To investigate the roles of neutral glycosphingolipids (N-GSLs) in the development of tumor, tumor antigen expression and immunological evasion, the regularities of the expression of N-GSLs in human embryonic and neoplastic tissue were studied, the influence of N-GSLs in the growth regulation, immunological evasion and multidrug resistance(MDR) of tumor cells were analyzed. The tumor embryonic associated antigen CDH, tumor multidrug resistance associated N-GSLs CMH and neoplasm inhi-bitor globoside were found. The significance of N-GSLs synthesis inhibition and de-N-glycosylation in the reversion of MDR and tumor therapy was also partly disclosed.

The author investigated the function of coenzyme NADH in increasing the level of energy metabolism, repairing cellular damage, improving cellular stress ability, decreasing cytotoxicity of chemotherapy drugs and radiation against normal tissue. The molecular regulation mechanism of NADH in cytoprotection was elucidated. A new prevention and cure way was provided in cytoprotection treatment for clinical disease.

Objective: To study the characterization of the cultured cervical cancer cell line transfected with anti- HPV16E6-ribozyme, and to investigate the effect of ribozyme on proliferation and apoptosis of cervical cancer cell. Metliods: Anti-HPV16E6-ribozyme had been designed to cleave the HPV16E6 gene. With the method of lipofectin transfec- tion, the anti-HPVI6E6-ribozyme and empty eucaryotic expressing plasmids were transfected into CaSKi cell, which named as CaSKi-R, CaSKi-P respectively. The amounts of E6 mRNA in the three kinds of cells were detected by northern blot. Cell cycle was detemined by flow cytometry, and cell apoptosis was examined by fluorescent (Hoechst) staining and TUNEL. The expression of some genes, including c-myc, bcl-2, p53, and fas, was also detected by flow cytometry analy- sis. Results: Northern blot showed that E6 mRNA was less in CaSKi-R than in CaSKi. In CaSKi-R cells, cycle was arres- ted in G1 phase, with decreasing in percentage of S phase cells. The apoptosis rate of CaSK1-R cell was much higher than those of CaSKi and CaSK1-P. Anti-HPV16E6-ribozyme could reduce the expression of E6, c-myc, bcl-2 genes on CaSKi- R cells, and increased the expression of p53. While this phenomenon was not found on the CaSK1-P cells. The expression of fas was similar in the three kinds of cells. Conclusion: Anti-HPVE6-rivozyme induces apoptosis of human cervical cancer CaSKi cells. The mechanisms may be the decrease of E6 gene's expression, and the succedent changing of some genes'expression.

R 5 was administered in vitro to observe its antitumor activity in human breast carcinoma cell line MCF 7. As the most useful and efficient anthracycline, epirubicin(EPI) was served as the positive chemical treatment control. MTT colorimetric assay was applied to detect cytotoxicity of R 5 to MCF 7 cells. Apoptotic rate of cells double marked with Annexin V FITC and PI was examined by flow cytometry. And, the alteration of wild type (WT) p53, bax and bcl 2 proteins expression was also observed. Ultrastructural change of MCF 7 cells was observed under a transmission electron microscope. The results showed that growth restrain was observed in MCF 7 cells when administered with different doses of R 5 or EPI. The effect was increased concomitantly with the increasing of R 5 or EPI concentration and culture time. Apoptosis was observed since 6 hours after the MCF 7 cells cultured with R 5 or EPI, and the effect was increased with the culture time extending and reached the highest peak at about 48 hours. However, the apoptotic rate decreased when cultured for 72 hours. Different doses of R 5 and EPI can all induce apoptosis in MCF 7 cell, and the apoptotic rate increased with their concentration, but decreased when the concentration was higher than 5?10 -6 mol/L. Ultrastructure of apoptotic MCF 7 cells, observed by transmission electron microscope, showed typical morphologic changes of apoptosis.

Objective: To investigate the diagnostic value of combined examination of copeptin and high sensitivity cardiac troponin T (hs-cTnT) in patients at the early stage of acute myocardial infarction (AMI) . Methods: A total of 272 patients were enrolled in this study, all of them suffered from chest pain and admitted within 4 hours. The patients were divided into 4 groups according to coronary artery angiography (CAG) results. Control group, the patients with normal CAG,n=64, UAP group (unstable angina pectoris),n=50, STEMI group,n=82, NSTEMI group,n=76. All patients received in-hospital observation, plasma levels of copeptin and hs-cTnT were examined at admission and at 6 hours after the chest pain respectively. Results: Within 4 hours of chest pain, combined examination of copeptin and hs-cTnT had the higher sensitivity for diagnosing AMI than a single detection of hs-cTnT with the cut-off point of hs-cTnT ≤ 14ng/L and Copeptin < 14pmol/L. In NSTEMI group, the AUC (area under curve) for combined examination was 0.97 (95% CI 0.88-0.99), AUC for single hs-cTnT detection was 0.75 (95% CI 0.62-0.87),P<0.05. In STEMI group, the AUC for combined examination was 0.97 (95% CI 0.88-0.99), AUC for single hs-cTnT detection was 0.74 (95% CI 0.60-0.88),P< 0.05. The AUC for combined examination of copeptin and hs-cTnT in diagnosing early AMI was 0.912 (95% CI 0.812-0.961) which was higher than single detection of hs-cTnT, AUC 0.851 (95% CI 0.713-0.936), Z=2.553,P<0.05. Conclusion: Combined examination of copeptin and hs-cTnT had the higher sensitivity and accuracy for diagnosing the patients at the early stage of AMI, it may help the risk stratiifcation of chest pain which is valuable in clinical practice.

Objective To establish a stable cell line expressing transmembrane form of human blood group A antigen mimotope vaccine by transfecting malignant melanoma cell line B16, and to detect the cytotoxicity of the vaccine against melanoma cells. Methods Cultured B16 cells were classified into 4 groups, i.e.,P/F-M-pIRES group [transfected with the recombinant plasmid mimotope peptide/Fas-macrophage inflammatory protein (Mip)-pIRES], P/F-pIRES group (transfected with the recombinant plasmid mimotope peptide/FaspIRES), M-pIRES group (transfected with the recombinant plasmid Mip-pIRES), and pIRES group (transfected with the empty plasmid pIRES). B 16 cells were transfected through Lipofectamine 2000. Subsequently, RT-PCR and Western blotting were performed to detect the mRNA and protein expressions of the mimotope peptide/Fas fusion gene and Mip3β in transfected B16 cells. Cell counting kit-8 (CCK-8) was used to evaluate the vaccinemediated complement dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC)against B16 cells. Results RT-PCR yielded specific DNA fragments with expected size. Western blotting revealed the anti-A antibody-binding activity of the recombinant mimotope peptide/Fas fusion protein. Factor analysis indicated significant differences in CDC (F = 244.522, P ＜ 0.01 ) and ADCC (F = 71.593, P ＜ 0.01 )against B16 cells between the 4 groups. Group comparisons demonstrated more intense CDC and ADCC in P/FM-pIRES and P/F-pIRES groups compared with M-plRES and pIRES groups, stronger ADCC in P/F-M-pIRES group in comparison with P/F-pIRES group (F = 15.42, P ＜ 0.05), but no significant difference in CDC was observed between M-pIRES and pIRES group. Conclusions The transmembrane form of human blood group A antigen mimotope vaccine could be stably expressed in B16 cells, and mediate ADCC and CDC against B16 cells in vitro.

BACKGROUND: Tissue-engineered artificial nerve was successfully constructed with the compound of acellular nerve graft and bone marrow mesenchymal stem cells, suggesting that it could promote peripheral neural regeneration.OBJECTIVE: To construct tissue-engineered artificial nerve, and to verify neural functional recovery of bridging rats following sciatic nerve defect.METHODS: A total of 60 adult male SD rats were used to induce sciatic nerve defect models (15 mm in length), and they were then randomly divided into three groups, with 20 rats in each group. Sciatic nerve defect group was treated with tissue-engineered artificial nerve; blank control group was treated with tissue-engineered nerve stent; autoallergic neural control group was treated with autoallergic neural transplantation. Twelve weeks after bridging, histology of sciatic nerve and neuralfunctional recovery were detected via gross observation, wet mass of tibialis anterior muscle, and histological analysis.RESULTS AND CONCLUSION: At 12 weeks after bridging surgery, rats in experimental group were able to stand on the floor,and withdrawal reflex was detected at plantar skin on the surgical side. S-100 protein of plantar skin was positive. There was no significant difference in wet mass of tibialis anterior muscle between experimental and autoallergic neural transplantation group (P ＞ 0.05). HRP retrograde tracing in the experimental group demonstrated that HRP-positive cells were observed in both spinalcord and posterior root ganglion. There was no significant difference in number of myetinated nerve fiber, thickness of myelin sheath, and area of nerve tissue between experimental and autoallergic neural transplantation group. The results demonstrated that the compound of acellular nerve graft and bone marrow mesenchymal stem cells could successfully construct tissue-engineered artificial nerve to repair sciatic nerve defect and promote neurohistological reconstruction and functional recovery.

To probe the effect of reduced form coenzyme Ⅰ(NADH) in antagonizing cardiac muscle toxicity induced by doxorubicin and its underlying mechanism. Primary culture of myocardial cells of SD rat and doxorubicin injury model were established. MTT assay, laser confocal microscopy, transmission electron microscopy and biological oxygen monitor were used to observe the morphology and function of mitochondria. The results showed that the killing rate was increased in the group of doxorubicin, and that in the group of NADH/doxorubicin was greatly decreased. Doxorubicin could induce ultrastructural damage of cardiomyocyte mitochondria, manifested as swelling, disintegration, disruption of cristae and fusion. Cardiac mitochondria were protected against injuries in the group treated with NADH/doxorubicin. There was a significant difference in the fluorescence intensity of mitochondria membrane potentional and ROS between the groups treated with doxorubicin and NADH/ doxorubicin. It is suggested that NADH can significantly antagonizing cardiac muscle toxicity induced by doxorubicin and can protect mitochondrial structure and function.

AIM To study effects of antioxidant NADH on damage of irradiated normal cell lines. METHODS L02 liver cells were cultured in RPMI 1640, exposed to X ray irradiation, and continued to culture in the presence or absence of NADH for 24 h. The cellular viability was determined by routine MTT method. Using fluorescence probe and confocal microscope, the level of cellular H 2O 2 was detected. Positive rate of bcl 2 and Bax protein expression in the L02 cells were analyzed by flow cytometry. RESULT NADH can not only antagonized growth inhibition of X ray irradiated L02 cells, decreased cellular H 2O 2 production, but also significantly increased positive rate of L02 cells expressed bcl 2 protein and decrease positive rate of L02 cells expressed Bax proteins ( P

BACKGROUND: Herterologous antigen has strong immunogenicity and easily induces immunological response. Introduction of herterologous antigen into tumor may induce a serial of immunological reactions in the tumor and may reverse the immunosuppression of tumor microenvironment to treat tumor.OBJECTIVE: To evaluate the antitumor efficacy of intratumoral injection of human erythrocyte membrane antigens in micebearing S180 sarcoma.METHODS: Kunming mice bearing S180 sarcoma model were established and treated with 5 g/L human erythrocyte membrane antigens suspension or normal saline for five days. Tumor volume was calculated before the first injection and 3, 7, and 14 days after the first injection. In addition, the tumor cells in combination with human erythrocyte membrane antigens group, the njectionof saline group (the control group), and the injection of human erythrocyte membrane antigens or saline group (pre-immunized by suspension of human erythrocyte of blood group type A). Another 60 mice bearing S180 sarcoma were established and subjected to the above pre-immunization and injection of saline or human erythrocyte membrane antigens. Six mice selected from each group were sacrificed 14 days after the first injection, and tumors were weighed, followed by histological examination. Survival of remainders in each group was observed.RESULTS AND CONCLUSION: Tumor volumes in each group increased gradually. Tumor volumes in the human erythrocyte membrane antigens injection group, the tumor cells in combination with human erythrocyte membrane antigens group, and the human erythrocyte membrane antigens injection group (immunized) were smaller than the control group, Intratumoral injection of human erythrocyte membrane antigens significantly reduced tumor weights. Tumor necrosis, infiltration of inflammatory cells such as lymphocytes were observed in tumor tissues section examination following the intratumoral injection of human erythrocyte membrane antigens. The mouse survival time showed no statistical difference among different groups. Intratumoral injection of heteroloaous ervthrocvte membrane antiaens can inhibit tumor arowth of S180 sarcoma bearina mice.