ECM430 Sigma-Aldrichα Integrin Blocking and IHC Kit

The overall mechanisms governing the role of laminins during osteogenic differentiation of human mesenchymal stem cells (hMSC) are poorly understood. We previously reported that laminin-332 induces an osteogenic phenotype in hMSC and does so through a focal adhesion kinase (FAK) and extracellular signal-related kinase (ERK) dependent pathway. We hypothesized that this is a result of integrin-ECM binding, and that it occurs via the known alpha3 LG3 integrin binding domain of laminin-332. To test this hypothesis we cultured hMSC on several different globular domains of laminin-332. hMSC adhered best to the LG3 domain, and this adhesion maximally activated FAK and ERK within 120 min. Prolonged culturing (8 or 16 days) of hMSC on LG3 led to activation of the osteogenic transcription factor Runx2 and expression of key osteogenic markers (osterix, bone sialoprotein 2, osteocalcin, alkaline phosphatase, extracellular calcium) in hMSC. LG3 domain binding did not increase matrix mineralization, demonstrating that the LG3 domain alone is not sufficient to induce complete osteogenic differentiation in vitro. We conclude that the LG3 domain mediates attachment of hMSC to laminin-332 and that this adhesion recapitulates most, but not all, of the osteogenic differentiation associated with laminin-5 binding to hMSC.

The development of the human fetal adrenal gland is characterized by a gradient of mitotic activity, cell migration, and cell apoptosis, all of which dictate its particular function. Such plasticity may possibly be under the control of the extracellular environment. The goal of this study was to identify components of the extracellular matrix in second-trimester fetal adrenal glands. Whereas collagen IV was expressed evenly throughout the gland, both fibronectin and laminin demonstrated a mirror-imaged distribution, with higher expression of fibronectin in the central portion and laminin at the periphery of the gland. The integrin subunit alpha1 was found mainly in the definitive zone and the alpha2-subunit mainly in the transitional zone, whereas integrin alpha3 (which binds both fibronectin and laminin) was detected only in the fetal zone. The beta2-subunit was observed solely in chromaffin cells. Such specific gradients of integrin and MEC component expression suggest that the extracellular environment does play a definite role during adrenal gland development. Indeed, compared with that in untreated plastic dishes, ACTH stimulation of dehydroepiandrosterone sulfate and cortisol was enhanced by collagen IV. In addition, fibronectin enhanced dehydroepiandrosterone sulfate but decreased cortisol secretion, compared with collagen IV substrates. These results provide fundamental insight into the contribution of the microenvironment in cellular processes leading to fetal adrenal gland development.