As one of the most sensitive cells of endoplasmic reticulum stress,pancreatic βcells have an abundance of endoplasmic reticulum .It is the adaptive response of pancreatic β cells under Endoplasmic reticulum stress conditions.Internal and external factors in the environment such as oxidative damage,lipid toxicity and the effect of cytokines in the endoplasmic reticulum could breake the steady-state,resulting in barriers to folding or misfolding of protein,thereby to trigger endoplasmic reticulum stress.Serious and lasting stress will lead to β cell apoptosis,involved in a variety of metabohc diseases as well as a variety of diseases.This article gives a review on the injury effects of multi-factor induced endoplasmic reticulum stress on the pancreatic β-cell

Objective To establish a stable and useful method for culturing human osteoclast-like cells in vitro,and investigate the effect of 1?,25-(OH)2D3,M-CSF and PGE2 on osteoclasts differentiation,proliferation and activation so as to lay the foundation for further study of the biological mechanism for tooth movement.Methods The HCMNC were isolated and cultured in 24-well plate with coverslips and human dentine slices.The experiment group was cultured with 1?,25-(OH)2D3,M-CSF and PGE2,respectively,while the control group was not.The liquid was changed every 3 days and the whole culture process lasted for 7 days.The phase contrast microscopy and TRAP staining were adopted to identify osteoclast-like cells.Results On the 3rd day the monocytes began to fuse and on the 7th day positive multinucleated cells could be seen with TRAP staining,but absorption pit was not formed on the dentin slices.The group with 1?,25-(OH)2D3 had the largest number of osteoclast-like cells.Conclusion After the monocytes in UCB are cultured by 1?,25-(OH)2D3,M-CSF,PGE2 induction,they can turn into TRAP(+) multinucleate osteoclast-like cells,the 1?,25-(OH)2D3 10-8mol/L being the most effective.

Objective To construct anti-IL-4R murine anti-human single-chain variable fragment (scFvs) antibodies through BL21 (DE3) prokaryotic expression system. Methods The anti-IL-4R scFv sequence was optimizated on the basis of previous findings. The optimized scFv sequence was analyzed. The recombinant plasmid pET-32a-scFv was constructed. The recombinant plasmid was detected through enzyme identification, and was turned into BL21 (DE3) prokaryotic expression bacteria to express the pET-32a-scFv recombinant protein in E.coli BL21 (DE3). The purification and renaturation were researched, and SDS-PAGE analysis was studied. The molecular weight of ScFv against IL-4R was analyzed by SDS-PAGE. The expression of the fusion protein was detected by Western-blot assay. Results The length of fusion gene scFv-MLT sequence was 761 bp. The molecular weight of the recombinant expression of proteins of anti-IL-4R single antibody was approximately 45 ku. The recombinant proteins showed high specificity with anti-6 × His-tag antibody. Conclusion This experiment successfully constructs pET-32a-scFv prokaryotic expression system of recombinant protein with high immune reactivity, which provides the basis for further study of anti-IL-4R single chain antibody as drug target.

BACKGROUND: Orthodontic tooth movement is dependent on reconstruction of periodontium. Osteoclastic bone resorption is the first step of tooth movement. The present study hotspots focus on signal transduction pathway regarding osteoclast differentiation and functional development under stress and on the relationship between periodontal ligament cells and osteoclasts. OBJECTIVE: To set up a simple method to in vitro culture human osteoclast-like cells and to observe the effects of bone resorption-stimulating factors on differentiation, proliferation, and function of osteoclast-like cells, DESIGN, TIME AND STTING: A cytological in vitro controUod observation was performed at the Central Laboratory,Stomatology Hospital, Xi'an Jiaotong University between October 2007 and May 2008. MATERIALS: Umbilical cord blood was sourced from the healthy puerperae who had not suffered from high-risk pregnancy. Freshly prepared fetal femur provided by Laboratory Animal Center, Xi'an Jiaotong University and were used for preparation of bone flaps at 100-200 μm thickness. 1α ,25-(OH)2D3, macrophage colony-stimulating factor (M-CSF), prostaglandin E2 were purchased from Sigma Company, USA.METHODS: Under aseptic condition, umbilical cord blood was collected. Following Ficoll solution separation and centrifugation, supematant was discarded. Umbilical cord blood-derived mononuclear cells were suspended with o -modified minimal essential medium (α-MEM) solution and then inoculated into a 24-well culture plate, in which, coverslips and femoral slices were pre-placed, at a density of 1×109/L, 1.0 mL per well. Five groups were set, blank control, 108 mol/L 1α ,25-(OH)2D3, 10-7 mol/L 1α ,25-(OH)2D3, macrophage colony stimulating factor (M-CSF), and 1α ,25-(OH)2D3+prostaglandin E2 (PGE2). Each group was cultured for 7 days. MAIN OUTCOME MEASURES: Cellular growth morphology was observed under an inverted microscope; osteoclast-like celt formation was examined by tartrate-resistant acid phosphatase (TRAP) staining; and osteoclastic Howship's lacuna was detected by toluidine blue staining. Any cell with TRAP-positive staining and more than two nuclei was considered osteoclast-like cell and counted. RESULTS: After 3 days of culture, cells from the blank control group did not exhibit apparent changes in morphology and quantity. In the remaining groups, mononuclear cells appeared with confluent tendency. After 7 days of culture, a small number of osteoclast- like cells with 2-3 nuclei were found in the blank control group; a great many of multinucleated osteoclast- like cells with 3-20 nuclei were present in the remaining groups. Through the use of optical microscope, osteoclast-like cells could be found for the presence of red cytoplasm, bright yellow nuclei, and TRAP-positive staining in each inducing factor-treated group, in particular in the 108 mol/L 1α ,25-(OH)2D3 group, which displayed osteoclast- like cells exhibiting 14 nuclei, strong TRAP-positive staining, and a relatively big cell body. But no osteoclastic Howship's lacuna was found in any group. Compared to the blank control group, the numbers of osteoclast-like cells were greater in each inducing factor-treated group (F = 9.78, P < 0.01). There was no significant difference in the numbers of osteoclast-like cells between the 108 mol/L 1α ,25-(OH)2D3 and the 10-7 mol/L 1 a ,25-(OH)2D3 groups (P0.05). The M-CSF group and the 1α ,25-(OH)2D3+PGE2 group exhibited significantly less numbers of osteoclast-like cells than the 108 mol/L 1α ,25-(OH)2D3 group (F= 7.46, P< 0.01). CONCLUSION: After in vitro culture of 1α ,25-(OH)2D3, M-CSF, and PGE2, umbilical cord blood-derived mononuclear cells can differentiate into TRAP-positive multinucleated osteoclast-like cells, the 10-8 mol/L 1α ,25-(OH)2D3 being the most effective.

Objective MiRNA-10b is an important member of the MiRNA family , which has been proven that miRNA-10b can promote the growth and invasion of a variety of tumor cells .The aim of this study was to to investigate the effect and mechanism of miR-NA-10b on proliferation and invasion of low metastasis of lung cancer cell line (95-C). Methods The recombinant of miRNA-10b was transfected into 95-C by lipofectin method .The experiment set up 3 groups:blank control group , negative control group and miRNA-10b expression plasmid transfected group .MiRNA-10b expression level and KLF4mRNA expression level were detected by real-time fluores-cence quantitative PCR ( RTFQ-PCR) .The cell proliferation was detected by cell proliferation assay .The invasive ability of cell was de-tected by Transwell experiment .The expression of KLF4 protein was assessed by Western blot . Results At 48 hours after transfection, compared with blank control group (1.01 ±0.08) and negative control group (0.86 ±0.07), the miRNA-10b expression level in miRNA-10b expression plasmid transfected group (1.61 ±0.12) increased significantly (P<0.05) and there was no statistical difference between blank control group and negative control group (P>0.05).From the growth curve, the cell proliferation rate was obviously increased in miRNA-10b expression plasmid transfected group ([188.0 ±15.1]/HP) compared with the other two groups ([151.0 ±11.3]/HP), ([136.0 ±10.8]/HP) (P <0.05) and there was no statistical difference between these two groups ( P >0.05 ).Transwell result showed more cells transferred to the other side of Transwell compared with the other two groups ( P <0.05 ) and there was no statistical difference between these two groups (P >0.05).The expression of KLF4 protein decreased in miRNA-10b expression plasmid transfected group compared with the other two groups ( P<0.05).KLF4mRNA expression decreased, but the difference had no statistical significance (P>0.05). Conclusion MiRNA-10b might promote the pro-liferation and invasion of 95-C through down-regulation of KLF4 protein expression .

Objective By culturing the osteoclasts together with the osteoblasts directly to investigate the effect of osteoblasts on the formation of mature osteoclasts.Methods The bone marrow mononuclear cells of rats were treated with 30?g/L M-CSF and 50?g/L RANKL and cultured for 6 days.Subsequently,the primary osteoblasts which were of the same quantity as the osteoclasts were co-cultured directly.In the co-culture system,we added the liquid containing 1,25-(OH)2D3 1?10-8mol/L and PGE2 1?10-6mol/L.The morphological observation,TRAP staining and pit staining were adopted to identify osteoclasts.Results When the osteoclasts were co-cultured with primary osteoblasts,the growth of osteoblasts had more preponderances.After staining,we could see more osteoblasts than osteoclasts.Conclusion The relationship between osteoblasts and osteoclasts is related to the relative quantities of the two cells.When osteoblasts outnumber osteoclasts,osteoblasts would inhibit the formation and differentiation of osteoclasts.

Objective To evaluate endoscopic ultrasonography (EUS) in assessing endoscopic re-sectability of early esophagogastric cancer. Methods Data of 65 consecutive cases suspected as having early esophagogastric cancer with EUS and confirmed by pathology after endoscopic mucosal resection or endoscop-ic submucosal dissection within 2 weeks were retrospectively analyzed. The invasion depth of the lesion de-tected by EUS was compared with that from pathology. Results The accuracy of EUS in diagnosis of depth of early cancer invasion was 93.8% (61/65), and the overall accuracy in assessment of endoscopic resectabili-ty was 89. 2%. Conclusion Endoscopic resectability of early esophagogastric cancer can be accurately eval-uated with EUS.

Objective To evaluate in vitro anti tumor effect of host lymphocyte primed by CalmetteGuerin bacillus (CGB) activated dendritic cells (DC) based PANC1 lysate. Methods DCs were obtained from peripheral blood mononuclear cells of healthy volunteer and cuitured by rhGM CSF and rhIL 4. DC vaccines for pancreatic cancer were loaded with PANC1 tumor lysate (TL) and were further maturated by CGB.CD1a, CD83, CD86 and HLA-DR phenotype was characterized by flow cytometer, and IL-12p70 and TNF-α concentration in DC culture supernatant were measured by ELISA. Autologous mixed lymphocyte proliferation and the cytotoxicity of cytotoxic T lymphocytes primed by activated DCs to PANC1, PaTu8988 and SCG7901 tumor cells was measured by CCK 8 test. Results When DCs based PANC1 lysate were activated by CGB,the expression rates of CD83 and CD86 were increased from (3.7±0.3)% and (38.6±5.0)% to (16.5±0.6)% and (76.6±2.5)% (P ＜0.05 ). The concentrations of cytokines ILpl2p70 and TNF-α were increased from (20.18±2.06 ) pg/mland (61.38±1.19) pg/mlto (62.48±3.80) pg/mland (592.53±17.96)pg/ml (P＜0. 01 ). When co-cultured with CGB activated DCs based PANC1 lysate in proportion of 0.40)% , (3.39±1.05)% , (2.82±0.39)% significantly increased to (55.38±3.58)% , (75.0±2.54) % , (77.07±3.4)% , (99.07±2.4)% (P＜0.01) , respectively. The killing effects of lymphocytes 2.77)%, (19.03±3.04) %; but the killing effects on PaTu8988 and SCG7901 were significantly decreased.Conclusions DC vaccines for pancreatic cancer could be more maturated when activated by CGB, and could show a high capability of anti-tumor in vitro.

Objective To analyze the characteristics of benign and malignant breast tumors with 2D and color Doppler ultrasound, and to assess the value of color Doppler ultrasound in the diagnosis of breast cancer. Methods A total of 674 patients (327 malignant and 347 benign) of breast tumor underwent 2D and color Doppler ultrasonogarphy. Two-dimensional ultrasound was used to observe the size, form, margin and internal echo of the tumors;color Doppler was performed to observe the degree of blood flow signal in the tumor, and to measure peak systolic velocity (PSV) and resistance index (RI). Results Totally 671 patients were diagnosed with ultrasound. The size of the tumors were from 0.50 cm×0.41 cm to 5.42 cm×4.10 cm. The ratio of vertical and transverse diameter of 69.11% (226/327) of the malignant tumors ≥1.0. Most tumors (266/327, 81.35%) presented with irregular margin like incised or feet of crab;61.47% (201/327) had microcalcification. Color Doppler found that 92.97% (304/327) of the tumors had blood flow signal;PSV was 15.34-39.76 cm/s, RI was 0.65-0.98, and 91.61% (262/286)≥0.70. Significant differences of the ratio of vertical and transverse diameter, the margin of the tumor, microcalcification and blood flow signal, PSV and RI (P＜0.01) were found between benign and malignant breast tumors. Conclusion The diagnosis and differential diagnosis of benign and malignant breast tumors can be significantly improved with comprehensive analysis of 2D ultrasound, blood flow signal, PSV and RI. Color Doppler ultrasound plays an important role in the diagnosis of breast cancer.

BACKGROUND:Diabetic foot ulcers threaten the patients’ health and even survival seriously. It is an international difficult problem and lacks an effective treatment. But gene therapy and stem celltherapy possess special advantages and potential in wound healing. OBJECTIVE:To assess the therapeutic effect of transplantation of bone marrow mesenchymal stem cells transfected by human vascular endothelial growth factor 165 (hVEGF165) gene on foot wound healing in diabetic rats. METHODS:Recombinant adenovirus was established in vitro which expressed hVEGF165 gene and transfected into the third generation of bone marrow mesenchymal stem cells. Total y 120 male Wistar rats were divided into five groups:group A (non-diabetic controls), group B (diabetic controls), group C (Ad-hVEGF165 therapy), group D (stem celltherapy) and group E (transplantation of bone marrow mesenchymal stem cells transfected by Ad-hVEGF165 gene). Rats in the latter four groups were intraperitoneal y injected with streptozotocin to induce diabetic models. In al rats, a 3 mm×7 mm rectangular ful-thickness skin sample was cut from the instep of the hind foot to make a model of foot wound. The rats were subcutaneously injected at equidistant six points 5 mm distal to the wound edge on the dorsum of the foot:50μL PBS per point for group A, 50μL adenovirus suspension (1×1013 pfu/L) per point for group C, 50μL stem cellsuspension (1×1010/L) per point for group D, and 50μL adenovirus suspension+50μL stem cellsuspension per point for group E. RESULTS AND CONCLUSION:After injection, the rate of wound healing, the expression of VEGF and the qualities of capil aries in group E were higher when compared with groups B, C, D (P<0.05), but were lower than those in group A (P<0.05). Transplantation of bone marrow mesenchymal stem cells transfected by hVEGF165 gene can promote foot wound healing, angiogenesis and expression of VEGF in diabetic rats.