Influence of Bushen Huoxue decoction on beta-catenin, MMP-7 of synoviocytes in rats with knee osteoarthritis / 中国骨伤

<p><b>OBJECTIVE</b>To observe beta-catenin expression of Wnt signaling pathway in rats with knee osteoarthritis, and influence of Bushen Huoxue decoction on beta-catenin and MMP-7 expression of synoviocytes in rats with knee osteoarthritis (OA).</p><p><b>METHODS</b>Rats model with knee osteoarthritis were established by Hulth method. Primary synoviocytes and OA synoviocytes were cultured with collagenase digestion method. The cultured synoviocytes were divided into normal group, OA model group and Bushen Huoxue decoction group. Western blotting method was used to detect beta-catenin, MMP-7 protein expression of synoviocytes after acting by Bushen Huoxue decoction for 48 h; ELISA method was used to detect MMP-7 expression of synovial supernatant.</p><p><b>RESULTS</b>OA synoviocytes were cultured successfully. Western blotting showed that beta-catenin, MMP-7 expression in OA synoviocytes was significantly higher than normal group, Bushen Huoxue decoction could significantly reduce beta-catenin, MMP-7 expression; ELISA results showed that MMP-7 expression of OA synovial supernatant was significantly higher than normal synoviocytes supernatant, Bushen Huoxue decoction significantly regulated the level MMP-7 down.</p><p><b>CONCLUSION</b>(1) High expression of beta-catenin in OA synoviocytes indicates that Wnt classical signal pathway is activated in rat with knee osteoarthritis; (2) High expression of MMP-7 expression in OA synoviocytes confirms the MMP-7 is downstream genes of Wnt/beta-catenin signal pathway; (3) Activation of Wnt signal pathway and increase of MMP-7 may cause degradation of articular cartilage, and promote the formation of osteoarthritis; (4) Bushen Huoxue decoction can reduce expression of MMP-7, and promote cartilage repair, which may be one of mechanisms of osteoarthritis.</p>

Manifestation of synovial lesions in osteoarthritis / 中国骨伤

Osteoarthritis is a degenerative joint disease,with the characters of degradation of articular cartilage, the formation of the joint marginal osteophyte and synovium lesions. Previous studies have focused on the treatment of articular cartilage lesions. In recent years, new research in shows synovial inflammation plays an important role in OA. Synovium lesions and synovial inflammation-related factors induced the degradation and destruction of articular cartilage, and promoted the development of osteoarthritis. The role of synovial lesions in osteoarthritis is increasingly prominent, and the treatment for synovial lesions will become a new target. So this paper reviews the various manifestations of synovial in osteoarthritis.

Experimental study on co-culture of early embryo and cancer cells in vitro / 生物医学工程学杂志

The co-culture system of early embryos and cancer cells is an important means to observe the biological behavior changes of embryos and cancer cells in vitro. In this study, we co-cultured the 3.5 dpc mouse embryo with malignant tumor cells, investigated the development of blastocyst by observing the hatchment, attachment and outgrowth, observed the biological behavior changes of cancer cells in the embryonic circumstances, and detected the proliferation and apoptosis of cancer cells. Compared with the control, the embryos developed normally in the tumor environments, and the rate of hatchment, attachment and outgrowth increased significantly (P<0.05). However, there was no significant change of cancer cells in morphology, proliferation and apoptosis in the co-culture system (P>0.05). Under the co-culture system, the early embryo developed normally, and the cancer cells also grew well. There may be similarities between the embryos and cancer cell's choice for living. Moreover, the growth of embryos could be promoted by cancer cells in the co-culture system. This might be related to the similarities of gene expression, growth factors and signal transduction mechanisms between embryos and cancer cells.

Synthesis and characterization of thermosensitive hydrogel based on PEG-PCL-PEG block copolymers / 生物医学工程学杂志

In this paper, a series of low-molecular-weight PEG-PCL-PEG triblock copolymers were successfully synthesized by ring-opening polymerization method, and were characterized using 1H-NMR and FTIR. The aqueous solution displayed specific thermosensitive gel-sol transition when the concentration was above corresponding critical gel concentration (CGC). The gel-sol phase diagram was recorded using test tube-inverting method, which was depended on the hydrophilic/hydrophobic balance in macromolecular structure, as well as heating history. As a result, the gel-sol transition temperature range could be altered, which might be very useful for its application as injectable drug delivery system.

Searching for genes interacting with human PCIA1 gene by using the bacterial two-hybrid system / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To search for the genes which could interact with newly found homo sapiens cross-immune reaction antigen (PCIA1) gene and accordingly to provide insights into the study of the gene function.</p><p><b>METHODS</b>The Stratagene's BacterioMatch Two-Hybrid System and BacterioMatch Fetal Kidney Library were adopted and the recombinant bait plasmid pBT-PCIA1 was cotransformated with the target plasmid pTRG-cDNA library DNA into the reporter stain. After screening and isolation of positive pTRG clones, the target genes were identified by DNA sequencing and bioinformation analysis.</p><p><b>RESULTS</b>Among all the seven detected target genes, three genes' function were not known, the other four genes had important functions. Their mutations or abberant expression resulted in severe diseases and overexpression of ACTN4 (actinin, alpha 4), PSAP (prosaposin) or EIF3S10 (eukaryotic translation initiation factor 3, subunit 10 theta) could promote tumor development and progression.</p><p><b>CONCLUSION</b>The bacterial two-hybrid system technique is an efficient method, which can provides insights into the study of novel genes' function by detecting protein-protein interactions. This study indicates that PCIA1 gene expression correlates with tumor formation, invasion and metastasis.</p>

Prokaryotic expression, purification, refolding and biological assays of recombinant human interleukin 4 inclusion body / 生物医学工程学杂志

A DNA fragment encoding human interleukin 4 was obtained by PCR from pORF-hIL4 plasmid. The amplified fragment was inserted into prokaryotic expression vector PQE60 and recombinant protein was expressed in E. Coli M15 by adding isopropyl-beta-D-thiogalactoside (IPTG). The hIL-4 protein was present as insoluble inclusion bodies in the bacterial extract. After denaturation of inclusion bodies with 5 mol/L guanidine hydrochloride, the supernate was diluted to get renaturized. Then dialysis and Ni chelating chromatography were used for purification. TF-1 proliferation assay of recombinant human interleukin 4 was performed, and then rhIL-4 was fit to be used for proliferation of human dendritic cells from monocyte in vitro.

Specific inhibition of Raf-1 gene expression in HNE1 by RNA interference / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To silence the expression of Raf-1 gene in HNE1 cells using vector-based RNA interference (RNAi) technique.</p><p><b>METHODS</b>The vector containing the human U6 promoter was used for targeted gene silencing when a dsDNA oligonucleotide encoding an appropriate shRNA was ligated into the vector, and 67nt oligonucleotide fragment was inserted into the downstream of the U6 promoter. Plasmids containing different Raf-1 target sequences [ (1) pshuttle-Raf-1-a( 225), (2) pshattle-Raf-1-b ( 358) and (3) pshuttle-Raf-1-c(474)], were transfected into HNE1 cells. Expression of Raf-1 mRNA was assayed by RT-PCR. Apoptosis were determined by cytometry.</p><p><b>RESULTS</b>Vector-based RNAi had advantages over antisense RNA because it could be delivered to the target cell more efficiently, and effect could last longer. Raf-1 expression could be inhibited by plasmid-expressed shRNA. Three different targeting sequences were selected from Raf-1 gene, and the inhibitory effect of pSIREN shuttle-Raf-1-b (358) was biggest.</p><p><b>CONCLUSION</b>Raf-1 expression in HNE1 cells can be inhibited significantly using plasmid-based RNAi.</p>

Construction of the recombinant adenovirus vector expressing human CD40 ligand gene / 华西口腔医学杂志

<p><b>OBJECTIVE</b>To construct a recombinant adenovirus vector expressing hCD40L gene and explore it in the use of anti-tumor gene therapy.</p><p><b>METHODS</b>1,900 bp gene fragment was obtained form plasmid pORF-hCD40L by Xho I/Swa I cutting and then cloned directionally into the pShuttle plasmid, finally, the resultant plasmid was digested by restriction endonnuclease PmeI and subsequently cotransformtion into BJ5183 cells with the adenoviral backbone pAdEasy-1 to obtain the homologous recombinant and then the recombinant was packaged in the 293 cells. Some methods such as PCR and endonulease digestion were employed to identify the recombinant adenovirus.</p><p><b>RESULTS</b>The evidences of endonulease digestion and PCR analysis confirmed that recombinant hCD40L gene was correctly inserted into adenovirus vector.</p><p><b>CONCLUSION</b>The adenoviral vector which expressed hCD40L gene was constructed. It provides an experimental basis for studies on it expression in the mammalian cells and in tumor gene therapy.</p>

Inhibition of mouse Lewis lung cancer via intravenous administration of recombinant mouse sFlt1 adenovirus / 中国肺癌杂志

<p><b>BACKGROUND</b>It has been known that the growth of solid tumors is dependent on angiogenesis, and neoangiogeneses of tumor become main target to control tumor growth. The aims of this study are to investigate the inhibition effect of replicate-deficient adenovirus encoding the soluble form of mouse vascular endothelial growth factor receptor 1 (sFlt1-Adv) on angiogenesis and tumor growth in established tumor model.</p><p><b>METHODS</b>Mouse Lewis lung cancer cells were inoculated subcutaneously into C57 mice. sFlt1-Adv, GFP-Adv and normal saline were injected twice intravenously after establishing Lewis cancer model. Diameters of tumors were measured every other day. Tumors were resected, weighed and fixed in 3% paraformadehyde. Microvessel density of tumors was determined by immunohistochemical staining with anti-CD31 antibody.</p><p><b>RESULTS</b>The planted tumor volume and weight in sFlt1-Adv group were significantly lower compared with the two controls (P < 0.01). Its inhibition rate was 71.8%. The microvessel density in sFlt1-Adv group decreased markedly compared with that of the control groups (P < 0.01).</p><p><b>CONCLUSIONS</b>sFlt1-Adv can inhibit the growth of tumor through the inhibition of tumor angiogenesis. sFlt1-Adv may be potentially valuable for clinical treatment of solid tumor.</p>

Screening and identification of stable transfectants of mouse soluble B lymphocyte stimulator / 生物医学工程学杂志

Mouse colon cancer cells CT26 were transfected with constructed plasmid expressing mouse soluble B lymphocyte stimulator (msBlyS). Single cell clones were selected with 100 microg/ml Zeosin and subcloned by serial limiting dilution. Eight resistant transfectants were isolated and expanded, and five of them displayed the desirable msBlyS cDNA band amplified by semi-quantitative RT-PCR assay. Western blot analysis showed that only msBlyS molecules of the expected size were detected in the cell lysates from transfectants. The supernatant of transfectants could costimulate B cell proliferation in standard costimulation assay. Thus we have successfully screened the stable transfectants expressing high levels of msBlyS in CT26 cells, which could be used as cancer vaccines for further anti-tumor immunotherapy.