Abstract
AimsAllogeneic bone marrow (BM) has been shown to support human islet survival and function in long-term culture by initiating human islet vascularization and β-cell regeneration. Various BM subpopulations may play different roles in human islet functions and survival. In this paper we investigated the effects of BM and its subpopulations, endothelial progenitor cells (E) and mesenchymal (M) cells on human islet's β-cell function and regeneration.Study DesignIsolation and identification of subpopulations from human bone marrow and culture with allogeneic human islet to investigate effects of different cell population on human islet function and regeneration.Place and Duration of StudyDepartment of Medicine, Center for Stem Cell & Diabetes Research, RWMC, Providence, RI, USA, between 2010 - 2014.MethodologyHuman islets were distributed from Integrated Islet Distribution Program (IIDP) and human bone marrow (BM) was harvested by Bone marrow transplantation center at Roger Williams Hospital. BM subpopulation was identified cell surface markers through Fluorescence-activated cell sorting, applied in flow cytometry (FACS), islet function was evaluated by human ELISA kit and β cell regeneration was evaluated by three methods of Cre-Loxp cell tracing, β cell sorting and RT-PCR for gene expression.ResultsFour different BM and seven different islet donates contributed human tissues. We observed islet β-cell having self regeneration capability in short term culture (3∼5 days) using a Cre-Loxp cell tracing. BM and its subtype E, M have similar benefits on β cell function during co-culture with human islet comparison to islet only. However, only whole BM enables to sustain the capability of islet β-cell self regeneration resulting in increasing β cell population while single E and M individual do not significantly affect on that. Mechanism approach to explore β-cell self regeneration by evaluating transcription factor expressions, we found that BM significantly increases the activations of β-cell regeneration relative transcription factors, the LIM homeodomain protein (Isl1), homologue to zebrafish somite MAF1 (MAFa), the NK-homeodomain factor 6.1 (NKX6.1), the paired box family factors 6 (PAX6), insulin promoter factor 1 (IPF1) and kinesin family member 4A (KIF4a).ConclusionThese results suggest that BM and its derived M and E cells enable to support human islet β-cell function. However, only BM can sustain the capability of β-cell self regeneration through initiating β-cell transcriptional factors but not individual E and M cells suggesting pure E and M cells less supportive for islet long-term survival in vitro.
Highlights
Islet β cell lost function after islet transplantation hinders successful islet transplantation [1,2]
Only whole bone marrow (BM) enables to sustain the capability of islet β-cell self regeneration resulting in increasing β cell population while single E and M individual do not significantly affect on that
Mechanism approach to explore β-cell self regeneration by evaluating transcription factor expressions, we found that BM significantly increases the activations of β-cell regeneration relative transcription factors, the LIM homeodomain protein (Isl1), homologue to zebrafish somite MAF1 (MAFa), the NKhomeodomain factor 6.1 (NKX6.1), the paired box family factors 6 (PAX6), insulin promoter factor 1 (IPF1) and kinesin family member 4A (KIF4a)
Summary
Islet β cell lost function after islet transplantation hinders successful islet transplantation [1,2]. Isolation process damage islet may result in β-cell lost function and caused apoptosis in vivo. BM and its derived stem cells have been used in the repair of heart [6,7], neuron [8,9] and muscle [10] injury. BM can support pancreatic growth in vivo and could potentially be manipulated to differentiate into β cells in vitro [11]. BM-derived stem cells migrate towards damaged islet site and differentiate into β cells under the influence of factors from the microenvironment (e.g. cell-cell, cellextracellular matrix interactions and growth factors) [12,13]
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