Transcriptome characterization of the effects of exosomes from different sources on testicular tissue of male mice
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ABSTRACT:
PROVIDER: PRJNA1035425 | ENA |
REPOSITORIES: ENA
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Project description:Collect the blood circulation exosomes from antibiotic treated or wild type mice, and then inject them into the recipient mice through tail vein, collect the testicular tissue of the recipient mice, conduct transcriptome sequencing, and analyze and compare gene expression differences
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Project description:Collect the blood circulation exosomes from antibiotic treated or wild type mice, and then inject them into the recipient mice through tail vein, collect the testicular tissue of the recipient mice, conduct transcriptome sequencing, and analyze and compare gene expression differences
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Project description:Single cell Transcriptome characterization of the effects of exosomes from different sources on testicular tissue of male mice
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Project description:Testicular injury is often observed in drug development. Serum hormones are usually used as non-invasive biomarkers for testicular injury; however, their sensitivity is low. Therefore, it is difficult to monitor testicular injury in pre-clinical and clinical drug developments. In recent years, molecules in body fluid exosomes are attracting attention as disease biomarkers such as cancer. In this study, small RNAs in serum exosomes were analyzed for identifying non-invasive biomarkers of testicular injury in rats, which are mainly used in a pre-clinical drug development. Testicular injury models in rats were prepared by a single oral administration of 2000 mg/kg ethylene glycol monomethyl ether in which spermatocytes degeneration and sertoli cells vacuolation were observed, or 400 mg/kg carbendazim in which sertoli cells vacuolation and seminiferous tubules dilation were observed. Serum exosome small RNA-seq was performed in these models. The analysis identified 3 small RNAs that fluctuated in common between the models, and miR-423-5p and miR-128-3p were selected as candidate markers. For qPCR validation of the candidates, testicular injury models were further prepared by a single oral administration of 60 mg/kg 1,3-dinitrobenzene or 500 mg/kg nitrofurazone in which spermatocytes degeneration and sertoli cells vacuolation were observed. In qPCR analysis, the selected two miRNAs in exosomes were upregulated in the all models except for 1,3-dinitrobenzene model in which severe hemolysis was observed. On the other hand, the miRNAs in serum did not significantly change in the any models. In conclusion, we identified miR-423-5p and miR-128-3p in serum exosome as non-invasive biomarkers for testicular injury in rats.
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Project description:Spermatogenomics: correlating testicular gene expression to human male infertility
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Project description:ITo derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine, we developed a method to produce a cytocompatible decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components. The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a wide variety of animal and human tissue sources. However, so far, testis scaffolds have never been considered for constructive remodelling purposes. We have therefore developed a protocol for the preparation of cell-free extracellular matrix and characterized the material extensively using a combination of proteomics, immunohistochemsitry, and cell population assays. To prepare cell-free matrix, human cadaveric testicular tissue was exposed for 24 h or 48 h to 1% Triton X-100 and/or 1% sodium dodecyl sulfate (SDS). The extent of decellularization was evaluated by histology. Confirmation of cell removal in DTM was done by a DNA quantification technique. The protein composition was analysed by LC-MS/MS. The retention of testicular tissue-specific characteristics was evaluated by immunohistochemistry, Alcian blue staining and scanning electron microscopy. Soluble toxicity and testicular cell attachment was assessed to check the cytocompatibility of DTM scaffolds. Histological analysis showed that DTM could be obtained by mechanical agitation in 1% SDS for 24 h. The resulting DTM was found to be clear of cells while retaining the typical three-dimensional structure. Proteomics analysis revelaed the presence of the major components of the native tissue scaffold, including collagen type I and IV, fibronectin, laminin and glycosaminoglycans. In addition, numerous additional ECM proteins in DTM were detected, indicating its complex nature. Importantly, we demonstrated that DTM scaffolds are not cytotoxic, as evidenced by MTT assay showing a normal fibroblast proliferation activity after indirect exposure, and support testicular cell attachment and infiltration.
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Project description:ITo derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine, we developed a method to produce a cytocompatible decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components. The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a wide variety of animal and human tissue sources. However, so far, testis scaffolds have never been considered for constructive remodelling purposes. We have therefore developed a protocol for the preparation of cell-free extracellular matrix and characterized the material extensively using a combination of proteomics, immunohistochemsitry, and cell population assays. To prepare cell-free matrix, human cadaveric testicular tissue was exposed for 24 h or 48 h to 1% Triton X-100 and/or 1% sodium dodecyl sulfate (SDS). The extent of decellularization was evaluated by histology. Confirmation of cell removal in DTM was done by a DNA quantification technique. The protein composition was analysed by LC-MS/MS. The retention of testicular tissue-specific characteristics was evaluated by immunohistochemistry, Alcian blue staining and scanning electron microscopy. Soluble toxicity and testicular cell attachment was assessed to check the cytocompatibility of DTM scaffolds. Histological analysis showed that DTM could be obtained by mechanical agitation in 1% SDS for 24 h. The resulting DTM was found to be clear of cells while retaining the typical three-dimensional structure. Proteomics analysis revelaed the presence of the major components of the native tissue scaffold, including collagen type I and IV, fibronectin, laminin and glycosaminoglycans. In addition, numerous additional ECM proteins in DTM were detected, indicating its complex nature. Importantly, we demonstrated that DTM scaffolds are not cytotoxic, as evidenced by MTT assay showing a normal fibroblast proliferation activity after indirect exposure, and support testicular cell attachment and infiltration.
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