Biocompatibility studies of low temperature nitrided and collagen-I coated AISI 316L austenitic stainless steel.

The biocompatibility of austenitic stainless steels can be improved by means of surface engineering techniques. In the present research it was investigated if low temperature nitrided AISI 316L austenitic stainless steel may be a suitable substrate for bioactive protein coating consisting of collagen-I. The biocompatibility of surface modified alloy was studied using as experimental model endothelial cells (human umbilical vein endothelial cells) in culture. Low temperature nitriding produces modified surface layers consisting mainly of S phase, the supersaturated interstitial solid solution of nitrogen in the austenite lattice, which allows to enhance surface microhardness and corrosion resistance in PBS solution. The nitriding treatment seems to promote the coating with collagen-I, without chemical coupling agents, in respect of the untreated alloy. For biocompatibility studies, proliferation, lactate dehydrogenase levels and secretion of two metalloproteinases (MMP-2 and MMP-9) were determined. Experimental results suggest that the collagen protection may be favourable for endothelial cell proliferation and for the control of MMP-2 release.

Biocompatibility evaluation of surface-treated AISI 316L austenitic stainless steel in human cell cultures.

The effects of AISI 316L austenitic stainless steel, tested in untreated state or subjected to glow-discharge nitriding (at 10 or 20 hPa) and nitriding + post-oxidizing treatments, on human umbilical vein endothelial cells (HUVEC) and on peripheral blood mononuclear cells (PBMC) were evaluated. All the treated samples showed a better corrosion resistance in PBS and higher surface hardness in comparison with the untreated alloy. In HUVEC put in contact for 72 h with the sample types, proliferation and apoptosis decreased and increased, respectively, in the presence of the nitrided + post-oxidized samples, while only slight differences in cytokine (TNF-alpha, IL-6, and TGF-beta1) release were registered. Intercellular adhesion molecule-1 (ICAM-1) increased in HUVEC incubated with all the treated samples, while vascular cell adhesion molecule-1 (VCAM-1) and E-selectin increased in the presence of all the sample types. PBMC incubated for 48 h with the samples showed a decrease in proliferation and an increase in apoptosis in the presence of the untreated samples and the nitrided + post-oxidized ones. All the sample types induced a remarkable increase in TNF-alpha and IL-6 release in PBMC culture medium, while only the untreated sample and the nitrided at 10 hPa induced an increase in ICAM-1 expression. In HUVEC cocultured with PBMC, previously put in contact with the treated AISI 316L samples, increased levels of ICAM-1 were detected. In HUVEC coincubated with the culture medium of PBMC, previously put in contact with the samples under study, a noteworthy increase in ICAM-1, VCAM-1, and E-selectin levels was always registered, with the exception of VCAM-1, which was not affected by the untreated sample. In conclusion, even if the treated samples do not show a marked increase in biocompatibility in comparison with the untreated alloy, their higher corrosion resistance may suggest a better performance as the contact with physiological environment becomes longer.

Sphingosine 1-phosphate induces myoblast differentiation through Cx43 protein expression: a role for a gap junction-dependent and -independent function.

Although sphingosine 1-phosphate (S1P) has been considered a potent regulator of skeletal muscle biology, acting as a physiological anti-mitogenic and prodifferentiating agent, its downstream effectors are poorly known. In the present study, we provide experimental evidence for a novel mechanism by which S1P regulates skeletal muscle differentiation through the regulation of gap junctional protein connexin (Cx) 43. Indeed, the treatment with S1P greatly enhanced Cx43 expression and gap junctional intercellular communication during the early phases of myoblast differentiation, whereas the down-regulation of Cx43 by transfection with short interfering RNA blocked myogenesis elicited by S1P. Moreover, calcium and p38 MAPK-dependent pathways were required for S1P-induced increase in Cx43 expression. Interestingly, enforced expression of mutated Cx43(Delta130-136) reduced gap junction communication and totally inhibited S1P-induced expression of the myogenic markers, myogenin, myosin heavy chain, caveolin-3, and myotube formation. Notably, in S1P-stimulated myoblasts, endogenous or wild-type Cx43 protein, but not the mutated form, coimmunoprecipitated and colocalized with F-actin and cortactin in a p38 MAPK-dependent manner. These data, together with the known role of actin remodeling in cell differentiation, strongly support the important contribution of gap junctional communication, Cx43 expression and Cx43/cytoskeleton interaction in skeletal myogenesis elicited by S1P.