Although in vivo studies have shown that low-magnitude, high-frequency (LMHF) vibration (LM: < 1 ×g; HF: 20-90 Hz) exhibits anabolic effects on skeletal homeostasis, the underlying cellular/molecular regulation involved in bone adaptation to LMHF vibration is little known.In this report, we tested the effects of microvibration (magnitude: 0.3 ×g, frequency: 40 Hz, amplitude: ±50 μm, 30 min/12 h) on proliferation and osteodifferentiation of bone marrow-derived mesenchymal stromal cells (BMSCs) seeded on human bone-derived scaffolds. The scaffolds were prepared by partial demineralisation and deproteinisation. BMSCs were allowed to attach to the scaffolds for 3 days. Morphological study showed that spindle-shaped BMSCs almost completely covered the surface of bone-derived scaffold and these cells expressed higher ALP activity than those cultured on plates. After microvibration treatment, BMSC proliferation was decreased on day 7 and 10; however, numbers of genes and proteins expressed during osteogenesis, including Cbfa1, ALP, collagen I and osteocalcin were greatly increased. ERK1/2 activation was involved in microvibration-induced BMSC osteogenesis. Taken together, this study suggests that bone-derived scaffolds have good biocompatibility and show osteoinductive properties. By increasing the osteogenic lineage commitment of BMSCs and enhancing osteogenic gene expressions, microvibration promotes BMSC differentiation and increase bone formation of BMSCs seeded on bone-derived scaffolds. Moreover, ERK1/2 pathway plays an important role in microvibrationinduced osteogenesis in BMSC cellular scaffolds.Keywords: Bone-derived scaffold, bone marrow-derived mesenchymal stromal cells, microvibration, osteogenesis, ERK1/2. *Address for correspondence: Haiyang Yu West China Hospital of Stomatology Sichuan University Chengdu, 610041, P.R. China Telephone/FAX Number: 86-028-85502869 E-mail: yhyang6812@scu.edu.cn IntroductionCurrent consensus for bone tissue engineering includes three essential elements, i.e., biomaterial scaffold, osteogenic cell lineage and bone inducing factors (e.g., mechanical stimulus, Ashammakhi and Ferretti, 2003; Khan et al., 2005;Mistry and Mikos, 2005). Scaffold materials should provide the support for cell attachment and have osteoinductive property (Langer and Vacanti, 1993;Ashammakhi and Ferretti, 2003). Due to the limited supply and donor-site morbidity of autogenous bone grafts, different physical structures and insuffi cient osteoinductive ability of synthetic materials (Ashammakhi and Ferretti, 2003;Silber et al., 2003), scaffolds derived from different individuals (allografts) and species (xenografts) provide a promising resource and approach to address the signifi cant drawbacks of existing scaffolds, because these scaffolds have similar structures to autogenous bone (Salkeld et al., 2001;Simion et al., 2004). Additionally, with the proper chemical and physical process on these bone materials, including demineralisation and deproteinisation (Tadjoedin et al., 2003;Xu et al., 2003...

The major histocompatibility complex (MHC) is a fundamental part of the vertebrate immune system, and the high variability in many MHC genes is thought to play an important role in the recognition of parasites. Baiji (Lipotes vexillifer) is one of the most endangered species in the world. Its wild population has declined to fewer than 100 individuals and has a very high risk of becoming extinct in the near future. In this study we present a first step in the molecular characterization of a DQB-like locus of baiji by nucleotide sequence analysis of the polymorphic exon 2 segments. In the examined 172 bp sequences from a group of 18 incidentally captured or stranded individuals, 48 variable sites were determined and 43 alleles were identified, many of which were represented by only one clone. Three to seven alleles were found in each individual, suggesting gene duplications. No deletion, insertion, or exceptional stop codon was detected, suggesting these alleles function in vivo. Phylogenetic reconstruction using neighbor joining grouped the 43 alleles into two distinct lineages, differing by seven nucleotides and four amino acids. Substitutions of amino acids tend to be clustered around sites postulated to be responsible for selective peptide recognition. In the peptide-binding region (PBR) of the DQB locus, the average number of nonsynonymous substitutions per site is greater than that of synonymous substitutions per site (0.1962 versus 0.0256, respectively). Nucleotide and amino acid sequences both showed a relatively high level of similarity (nucleotides 90.6%; amino acids 80.6%) to those of beluga whale (Delphinapterus leucas) and narwhal (Monodon monoceros). The high level of baiji MHC polymorphism revealed in the present study has not been reported in other cetaceans and could be a consequence of the small baiji population adapting to freshwater with a relatively high level of pathogens.

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Wnt signaling pathway is essential for development and tumorigenesis, however, this signaling pathway in the progress of hepatocellular carcinoma (HCC) remains unclear. In this paper, we studied the function of human T-cell transcription factor-4 (TCF4), a key factor of Wnt signaling pathway, on the proliferation of HCC cell line. We showed that the expression of TCF4 mRNA in HCC cell line BEL-7402 was higher than that in immortalized normal liver cell line L02. Blockage of Wnt pathway by ∆NTCF4, a dominant negative TCF4, could suppress BEL-7402 cells growth and decrease the expression of cyclin D1 and c-myc, two of target genes of Wnt pathway. On the other hand, stimulating Wnt pathway by introducing a degradation-resistant β-catenin S37A could increase BEL-7402 cells proliferation. But all the treatments had no effect on L02 cells. Our data indicated that TCF4 might be another key factor in Wnt pathway involved in HCC cells proliferation and TCF4 could be an effective therapeutic target for suppressing the growth of hepatocellular cancers.

BackgroundInterleukin-7 receptor (IL-7R) is involved in the abnormal function of solid tumors, but the role and regulatory mechanisms of IL-7R in HBV-related hepatocellular carcinoma (HCC) are still unclear.MethodsGene and protein expression levels of IL-7R were examined in hepatoma cells transfected with hepatitis B virus (HBV) plasmids and in hepatoma cells transfected with the multifunctional nonstructural protein X (HBX). The expression of HBX and IL-7R was measured by immunohistochemical analysis in HBV-related HCC tissues. The role of NF-κB and Notch1 pathways in HBX-mediated expression of IL-7R in hepatoma cells was examined. Activation of IL-7R downstream of intracellular signaling proteins AKT, JNK, STAT5, and the associated molecules CyclinD1 and matrix metalloproteinase-9 (MMP)-9, was assessed in HBX-positive cells with or without treatment with IL-7R short hairpin RNA (shRNA). Additionally, the role of IL-7R in HBX-mediated proliferation and migration of hepatoma cells was investigated.ResultsThe expression of IL-7R was increased in hepatoma cells transfected with HBV plasmids; HBX was responsible for the HBV-mediated upregulation of IL-7R. Compared to adjacent tissues, the expression of HBX and IL-7R was increased in HBV-related HCC tissues. Additionally, the relative expression levels of HBX were associated with IL-7R in HBV-related HCC tissues. The activation of NF-κB pathways and expression of Notch1 were increased in hepatoma cells transfected with HBX, and inhibition of NF-κB and Notch1 pathways significantly decreased HBX-mediated expression of IL-7R. The activation of AKT and JNK and the expression of CyclinD1 and MMP-9 were increased in HBX-positive cells. When cells were treated with IL-7R shRNA, the activation of AKT and JNK, as well as the expression of CyclinD1 and MMP-9, were significantly inhibited. Additionally, IL-7R was responsible for HBX-induced proliferation and migration ability of hepatoma cells.ConclusionsOur data demonstrate that HBX can upregulate IL-7R via NF-κB and Notch1 pathways to facilitate the activation of intracellular pathways and expression of associated molecules, and contribute to proliferation and migration of hepatoma cells.

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Objectives Interleukin‐34 (IL‐34) is associated with hepatitis B virus (HBV) infection and hepatocellular carcinoma (HCC). However, the role and associated mechanisms of IL‐34 in HBV‐related HCC remain unclear. In this study, the expression, biological function and associated mechanisms of IL‐34 in HBV‐related HCC cells were investigated. Methods IL‐34 expression induced by HBV and HBV X (HBX) gene was measured in hepatoma cells. The role of CCAAT/enhancer‐binding protein α (CEBP/α) in HBX‐induced IL‐34 expression was examined. The signal pathways involved in the expression of CEBP/α and IL‐34 induced by HBX were assessed. The role of IL‐34 in the proliferation and migration of HCC cells, and related mechanisms were explored. Results Dependent on HBX, HBV increased IL‐34 expression in hepatoma cells, and HBX upregulated and interacted with CEBP/α to enhance the activity of IL‐34 promoters. CEBP/α mediated by HBX was associated with the activation of PI3‐K and NF‐κB pathways to promote IL‐34 expression. Via CSF1‐R and CD138, IL‐34 promoted the proliferation and migration of hepatoma cells, and contributed to the activation of ERK and STAT3 pathways and the upregulation of Bcl‐xl and c‐Myc mediated by HBX. Conclusion We demonstrate that IL‐34 contributes to HBX‐mediated functional abnormality of HCC cells and provides a novel insight into the molecular mechanism of carcinogenesis mediated by HBX.

We tested the ability of retrocyclin 2, a type of theta defensin, to protect cells and chicken embryos from infection by H5N1 highly pathogenic avian influenza virus. A gene fragment of retrocyclin 2 was designed based on the protein sequence of retrocyclin 2 and cloned into the eukaryotic expression vector pcDNA4.01 (HismaxA), named pcDNA4-RC2. The expression vector pcDNA4-RC2 protected MDCK cells and chicken embryos from infection by the H5N1 virus through inhibition of virus replication and viral mRNA transcription. Retrocyclin 2 is therefore effective in preventing H5N1 virus infection in vivo and in vitro and could be considered as a new therapy for H5N1 influenza and other diseases.