Objective To establish human renal cell carcinoma (RCC) cell lines,and to investigate the cell phenotypes and expression of tumor associated antigens. Methods Fresh tumor tissues from pathologically proven human RCCs were primary cultured and passed generation to generation until a stably grow in vitro.The cell cloning form,chromosome and graft into nude mice in vivo were examined for cell lines.Immunofluorescent stain and flow cytometric analysis were carried out for cell's phenotype,RT PCR examination for MN/CA9 gene expression. Results Six human RCC cell lines have been established,including four of the cell lines derived from clear cells,one from mixed clear granular cells and one from papillary cells.All the cell lines showed the characteristics of malignant cells.All the lines highly expressed HLA ClassⅠ and HER2/neu.Three lines weakly expressed HLA ClassⅡ and one cell line highly expressed CD86 but all the lines did not express CD80.RT PCR showed that three cell lines have the expression of MN/CA9 gene. Conclusions These newly established RCC cell lines would provide a useful in vitro model for studies related to biological characteristics,tumor associated antigen,immunogenity and immunotherapy of human RCC.

ObjectiveTo provide experimental proof for the specific immunotherapy in patients with renal cell carcinoma (RCC).MethodsThe expression of c-erbB-2 and HLA-A2 molecules in RCC were examined by flow cytometery and HLA-A2 cDNA RT-PCR.Tumor lysatic antigens (Tuly) was used to load DCs (DC-Tuly) to induce the generation of c-erbB-2 specific CTLs derived from peripheral blood mononuclear cells of RCC patients in vitro and CTLs was used to kill RCC cells which express or not express c-erbB-2 and HLA-A2.The cytotoxic activity of induced CTLs was further studied by antibodies (anti-c-erbB-2 and anti-CD 8) blocking assay.Resultsc-erbB-2 protein is tumor associated antigens for RCC.DC-Tuly induced specific CTLs highly killed c-erbB-2+ HLA-A2+ autologous and allogenic RCC,but non-specific ones lowly killed c-erbB-2- HLA-A2+ or c-erbB-2+ HLA-A2-RCC cells. The cytotoxic activity against tumor cells was blocked by anti-c-erbB-2 and anti-CD 8 monoclonal antibodies.ConclusionsThese results suggest that this specific CTLs adoptive immunotherapy for RCC patients with c-erbB-2+ HLA-A2+ could be a novel approach for clinical use.

Objective To investigate the secrection of Polytuftsin, a macrophage activator in Pichia pastoris expression system. Methods The recombinant plasmid encoding polytuftsin was transformed into the yeast by chemical method, and the positive clones were screened and expressed under the induction of methanod. It was observed that the influence of polytuftsin on macrophage in vivo and in vitro. Results It was verified that the recombinant products was confirmed to be our target protein by SDS-PAGE analysis and PCR assay. The expressed product could activate macrophage to secret cytokine and enhance the capacity of macrophages to kill L1210 tumor cells, the maximum killing rate is up to 40 %. In vivo the leukemia therapeutic trial, the time of tumor arise was delayed; the survival time of L1210 tumor-bear mice treated with Polytuftsin was extended, which is about 20 day. Conclusion The recombination protein polytuftsin, which could activate macrophage and has anti-tumor activity.

This study was aimed to explore the mechanism of Xin-Feng capsule (XFC) on pulmonary function of Sjogren's syndrome (SS) rat model based on the TGF-β1-ERK1 signal pathway. A total of 50 SD rats were ran-domly divided into the normal control (NC) group, model control (MC) group, hydroxychloroquine sulfate (HCQ) group, radix paeoniae alba (total glucosides of paeony capsules, TGP) group, and the XFC treatment group, with 10 rats in each group. Except the NC group, the complete Freund's adjuvant (CFA) and the same rat sub-mandibular gland antigen induced method were applied to each rat after metatarsus injection on two feet and CFA after sufficient emulsification of 0.2 mL submandibular gland protein mixed antigen induced SS. The water drink-ing quantity, body weight change, pulmonary function, ERK1, TGF-β1 protein expression and serum cell factor (IL-17, IL-4) changes were detected in each group. The results showed that compared with the NC group, the body quality and serum IL-4 of MC rats were obviously decreased, and the water drinking quantity, submandibu-lar gland / lung index, submandibular gland pathological score, ERK1, TGF-β1 integral and IL-17 were increased (P < 0.01 or P < 0.05), and the pulmonary function parameter was decreased (P < 0.01 or P < 0.05). Compared with the MC group, the body quality, pulmonary function parameters FEF50 and MMF of the XFC group were in-creased, the IL-4 expression was increased, the water drinking quantity, submandibular gland / lung index, sub-mandibular gland pathological score, serum IL-17 expression, ERK1 and TGF-β1 integral were decreased (P <0.01 or P < 0.05). Compared with the HCQ group, the body quality and IL-17 were significantly decreased in the XFC group (P < 0.01), FEF25, FEF75 and MMF were increased (P < 0.01 or P < 0.05). Compared with the TGP group, the lung index and IL-17 were decreased in the XFC group (P < 0.01 or P < 0.05). It was conclud-ed that the lung function of SS rats was declined, which is closely related to the activation of TGF-β1-ERK1 sig-nal pathway. Traditional Chinese medicine of XFC is able to downregulate the TGF-β1 expression and to restrain epithelium-mesenchymal cell transformation, in order to inhibit ERK1 phosphorylation activation and proliferation, decrease immune inflammation, in order to improve SS and lung function.

Objective To observe the clinical effect and comfortable degree of the mode of super hair removal. Methods The mode of super hair removal was used to depilate the hair nearby the hair line, cheeks, upper lip, beard, ventrum, areola of breast, axillary cavity, extremities, bikini area and so on. The total number of sites was 1 000. Some sites that were especially susceptible to pain, for example, upper lip and buccal region, were smeared with compound lidocaine cream for 1 hour at least before treatment. Results Hairs in the areas of extremities, ventrum, back and axillary cavity generally needed 4 to 5 times to eradicate, and the patients had no evident discomfortableness; hairs near to the upper lip and lower mandible generally needed 5 to 7 times to reach the effect which the patient was content, and anesthetics was indispensable, or the patients would present discomfortableness. Conclusions The mode of super hair removal is more effective, quicker and more comfortable in comparison with conventional methods. Therefore, it deserves to be spread.

Objective To observe the changes of lung parameters,the ratios of B and T lymphocyte attenuator(BTLA) and serum cytokines in patients with knee osteoarthritis (KOA),and explore possible molecular mechanism of them.Methods Forty-seven cases of knee osteoarthritis from the First Affiliated Hospital of Anhui University of Traditional Chinese Medicine from 2011 October to 2012 July were analyzed in this study.Pulmonary parameters were detected by spirometer; B and T lymphocyte attenuator(BTLA) was detected by flow cytometry ; Interleukin (IL)-1β,IL-10,matrix metalloproteinase-9 (MMP-9),tissue inhibitor of matrix metalloprotease-1 (TIMP-1) were detected by ELISA;ESR was determined by Westergren method ; hs-CRP was determined by the automatic biochemistry analyzer.Results (1) Compared with NC group,levels of FVC,FEV1,FEV1/FVC,PEF,MEF25.75,MEF50,MEF25,CD3 + BTLA+ T cell,CD4+ BTLA+ Tcell,IL-10,TIMP1 were significantly decreased,IL-1 β,MMP9 were significantly increased.(2)Correlation analysis showed FVC was negatively correlated to Lequesne MG,symptom classify quantization scores,course,MMP9,while positively related to CD3+ BTLA+T cell,IL-10,TIMP1 ;FEV1 was positively correlated to CD3 + BTLA+T cell,CD4+ BTLA+T cell,TIMP1,while negatively correlated to course ; MEF50 was positively related to CD3+BTLA+T cell,CD4+ BTLA+T cell.Conclusion While articular cartilage lesions occurred in KOA patients,the lung function was also declined.The mechanism may be associated with the declination of expression of BTLA,which can cause up-regulating of IL-1 β,MMP9 and down-regulating of IL-10,TIMP1,thus leading to immune dysfunction and abnormal immune response.Those may induce airway injuries and result in lung function decline finally.

Objective To establish a human adrenocortical carcinoma (ACC)cell line ACC-LWL, and investigate its cell phenotypes and expression of tumor associated antigens, to detect sensitivity of ACC-LWL against adoptive immune cells. Methods Fresh tumor tissues from the resection of a human ACC were primary cultured and passed generation to generation so as to achieve stable growth in vitro. For analyzing its biological characteristic, the cell cloning formation in soft agar, chromosome and tumorigenesis were tested. Flow cytometric analysis was carried out for phenotype analysis and RT-PCR examination for MN/CA9 and HLA-A2 genes expression. PBMC from 4 healthy donors (2 HLA-A2+ or 2 HLA-A3+) co-cultured with IL-2 (200 U/ml)and the tumor cell lysate of ACC-LWL (20 μg/ml) in vitro to generate CTL. After 14 day stimulation, CTLs were incubated with ACC-LWL with a E/T ratio of 10:1 for 4 hours at 37 ℃. Cytotoxicity was measured with MTr assay. Results One human ACC cell line had been established that showed the characteristics of malignant cells. The cells grew a solid tumor in nude mice. ACC-LWL had a high expression of MHC-Ⅰ, weakly expression of Her2/neu and MHC-Ⅱ, no expression of CD80 or CD86. RT-PCR showed that ACC-LWL expressed MN/CA9 and HLA-A2 genes. CTL, either HLA-A2+ or HLA-A3+, had the capability of killing ACC-LWL in vitro. Conclusion This newly established ACC would provide a useful target in vitro and in vivo for investigations related to human ACC.

Objective To explore the molecular pathological mechanism of gout, and to explore the mechanism of how interleukin (IL)-37 influencing PDZK1 protein in gout through nuclear factor κB (NF-κB) pathway. Methods HK-2 cells were stimulated with inflammatory signal IL-37. The expression of PDZK1 and its subcellular localization were detected by real-time fluorescence quantitative polymerase chain reaction (real-time PCR) at different concentrations of IL-37 (defined as group A), PDTC+IL-37 (defined as group B), Wortmannin+IL-37 (defined as group C), respectively.The changes of PDZK1 protein expression in HK-2 cells were detected by adding inhibitor PDTC (NF-κB pathway inhibitor) or Wortmannin (PI3K pathway inhibitor) and inflammatory signal stimulating protein imprinting method. The comparative t test was used for statistical analysis between groups A, B and A and C. Results The average levels of PDZK1 mRNA were as follows:(group A: 28.71 ±0.35, 28.57 ±0.31, 28.78 ±0.28, 28.63 ±0.29, 28.62 ±0.19; group B: 28.71 ±0.31, 28.83 ±0.27, 28.58±0.26, 28.73±0.36, 28.68±0.35;group C:28.81±0.32, 28.91±0.29, 28.72±0.24, 28.59±0.18, 28.58±0.22). There was no significant change in PDZK1 mRNA level in group A and B. The same IL-37 was found in group A and B. The values of T were 5.73, 4.72, 4.69, 5.86 and 6.79, respectively. The P values were all greater than 0.05, and there was no significant difference between the two groups. The values of T were 6.78, 7.13, 7.47, 6.38 and 5.98 in the same IL-37 concentration groups of A and C, respectively, and the P values were all greater than 0.05, with no significant difference. The levels of PDZK1 protein in the three groups by Western blot analysis were as follows: (group A: 0.200±0.082, 0.412±0.032, 0.723±0.063, 1.202±0.021, 1.222±0.023;group B: 0.124±0.064, 0.303±0.081, 0.325±0.062, 0.223±0.071, 0.343±0.052; group C: 0.017±0.022, 0.204± 0.015, 0.187±0.053, 0.302±0.051, 0.404±0.051), The levels of PDZK1 protein in group A treated with different concentrations of IL-37 followed by the concentration of IL-37. The level of PDZK1 protein in group B increased with the increase of IL-37 concentration, but the level of PDZK1 protein in group B was lower than that in the group treated with IL-37 only, and decreased when the concentration of IL-37 was 40 ng/ml.The level of PDZK1 protein in group C increased with the increase of IL-37 concentration, but the level of PDZK1 protein in group C was lower than that in the group treated with IL-37 only, and the same concentration of IL-37 in group A and B. The values of T were 1.83, 1.37, 1.64, 1.57 ,1.49, with P values greater than 0.05. There was no significant difference between the two groups. The values of T were 1.28, 1.37, 1.26, 1.42, 1.39 in the same concentration of IL-37 in group A and C, with P values greater than 0.05, with no significant difference.The results of immunofluorescence analysis showed that the trend of the three groups was basically consistent with that of Western-Blot. Conclusion The pathogenesis of gout induced by IL-37 through PDZK1 protein may not occur at the transcriptional level, but may occur at the level of protein translation.