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HOW TO CULTURE BONE CELLS ON BIOLAMININ SUBSTRATES
High expression of laminin isoforms in the bone microenvironment
Bone cells, including include osteocytes, osteoblasts, osteoclasts, osteogenic cells (stem cells), and lining cells, reside in the bone marrow. There are several laminin family proteins expressed in human bone marrow microenvironment where laminin 411/421 and laminin 511/521 are the most abundant isoforms, synthesized by human bone marrow stromal cells. Laminin isoforms 111, 331 and 332 are also expressed in the human bone marrow microenvironment (Siler, 2000). Laminin 332 is specifically expressed around primary osteoblasts and osteoblast-like cells localized on the bone surface (Uehara, 2017)
LN332 negatively regulates osteoclastogenesis and promote osteogenic differentiation
A publication by German researchers show that laminin 332 promote attachment, and compared to plastic, osteogenic differentiation was significantly increased by laminin 332 (Mittag, 2012). In addition, laminin 332 (has been shown to markedly inhibit RANKL induced osteoclastogenesis (Uehara, 2017). The suppression is thought to be mediated through laminin binding to integrin receptors α3β1, α6β1, and α6β4 (Uehara, 2017; Hashimoto, 2006). Laminin 332 is expressed by primary osteoblasts in culture and the laminin g2 chain is transiently expressed in chondrocytes during development (Uehara, 2017; Hashimoto, 2006). Laminin 332 expression is negatively regulated by osteoclastogenic factors, suggesting that LN332 is a novel negative regulator that may define osteoclastogenesis spatiotemporally in bone tissues (Uehara, 2017). Laminin 332 has also been shown to suppresses the chondrogenic differentiation of BM-MSC without inducing apoptosis or inhibiting cell growth (Hashimoto, 2005; Hashimoto, 2006). However, laminin 332 had no effect on the osteogenic differentiation of MSCs (Hashimoto, 2006). These results suggest that laminin 332 may contribute to the development of bone tissues by promoting the proliferation and by suppressing the chondrogenic differentiation of MSCs.
Best adhesion, growth, and proliferation of BM-MSC on LN511 and LN521
Laminin 511 and 521 are the most abundant isoforms in the bone marrow and bone marrow-derived mesenchymal stem cells (BM-MSCs) cultured in vitro has been shown to synthesize α5, α4, α3, α1 and β2 at the significant amount (Seeger, 2015; Siler, 2000; Hashimoto, 2006). Naturally, laminin 511 and 521 have been shown to have strong adhesive interactions with human CD34+ cell lines (Siler, 2000). However, MSCs does not do not attach well to laminin 111, 211 and 221(Sun, 2017). Laminin 511, 521 and 332 promote the strongest BM-MSC growth and proliferation rate and affect the mitogenic activity and migration of these cells via binding to integrin α6β1 and α3β1 (Siler, 2000; Sun, 2017, Hashimoto, 2005; Hashimoto, 2006). In a recent publication by Yang and Xiao, the authors present a protocol for the culture of bone marrow MSC (BM-MSCs) on laminin 521 and laminin 511. Both laminin isoforms show a significantly faster and stronger attachment compared to uncoated wells and support the seeding of a lower cell number compared to uncoated plats (Yang and Xiao, 2016). When investigating the effects of different ECM coatings for the formation of cell sheets, the result showed the highest success for laminin 521 (Jiang, 2016). Bone marrow mesenchymal stem cells (BMSC) cultured on laminin 521 coated TiO2 nanodot films rapidly attached and spread and formed an intact cell with good viability improved osteogenesis. Compared with mechanical scraping, this light-induced method provides a more robust strategy to fabricate BMSC sheet−implant complexes with better survival and enhanced osteogenesis (Jiang, 2016).
BIOLAMININ KEY ADVANTAGES
Laminin 511and 521 are the most abundant isoforms in the bone marrow. Laminin isoforms 111, 331 and 332 are also expressed.
Laminin 511 and 521 have been shown to have strong adhesive interactions with human CD34+ cell lines and promote BM-MSC growth and proliferation
Laminin 332 negatively regulates osteoclastogenesis and promote attachment and osteogenic differentiation
Defined and animal component-free culture
No lot-to-lot variability for standardized experiments with less variation