Injectable oligomer-cross-linked gelatine hydrogels via anhydride-amine-conjugation.

Injectable gelatine-based hydrogels are valuable tools for drug and cell delivery due to their extracellular matrix-like properties that can be adjusted by the degree of cross-linking. We have established anhydride-containing oligomers for the cross-linking of gelatine via anhydride-amine-conjugation. So far, this conversion required conditions not compatible with cell encapsulation or in vivo injection. In order to overcome this limitation, we developed an array of quarter-oligomers varying in comonomer composition and contents of reactive anhydride units reactive towards amine groups under physiological conditions. The oligomers were of low molecular weight (Mn < 5 kDa) with a high degree of chemically intact anhydrides. Chemical comonomer composition was determined by 1H-NMR. Dissolutions experiments confirmed improved hydrophilicity of the synthesized oligomers over our established compositions. Injectable formulations are described utilizing cytocompatible concentrations of constituent materials and proton-scavenging base. Degree of cross-linking and stiffness of injectable hydrogels were controlled by composition. The gels hold promise as injectable drug or cell carrier and as bioink.

Co(II)-mediated effects of plain and plasma immersion ion implanted cobalt-chromium alloys on the osteogenic differentiation of human mesenchymal stem cells.

Medical CoCr is one of the main alloys used for metal-on-metal prosthesis in patients with total hip arthroplasty. CoCr surfaces modified by nitrogen plasma immersion ion implantation (PIII) are characterized by improved wear resistance but also showed increased Co(II) ion release under in vitro conditions. For the first time, CoCr modified by nitrogen PIII was evaluated with regard to its effect on the osteogenic differentiation of MSC. The activity of alkaline phosphatase, the expression of the osteogenic genes Runt-related transcription factor 2, osteopontin as well as integrin-binding bone sialoprotein and the production of osteocalcin and hydroxyapatite were determined. The results of our study demonstrate that Co(II) ions released from the alloy affected the osteogenic differentiation of MSC. Distinct differences in differentiation markers were found between pristine and modified alloys for osteocalcin but not for integrin-binding sialoprotein and hydroxyapatite. Interestingly, osteopontin was upregulated in naive and differentiated MSC by Co(II) ions and modified CoCr, likely through the induction of a cellular hypoxic response. The findings of this study contribute to a better understanding of possible risk factors with regard to a clinical applicability of surface modified CoCr implant materials.

Cytocompatibility of nitrogen plasma ion immersed medical cobalt-chromium alloys.

Wear particles and ion release from medical CoCr contribute to the risk for aseptic loosening. Nitrogen plasma immersion ion implantation (PIII) has been shown to reduce wear of CoCr but is associated with increased Co ion release. This work investigated the cytocompatibility of CoCr modified by nitrogen PIII at different temperatures (mCoCr). The osteosarcoma cell line Saos-2, mesenchymal stem cells (MSCs), and mononuclear cells (MNCs) were grown directly on CoCr/mCoCr discs or treated with CoCl2. Proliferation and metabolic activity of Saos-2 and MSC were decreased on mCoCr in correlation with increasing implantation temperature. The elevated Co ion release seemed to play a decisive role since analog effects were observed by treatment of cells with CoCl2. Proliferation of phytohemagglutinin-stimulated MNC was reduced in the presence of CoCr discs or CoCl2. For MNC-alloy cultures we observed an increase in interleukin 2 (IL-2) and a decrease in interferon γ (IFN-γ) and IL-10 secretion compared to cultures without alloys. The results of this study demonstrate that PIII process temperature and corresponding Co ion release correlate with material cytocompatibility. Therefore, the two competing parameters, reduction of wear and increase in Co ions, have to be taken into consideration for the evaluation of the clinical applicability of nitrogen-implanted CoCr.

Visfatin is a positive regulator of MCP-1 in human adipocytes in vitro and in mice in vivo.

Visfatin is a proinflammatory and potentially insulin-mimetic adipokine contributing to whole body glucose and lipid metabolism, as well as atherosclerosis. Monocyte chemoattractant protein (MCP)-1 is an adipocyte-secreted protein which might play a crucial role in metabolic and vascular disease. MCP-1 expression and secretion after visfatin treatment were determined by quantitative real-time reverse transcription-PCR and enzyme-linked immunosorbent assay (ELISA) in fully differentiated human mesenchymal stem cell-derived adipocytes (hMSC-Ads) in vitro. In addition, circulating levels of MCP-1 and visfatin were quantified by ELISA in 60 patients (30 nondiabetic, 30 diabetic) and MCP-1 serum levels in mice were determined after visfatin treatment in vivo. Interestingly, protein secretion and mRNA production of MCP-1 were induced significantly in hMSC-Ads after visfatin stimulation. Visfatin-induced MCP-1 secretion 1.9-fold after 8 h and 2.5-fold after 24 h relative to untreated cells (P < 0.05). MCP-1 mRNA synthesis was significantly stimulated by visfatin with maximal upregulation detectable at 250 ng/ml visfatin and after 4 h of treatment. Signaling studies suggested that p44/42 mitogen-activated protein (MAP) kinase is involved in visfatin-induced MCP-1 mRNA expression in hMSC-Ads. Detectability of visfatin in serum predicted circulating MCP-1 independent of age and gender in humans in vivo. MCP-1 serum levels were significantly increased more than twofold after visfatin treatment in mice in vivo. Taken together, our results demonstrate that visfatin upregulates MCP-1 supporting a possible role of MCP-1 in mediating the proinflammatory effects of visfatin.

Interleukin-1beta is a positive regulator of TIARP/STAMP2 gene and protein expression in adipocytes in vitro.

The impact of interleukin (IL)-1beta on tumor necrosis factor alpha-induced adipose-related protein (TIARP)/six-transmembrane protein of prostate 2 (STAMP2) was determined in adipocytes. TIARP/STAMP2 mRNA synthesis was significantly stimulated by IL-1beta in a dose- and time-dependent fashion in 3T3-L1 adipocytes. Signaling studies suggested that janus kinase 2, nuclear factor kappaB, and p44/42 mitogen-activated protein kinase are involved in IL-1beta-induced TIARP/STAMP2 mRNA expression. Furthermore, IL-1beta, TNFalpha, and IL-6 showed synergistic stimulatory effects on TIARP/STAMP2 gene expression. Moreover, both TIARP/STAMP2 mRNA synthesis and protein expression were induced by IL-1beta in fully differentiated human mesenchymal stem cell-derived adipocytes (hMSC-Ad). Taken together, TIARP/STAMP2 is highly upregulated in 3T3-L1 cells and hMSC-Ad by IL-1beta and might, therefore, modulate proinflammatory and insulin resistance-inducing effects of IL-1beta.