RESUMEN

BACKGROUND AND AIM: The pathological features of non-alcoholic steatohepatitis (NASH) have not been determined, so fundamental treatment has not been established. Adipose-tissue-derived stromal/stem cells (ADSCs) are beneficial for repair/regenerative therapy of impaired organs because of their immuno-modulatory capability. In this study, we assessed how liver damage progresses during the early development phase of the murine NASH model and investigated whether ADSCs are preventatively efficacious against the fibrosis progression of NASH. METHODS: C57BL/6J mice were fed with atherogenic high fat or high-fat diet 60 developing into NASH or simple steatosis. Their hepatic inflammatory cells (HICs) were analyzed by cDNA microarray. NASH mice were treated with ADSCs injected into spleen when hepatic inflammation was initially observed, and liver samples were analyzed. The effect of ADSCs on the mice hepatic stellate cell (HSC) line stimulated by recombinant IL-17 and HICs from NASH mice was analyzed. RESULTS: The gene expression features of HICs implicated as humoral cytokine mediators of lymphoid cells during NASH development, compared with a simple steatosis model. One of the featured cytokines was IL-17. The development of hepatic fibrosis was alleviated when NASH mice were treated with ADSCs as well as treated with anti-IL-17 antibody, and the frequency of IL-17-secreting HICs decreased. NASH-HICs enhanced proliferation of HSCs, in which proliferation was sensitive to IL-17 stimulation. The stimulatory effect of NASH-HICs on the activation of HSCs was attenuated by co-culture with ADSCs. CONCLUSION: ADSCs treatment prevented progression of NASH fibrosis by suppressing IL-17-mediated inflammation, which was associated with HSCs activation.

Asunto(s)

Tejido Adiposo/trasplante , Mediadores de Inflamación/metabolismo , Interleucina-17/metabolismo , Cirrosis Hepática Experimental/prevención & control , Hígado/metabolismo , Enfermedad del Hígado Graso no Alcohólico/prevención & control , Trasplante de Células Madre , Células Madre/metabolismo , Tejido Adiposo/citología , Animales , Línea Celular , Proliferación Celular , Técnicas de Cocultivo , Dieta Alta en Grasa , Progresión de la Enfermedad , Femenino , Células Estrelladas Hepáticas/inmunología , Células Estrelladas Hepáticas/metabolismo , Células Estrelladas Hepáticas/patología , Mediadores de Inflamación/inmunología , Interleucina-17/inmunología , Interleucina-17/farmacología , Hígado/inmunología , Hígado/patología , Cirrosis Hepática Experimental/inmunología , Cirrosis Hepática Experimental/metabolismo , Cirrosis Hepática Experimental/patología , Ratones Endogámicos C57BL , Enfermedad del Hígado Graso no Alcohólico/inmunología , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Enfermedad del Hígado Graso no Alcohólico/patología , Células Madre/efectos de los fármacos

RESUMEN

Stromal cells in adipose tissue are useful for repair/regenerative therapy as they harbor a substantial number of mesenchymal stem cells; therefore, freshly isolated autologous uncultured adipose tissue derived stromal cells (u-ADSCs) are useful for regenerative therapy, and obviate the need for mesenchymal stem cells. We evaluated the therapeutic effect of murine u-ADSCs and sorted subsets of u-ADSCs in a concanavalin A (ConA) induced murine model of hepatitis, as well as their characteristics. We found that 10-20% of u-ADSCs expressed the CD45 leukocyte-related antigen. CD68, which is a marker of macrophages (MΦs), was expressed by 50% of CD45+ u-ADSCs. About 90% of CD68+ CD45+ cells expressed CD206 antigen, which is a marker of inhibitory M2-type MΦs. Genes related to M2-type MUs were especially more highly expressed by CD45+ CD206+ u-ADSCs than by CD45- u-ADSCs. CD45+ u-ADSCs inhibited the expression of cytokines/chemokines and suppressed the proliferation of splenocytes stimulated with ConA. We observed that not only whole u-ADSCs, but also the CD45+ subset of u-ADSCs ameliorated the ConA-induced hepatitis in mice. In conclusion, we show that freshly isolated murine u-ADSCs were effective against acute hepatitis, and CD45+ u-ADSCs acting phenotypically and functionally like M2-type MΦs, contributed to the repair of liver tissue undergoing inflammation.

Asunto(s)

Tejido Adiposo/inmunología , Antígenos Comunes de Leucocito/inmunología , Células Madre Mesenquimatosas/inmunología , Células del Estroma/inmunología , Tejido Adiposo/citología , Tejido Adiposo/metabolismo , Animales , Células Cultivadas , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Enfermedad Hepática Inducida por Sustancias y Drogas/inmunología , Enfermedad Hepática Inducida por Sustancias y Drogas/terapia , Concanavalina A/toxicidad , Citometría de Flujo , Perfilación de la Expresión Génica/métodos , Interleucina-10/inmunología , Interleucina-10/metabolismo , Antígenos Comunes de Leucocito/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Masculino , Trasplante de Células Madre Mesenquimatosas/métodos , Células Madre Mesenquimatosas/metabolismo , Ratones Endogámicos C57BL , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células del Estroma/metabolismo , Células del Estroma/trasplante

RESUMEN

INTRODUCTION: Adipose tissue stromal cells contain a substantial number of mesenchymal stem cells. As such, their application to regeneration of miscellaneous impaired organs has attracted much attention. METHODS: We designed a clinical study to investigate freshly isolated autologous adipose tissue-derived stromal/stem (regenerative) cell (ADRC) therapy for liver cirrhosis and conducted treatment in four cirrhotic patients. ADRCs were isolated from autologous subcutaneous adipose tissue obtained by the liposuction method, followed with use of the Celution system adipose tissue dissociation device. The primary endpoint is assessment of safety one month after treatment. We also characterized the obtained ADRCs. RESULTS: Two patients had type C cirrhosis, one had nonalcoholic steatohepatitis-cirrhosis, and one had type B cirrhosis. No serious adverse events were observed during the 1-month study period after freshly isolated ADRC infusion. Serum albumin concentrations were maintained or improved during this period as well as during the succeeding follow-up of approximately 1 year in two patients and 6 months in another patient. Liver regeneration-related factors, namely hepatocyte growth factor and interleukin-6, were elevated 1 day after ADRC treatment in all patients. The obtained freshly isolated ADRCs were expanded in culture and found to express mesenchymal stem cell markers. Gene expression profile analysis of ADRCs was shown to involve inflammatory features, suggesting that characteristics of the obtained ADRCs were related to immunomodulatory biological effects. CONCLUSION: This clinical study treatment for liver cirrhosis using ADRCs was proven to be safely conductible, and can be further investigated in future for regeneration/repair of liver cirrhosis.