Background Thyroid associated ophthalmopathy (TAO) is an autoimmune disease.Current research on the pathogenesis focuses on common autoantigen.Insulin-like growth factor-1 receptor (IGF-1R) is necessary for the function of IGF-1,also IGF-1 plays an important role in signaling pathway of thyroid stimulating hormone receptor (TSHR).Objective This study was to investigate the effects of IGF-1 on the proliferation,expression of IGF-1R and TSHR on cultured orbital fibroblasts (OFs) derived from TAO.Methods Human orbital tissue was obtained from 17 TAO patients who received orbital adipectomy and 4 normal controls who received cosmetic surgery in West China Hospital from March 2016 to June 2016.OFs were cultured by explant culture with DMEM/F12 containing 5％ fetal bovine serum and identified by immunochemistry.The OFs were treated with different concentrations of IGF-1.IGF-1 at different concentrations (0,50,100,125 μg/L) was added into the medium,respectively,and the proliferation of the cells (absorbancy) was detected by MTS.The percentages of IGF-1R and TSHR expressions in the cells were assayed by flow cytometry.Results Cultured cells appeared to be spindle-like in shape and grew well with abundant cytoplasm.The characteristics of the cells derived from TAO patients were consistant with normal ones.The cells showed the positive response for vimentin and absent respose for desmin,S-100,myoglobin and cytokeratin.The proliferative values of OFs were gradually elevated with the increase of IGF-I dose in both TAO group and normal group (Fgroup =219.639,P＜0.001;F ion =17.752,P＜0.001) with the optimal effects in 100 μg/L IGF-1.The expression levels of IGF-1R in the OFs were (0.009 1 ±0.008 7)％,(0.095 3±0.023 3) ％,(0.083 7±0.022 7) ％ and (0.070 9 ± 0.024 1) ％ in the TAO group,and those in the normal group were (0.0023± 0.0006)％,(0.0093±0.0012)％,(0.0073±0.0015)％ and (0.0083±0.0012)％ after treatment of 50,100,125 μg/L IGF-1.The expression levels of IGF-1 R were significantly higher after treatment of 50,100 and 125 μtg/L IGF-1 than those treatment of 0 μg/L IGF-1 in both TAO group and normal group,and the expression levels of IGF-1R in the OFs were significantly increased in the TAO group compared with the normal group (all at P＜0.05).No statistical difference was seen in the TSHR expression between the TAO group and normal group after treatment of 0,50,100 and 125 μg/L IGF-1 (Fgroup =0.133,P ＞ 0.05;F ion =0.004,P ＞ 0.05).Conclusions IGF-1 can promote the proliferation of OFs and up-regulate the expression of IGF-1R in OFs.However,IGF-1 dose not play a regulating effect on the expression of TSHR in OFs.

AIM: To observe whether arachidonic acid (AA) could induce apoptosis in mouse fibroblast cell line L929 and the potential mechanism involved. METHODS: The viability and damaged degree of L929 was monitored by MTT and the release of lactate dehydrogenase (LDH). Lipid peroxidation in L929 was measured as malondialdehyde (MDA) content by colorimetric assay. Hoechst 33258 staining was used to observe AA-induced morphological changes. Agarose gel electrophoresis was used to detect DNA fragmentation. RESULTS: Treatment of L929 cell with AA for 24 h, in the range of 40-160 ?mol/L, caused a great decrease in cell survival and increased MDA contents and the release of LDH simultaneously( P

OBJECTIVE To describe the clinical features of the parapharyngeal space tumors and assess the postoperative complications and outcomes in our hospital.METHODS The clinical data of 135 cases with parapharyngeal space tumor treated from Jan.1995 to Dec.2005 in our hospital were retrospectively studied.RESULTS It included 24 heterogeneous histologies in this group.Neurogenic tumors(72.6 %) were the most common tumors,next were salivary gland tumors(15.6 %),and others 11.8 % tumors were miscellaneous tumors.There were 121(89.6 %) patients with benign lesions and 14(10.4 %) with malignant tumors.Transcervical approach was the most commonly applied route.Only 4 cases recurred in 113 operated benign patients.At end of the follow-up,of 14 patients with malignant tumors,4(28.6 %) were alive with no evidence of disease,5(35.7 %) were alive with disease,5(35.7 %) died of the diseases.CONCLUSION Primary parapharyngeal space neoplasms are rare and the majority of these tumors are benign.Surgery is the mainstay of treatment for parapharyngeal space tumors.Most benign cases with a low rate of complication and recurrence after operation,but malignant neoplasms have a poor prognosis.

Rv2742 is a novel gene identified from Mycobacterium tuberculosis H37Rv by the proteogenomics strategy. The aim of this study was to establish a system of soluble expression and purification of the missing protein Rv2742 in M. tuberculosis H37Rv, to provide reference for further research on the biological function of Rv2742. The soluble protein was not successfully induced by prokaryotic expression vectors pGEX-4T-2-Rv2742, pET-32a-Rv2742, pET-28a-Rv2742 and pMAL-c2X-Rv2742. After the codon of novel gene Rv2742 was optimized according to E. coli codon usage frequency, only the recombinant strain containing plasmid pMAL-c2X-Rv2742 could produce soluble products of Rv2742 encoding gene. In addition, the expression effects of the desired fusion protein were also analyzed under different conditions including hosts, culture temperatures and IPTG concentrations. The optimum expression conditions were as follows: Rosetta (DE3) host, 16 °C culture temperature and 0.5 mmol/L IPTG. After being purified by affinity chromatography with amylose resin, the fusion protein sequence was confirmed by LC-MS/MS. These results indicated that the novel gene Rv2742 product could be successfully induced and expressed in a soluble form by the expression system pMAL-c2X with MBP tag. Our findings provide reference for studies on potential interaction and immunogenicity.
Chromatography, Liquid ; Cloning, Molecular ; Escherichia coli ; Mycobacterium tuberculosis ; genetics ; Recombinant Fusion Proteins ; Tandem Mass Spectrometry