Changes in actin microfilaments are mediated
Changes in 134 4 microfilaments are mediated by changes in extracellular matrix, mechanical stimulation or growth factors intracellular signaling. We observed significant changes in a number of intracellular signaling pathways associated with changes in actin dynamics during hMSCs differentiation. During adipocyte differentiation, decreased in actin assembly was associated with significant changes in Smad2 and ERK MAPK activation. Smad2 has been reported as a transcriptional modulator that represses CEBP activation which is an adipocyte master transcriptional factor. Consistent with this notion, we observed inhibition of Smad2 activation by CytoD and the enhancement of Smad2 activation by Phalloidin, suggested Smad2 signaling might be involved in the regulation of adipogenic differentiation of actin assembly. While SB431546, a TGFR inhibitor that inhibits TGF/Smad2 activation in cells, did not rescued the inhibition of adipogenic differentiation by Phalloidin, either the activation of phos-Smad2 activation by Phalloidin and adipogenic induction (data not shown), suggested the Smad2 activation by Phalloidin (actin polymerization) during adipogenic induction might not related with TGFβ-TGFR pathways. Actin cytoskeleton has been reported to affect ERK activation in cell type dependent manner (Ailenberg and Silverman, 2003). In MSC, Muller et al. reported that CytoD enhances adipocyte differentiation in presence of mechanical stimulation and that these effects were associated with Akt but not ERK activation, while another actin depolymerizing reagent latrunculin A (LatA) blocked both Akt and ERK activation (Muller et al., 2013). We did not detected clear changes at activation of AKT, p38 or JNK by CytoD or Phalloidin in hMSCs during adipogenic differentiation (data not shown). These results indicate there are multiple signaling pathways associated with actin cytoskeletal changes dependent on cell type and culture conditions.
Changes in actin microfilament assembly and disassembly are associated with several cellular functions including cell cycle, morphological maintenance, cell attachment and adhesion, and cell locomotion which are determining factors for cellular differentiation fate (McBeath et al., 2004). While changing cell shape of hMSCs influences differentiation potential of the cells as demonstrated in a number of studies where MSC were cultured on substrates of different stiffness (Engler et al., 2006). This approach has been utilized in tissue engineering protocols with the aim of controlling differentiation of MSC through engineering physical cues on cell culture surfaces or through exposing the cells to mechanical stimulation (Kshitiz et al., 2012). Our current study and previous study (Chen et al., 2015) demonstrate that by targeting the molecular mechanisms responsible for actin microfilaments assembly, it is possible to direct the differentiation fate of MSC into osteoblast or adipocytes. Thus, targeting intracellular molecular machinery responsible for cytoskeletal homoeostasis is a plausible approach for controlling differentiation fate of MSC and is relevant approach for regenerative medicine applications (Fig. 6).
Acknowledgment This study is supported by a grant from the Novo Nordisk Foundation (R179-A15165, L.C.), ECTS Postdoctoral Fellowship 2012 (L.C.) and Novo Nordisk Foundation (NNF15OC0016284, M.K.). All authors have no conflicts of interest.
Lactobacilli are renowned and prevalent in food industry for their probiotic functions. General contributions of lactobacilli such as expelling pathogens and virus , , , metabolizing/producing prebiotics , , regulating intestinal permeability , , and enhancing immune system rely on bacterial colonization in host , , , . The ability to adhere to host cells is the prerequisite for probiotics to manifest biological functions. Actin as the basic structure of cytoskeleton offers a wide-surfaced and well-knit arena for bacterial adhesion and colonization . Many studies on host-pathogen interaction have highlighted actin as an important vehicle for bacterial adhesion and translocation , . This study endeavors to isolate adhesion proteins in lactobacilli, which target actin as receptor. Binding domain between an adhesion protein and actin was also investigated.