Objective To investigate the effect and mechanism of high concentration glucose on cholesterol absorption of human colon cancer epithelial Caco-2 cells.Methods The experimental study was used.(1) CCK-8 detected cell proliferation:the proliferation rate changes of Caco-2 cells were detected by CCK-8 when different concentrations (12.5,100.0,300.0,700.0,1 000.0,1 388.0 mmol/L) of glucose solution effects on Caco-2 cells in order to ensure the half hindering concentration of glucose concentration on Caco-2 cells.(2)Cholesterol absorption of Caco-2 cells was detected:Caco-2 cells were divided into the cholesterol group,cholesterol plus ezetimibe (cholesterol inhibitor) group and blank control group.Cholesterol group:100 μmol/L cholesterol solution and different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added.Cholesterol plus ezetimibe group:100 μmol/L ezetimibe,100 μmol/L cholesterol solution and different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added.Blank control group:DMEM culture medium and corresponding concentrations of DMSO were added.The cholesterol absorption amounts of Caco-2 cells were measured.(3) The relative expressions of ATP binding cassette G8 (ABCG8),ATP binding cassette G5 (ABCG5),Nickman-Pick CI Like 1 (NPC1L1) and scavenger receptor class B type Ⅰ (SR-B Ⅰ) were examined by Western blot in the different groups.Caco cells were divided into the glucose group,glucose plus ezetimibe group and control group.The different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution were added into the glucose group,different concentrations (5.0,25.0,50.0 mmol/L) of glucose solution and 100 μmol/L ezetimibe were added into the glucose plus ezetimibe group,and 100 μmol/L ezetimibe were added into the control group.The relative expressions of ABCG8,ABCG5,NPC1L1 and SR-B Ⅰ were detected by Western blot.Measurement data were presented as (x) ±s,repeated measure variance analysis was used to perform variation trend test,and t test was utilized to conduct comparisons among groups.Results (1) CCK-8 results showed:proliferation rates of Caco-2 cells with the glucose solution concentration of 12.5,100.0,300.0,700.0,1 000.0 and 1 388.0 mmol/L were 1.380 ±0.043,1.238 ±0.072,0.736 ±0.035,0.336 ±0.021,0.316 ±0.020 and 0.288 ±0.010,respectively,with a statistically significant difference in the proliferation rates (F =11.019,P ＜ 0.05).The half hindering concentration of glucose solution on Caco-2 cells was 283.54 mmol/L.(2)Cholesterol absorption of Caco-2 cells:① the cholesterol absorption amounts of Caco-2 cells with the glucose solution concentration of 5.0,25.0 and 50.0 mmol/L were 0.282 ± 0.042,0.380 ± 0.063,0.390 ± 0.060 in the cholesterol group and 0.042 ± 0.012,0.197 ± 0.015,0.277 ± 0.029 in the cholesterol plus ezetimibe group,respectively,showing a statistically significant difference between the 2 groups (F =55.566,P ＜ 0.05).②There was a statistically significant difference in cholesterol absorption amounts of Caco-2 cells with different glucose solution concentration in the cholesterol group (F =79.117,P ＜ 0.05).The cholesterol absorption amounts of Caco-2 cells with the glucose solution concentrations of 5.0 mmol/L was lower than that with the glucose solution concentrations of 25.0 mmol/L and 50.0 mmol/L,respectively (t =11.207,11.532,P ＜0.05).There was no statistically significant difference in the cholesterol absorption amounts of Caco-2 cells between the glucose solution concentrations of 25.0 mmol/L and 50.0 mmol/L (t =12.389,P ＞ 0.05).③There were statistically significant differences in cholesterol absorption amounts of Caco-2 cells with the glucose concentration of 5.0 mmol/L and 25.0 mmol/L between cholesterol group and cholesterol plus ezetimibe group (t =10.908,10.644,P ＜ 0.05).(3) The results of Western blot showed:① the relative expression of NPC1L1 protein in Caco-2 cells with the glucose solution concentrations of 5.0,25.0 and 50.0 mmol/L were respectively 0.277 ±0.019,0.558 ±0.015,0.576 ±0.003 in the glucose group and 0.057 ±0.002,0.054 ±0.005,0.077 ±0.005 in the glucose plus ezetimibe group,showing a statistically significant difference (F =482.207,P ＜0.05).② The relative expression of NPC1L1 protein of Caco-2 cells with the different concentration of glucose solution in the glucose group were compared,with a statistically significant difference (F =8.112,P ＜ 0.05).There was a statistically significant difference in the relative expression of NPC1L1 protein in Caco-2 cells with the different concentration of glucose solution in the glucose plus ezetimibe group (F =11.708,P ＜ 0.05).③ The relative expression of NPC1L1 protein in Caco-2 cells with the glucose solution concentrations of 5.0,25.0 and 50.0 mmol/L in the glucose group was statistically different from that in the glucose plus ezetimibe group (t =8.112,11.708,13.920,P ＜ 0.05).Conclusion High concentration glucose solution could promote the reabsorption of cholesterol through increasing NPC1L1 protein expression in Caco-2 cells,and increase the risk of suffering from cholelithiasis in diabetes patients.

Objective To explore the relationship between X - linked spondyloepiphyseal dysplasia tarda (SEDL) gene escaping X chromosome inactivation( XCI) and SEDL phenotype. Methods RT - PCR was performed on total RNA which was isolated from blood samples of patients, female carriers and controls. Patients and female carriers were selected from the pedigree with SEDL caused by the mutation (IVS2 - 2A→C) of the gene. cDNA was analyzed by polyacrylamide gelelectrophoresis(PAGE). Results PAGE data indicateed that female carriers expressed both normal and mutant SEDL mRNA,meaning the SEDL gene escaping XCI. Family investigation showed carrier females in the SEDL pedigree presented no symptoms. Conclusions The SEDL gene escaping X chromosome in-activation is firstly identified from human body. This may explain that carrier females present no symptoms.

Objective To establish a diabetic rat model of hindlimb ischemia, and provide a test method for dia-betic limb ischemia and diabetic foot study.Methods Twenty-five Wistar rats were divided into three groups:the model group and control group ( n=10) , and sham operated group ( n=5) .The rats of model group were induced by intraperito-neal injection of streptozotocin ( STZ, 60 mg/kg) , with the blood glucose level over 16.8 mmol/L, while the rats of control group were injected with normal saline.All the rats had the left femoral artery and its branches ligated, and then Doppler scan blood flow analysis was performed for the two hind limbs after operation at different time points.In the meantime, the changes of body weight and blood glucose were observed.The animals were sacrificed at 21days after operation.HE stai-ning was used to observe the pathological changes of gastrocnemius and femoral arteries.Capillary density and intima hyper-plasia were examined using immunostaining for CD31 and a-SMA.Results Blood glucose of the model group rats was sig-nificantly increased as well as the quantity of urine and feces,and associated with weight loss, which were maintained for more than 21 days.The blood flow of control rats was markedly decreased immediatly after operation, reached to the lowest point after surgery, and recovered from 7 -14 days after operation.In contrast to the control group, the model group showed a marked reduction in blood flow in the ischemic hind limb.Pathological examination revealed remarkable vascular atrophy and a significantly reduced number of vessels per high power field in the gastrocnemius muscle of model group rats with respect to the control group, and the femoral arteries of model rats were more narrowed than that of the control rats. Conclusions In this study, an effective, convenient diabetic rat model of hindlimb ischemia is successfully established which will facilitate the studies of drug intervention for diabetic limb ischemia and diabetic foot in the future.

Objective To find out the potential risk factor of plague in Wanzhou section of the Three Gorges Reservoir area, and to provide scientific basis for prevention and control of plague. Methods Rodents were captured by rat traps/cages at night and identified into species in Wanzhou section of the Three Gorges Reservoir area from 2001 to 2009. Flea was counted and serum antibodies against plague F1 of rats, cats and dogs were detected by indirect hemagglutination (IHA). Plague surveillances were performed in human beings and rats. Results The rodents captured belonged to 9 species, 2 families, 2 orders and 1 classes. The average indoor rodent density was 1.16% (961/82 558), and was 1.12% (1345/119 671) outdoors. Rattus norvegicus was the dominant species,accounting for 50.37%. The proportion of R. Flavipectus was 3.80% in 2004, 4.50% in 2008 and 10.12% in 2009,showing an increasing trend year by year. There were three kinds of mice infected fleas in Wanzhou, which including Xenopsylla cheopis, Leptopsylla segnis and Ctenocephalides felis. The average rate of flea infected mice was 1.18%(82/6959) and the total flea index was 0.036. No F1 antibody against plague was detected in 6959 dogs and 160 cats serum samples. Conclusions No plague is found in Wanzhou section of the Three Gorges Reservoir area. But R.Flavipectus, Xenopsylla cheopis and Leptopsylla segnis are dominant species in Wanzhou section, and the proportion of which shows an increasing trends year by year. There is a potential risk of plague outbreaks in Wanzhou section of the Three Gorges Reservoir area.

AIM:To investigate whether serum microRNA(miR)-103b plays a critical role in the pathogene-sis of type 2 diabetes mellitus(T2DM)and pre-diabetic syndrome.METHODS:Bioinformatic analysis was used for iden-tification of miR-103b and its targets,and the results were assessed by real-time PCR and receiver-operating characteristic (ROC)curve analysis in 48 patients with pre-diabetes mellitus(pre-DM),47 patients with noncomplicated diabetes melli-tus(NCDM),and 50 healthy individuals.RESULTS:miR-103b was significantly down-regulated in serum from the pa-tients with pre-DM and NCDM compared with healthy individuals.The ROC curve analysis found that the area under the curve(AUC)of miR-103b was 0.887(95% CI 0.809~0.944).The bioinformatic analysis has demonstrated that miR-103b has a high degree of site conservation among different mammalian species,such as Homo sapiens,Mus musculus,Rat-tus norvegicus,Pongo pygmaeus,Sus scrofa,etc.Fifty-three potential targets of miR-103b were predicted, most of which were involved in MAPK,Wnt,insulin and Ras signaling pathways,and enriched in various biological processes(such as phosphoprotein,DNA regulation transcription,cell growth and proliferation,apoptosis, cell cycle, etc), molecular func-tions(such as protein binding)and cell component(such as filamentous actin).CONCLUSION:Serum miR-103b can be used as an objective complement to traditional diagnosis of pre-diabetes,indicating important implications regarding the distinguish of the undiagnosed cases between diabetes and pre-diabetes by circulating miRNA.

OBJECTIVETo study the anti-tumor immunotherapeutic effect induced by the suicidalcancer vaccine FC/TK, and to evaluate the safety of this vaccine.

METHODSThe suicidal cancer vaccine, named FC/TK, was prepared by fusion of suicide gene (HSVI,-TK gene) -modified ovarian carcinoma NuTu-19 cells with rat bone marrow-derived dendritic cells (DCs). The morphology of FC/TK was evaluated by scanning electron microscopy. The stimulatory effect of FC/TK on T cells was determined by T cell proliferation assay. In immunotherapeutic studies in vivo, Fischer344 rats were injected subcutaneously with NuTu-19 cells, followed by treatment of FC/TK on days 7 and 14, compared to controls treated with irradiated FC/TK, FC or PBS, respectively. Tumor incidence and volume were measured in 90 days after challenge. To determine the killing effect of FC/TK in vivo, TUNEL assays were applied to detect apoptotic cell death in spleen of vaccinated rats with prodrug ganciclovir administration.

RESULTSFC/TK cells were of irregular shape with surface membrane processes. Compared to the control groups, FC/TK significantly promoted T cell proliferation (P <0.01). The rats vaccinated with FC/TK and FC significantly inhibited the tumor growth compared to rats vaccinated with irradiated FC/TK (P <0.05) or with PBS ( P <0.01). The immunotherapeutic effect induced by FC/TK was similar to that using FC. Fluorescence microscopy showed that fluorescein-stained FC/TK cells migrated into spleen also showed to be TUNEL-positive, suggesting that the FC/TK cells were killed by ganciclovir in vivo.

CONCLUSIONOur data indicate that suicidal cancer vaccine is an effective and safe therapy for ovarian carcinoma and may serve as a broadly applicable approach for other cancer vaccines in the future.

Animals ; Apoptosis ; drug effects ; Cancer Vaccines ; immunology ; Cell Fusion ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dendritic Cells ; cytology ; immunology ; Female ; Ganciclovir ; pharmacology ; Genes, Transgenic, Suicide ; Herpesvirus 1, Human ; enzymology ; genetics ; Immunotherapy ; methods ; Microscopy, Electron, Scanning ; Microscopy, Fluorescence ; Neoplasms, Experimental ; enzymology ; pathology ; therapy ; Ovarian Neoplasms ; enzymology ; pathology ; therapy ; Rats ; Rats, Inbred F344 ; Survival Analysis ; T-Lymphocytes ; drug effects ; metabolism ; pathology ; Thymidine Kinase ; genetics ; metabolism ; Transfection

OBJECTIVETo compare the efficacy and feasibility of neoadjuvant chemoradiotherapy with docetaxel plus cisplatin or with cisplatin plus fluorouracil in the treatment of local advanced esophageal squamous cell carcinoma.

METHODSA total of 154 cases in the stage of cT3N0-1M0 were randomly assigned to two arms. The arm A received 2 cycles of doctaxel 75 mg/m(2) plus cisplatin 25 mg/m(2) d1-3 and 40 Gy of radiation therapy, and the arm B received 2 cycles of cisplatin 25 mg/m(2) d1-3 plus fluorouracil 600 mg/m(2) d1 ∼ 5 and 40 Gy of radiation therapy. The surgery was performed 3 - 4 weeks later.

RESULTSGrade 3/4 toxicities occurred in 53.2% of the patients in arm A and in 36.4% of the patients in arm B (P = 0.035). Neutropenia occurred in 20.7% of the patients in arm A and 5.6% of the patients in arm B (P = 0.004). Nine patients aborted surgery due to tumor progression. 71 patients underwent resection in 73 cases of the arm A and 69 patients underwent complete resection, 70 patients underwent resection in 72 cases and 70 complete resection of the arm B, respectively (P > 0.05). No mortality was noted. The overall complication rate was similar in the two arms (21.9% vs. 23.6%). Pathological complete response was achieved in 27 patients (35.1%) in the arm A and 16 patients (20.8%) in the arm B (P = 0.048).

CONCLUSIONSNeoadjuvant chemoradiotherapy with docetaxel plus cisplatin can be well tolerated and achieves a higher pathological complete response rate than with cisplatin plus fluorouracil.

Adult ; Aged ; Antineoplastic Combined Chemotherapy Protocols ; adverse effects ; therapeutic use ; Carcinoma, Squamous Cell ; pathology ; surgery ; therapy ; Chemoradiotherapy ; Cisplatin ; administration & dosage ; adverse effects ; Esophageal Neoplasms ; pathology ; surgery ; therapy ; Female ; Fluorouracil ; administration & dosage ; adverse effects ; Humans ; Male ; Middle Aged ; Neoadjuvant Therapy ; Neoplasm Staging ; Neutropenia ; chemically induced ; Radiotherapy, High-Energy ; Remission Induction ; Taxoids ; administration & dosage ; adverse effects ; Vomiting ; chemically induced

AIM:To explore the changes and significance of Kupffer cells in the process of tree shrew chroni -cally infected with hepatitis B virus (HBV).METHODS:The animals were divided into 3 groups.Group A consists of 6 tree shrews that were identified as persistently infected with HBV;group B consists of 3 tree shrews that were suspected as persistently infected with HBV;group C consists of 4 tree shrews that were not inoculated with HBV and were applied as normal controls.Liver biopsies were collected regularly from all animals , and the Kupffer cells were isolated , purified and primarily cultured.The techniques of flow cytometry , immunohistochemistry, lysosomal fluorescent probe staining and real-time RT-PCR were applied to determine the number and function of these Kupffer cells .RESULTS: The result showed that the count and proportion of CD 163+cells in group A were significantly higher than those in group B and group C ( P<0.05).Meanwhile, the fluorescence intensity levels of lysosomal , the number of lysozyme-positive cells and the mRNA ex-pression level of TNF-αin the Kupffer cells in group A were significantly lower than those in group B and group C ( P<0.05).CONCLUSION:Kupffer cells may play a regulatory role during host’s chronic HBV infection.

Objective To investigate the reliability of ultrasound tissue characterization (UTC) for quantifing the integrity of achilles ten-don and patellar tendon in subjects without tendinopathy. Methods From August, 2016 to April, 2017, testers A and B quantified integrity of achilles tendon and patellar tendon of dominant legs from 43 subjects without tendinopathy by UTC in random order respectively. Tester B quantified integrity of patellar tendon of dominant legs from other 22 subjects without tendinopathy by the same way, and repeated the mea-surements the next day. Interclass correlation coefficient (ICC) and the minimum detectable different (MDD) were calculated. Results The ICC of inter-rater reliability of achilles tendon assessment was 0.492 to 0.735, MDD was 0.3%to 3.0%. The ICC of inter-rater reliability of patellar tendon assessment was 0.383 to 0.678, MDD was 0.3%to 4.4%. The ICC of intra-rater reliability of patellar tendon assessment was 0.525~0.760, MDD was 0.6%to 5.3%. Conclusion The test-retest reliability of UTC for the quantification of integrity structure of achilles tendon and patellar tendon is satisfactory with little error, the inter-rater reliability need to be improved. which is a accurate and feasible tool to quantify tendon by the same tester.