Laminin Production and Basement Membrane Deposition by Mesenchymal Stem Cells upon Adipogenic Differentiation

Abstract

The aim was to study laminin (LM) synthesis, integration, and deposition into the basement membrane (BM) during adipogenesis. Human bone marrow-derived mesenchymal stromal cells (MSCs) were induced along the adipogenic lineage. LM chain mRNA and protein levels were followed using quantitative real-time polymerase chain reaction (qRT-PCR), immunofluorescence (IF) staining, transmission electron microscopy (TEM), and immunoprecipitation. MSCs produced low levels of LM mRNAs but were not surrounded by BM in IF and TEM imaging. LM-α4, LM-β1, and LM-γ1 mRNAs increased during adipogenesis 3.9-, 5.8-, and 2.8-fold by day 28. LM-411 was immunoprecipitated from the ECM of the differentiated cells. Immunostaining suggested deposition of LM-411 and some LM-421. BM build-up was probably organized in part by integrin (Int) α6β1. At day 28, TEM images revealed BM-like structures around fat droplet-containing cells. The first signs of BM formation and Int α6β1 were seen using IF imaging at day 14. Laminin-411 and Int α6β1 were expressed in vivo in mature human subcutaneous fat tissue. Undifferentiated human MSCs did not organize LM subunits into BM, whereas LM-411 and some LM-421 are precipitated in the BM around adipocytes. This is the first demonstration of LM-411 precipitation during hMSC adipogenesis around adipocytes as a structural scaffold and Int-regulated signaling element.

Laminin-Alginate Beads as Preadipocyte Carriers to Enhance Adipogenesis In Vitro and In Vivo

“…Moreover, interactions between laminin and its cellular receptors are required to establish cell polarity and to trigger differentiation in embryoid body [18] . For mesenchymal stem cells, adipogenic induction upregulates laminin expression [21] , and stem cell adhesion to the ECM strongly correlates with adipogenic differentiation [25] . These evidence supports that laminin is a critical component of the ECM for stem cell differentiation.…”

“…The final published version may differ from this proof. binding to integrins [21] . Laminin can also promote the aggregation and vascularization of endothelial cells, as blocking laminin in matrigels is sufficient to inhibit tube formation by endothelial cells [22] .…”

Evolution of the Marrow Adipose Tissue Microenvironment

“…Fibronectin is thought to restrain adipocyte cell shape, furthermore, assembly of a fibronectin rich matrix impedes adipocyte differentiation [68–70]. Lastly, laminin is an adipocyte basement membrane protein which in WAT appears to be expressed not by the adipocyte precursor, but the mature adipocyte itself [71]. …”

Vitronectin-Based, Biomimetic Encapsulating Hydrogel Scaffolds Support Adipogenesis of Adipose Stem Cells

“…2,3,32 Changes in ECM expression of adipogenically differentiated mesenchymal cells isolated from bone marrow have been studied previously. 4,5,[27][28][29]33 However, little work has been done examining adipogenesis in the more readily abundant cell population from fat lipoaspirate.…”

“…There are at least eight known integrins that can bind to laminin 28,54 and adipocytes use a small number of these. 33 While integrin expression of BM-MSC-derived adipocytes has been examined previously, 29 the expression of integrins in adipocytes derived from lipoaspirate, compared to their undifferentiated counterparts, has yet to be examined fully. Using immunocytochemistry, we characterized the expression of integrin subunits in undifferentiated ASCs and ASC-derived adipocytes that had been differentiated for 21 days (Fig.…”