57 results about "Testicular Interstitial Cells" patented technology

The invention discloses a method and application for inducing human umbilical cord mesenchyme stem cells to be differentiated into testicular interstitial cells. The method comprises the following step of culturing human umbilical cord mesenchyme stem cells of patients suffering from adenovirus and carrying mice steroidogenic factor-1 genes in a DMEM-F12 culture solution containing 0.3-3ng / ml of luteinizing hormone, 200-800mu M of dibutyryl cyclic adenosine monophosphate, 5*10<-6>-5*10<-4>M of all-trans retinoic acid (ATRA), 10mU / ml of human chorionic gonadotropin and 2.4uM of adrenocorticotrophic hormone for a week. Induced by the method in the invention, the human umbilical cord mesenchyme stem cells can be differentiated into testicular interstitial cells in vitro and provides important sources of cells for treating testosterone shortage by the cell replacing method or the genetic method.

A cell composition composed of spermatogonial stem cells, Sertoli cells, Leydig cells and optionally peritubular cells, is provided, as is a culture composition, artificial testicular construct, hydrogel composition, and device containing the same. A method for using the device as a physiologically relevant in vitro model of human testicular function to screen compounds for pharmacological or toxicological activity is also provided.

The invention provides a method for constructing a model of oriented differentiation of embryonic stem (ES) cells into testicular interstitial cells. The method comprises the following steps of: firstly, cotransfecting a steroidogenic factor 1 and a luteinizing hormone receptor plasmid into an ES cell; then inducing the ES cell by a compound or growth factor to directionally differentiate the ES cell into the testicular interstitial cell. In the invention, the construction and the application of the model for in-vitro oriented differentiation of the stem cells into the testicular interstitialcells are demonstrated at the cellular level; a great deal of biology information is discussed initially; the constructed cell model is a substituted model for researching the influencing factor of the secretory regulation function of the testicular interstitial cell and can be used for initially selecting and evaluating the medical effect of the pharmaceutical drug using the testicular interstitial cell as the target cells; the transplanting of the testicular interstitial cell differentiated from the stem cells is used as a new method for supplying testosterone, and the application prospect of the renewable alternative treatment of the testicular interstitial cell is improved.

The invention provides an external directional induced differentiation method for differentiating human induced pluripotent stem (iPS) cells into leydig cells (LCs) through neural crest stem cells (NCSCs). By the utilization of an animal model, it is verified that the LCs from the iPS cells have the capacity of regenerating aged or damaged LCs, and a new testosterone-supplying scheme is provided for a patient with hypogonadism and particularly an LOH patient.

The invention relates to a method for inducing transdifferentiation of fibroblast into similar testicular interstitial cells. The method is characterized in that required transcription factors are respectively cloned to lentiviral vectors, and packaged to be virus particles for transfection to fibroblast; by utilizing an overexpression transcription factor method, inducing transdifferentiation of the fibroblast into inducible testicular interstitial cells, wherein the transcription factors are sourced from human beings, mice or rats, and the transcription factors are a part selected from a group of Dmrt1, Nr0b1, Sp1, Wt1, Nr4a1, Nr5a2, AP1, Creb1, Smad3, Nr5a1 and Gata4,or a combination of all transcription factors. The cells after transdifferentiation have the androgen synthesis related gene expression profile similar to that of mature testicular interstitial cells, and can synthesize and secrete testosterone. The similar testicular interstitial cells prepared with the method can be used for treating male hypogonadism syndrome.

The invention discloses a method for induced differentiation of testicular interstitial cells and applications of the testicular interstitial cells in sexual function recovery, belonging to the technical field of biological medicines. Mesenchymal stem cells are utilized for induced differentiation of testicular interstitial cells, the testicular interstitial cells and other ingredients with a certain ratio form a medicine for sexual function recovery, the medicine for sexual function recovery is prepared into an injection, the injection is injected to the focus area in the injection form, thetestosterone level in the body is recovered in a manner that the testicular interstitial cells secrete testosterone, and thus the sexual function of the male is effectively recovered. In short, the method is safe and effective, and is strong in practicability, testicular interstitial cells are efficiently obtained through induced differentiation, and the medicine can effectively recover the sexualfunction of the male.

The invention relates to use of a nerve growth factor in preparation of a medicine for treating sexual debility syndrome of medium-elderly men. The nerve growth factor is mainly used for preparing a medicine for treating testosterone content decline of human body caused by decrease of secretion of interstitial cell-stimulating hormone increased by hypophysis or aging of testicular interstitial cells. The nerve growth factor used is originated from humans, mice and rats, preferably humans. The amino acid sequences of the nerve growth factor are shown in SEQ ID No:1-3.

This invention pertains to the discovery that stem cells (e.g., bone marrow stem cells) transplanted directly into a testicular environment are transdifferentiated into bona fide Sertoli cells, and / or Leydig cells, and / or and germ cells. This provides a mechanism for the treatment of male infertility and / or testosterone deficiency. Thus, in one embodiment, this invention provides a method of treating infertility or testosterone deficiency in a male mammal. The method typically involves implanting stem cells into the testes of the mammal whereby the stem cells differentiate into germ cells and / or Sertoli cells and / or Leydig cells thereby reducing infertility and / or testosterone deficiency.

The invention discloses a method for differentiating inducible pluripotent stem cells into testicular interstitial cells by small molecule induction. The method comprises the following steps: firstly,obtaining human inducible pluripotent stem cells; secondly, carrying out pretreatment culture on the human inducible pluripotent stem cells by using an E7 culture medium without basic fibroblast growth factors; thirdly, inducing the human inducible pluripotent stem cells to differentiate by using a differential medium and combining with a small molecule inducer, wherein the small molecule inducercomprises a DHH agonist SAG, 22R-OHC, lithium chloride, human platelet-derived growth factor AA, fibroblast growth factor 2, insulin-like growth factor 1, androgen, luteinizing hormone, retinol and 8-bromo-cyclic adenosine monophosphate; fourthly, manually removing clonal cluster-like parenchyma cells; fifthly, continuously culturing the remaining cells after the parenchyma cells are removed by using an enriched medium, replacing the enrichment medium regularly, and finally obtaining the target testicular interstitial cells.

The invention discloses a method for inducing human umbilical cord mesenchymal stem cells into testicular interstitial cells and application thereof. The method comprises the following steps: cloning mouse steroidogenic factor-1 (SF-1) gene into an adenovirus shuttle plasmid pAdtrack-CMV-EGFP to construct a recombinant vector pAdtrack-CMV-SF-1; recombining the recombinant vector pAdtrack-CMV-SF-1 and an adenovirus carrier plasmid pAdEasy-1 to obtain an adenovirus plasmid pAd-SF-1 carrying SF-1 gene; digesting and linearizing the adenovirus plasmid pAd-SF-1, and packaging with human embryonic kidney cells AD-293 to obtain adenovirus; adding the adenovirus into an induction culture medium to infect human umbilical cord mesenchymal stem cells, and carrying induced differentiation of the human umbilical cord mesenchymal stem cells to testicular interstitial cells. Through the induction using the method of the invention, human umbilical cord mesenchymal stem cells can in vitro differentiate into testicular interstitial cells only in one week, so as to provide an important cell source for treatment of testosterone deficiency using cell replacement or genetic method.

The invention discloses a method for obtaining Leydig cells and purposes of the method. The method disclosed by the invention has the advantages of quickness, simpleness and convenience, and capability of obtaining a large quantity of Leydig cells with high purity and good functions, and is used for establishing seed cells of tissue engineered androgen-secreting tissue and other purposes.

The invention discloses a method for inducing human ADSC (Adipose-Derived Stem Cells) to be differentiated into LCs (Leydig Cells) by micromolecules. The method comprises the following steps: firstly,separating and extracting the human ADSC, and completing identification; then, inducing differentiation of the human ADSC by adopting a specific culture medium, and simultaneously adding a certain amount of micromolecules such as SAG, 22OHC, Li, PDGF-AA, FGF2, Androgen and LH in the special culture medium at different time points, thus promoting differentiation; finally, manually removing ALCs (Adult Leydig Cells), enriching and differentiating target cells ADSC-ALCs, and completing the identification. By inducing according to the method, the human ADSC can be successfully differentiated intothe LCs, so that a cell source is provided for treating testosterone deficiency by adopting cell transplantation.

The invention provides a method for inducing umbilical cord mesenchymal stem cells to be differentiated into leydig cells through gold nanoparticle medicated gene transfection. Plasmids with SF-1 (seroidogenic factor 1) genes and LHR (luteotropic hormone receptor) genes are cotransfected into the umbilical cord mesenchymal stem cells through gold nanoparticles, then the umbilical cord mesenchymal stem cells are induced by compounds or growth factors to be directionally differentiated into the leydig cells. The leydig cell differentiation method is efficient and safe, the differentiated leydig cells can serve as a research substitution model and can be used as seed cells of clinical cell treatment of the leydig cells, and an application prospect of leydig cell regeneration replacement treatment is widened.

The invention relates to the field of cell cultivation and in particular relates to a separation and purification method of testicular interstitial cells of a naked mole rat. The interstitial cells of the naked mole rat are separated and purified by adopting enzymatic digestion and a Percoll continuous density gradient method. The method provided by the invention can obtain a lot of testicular interstitial cells of the naked mole rat with normal functional activity simply, efficiently and economically, and cultivation under a low oxygen condition can ensure that the cells still can keep the biological characteristics in an in-vivo state in an in-vitro environment, so that the special physiological functions of the testicular interstitial cells of the naked mole rat can be conveniently and directly researched in a pure in-vitro cell cultivation model further, thereby providing an important theoretical basis for exploring the biological mechanism and applying the same to the clinical related fields.

The invention provides a preparation method and application of a eucommia ulmoides male flower extract with an anti-fatigue effect. The preparation method comprises the following steps of: taking a certain amount of eucommia ulmoides male flowers, extracting the eucommia ulmoides male flowers with water of which the mass is 15 times that of the eucommia ulmoides male flowers, refluxing for 2 hoursat 100 DEG C, filtering and taking filtrate for later use, adding water of which the mass is 15 times that of the eucommia ulmoides male flowers into the eucommia ulmoides male flowers, boiling and refluxing for the second time, filtering and collecting filtrate, mixing the filtrate obtained in the two times, concentrating and performing spray drying to obtain the eucommia ulmoides male flower extract. According to the preparation method, the eucommia ulmoides male flowers are extracted with water, filtered and concentrated to obtain the extract; and experiments prove that the eucommia ulmoides male flower extract and monomer components thereof can promote secretion of testosterone in testicular interstitial cells, and can be used for health-care food with the main anti-fatigue function.

The invention discloses a method for testing influence of midazolam on development of testicular interstitial cells cultured in vitro. Comprising the following steps: culturing rats, separating CD90 positive cells, culturing testicular mesenchymal stem cells (SLC), detecting the influence of midazolam on proliferation and differentiation, amplifying the SLCs and detecting the cell activity, carrying out DAPI dyeing and counting, analyzing cells by a flow cytometer, analyzing testosterone (T) and progesterone (P4), carrying out mRNA and protein expression detection through QPCR (Quantitative Polymerase Chain Reaction) and WB (White Brown) experiments, and determining significant differences and drawing a chart through SNK (Sodium Nitrogen Kinase) detection. The steps are simple, and the influence of midazolam medicine on proliferation and differentiation of the testicular mesenchymal stem cells can be effectively verified.

The invention discloses a method for constructing a machin progenitor leydig cell immortal line. The method comprises the following steps: by taking machin testicular tissues, obtaining leydig cells of a high ratio, culturing in a complete medium for 20-28 hours, replacing fresh culture solution, continuously culturing, and performing cell passage to the leydig cells presenting fibroblast growth in uniform form; infecting the leydig cells by utilizing slow virus solution carrying Large T and Egfp, performing fluorescence labeling cell screening, and performing cell passage to a cell strain for stably expressing Egfp; and screening the cell strain for stably expressing Egfp, identifying the progenitor leydig cells, continuously performing cell passage for over 50 generations, and finally constructing the machin progenitor leydig cell immortal line. The in-vitro cell passage of the progenitor leydig cell immortal line with the transferred Large T exceeds 62 generations, the leydig cell labeled proteins such as CYP11A1, CYP17A1 and STAR are expressed, marker genes of mesenchymal stem cells such as Pdgfral and Lifr are expressed, the cell shape is maintained into the fiber shape, and a characteristic progenitor leydig cell immortal line is obtained.

The invention discloses a method for testing that SLC can maintain homeostasis of Leydig and muscle-like cells in adult testis. Stem cells related to testicular seminiferous tubules of adult SD rats are called as Leydig stem cells (SLC) and can be differentiated into Leydig cells in vitro and in vivo. Proliferation and differentiation of the SLC are regulated by locally generated factors, including PDGF family members PDGF-AA and PDGF-BB. Both members are able to stimulate proliferation of SLC, while only PDGF-AA is able to stimulate differentiation of SLC into Leydig cells producing testosterone (T), while PDGF-BB inhibits the process. In the method, it is verified that the SLC can actually form Leydig cells or myocyte-like cells under different differentiation conditions. PDGF-AA + LH can enable the SLC to be differentiated into Leydig cells capable of generating T, and PDGF-BB + TGFB induces the cells to become muscle-like cells for expressing smooth muscle actin. The method can effectively verify pluripotency of seminiferous tubule-associated stem cells (SLC), and test important role of SLC in maintaining homeostasis of Leydig and Myoid populations in vivo.

A cell composition composed of spermatogonial stem cells, Sertoli cells, Leydig cells and optionally peritubular cells, is provided, as is a culture composition, artificial testicular construct, hydrogel composition, and device containing the same. A method for using the device as a physiologically relevant in vitro model of human testicular function to screen compounds for pharmacological or toxicological activity is also provided.

The invention relates to a method for inducing transdifferentiation of fibroblast into similar testicular interstitial cells. The method is characterized in that required transcription factors are respectively cloned to lentiviral vectors, and packaged to be virus particles for transfection to fibroblast; by utilizing an overexpression transcription factor method, inducing transdifferentiation of the fibroblast into inducible testicular interstitial cells, wherein the transcription factors are sourced from human beings, mice or rats, and the transcription factors are a part selected from a group of Dmrt1, Nr0b1, Sp1, Wt1, Nr4a1, Nr5a2, AP1, Creb1, Smad3, Nr5a1 and Gata4,or a combination of all transcription factors. The cells after transdifferentiation have the androgen synthesis related gene expression profile similar to that of mature testicular interstitial cells, and can synthesize and secrete testosterone. The similar testicular interstitial cells prepared with the method can be used for treating male hypogonadism syndrome.

The invention relates to a method for inducing adipose tissue-derived stem cells to differentiate into testicular interstitial-like cells by a single transcription factor, and belongs to the fields of stem cell biology and regenerative medicine. According to the method, chemically modified mRNA of a transcription factor NR5A1 is synthesized in vitro, exosomes extracted from autologous urine of a patient are used as a delivery carrier to form an exosome-modNR5A1 compound, and then the exosome-modNR5A1 compound and ADMSCs derived from autologous tissue of the patient are co-cultured to induce the ADMSCs to be differentiated into Leydig-like cells with a testosterone secretion function. According to the method disclosed by the invention, the Leydig-like cells with the testosterone secretion function can be quickly and efficiently obtained, meanwhile, the cells can effectively react to the stimulation of luteinizing hormone, and the cells have a gene expression profile related to the androgen synthesis of mature Leydig cells. The Leydig cell obtained by the invention has a good application prospect in the aspect of treatment of male hypogonadism.

The use of an aromatase blocker or an estrogen blocker is described in a method for increasing spermatogenesis and Seritolli cell function, and / or improving Leydig cell function, in order to to increase endogenous testosterone levels in a male mammal. The levels of active materials used are significantly lower than the levels of these materials used to treat female estrogen sensitive tumors.

The partial androgen deficiency of the aging male (PADAM) and the supplement of synthetic testosterone is an important research subject in the American biological science. The activity of leydigs cells of testis and the synthesizing and excreting state of testosterone has aroused wide concern of the medical circles. Taking synthetic testosterone may cause the cancer of genital system, which makes the middle-aged and old male people do not dare to taking the drugs of the synthetic testosterone. The Compound Recipe Product of 4 Kinds of Plants to Increase Testosterone is made by following procedures: Under the carefully and strictly controlled condition, use the technology of absorption and elution of WLD resin to extract and purify it and measured by the gas chromatograph and make a fingerprinting chromatograph quantitative determination so as to guarantee the product has a stable technological process and a high quality. The experiment of pharmacological activity shows that, by activating the leydigs cells of testis to enhance its ability to excrete testosterone, the compound recipe product has a remarkable function to obviously raise the quantity of testosterone blood serum of the normal and injured genital system and is used for treating the partial androgen deficiency of the aging male (PADAM) in clinical treatment.

The invention discloses a method for culturing high-purity primary Leydig cells of Guizhou fragrance pigs. The method is characterized by comprising the following steps: (1) digesting testicular tissue blocks of fragrance pigs through collagenase to obtain a cell suspension solution; (2) repeatedly digesting with 0.25 percent Trypsin-EDTA to obtain Leydig cells; (3) enriching the Leydig cells through a differential centrifugation method, wherein the differential centrifugation method at least comprises three times of centrifugation, the speed of the first-time centrifugation is 800 to 1200 rpmand the time is 3 to 8 min, and the speed of the second-time and third-time centrifugation is 600 to 1000 rpm and the time is 3 to 8 min; (4) purifying the Leydig cells through a differential attachment method. Compared with an existing primary culture and purification method of pig Leydig cells, the method has the advantages that the operation is simpler and more efficient, and the Leydig cellsof the Guizhou fragrance pigs, which have a great quantity, strong activity and high purity, are obtained. The method disclosed by the invention can provide effective technological guidance for deeplyresearching sexual precocity properties of the Guizhou fragrance pigs and related functions of the Leydig cells.

The invention belongs to the field of cell separation and purification and more specifically relates to a method for preparing poultry testicular interstitial cells. The method comprises the followingsteps: digesting a testicular tissue block by utilizing collagenase to obtain a cell suspension solution; centrifuging the cell suspension solution obtained by step a) by utilizing a differential centrifugation method and separating to obtain the testicular interstitial cells; culturing the testicular interstitial cells obtained by step b) by utilizing a differential attachment method, so as to enrich the testicular interstitial cells. According to the method for preparing the poultry testicular interstitial cells, provided by the invention, the testicular interstitial cells with great cell amount and high purity can be obtained; an operation process is simple and standard, damages to the cells are relatively small and cell pollution is effectively reduced.

The invention discloses a long-term culture method for swine testicle mesenchymal progenitor cells. The long-term culture method includes separation, detection and differentiation in vitro, and specifically includes the steps of culturing piglet testicle fibroblasts for 5-8 days by means of an in-vitro proliferation culture medium of swine PLCs (progenitor leydig cells) to acquire a PLCs clone colony, culturing by means of a differentiation medium of swine PLCs so that PLCs begin to differentiate into mature testicle mesenchymal cells in the third day, then culturing in the proliferation culture medium, and maintaining in vitro for six month at most. According to the long-term culture method, a mature process of separation, culture, detection and differentiation of swine PLCs is created initially, and proliferation and differentiation maintained in vitro of PLCs can be achieved and controlled by different in-vitro culture systems determined finally. The long-term culture method has high application value in research on biological property of PLCs in vitro, research and development of drugs and vaccine production and the like.

The invention discloses a biomarker of perimuscular cells of testicular tubes and application of the biomarker. The biomarker comprises an ITGA9 protein. On the basis of the ITGA9 protein, the kit for identifying the testicular perimuscular cells is prepared, the anti-ITGA9 protein antibody is used for specifically recognizing and combining the perimuscular cells, then the testicular perimuscular cells can be effectively identified through methods such as flow analysis and the like, in addition, after the testicular perimuscular cells are identified, the testicular perimuscular cells can be distinguished from Leydig cells, and therefore the identification accuracy of the testicular perimuscular cells is improved. Therefore, the two cells can be effectively separated, the problem that the two cells are difficult to separate by conventional stepwise digestion, differential adherence, gradient centrifugation and other methods is solved, and high-purity Leydig cells and high-purity peritubular muscle-like cells can be obtained through separation and purification.