assessed by 2-way ANOVA with Tukey’s post-hoc test (p<0.05).

[0069] Results.

[0070] Magnetic microgels were successfully synthesized using water-in-oil emulsions (FIG. 6). FIG. 6 shows images of control, low FesCU, and high FesCh microgels made with mineral and synthetic oil at high speed for 3 seconds. Microgel size and shape were dependent on oil type, such that microgels synthesized with mineral oil exhibited a 4-fold increase in diameter and lower circularity values than microgels generated with synthetic oil (FIG. 7A, p<0.05). Incorporating high iron oxide content increased the diameter of microgels fabricated with mineral oil and reduced the circularity of microgels fabricated with synthetic oil (FIG. 7A, p<0.05). In this Example, opacity increased proportionally with the concentration of iron oxide particles (FIG. 7A, p<0.05). FIG. 7B illustrates a schematic of the experimental setup to evaluate microgel translocation. In the absence of a magnetic field, microgels from all groups remained stationary. However, upon exposure to an external magnetic field, only low and high iron oxide microgels exhibited directed movement towards the magnetic source (FIGs. 7C, 7D, p<0.05). The magnetic sensitivity of the microgels synthesized using mineral oil was, in this Example, directly proportional to the concentration of embedded iron oxide particles, such that microgels with high iron oxide content moved significantly faster than the low group (FIG. 7C, p<0.05). In contrast, low and high iron oxide microgels synthesized with synthetic oil displayed no significant difference in velocity (FIG. 7D, p>0.05).

[0071] The effects of vortex mixing speed and time on microgel size were investigated (FIG. 8), continuing with synthetic oil due to its ability to produce smaller microgels. Based on determining that high Fe304microgels reached saturation in iron oxide particle incorporation, the low Fe304group was optimized. Various vortex mixing speeds were investigated: low, medium, and high, corresponding to 1900 rotations per minute (RPM), 2550 RPM, and 3200 RPM, respectively. Diameter analysis revealed that medium mixing speed produced the smallest microgels, with both control and Fe3O4-loaded groups showing reduced diameters compared to low and high speeds.

[0072] For mixing time, control microgels decreased in size as time increased, whereas low Fe304microgels remained unchanged between 3 and 7 seconds. These findings indicate that both mixing speed and duration influence microgel size, supporting the use of synthetic oil for optimized fabrication.

[0073] Discussion.

[0074] This Example describes a controlled delivery system aimed for the spatiotemporal release of multiple growth factors. Both non-magnetic and magnetically sensitive microgels were synthesized, and it was demonstrated that their morphology can be modulated by selecting different oils for the water-in-oil emulsion (Garcia-Herreros, supra). Smaller microgels may facilitate more rapid protein release due to an increased surface area-to-volume ratio and translocate more readily through a dense extracellular matrix. Furthermore, it was found that magnetic sensitivity could be tuned by adjusting the iron oxide concentration. In various implementations, varying the polymer crosslinking density and incorporation of protease-degradable crosslinkers can enable control over growth factor release (Han, et al. Acta Biomater. 2019;173:495-508). This Example describes the development of a clinically translatable strategy to promote composite musculoskeletal tissue regeneration after digit amputation and other traumatic limb injuries.EXPERIMENTAL EXAMPLE 3

[0075] This Example describes the assessment of magnetic microgel movement within a 0.5 mg / mL collagen hydrogel under a magnetic field. This Example also describes the attachment of cells to the microgels using cell-adhesive peptides.

[0076] To assess magnetic microgel translocation in a three-dimensional (3D) biological environment, the microgels were embedded in a collagen hydrogel and a magnetic field was applied (FIG. 9A). Collagen without microgels was first injected into a silicone well and crosslinked at 37°C for 10 minutes Magnetic microgel-containing collagen was then layered on top and altogether crosslinked for another 40 minutes. Once fully set, a magnet was applied on the side of the collagen gel for 5 minutes.

[0077] Time-lapse imaging over 5 minutes showed microgel translocation, with microgels staying stationary without magnet and shifting from their initial positions (t = 0s) to new locations (t = 300s) in response to a magnet (FIG. 9B, FIG. 9C).

[0078] To assess whether cells could be attached to the microgels, HEK 293T cells were adhered to the microgels using the cell-adhesive peptide arginylglycylaspartic acid (RGD) (FIG. 10).EXAMPLE CLAUSES1. A composition, including: magnetic microgels that include: a hydrogel including at least one polymer; magnetic particles; and a therapeutic agent.2. The composition of clause 1 , wherein the magnetic microgels have a diameter in a range of about 100 nanometers (nm) to about 500 micrometers (pm).3. The composition of clause 1 or 2, wherein the magnetic microgels have a diameter in a range of about 30 pm to about 150 pm.4. The composition of any of clauses 1-3, wherein the at least one polymer includes at least one of polyethylene glycol) (PEG), norbornene, PEG-norbornene, PEG-dithiol, or biotin-PEG-thiol.5. The composition of any of clauses 1-4, wherein the magnetic particles include an iron oxide.6. The composition of any of clauses 1-5, wherein the therapeutic agent includes BMP-2 and / or BMP-9.7. The composition of any of clauses 1-6, wherein the magnetic microgels further include a photoinitiator and / or at least one of a fluorescent particle, a fluorescent tag, a protease-sensitive peptide, or a cell-adhesive peptide.8. The composition of clause 7, further including: cells bound to the cell-adhesive peptide.9. The composition of clause 7 or 8, wherein the protease-sensitive peptide includes a sequence having 95% sequence identify to SEQ ID NO: 1.10. The composition of any of clauses 7-9, wherein the protease-sensitive peptide includes SEQ ID NO: 1.11 . The composition of any of clauses 7-10, wherein the photoinitiator includes lithium phenyl-2, 4,6- trimethylbenzoylphosphinate.12. A formulation including the composition of any of clauses 1-11 and a pharmaceutically acceptable carrier.13. The formulation of clause 12, wherein the formulation is suitable for at least one of subcutaneous injection, intradermal injection, intramuscular injection, or intraperitoneal injection.14. A method of treatment, including: administering a therapeutically effective amount of the formulation of clause 12 or 13 to a subject; and applying a magnetic field to the subject, thereby moving the magnetic microgels within the subject.15. The method of clause 14, further including: in response to applying the magnetic field to the subject, administering at least one protease to the subject.16. The method of clause 15, wherein the at least one protease includes MMP1 and / or MMP2.17. The method of any of clauses 14-16, wherein applying the magnetic field to the subject includes moving the magnetic microgels toward a distal end of an amputated bone of the subject.18. A method of treating a digit amputation injury, the method including: administering, to a subject, a therapeutically effective amount of a first formulation including non-magnetic microgels, the non-magnetic microgels including a first therapeutic agent; administering, to the subject, a therapeutically effective amount of a second formulation including magnetic microgels, the magnetic microgels including a second therapeutic agent; and applying a magnetic field to the subject, thereby causing the magnetic microgels to move away from the non-magnetic microgels.19. The method of clause 18, wherein the first therapeutic agent includes BMP-2.20. The method of clause 18 or 19, wherein the second therapeutic agent includes BMP-9.21 . The method of any of clauses 18-20, wherein the first formulation and the second formulation are administered as a third formulation including the first formulation and the second formulation.22. The method of any of clauses 18-21 , wherein the first formulation and the second formulation are administered to the same physical location on or within the subject.23. The method of any of clauses 18-22, wherein applying the magnetic field to the subject causes the magnetic microgels to move toward a distal end of an amputated limb of the subject.24. The method of any of clauses 18-23, wherein a crosslinking density of the non-magnetic microgels is different than a crosslinking density of the magnetic microgels.25. The method of any of clauses 18-24, wherein an amount of a protease-degradable crosslinker in the nonmagnetic microgels is different than an amount of the protease-degradable crosslinker in the magnetic microgels.26. The method of any of clauses 18-25, wherein an average size of the non-magnetic microgels is different than an average size of the magnetic microgels.27. A method for generating magnetic microgels, the method including: generating a precursor solution including: at least one macromer; at least one crosslinker; a photoinitiator activated by light having a predetermined wavelength; and at least one therapeutic agent; generating a water-in-oil emulsion by mixing the precursor solution with an oil; and applying, to the water-in-oil emulsion, a light having the predetermined wavelength, thereby forming the magnetic microgels from the precursor solution.28. The method of clause 27, wherein the oil includes a synthetic oil and / or a mineral oil.29. The method of clause 27 or 28, further including: isolating the magnetic microgels by separating the oil from the magnetic microgels.30. The method of clause 29, wherein the precursor solution further includes a surfactant, the method further including: in response to isolating the magnetic microgels, washing the magnetic microgels, thereby removing the surfactant from the magnetic microgels.31. The method of clause 30, wherein the surfactant includes a fluorinated synthetic oil.32. The method of clause 30 or 31, wherein the magnetic microgels are washed with a washing oil including a surfactant remover.33. The method of clause 32, wherein the surfactant remover includes 1 H,1 H,2H,2H-perfluoro-1 -octanol.34. An apparatus for generating magnetic microgels, the apparatus including: a first inlet configured to receive a precursor solution including: at least one macromer; at least one crosslinker; a photoinitiator activated by light having a predetermined wavelength; and at least one therapeutic agent; a second inlet configured to receive an oil; a nozzle configured to output droplets of the precursor solution into the oil; a serpentine channel configured to move the droplets in the oil; and a light source configured to output the light having the predetermined wavelength onto the serpentine channel.35. The apparatus of clause 34, wherein the at least one macromer includes at least one of PEG, norbornene, PEG- norbornene, PEG-dithiol, or biotin-PEG-thiol, wherein the at least one crosslinker includes a protease-sensitive peptide, and wherein the at least one therapeutic agent includes BMP-2 and / or BMP-9.36. The apparatus of clause 34 or 35, wherein the precursor solution further includes magnetic particles, fluorescent particles, fluorescent tags, and / or a cell-adhesive peptide.37. The apparatus of any of clauses 34-36, wherein the apparatus includes a microfluidic device.38. A composition, including: magnetic microgels that include: a hydrogel including at least one cell-adhesive peptide; and magnetic particles; and cells bound to the at least one cell-adhesive peptide.39. A method of generating the composition of clause 38, including: incubating the cells with the magnetic microgels.40. A method of treatment including: administering a therapeutically effective amount of a formulation including the composition of clause 38 to a subject.

[0079] The features disclosed in the foregoing description, or the following claims, or the accompanying drawings, expressed in their specific forms or in terms of a means for performing the disclosed function, or a method or process for attaining the disclosed result, as appropriate, may, separately, or in any combination of such features, be used for realizing implementations of the disclosure in diverse forms thereof.

[0080] As will be understood by one of ordinary skill in the art, each implementation disclosed herein can comprise, consist essentially of or consist of its particular stated element, step, or component. Thus, the terms "include” or “including” should be interpreted to recite: “comprise, consist of, or consist essentially of.” The transition term “comprise” or “comprises" means has, but is not limited to, and allows for the inclusion of unspecified elements, steps, ingredients, or components, even in major amounts. The transitional phrase “consisting of” excludes any element, step, ingredient or component not specified. The transition phrase “consisting essentially of limits the scope of the implementation to thespecified elements, steps, ingredients or components and to those that do not materially affect the implementation. As used herein, the term “based on” is equivalent to “based at least partly on,” unless otherwise specified.

[0081] Unless otherwise indicated, all numbers expressing quantities, properties, conditions, and so forth used in the specification and claims are to be understood as being modified in all instances by the term “about.” Accordingly, unless indicated to the contrary, the numerical parameters set forth in the specification and attached claims are approximations that may vary depending upon the desired properties sought to be obtained by the present disclosure. At the very least, and not as an attempt to limit the application of the doctrine of equivalents to the scope of the claims, each numerical parameter should at least be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. When further clarity is required, the term “about” has the meaning reasonably ascribed to it by a person skilled in the art when used in conjunction with a stated numerical value or range, i.e. denoting somewhat more or somewhat less than the stated value or range, to within a range of ±20% of the stated value; ±19% of the stated value; ±18% of the stated value; ±17% of the stated value; ±16% of the stated value; ±15% of the stated value; ±14% of the stated value; ±13% of the stated value; ±12% of the stated value; ±11 % of the stated value; ±10% of the stated value; ±9% of the stated value; ±8% of the stated value; ±7% of the stated value; ±6% of the stated value; ±5% of the stated value; ±4% of the stated value; ±3% of the stated value; ±2% of the stated value; or ±1 % of the stated value.

[0082] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the disclosure are approximations, the numerical values set forth in the specific examples are reported as precisely as possible. Any numerical value, however, inherently contains certain errors necessarily resulting from the standard deviation found in their respective testing measurements.

[0083] The terms “a,” “an,” “the" and similar referents used in the context of describing implementations (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context. Recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g., “such as”) provided herein is intended merely to better illuminate implementations of the disclosure and does not pose a limitation on the scope of the disclosure. No language in the specification should be construed as indicating any non-claimed element essential to the practice of implementations of the disclosure.

[0084] Groupings of alternative elements or implementations disclosed herein are not to be construed as limitations. Each group member may be referred to and claimed individually or in any combination with other members of the group or other elements found herein. It is anticipated that one or more members of a group may be included in, or deleted from, a group for reasons of convenience and / or patentability. When any such inclusion or deletion occurs, the specification is deemed to contain the group as modified thus fulfilling the written description of all Markush groups used in the appended claims.

[0085] Variants of the sequences disclosed and referenced herein are also included. Guidance in determining which amino acid residues can be substituted, inserted, or deleted without abolishing biological activity can be found using computer programs well known in the art, such as DNASTAR™ (Madison, Wisconsin) software. Preferably, amino acid changes in the protein variants disclosed herein are conservative amino acid changes, i.e., substitutions of similarly charged or uncharged amino acids. A conservative amino acid change involves substitution of one of a family of amino acids which are related in their side chains.

[0086] In a peptide or protein, suitable conservative substitutions of amino acids are known to those of skill in this art and generally can be made without altering a biological activity of a resulting molecule. Those of skill in this art recognize that, in general, single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al. Molecular Biology of the Gene, 4th Edition, 1987, The Benjamin / Cummings Pub. Co., p. 224). Naturally occurring amino acids are generally divided into conservative substitution families as follows: Group 1 : Alanine (Ala), Glycine (Gly), Serine (Ser), and Threonine (Thr); Group 2: (acidic): Aspartic acid (Asp), and Glutamic acid (Glu); Group 3: (acidic; also classified as polar, negatively charged residues and their amides): Asparagine (Asn), Glutamine (Gin), Asp, and Glu; Group 4: Gin and Asn; Group 5: (basic; also classified as polar, positively charged residues): Arginine (Arg), Lysine (Lys), and Histidine (His); Group 6 (large aliphatic, nonpolar residues): Isoleucine (lie), Leucine (Leu), Methionine (Met), Valine (Vai) and Cysteine (Cys); Group 7 (uncharged polar): Tyrosine (Tyr), Gly, Asn, Gin, Cys, Ser, and Thr; Group 8 (large aromatic residues): Phenylalanine (Phe), Tryptophan (Trp), and Tyr; Group 9 (non-polar): Proline (Pro), Ala, Vai, Leu, lie, Phe, Met, and Trp; Group 11 (aliphatic): Gly, Ala, Vai, Leu, and lie; Group 10 (small aliphatic, nonpolar or slightly polar residues): Ala, Ser, Thr, Pro, and Gly; and Group 12 (sulfur-containing): Met and Cys. Additional information can be found in Creighton (1984) Proteins, W.H. Freeman and Company.

[0087] In making such changes, the hydropathic index of amino acids may be considered. The importance of the hydropathic amino acid index in conferring interactive biologic function on a protein is generally understood in the art (Kyte and Doolittle, 1982, J. Mol. Biol. 157(1), 105-32). Each amino acid has been assigned a hydropathic index on the basis of its hydrophobicity and charge characteristics (Kyte and Doolittle, 1982). These values are: lie (+4.5); Vai (+4.2); Leu (+3.8); Phe (+2.8); Cys (+2.5); Met (+1.9); Ala (+1.8); Gly (-0.4); Thr (-0.7); Ser (-0.8); Trp (-0.9); Tyr (-1.3); Pro (-1.6); His (-3.2); Glutamate (-3.5); Gin (-3.5); aspartate (-3.5); Asn (-3.5); Lys (-3.9); and Arg (-4.5).

[0088] It is known in the art that certain amino acids may be substituted by other amino acids having a similar hydropathic index or score and still result in a protein with similar biological activity, i.e., still obtain a biological functionally equivalent protein. In making such changes, the substitution of amino acids whose hydropathic indices are within ±2 is preferred, those within ±1 are particularly preferred, and those within ±0.5 are even more particularly preferred. It is also understood in the art that the substitution of like amino acids can be made effectively on the basis of hydrophilicity.

[0089] As detailed in US 4,554,101 , the following hydrophilicity values have been assigned to amino acid residues: Arg (+3.0); Lys (+3.0); aspartate (+3.0±1); glutamate (+3.0±1); Ser (+0.3); Asn (+0.2); Gin (+O.2); Gly (0); Thr (-0.4); Pro (-0.5±1 ); Ala (-0.5); His (-0.5); Cys (-1.0); Met (-1.3); Vai (-1.5); Leu (-1.8); lie (-1.8); Tyr (-2.3); Phe (-2.5); Trp(-3.4). It is understood that an amino acid can be substituted for another having a similar hydrophilicity value and still obtain a biologically equivalent, and in particular, an immunologically equivalent protein. In such changes, the substitution of amino acids whose hydrophilicity values are within ±2 is preferred, those within ±1 are particularly preferred, and those within ±0.5 are even more particularly preferred.

[0090] As outlined above, amino acid substitutions may be based on the relative similarity of the amino acid side-chain substituents, for example, their hydrophobicity, hydrophilicity, charge, size, and the like. As indicated elsewhere, variants of gene sequences can include codon optimized variants, sequence polymorphisms, splice variants, and / or mutations that do not affect the function of an encoded product to a statistically-significant degree.

[0091] Variants of the protein, nucleic acid, and gene sequences disclosed herein also include sequences with at least 70% sequence identity, 80% sequence identity, 85% sequence, 90% sequence identity, 95% sequence identity, 96% sequence identity, 97% sequence identity, 98% sequence identity, or 99% sequence identity to the protein, nucleic acid, or gene sequences disclosed herein.

[0092] “% sequence identity” refers to a relationship between two or more sequences, as determined by comparing the sequences. In the art, "identity" also means the degree of sequence relatedness between protein, nucleic acid, or gene sequences as determined by the match between strings of such sequences. "Identity" (often referred to as "similarity") can be readily calculated by known methods, including those described in: Computational Molecular Biology (Lesk, A. M., ed.) Oxford University Press, NY (1988); Biocomputing: Informatics and Genome Projects (Smith, D. W., ed.) Academic Press, NY (1994); Computer Analysis of Sequence Data, Part I (Griffin, A. M., and Griffin, H. G., eds.) Humana Press, NJ (1994); Sequence Analysis in Molecular Biology (Von Heijne, G., ed.) Academic Press (1987); and Sequence Analysis Primer (Gribskov, M. and Devereux, J., eds.) Oxford University Press, NY (1992). Preferred methods to determine identity are designed to give the best match between the sequences tested. Methods to determine identity and similarity are codified in publicly available computer programs. Sequence alignments and percent identity calculations may be performed using the Megalign program of the LASERGENE bioinformatics computing suite (DNASTAR, Inc., Madison, Wisconsin). Multiple alignment of the sequences can also be performed using the Clustal method of alignment (Higgins and Sharp CABIOS, 5, 151-153 (1989) with default parameters (GAP PENALTY=10, GAP LENGTH PENALTY=10). Relevant programs also include the GCG suite of programs (Wisconsin Package Version 9.0, Genetics Computer Group (GCG), Madison, Wisconsin); BLASTP, BLASTN, BLASTX (Altschul, et al., J. Mol. Biol. 215:403-410 (1990); DNASTAR (DNASTAR, Inc., Madison, Wisconsin); and the FASTA program incorporating the Smith-Waterman algorithm (Pearson, Comput. Methods Genome Res., [Proc. Int. Symp.] (1994), Meeting Date 1992, 111-20. Editor(s): Suhai, Sandor. Publisher: Plenum, New York, N.Y.. Within the context of this disclosure it will be understood that where sequence analysis software is used for analysis, the results of the analysis are based on the "default values" of the program referenced. As used herein "default values" will mean any set of values or parameters, which originally load with the software when first initialized.

[0093] Variants also include nucleic acid molecules that hybridizes under stringent hybridization conditions to a sequence disclosed herein and provide the same function as the reference sequence. Exemplary stringent hybridizationconditions include an overnight incubation at 42 °C in a solution including 50% formamide, 5XSSC (750 mM NaCI, 75 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5XDenhardt's solution, 10% dextran sulfate, and 20 g / ml denatured, sheared salmon sperm DNA, followed by washing the filters in 0.1XSSC at 50 °C. Changes in the stringency of hybridization and signal detection are primarily accomplished through the manipulation of formamide concentration (lower percentages of formamide result in lowered stringency); salt conditions, or temperature. For example, moderately high stringency conditions include an overnight incubation at 37°C in a solution including 6XSSPE (20XSSPE=3M NaCI; 0.2M NaH2PO4; 0.02M EDTA, pH 7.4), 0.5% SDS, 30% formamide, 100 g / ml salmon sperm blocking DNA; followed by washes at 50 °C with 1XSSPE, 0.1 % SDS. In addition, to achieve even lower stringency, washes performed following stringent hybridization can be done at higher salt concentrations (e.g. 5XSSC). Variations in the above conditions may be accomplished through the inclusion and / or substitution of alternate blocking reagents used to suppress background in hybridization experiments. Typical blocking reagents include Denhardt's reagent, BLOTTO, heparin, denatured salmon sperm DNA, and commercially available proprietary formulations. The inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility.

[0094] "Specifically binds" refers to an association of a binding domain (of, for example, a CAR binding domain or a nanoparticle selected cell targeting ligand) to its cognate binding molecule with an affinity or Ka(i.e., an equilibrium association constant of a particular binding interaction with units of 1 / M) equal to or greater than 106MA while not significantly associating with any other molecules or components in a relevant environment sample. “Specifically binds” is also referred to as “binds” herein. Binding domains may be classified as "high affinity" or "low affinity". In particular implementations, "high affinity" binding domains refer to those binding domains with a Ka of at least 107MA at least 108MA at least 109M-1, at least 1010M-1, at least 1011M-1, at least 1012M-1, or at least 1013MA In particular implementations, "low affinity" binding domains refer to those binding domains with a Kaof up to 107M-1, up to 106M-1, up to 105MA Alternatively, affinity may be defined as an equilibrium dissociation constant (Kd) of a particular binding interaction with units of M (e.g., 10-5M to 1CF13M). In certain implementations, a binding domain may have "enhanced affinity," which refers to a selected or engineered binding domains with stronger binding to a cognate binding molecule than a wild type (or parent) binding domain. For example, enhanced affinity may be due to a Ka(equilibrium association constant) for the cognate binding molecule that is higher than the reference binding domain or due to a Kd (dissociation constant) for the cognate binding molecule that is less than that of the reference binding domain, or due to an off-rate (KOff) for the cognate binding molecule that is less than that of the reference binding domain. A variety of assays are known for detecting binding domains that specifically bind a particular cognate binding molecule as well as determining binding affinities, such as Western blot, ELISA, and BIACORE® analysis (see also, e.g., Scatchard, et al., 1949, Ann. N.Y. Acad. Sci. 51 :660; and US 5,283,173, US 5,468,614, or the equivalent).

[0095] Unless otherwise indicated, the practice of the present disclosure can employ conventional techniques of immunology, molecular biology, microbiology, cell biology and recombinant DNA. These methods are described in the following publications. See, e.g., Green and Sambrook, Molecular Cloning: A Laboratory Manual, 4nd Edition (2012); F. M. Ausubel, et al. eds., Current Protocols in Molecular Biology, (2003); the series Methods In Enzymology (AcademicPress, Inc.); Behlke, et al., Polymerase Chain Reaction: Theory and Technology (2019); Greenfield, ed. Antibodies, A Laboratory Manual, Second Edition (2014); and Capes-Davis and R. I. Freshney, eds. Freshney's Culture of Animal Cells 8th Edition (2021).

[0096] Certain implementations are described herein, including the best mode known to the inventors for carrying out implementations of the disclosure. Of course, variations on these described implementations will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects skilled artisans to employ such variations as appropriate, and the inventors intend for implementations to be practiced otherwise than specifically described herein. Accordingly, the scope of this disclosure includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the abovedescribed elements in all possible variations thereof is encompassed by implementations of the disclosure unless otherwise indicated herein or otherwise clearly contradicted by context.

Claims

CLAIMS1. A composition, comprising: magnetic microgels that comprise: a hydrogel comprising at least one polymer; magnetic particles; and a therapeutic agent.

2. The composition of claim 1 , wherein the magnetic microgels have a diameter in a range of about 100 nanometers (nm) to about 500 micrometers (pm).

3. The composition of claim 1 , wherein the magnetic microgels have a diameter in a range of about 30 pm to about 150 pm.

4. The composition of claim 1 , wherein the at least one polymer comprises at least one of poly (ethylene glycol) (PEG), norbornene, PEG-norbornene, PEG-dithiol, or biotin-PEG-thiol.

5. The composition of claim 1 , wherein the magnetic particles comprise an iron oxide.

6. The composition of claim 1 , wherein the therapeutic agent comprises BMP-2 and / or BMP-9.

7. The composition of claim 1 , wherein the magnetic microgels further comprise a photoinitiator and / or at least one of a fluorescent particle, a fluorescent tag, a protease-sensitive peptide, or a cell-adhesive peptide.

8. The composition of claim 7, further comprising: cells bound to the cell-adhesive peptide.

9. The composition of claim 7, wherein the protease-sensitive peptide comprises a sequence having 95% sequence identify to SEQ ID NO: 1.

10. The composition of claim 7, wherein the protease-sensitive peptide comprises SEQ ID NO: 1.

11. The composition of claim 7, wherein the photoinitiator comprises lithium phenyl -2, 4,6- trimethylbenzoylphosphinate.

12. A formulation comprising the composition of claim 1 and a pharmaceutically acceptable carrier.

13. The formulation of claim 12, wherein the formulation is suitable for at least one of subcutaneous injection, intradermal injection, intramuscular injection, or intraperitoneal injection.

14. A method of treatment, comprising: administering a therapeutically effective amount of the formulation of claim 12 to a subject; and applying a magnetic field to the subject, thereby moving the magnetic microgels within the subject.

15. The method of claim 14, further comprising: in response to applying the magnetic field to the subject, administering at least one protease to the subject.

16. The method of claim 15, wherein the at least one protease comprises MMP1 and / or MMP2.

17. The method of claim 14, wherein applying the magnetic field to the subject comprises moving the magnetic microgels toward a distal end of an amputated bone of the subject.

18. A method of treating a digit amputation injury, the method comprising:administering, to a subject, a therapeutically effective amount of a first formulation comprising non-magnetic microgels, the non-magnetic microgels comprising a first therapeutic agent; administering, to the subject, a therapeutically effective amount of a second formulation comprising magnetic microgels, the magnetic microgels comprising a second therapeutic agent; and applying a magnetic field to the subject, thereby causing the magnetic microgels to move away from the nonmagnetic microgels.

19. The method of claim 18, wherein the first therapeutic agent comprises BMP-2.

20. The method of claim 18, wherein the second therapeutic agent comprises BMP-9.

21. The method of claim 18, wherein the first formulation and the second formulation are administered as a third formulation comprising the first formulation and the second formulation.

22. The method of claim 18, wherein the first formulation and the second formulation are administered to the same physical location on or within the subject.

23. The method of claim 18, wherein applying the magnetic field to the subject causes the magnetic microgels to move toward a distal end of an amputated limb of the subject.

24. The method of claim 18, wherein a crosslinking density of the non-magnetic microgels is different than a crosslinking density of the magnetic microgels.

25. The method of claim 18, wherein an amount of a protease-degradable crosslinker in the non-magnetic microgels is different than an amount of the protease-degradable crosslinker in the magnetic microgels.

26. The method of claim 18, wherein an average size of the non-magnetic microgels is different than an average size of the magnetic microgels.

27. A method for generating magnetic microgels, the method comprising: generating a precursor solution comprising: at least one macromer; at least one crosslinker; a photoinitiator activated by light having a predetermined wavelength; and at least one therapeutic agent; generating a water-in-oil emulsion by mixing the precursor solution with an oil; and applying, to the water-in-oil emulsion, a light having the predetermined wavelength, thereby forming the magnetic microgels from the precursor solution.

28. The method of claim 27, wherein the oil comprises a synthetic oil and / or a mineral oil.

29. The method of claim 27, further comprising: isolating the magnetic microgels by separating the oil from the magnetic microgels.

30. The method of claim 29, wherein the precursor solution further comprises a surfactant, the method further comprising:in response to isolating the magnetic microgels, washing the magnetic microgels, thereby removing the surfactant from the magnetic microgels.31 . The method of claim 30, wherein the surfactant comprises a fluorinated synthetic oil.

32. The method of claim 30, wherein the magnetic microgels are washed with a washing oil comprising a surfactant remover.

33. The method of claim 32, wherein the surfactant remover comprises 1 H,1 H,2H,2H-perfluoro-1-octanol.

34. An apparatus for generating magnetic microgels, the apparatus comprising: a first inlet configured to receive a precursor solution comprising: at least one macromer; at least one crosslinker; a photoinitiator activated by light having a predetermined wavelength; and at least one therapeutic agent; a second inlet configured to receive an oil; a nozzle configured to output droplets of the precursor solution into the oil; a serpentine channel configured to move the droplets in the oil; and a light source configured to output the light having the predetermined wavelength onto the serpentine channel.

35. The apparatus of claim 34, wherein the at least one macromer comprises at least one of PEG, norbornene, PEG-norbornene, PEG-dithiol, or biotin-PEG-thiol, wherein the at least one crosslinker comprises a protease-sensitive peptide, and wherein the at least one therapeutic agent comprises BMP-2 and / or BMP-9.

36. The apparatus of claim 34, wherein the precursor solution further comprises magnetic particles, fluorescent particles, fluorescent tags, and / or a cell-adhesive peptide.

37. The apparatus of claim 34, wherein the apparatus comprises a microfluidic device.

38. A composition, comprising: magnetic microgels that comprise: a hydrogel comprising at least one cell-adhesive peptide; and magnetic particles; and cells bound to the at least one cell-adhesive peptide.

39. A method of generating the composition of claim 38, comprising: incubating the cells with the magnetic microgels.

40. A method of treatment comprising: administering a therapeutically effective amount of a formulation comprising the composition of claim 38 to a subject.

Images