Effect Of MicroRNA10b On The Directed Migration Of MSCs Indcuced By HGF

Multipotent mesenchymal stromal cells(MSCs) are a group of clonogenic cellspresent among the bone marrow stroma and capable of multilineage differentiation intomesoderm-type cells such as osteoblast, adipocyte and chondrocyte. Due to their ease ofisolation and their differentiation potential, MSC are being introduced into clinicalmedicine in variety of applications and through different ways of administration. Recentdata have pointed to the unexpected ability of both human and rodent MSCs todifferentiate into neural cells. The evidence has suggested their therapeutic potential toregenerate neural cells in the injured or diseased brain. And mesenchymal stem cells(MSCs) can also be used as a delivery vehicle for gene therapy against brain tumors,because these cells have a migratory capacity toward glioma cells. The precisemechanisms underlying migration of MSCs into the injured tissue are still not fullyunderstood, although multiple signaling pathways and molecules were reported. Solublefactors including chemokines or growth factors expressed and released by glioma cellsmediate the tropism of MSCs for gliomas. In vitro, HGF can induce the directionalmigration of MSCs which express both HGF and its receptor c-Met. It is show thathuman neural stem cells (NSCs) lines responded to multiple tumor-derived growthfactors and that hepatocyte growth factor (HGF) induced the strongest chemotacticresponse.MicroRNAs (miRNAs) are20-25nucleotide(nt) non-coding RNAs that bind to the3′untranslated region (UTR) of target mRNAs through an imperfect match to represstheir translation and stability. Recently, miRNAs have emerged as important players inpost-translational regulation and have implications in controlling stem cell fate andbehaviour. For example, microRNA17/20, microRNA10b, miRNA151, microRNA146band microRNA23modulate tumor cells migration by targeting different key proteins.MicroR-9and microR-124which specifically expressed in neuronal lineage can alsoregulate these cells differentiation and migration, whereas microRNA145suppressestumor cell proliferation, migration and invasion and play crucial roles in balancing self-renewal and differentiation of embryonic stem (ES) cells. Above evidences suggestthat these microRNA might have an important role in modulating MSCs directionalmigration induced by HGF.In this study, we evaluate the differential expression of nine microRNA involved incell migration after MSCs treated with5ng/ml HGF for30minute through quantitativeRT-PCR. We show that after the HGF treatment microRNA145, microRNA17-5p,microRNA181, microRNA9were significant upregulated, whereas microRNA10b,microRNA124,microRNA146were remarkable downregulated. It is report thatmicroRNA10b suppress cell migration by targeting Tiam1and HoxD10which canactivate Rac1and RhoC respectively. Thus we further investigate the mechanisms ofmicroRNA10b and its downstream signal molecular Rac1regulation on HGF induceddirectional migration of MSCs.In order to study the effect of microRNA10b on the directed migration of MSCsindcuced by HGF, we used directed-viewing Dunn chamber to detail the MSCs whichtransfected with microRNA10b vector or its inhibitor vector migratory responses toHGF. Quantitative analyses revealed that overexpression of microRNA10b significantlydecreased the migration efficiency of MSCs, whereas no significant changed inmigration speed; Both the migration speed and migration efficiency were increased bysuppression of microRNA10b.Rac1which is the downstream signaling molecule of microRNA10b can modulatescell spreading, cell polarity and cell migration through regulating actin cytoskeletonrearrangement, focal complex/adhesion turnover and lamellipodium extension. In orderto study the effect of Rac1on the directed migration of MSCs, we constructed therecombinant adenovirus vector of Rac1and its mutants Rac1Q61L, Rac1G12V,Rac1T17N. About107pfu/ml of high titer recombinant virus was successfully obtained,the optimal virus concentration for MSCs was150MOI.Dunn chamber was used to detch migration speed and FMI of MSCs induced byHGF at single cell level. These results show that MSCs infected with Ad-Rac1Q61Lexhibits an incresed migration speed, but no change is observed in the migration FMI.Ad-Rac1T17N infected MSCs both migration speed and FMI are decreased, whereasAd-Rac1infected MSCs shows no change in both migartion speed and FMI. Meanwhile,boyden chamber was used to assess cells migration efficients at the population level. Wefind that both Ad-Rac1and Ad-Rac1Q61L can promote MSCs migration toward HGF, whereas Ad-Rac1T17N inhibits this chemotactic movement.In conclusion, Rac1and microRNA10b regulates the HGF-induced directedmigration of MSCs. In further study will continue to investigate the function ofdifferentially expressed microRNAs which detected by real-time quantitative PCR,expecting to find the crosstalk between the microRNA pathways. This study is requiredto delineate the molecular mechanisms that govern the directed migration of MSCs,thereby allowing for optimization of the therapeutic potential of MSCs to be employedfor neural regeneration after injury.