Influence of sodium fluoride on alkaline phosphatase activity and bone gla protein synthesis in human yellow ligament cells in vitro

Objective To investigate the influence of sodium fluoride(NaF)on alkaline phosphatase(ALP)activity and bone gla protein(BGP)synthesis in yellow ligament cells from different surgical simples in vitro.Methods The human ligament cells

Recombinant human bone morphogenetic protein-7 expressed from CHO cells possessing the activity of bone-induced in vitro

Objective To express the recombinant human bone morphogenetic protein-7 (rhBMP-7) in Chinese hamster ovary (CHO) cells and to establish the in vitro biological activity assay of rhBMP-7. Methods Human BMP-7 cDNA was subcloned into pcDNA3.1 mammalian expression vector and transfected to CHO cells by using the lipofectin transfection method. BMP-7 expression cell culture supernatants were harvested and purified for target protein. To analyze the bioactivity of the secreted rhBMP-7, a novel in vitro assay was established by measuring its alkaline phosphatase (ALP) stimulating of osteoblast cell line, W-20-17. Results BMP-7 stably expressing cell clone was selected, which secreted mature disulfide-linked homodimer form of hBMP-7 and had an apparent molecular weight of 36kDa. rhBMP-7 with >95% purity was obtained using 3 step chromatography method. Bioactivity assay showed that the purified protein specifically stimulated W-20-17 cell producing ALP, with a 4-fold increase of ALP activity at 100ng/ml or more, and the EC50 of 15.6ng/ml. Conclusion Purified rhBMP-7 from this CHO expression system has significant biological activity in induction of osteoblast phenotype, which demonstrates potential bone regeneration activity.

Microbiota-host interplay at the gut epithelial level,health and nutrition

Growing evidence suggests the implication of the gut microbiota in various facets of health and disease. In this review, the focus is put on microbiota-host molecular cross-talk at the gut epithelial level with special emphasis on two defense systems： intestinal alkaline phosphatase（IAP） and inducible heat shock proteins（iHSPs）. Both IAP and iHSPs are induced by various microbial structural components（e.g. lipopolysaccharide, flagellin, CpG DNA motifs）,metabolites（e.g. n-butyrate） or secreted signal molecules（e.g., toxins, various peptides, polyphosphate）. IAP is produced in the small intestine and secreted into the lumen and in the interior milieu. It detoxifies microbial components by dephosphorylation and, therefore, down-regulates microbe-induced inflammation mainly by inhibiting NF-κB pro-inflammatory pathway in enterocytes. IAP gene expression and enzyme activity are influenced by the gut microbiota. Conversely, IAP controls gut microbiota composition both directly, and indirectly though the detoxification of pro-inflammatory free luminal adenosine triphosphate and inflammation inhibition. Inducible HSPs are expressed by gut epithelial cells in proportion to the microbial load along the gastro-intestinal tract. They are also induced by various microbial components, metabolites and secreted molecules. Whether iHSPs contribute to shape the gut microbiota is presently unknown. Both systems display strong anti-inflammatory and anti-oxidant properties that are protective to the gut and the host. Importantly, epithelial gene expressions and protein concentrations of IAP and iHSPs can be stimulated by probiotics, prebiotics and a large variety of dietary components, including macronutrients（protein and amino acids, especially L-glutamine, fat, fiber）, and specific minerals（e.g. calcium）and vitamins（e.g. vitamins K1 and K2）. Some food components（e.g. lectins, soybean proteins, various polyphenols） may inhibit or disturb these systems. The general cel ular and molecular mechanisms involved in the microbiota-host epithelial crosstalk and subsequent gut protection through IAP and iHSPs are reviewed along with their nutritional modulation.Special emphasis is also given to the pig, an economically important species and valuable biomedical model.