RESUMEN
The use of stem cells capable of multilineage differentiation in treating Pelvic Floor Dysfunction (PFD) holds great promise since they are susceptible to entering connective tissue of various cell types and repairing damaged tissues. This research investigated the effect of microRNA-181a-5p (miR-181a-5p) on Bone Marrow Mesenchymal Stem Cells (BMSCs) in rats with PFD. BMSCs were transfected and analyzed for their fibroblast differentiation ability. miR-181a-5p, MFN1, and fibroblast-related genes were quantitatively analyzed. Whether MFN1 is a target gene of miR-181a-5p was predicted and confirmed. The efficacy of BMSCs in vivo rats with PFD was evaluated by measuring Leak Point Pressure (LPP), Conscious Cystometry (CMG), hematoxylin and eosin staining, and Masson staining. The present results discovered that miR-181a-5p was up-regulated and MFN1 was down-regulated during the differentiation of BMSCs into fibroblasts. Fibroblast differentiation of BMSCs was promoted after miR-181a-5p was induced or MFN1 was suppressed, but it was suppressed after miR-181a-5p was silenced. miR-181a-5p improved LPP and conscious CMG outcomes in PDF rats by targeting MFN1 expression, thereby accelerating fibroblast differentiation of BMSCs. In brief, miR-181a-5p induces fibroblast differentiation of BMSCs in PDF rats by MFN1, potentially targeting PDF therapeutics.
Asunto(s)
Diferenciación Celular , Fibroblastos , Células Madre Mesenquimatosas , MicroARNs , Animales , Células Madre Mesenquimatosas/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , Femenino , Ratas Sprague-Dawley , Trastornos del Suelo Pélvico/genética , Trastornos del Suelo Pélvico/terapia , Ratas , Regulación hacia Arriba , Modelos Animales de Enfermedad , Regulación hacia Abajo , Células CultivadasRESUMEN
Abstract The use of stem cells capable of multilineage differentiation in treating Pelvic Floor Dysfunction (PFD) holds great promise since they are susceptible to entering connective tissue of various cell types and repairing damaged tissues. This research investigated the effect of microRNA-181a-5p (miR-181a-5p) on Bone Marrow Mesenchymal Stem Cells (BMSCs) in rats with PFD. BMSCs were transfected and analyzed for their fibroblast differentiation ability. miR-181a-5p, MFN1, and fibroblast-related genes were quantitatively analyzed. Whether MFN1 is a target gene of miR-181a-5p was predicted and confirmed. The efficacy of BMSCs in vivo rats with PFD was evaluated by measuring Leak Point Pressure (LPP), Conscious Cystometry (CMG), hematoxylin and eosin staining, and Masson staining. The present results discovered that miR-181a-5p was up-regulated and MFN1 was down-regulated during the differentiation of BMSCs into fibroblasts. Fibroblast differentiation of BMSCs was promoted after miR-181a-5p was induced or MFN1 was suppressed, but it was suppressed after miR-181a-5p was silenced. miR-181a-5p improved LPP and conscious CMG outcomes in PDF rats by targeting MFN1 expression, thereby accelerating fibroblast differentiation of BMSCs. In brief, miR-181a-5p induces fibroblast differentiation of BMSCs in PDF rats by MFN1, potentially targeting PDF therapeutics.
RESUMEN
BACKGROUND: Survival and therapeutic actions of bone marrow-derived mesenchymal stem cells (BMMSCs) can be limited by the hostile microenvironment present during acute spinal cord injury (SCI). Here, we investigated whether BMMSCs overexpressing insulin-like growth factor 1 (IGF-1), a cytokine involved in neural development and injury repair, improved the therapeutic effects of BMMSCs in SCI. METHODS: Using a SCI contusion model in C57Bl/6 mice, we transplanted IGF-1 overexpressing or wild-type BMMSCs into the lesion site following SCI and evaluated cell survival, proliferation, immunomodulation, oxidative stress, myelination, and functional outcomes. RESULTS: BMMSC-IGF1 transplantation was associated with increased cell survival and recruitment of endogenous neural progenitor cells compared to BMMSC- or saline-treated controls. Modulation of gene expression of pro- and anti-inflammatory mediators was observed after BMMSC-IGF1 and compared to saline- and BMMSC-treated mice. Treatment with BMMSC-IGF1 restored spinal cord redox homeostasis by upregulating antioxidant defense genes. BMMSC-IGF1 protected against SCI-induced myelin loss, showing more compact myelin 28 days after SCI. Functional analyses demonstrated significant gains in BMS score and gait analysis in BMMSC-IGF1, compared to BMMSC or saline treatment. CONCLUSIONS: Overexpression of IGF-1 in BMMSC resulted in increased cell survival, immunomodulation, myelination, and functional improvements, suggesting that IGF-1 facilitates the regenerative actions of BMMSC in acute SCI.
Asunto(s)
Factor I del Crecimiento Similar a la Insulina/genética , Trasplante de Células Madre Mesenquimatosas , Células-Madre Neurales/trasplante , Traumatismos de la Médula Espinal/terapia , Animales , Células de la Médula Ósea/citología , Diferenciación Celular/genética , Modelos Animales de Enfermedad , Humanos , Células Madre Mesenquimatosas/citología , Ratones , Vaina de Mielina/genética , Vaina de Mielina/patología , Células-Madre Neurales/citología , Recuperación de la Función , Regeneración/genética , Traumatismos de la Médula Espinal/genética , Traumatismos de la Médula Espinal/patologíaRESUMEN
RESUMEN: Las células madre de la línea germinal masculina son factores clave para la espermatogénesis masculina y la fertilidad. Las células sustentaculares (células de Sertoli) como células somáticas juegan un papel fundamental en la creación de un microambiente esencial para la auto-renovación y diferenciación de las células de la línea germinal masculina. Las células madre mesenquimales son reconocidas como células auto-renovables y multipotentes capaces de diferenciarse en múltiples tipos de células. La generación de células germinales masculinas a partir de células madre mesenquimales puede proporcionar un método terapéutico para tratar la infertilidad masculina. En este estudio, las células mesenquimales derivadas de la médula ósea (BMMSCs) se recuperaron de la médula ósea de ratones de 6-8 semanas de edad del Instituto de Investigación Médico Naval (NMRI). En el estudio se aislaron las células sustentaculares y se enrriquecieron usando placas revestidas con lectina. Se obtuvo el medio de condición celular después de diferentes intervalos de tiempo. Posteriormente se cultivaron las BMMSC con diferentes concentraciones de SCCM y medio de Eagle modificado por Dulbecco (DMEM) en diversos momentos. Se evaluaron marcadores específicos de células de línea germinal usando la reacción en cadena de polimerasa transcriptasa inversa (RT-PCR) e inmunocitoquímica. Los resultados mostraron que las BMMSCs cultivadas con SCCM durante 48h exhibieron transcritos específicos de línea germinal (Mvh, Iid4, piwil2) (p <0,05) y marcadores (Mvh, Scp3). Nuestros resultados indican que el cultivo de BMMSCs con SCCM puede conducir a la diferenciación efectiva de BMMSCs en células germinales y proporcionar una estrategia de tratamiento para la infertilidad masculina.
SUMMARY: Male germ line stem cells are key factors for male spermatogenesis and fertility. Sustentacular cells (Sertoli cells) as somatic cells play a pivotal role in creating essential microenvironment for the self-renewal and differentiation of the male germ line cells. Mesenchymal stem cells are recognized as self-renewing and multipotent cells able to differentiate into multiple cell types. The generation of male germ cells from mesenchymal stem cells may provide a therapeutic method to treat male infertility. In this study, Bone marrow derived mesenchymal cells (BMMSCs) were retrieved from the bone marrow of 6-8-week old Naval Medical Research Institute (NMRI) mice. Sustentacular cells (Sertoli cells) were isolated and made rich using lectin coated plates. Sustentacular cell condition medium (SCCM) was collected after different time intervals. Then the BMMSCs were cultured with different concentration of SCCM and Dulbecco's Modified Eagle's medium (DMEM) at various times. Specific markers of Germ line cells were evaluated by using Reverse transcriptase polymerase chain reaction (RT-PCR) and immunocytochemistry. The results showed that BMMSCs cultured with SCCM for 48h exhibited germ line specific transcripts (Mvh, Iid4, piwil2) (p< 0.05) and markers (Mvh, Scp3). Our findings represent that culturing BMMSCs with SCCM may lead to effective differentiation of BMMSCs into germline cells and provide a treatment strategy for male infertility.
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Animales , Masculino , Ratones , Células de Sertoli/citología , Células Madre Mesenquimatosas/citología , Células de Sertoli/ultraestructura , Testículo/citología , Médula Ósea , Inmunohistoquímica , Diferenciación Celular , Medios de Cultivo Condicionados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Citometría de FlujoRESUMEN
Adult stem cells have beneficial effects when exposed to damaged tissue due, at least in part, to their paracrine activity, which includes soluble factors and extracellular vesicles (EVs). Given the multiplicity of signals carried by these vesicles through the horizontal transfer of functional molecules, human mesenchymal stem cell (hMSCs) and CD133+ cell-derived EVs have been tested in various disease models and shown to recover damaged tissues. In this study, we profiled the protein content of EVs derived from expanded human CD133+ cells and bone marrow-derived hMSCs with the intention of better understanding the functions performed by these vesicles/cells and delineating the most appropriate use of each EV in future therapeutic procedures. Using LC-MS/MS analysis, we identified 623 proteins for expanded CD133+-EVs and 797 proteins for hMSCs-EVs. Although the EVs from both origins were qualitatively similar, when protein abundance was considered, hMSCs-EVs and CD133+-EVs were different. Gene Ontology (GO) enrichment analysis in CD133+-EVs revealed proteins involved in a variety of angiogenesis-related functions as well proteins related to the cytoskeleton and highly implicated in cell motility and cellular activation. In contrast, when overrepresented proteins in hMSCs-EVs were analyzed, a GO cluster of immune response-related genes involved with immune response-regulating factors acting on phagocytosis and innate immunity was identified. Together our data demonstrate that from the point of view of protein content, expanded CD133+-EVs and hMSCs-EVs are in part similar but also sufficiently different to reflect the main beneficial paracrine effects widely reported in pre-clinical studies using expanded CD133+ cells and/or hBM-MSCs.