P4HA2 promotes occurrence and progression of liver cancer by regulating the PI3K/Akt/mTOR signaling pathway / 南方医科大学学报

OBJECTIVE@#To investigate the role of proline 4-hydroxylase Ⅱ (P4HA2) in the occurrence and progression of liver cancer.@*METHODS@#GEPIA and Human Protein Atlas database were used to predict the expression of P4HA2 in hepatocellular carcinoma (HCC), and K-M plotter online database was used to analyze the relationship between P4HA2 expression and the prognosis of HCC. We also examined the expressions of P4HA2 in HCC cells and normal hepatocytes using qRT-PCR and Western blotting. With lentivirus-mediated RNA interference, P4HA2 expression was knocked down in hepatoma SNU-449 and Hep-3B cells, and the changes in cell proliferation, migration and invasion were assessed using cell counting kit-8 (CCK-8) assay, colony formation test, scratch test and Transwell assay. The changes in the expressions of epithelial-mesenchymal transition (EMT) and PI3K/Akt/mTOR signal pathway-related proteins were detected using Western blotting.@*RESULTS@#Online database analysis showed that the expression of P4HA2 was significantly higher in HCC tissues than in normal liver tissues (P < 0.05). The expression levels of P4HA2 mRNA and protein were also significantly higher in HCC cell lines than in normal hepatocytes (P < 0.01). Lentivirus-mediated RNA interference of P4HA2 significantly lowered the expression levels of P4HA2 mRNA and protein in the hepatoma cells (P < 0.05) and caused obvious inhibition of cell proliferation, migration and invasion. P4HA2 knockdown significantly increased the expression of E-cadherin protein, lowered the expressions of N-cadherin and Snail, and obviously decreased the expressions of phosphorylated PI3K, AKT and mTOR (P < 0.05).@*CONCLUSION@#P4HA2 enhances the proliferation, migration, invasion, and EMT of hepatoma cells by activating the PI3K/Akt/mTOR signaling pathway to promote the occurrence and progression of liver cancer.

LY294002 Enhaces Chemosensitivity of K562 Cells to Daunorubicin / 中国实验血液学杂志

OBJECTIVE@#To investigate the proliferation inhibition and pro-apoptosis effect of LY294002 (PI3K/AKT inhibtor) combined with daunorubicin (DNR) on the chronic myeloid leurenia cell line K562 and its possible mechanisms.@*METHODS@#The effect of LY294002 and DNR on the proliferation of K562 cells in different treating time and concentration were measured by MTT assay. The cell cycle was determined by flow cytometry, the mRNA and protein expression of SKP2 , P27, BCL-2 and BAX were determined by RT-PCR and Western blot.@*RESULTS@#LY294002 and DNR were able to inhibit the growth of K562 cells and promote apoptosis in time- and concentration-dependent manner (P＜0.05), both the cell proliferation-inhibiting rate and apoptosis rate in combination therapy group were higher than that in DNR-monotherapy group (P＜0.05). After K562 cells treated by LY294002 combined DNR for 36 h, the cells were statistically significantly reduced in G/M phase (P＜0.05), as compared with control group and DNR group. Compared with DNR group, the cell level of G/G phase rased (P＜0.05) and cell level of S phase decreased (P＞0.05). Compared with DNR group, the expresson of SKP2 and BCL-2 mRNA decreased, and the expression of P27 mRNA increased in the combination therapy group (P＜0.05). The expression of BAX mRNA was not significantly different between different groups. The same result was found in the protein expression.@*CONCLUSION@#LY294002 has the sensibilizative effect on DNR chemotherapy, which may be relative with blocking the cell cycle and inducing cell apoptosis.

Establishment and application of loop-mediated isothermal amplification for Candida albicans / 临床检验杂志

Objective@#To establish a simple and rapid loop-mediated isothermal amplification (LAMP) to detect Candida albicans (CA) and explore its clinical value.@*Methods@#The Primer Explorer 5.0 software was used to design 4 primers for amplification of CA by LAMP. The system and conditions of LAMP reaction were optimized to evaluate its specificity and the minimum limit in the detection. The vaginal swabs were collected from 123 vulvovaginal candidiasis (VVC) patients and 42 healthy individuals. Fungal culture, LAMP test, PCR test and 1.79 mol/L KOH microscopy were conducted in parallel. Fungal culture was used as the reference method for VVC diagnosis. The positive rates between two groups were compared by Pearson χ² test. The consistency of the results from LAMP, PCR, microscopy and culture were analyzed by Kappa test and P＜0.05 was considered as statistically significant difference. ROC curve analysis was used to evaluate LAMP and PCR test for diagnosis of VVC.@*Results@#The optimum reaction temperature of LAMP was 61 ℃ with high specificity. No cross reaction with other strains was found. The minimum detection limit was 10³ copies/ul. The positive rates of LAMP, PCR and microscopy between VVC and healthy group showed statistically significant difference（LAMP: χ²=68.576；PCR: χ²=64.918；microscopy: χ²=50.076，P＜0.01）. LAMP detection and PCR showed good consistency (κ=0.744, 0.720), but microscopy examination showed poor consistency (κ=0.533). LAMP showed diagnostic sensitivity of 87.62%, specificity of 88.33%, positive predictive value of 92.93% and negative predictive value of 80.30%. The area under the curves of LAMP and PCR were 0.873 and 0.888, respectively. No difference in efficacies between LAMP and PCR was found (Z=0.849, P=0.395 6), but the lowest detection time of LAMP was shorter than 1 hour.@*Conclusion@#A rapid, reliable, sensitive and specific LAMP technique for detecting CA was established. The comprehensive screening performance should be superior to the routine method in laboratories, so LAMP could be used for the supplementary diagnosis and monitoring therapeutic effects in CA infection.

Influence of BMP4 on Regulation of Cell Cycle and Apoptosis of Hematopoietic Stem Cells/Progenitor Cells and Its Mechanism in Chemotherapy-Induced Myelosuppression / 中国实验血液学杂志

OBJECTIVE@#To explore the effect of bone morphogenetic protein 4(BMP4) on the cell cycle and apoptosis of hemaropoictic stem and progenitor cells (HSPC) in conditions of 5-fluorouracil (5-FU)-inducing bone marrow suppression and stress hemogenesis, and its possible mechanism.@*METHODS@#The C57BL transgenic mice with BMP4 overexpression were established and were enrolled in transgenic group (BMP4 group), at the same time the wild type mice matching in age, sex and body weight were selected and were enrolled in control group (WT group). The bone marrow suppression was induced by injection with 5-FU in dose of 150 mg/kg, then the nucleated cells were isolated from bone marrow. After the HSPCs were markered with C-kit/sca-1 fluorescent antibodies, the changes of cell cycle and apoptosis of HSPC were detected by Aunexin V/PI and Ki67/DAPI double staining; the cell cycle-essociated hemotopoietic regulatory factors were detected by RT-qPCR.@*RESULTS@#Under physiologic status, there were no significant differences in cell cycle and apoptotic rate of HSPC between WT group and BMP-4 group. After the bone marrow was suppressed, the ratio of HSPC at G0 phase in BMP4 group significantly decreased(P＜0.05); the apoptosis rate of HSPC significantly increased(P＜0.05); the mRNA expression levels of hypoxia-inducing factor Hif-1α and chemotactic factor CXCL12 in stroma of BMP4 group were down-regulated significanfly(P＜0.05).@*CONCLUSION@#Under non-physiologic conditions such as stress hemogenesis or bone marrow suppression, the up-regulation of BMP4 can promote HSPC into cell cycle and apoptosis of HSPC, moreover, the BMP4 may play a regulatory role for cell cycle of HSPC through direct or indirect down-regulation of Hif-1α and CXCL-12 expressions.

Research on a new NBC casualty decontamination shelter / 医疗卫生装备

Objective To develop a new NBC casualty decontamination shelter,which can be used to decontaminate NBC casualties. Methods The shelter integrated the technologies of plate shelter expansion, auto waterway control, micro-environment control, rapid sewage collection and etc, and took considerations on decontamination requirements of the ambulatory and non-ambulatory casualties.It was designed combining fixed and extendable shelters,which realized double-side expansion by manual push and pull. The shelter was composed of the plate shelter, decontamination devices and facilities, water supply & heating system, waterway system, power supply and distribution system, lighting system, air conditioning and heating system and other accessories.Results The shelter could decontaminate the ambulatory and non-ambulatory casualties simultaneously,the decontamination water temperature ranged from the temperature of water source to 50 ℃, the mixing ratio of the decontamination water was from 0.2% to 2%, and the time consumed for deployment and withdrawal was not more than 20 min. Conclusion The shelter gains advantages in layout and technology, and enhances decontamination ability for NBC medical rescue.

Participatory Teaching Class for MBBS Students / 昆明医科大学学报

Objective To determine the effect of participatory teaching method in the Clinic teaching of internal medicine for MBBS students. Methods The 60 MBBS students were divided into group A and B, 30 students in each group. Participatory teaching was done in group A, and traditional teaching was carried out in group B,and the effects of the two types of teaching methods were compared. Results In the clinic teaching of internal medicine, the effects in improving students' clinical thinking and practical ability, creativeness, initiative and efficiency were better in the participatory teaching method than in traditional teaching method (P＜0.05) . Conclusion The effects of participatory teaching in the clinic teaching of internal medicine for MBBS students was better than the traditional teaching.

The Value of Using sPESI Score Combined with hs-TnT in the Evaluation of Short-term Prognosis in Hemodynamically Stable Pulmonary Embolism / 昆明医科大学学报

Objective To explore the value of using sPESI score and hs-TnT in the evaluation of short-term prognosis in hemodynamically stable pulmonary embolism. Methods We collected 99 patients with hemodynamically stable PE from our department. According to the sPESI score and hs-TnT, patients were divided into high risk group (sPESI score≥1) and low risk group (sPESI score=0), positive group (hs-TnT≥0.014 ng/mL) and negative group (hs-TnT＜0.014 ng/mL) . Then all patients were treated and followed up for 30 days. The sensitivity, specificity, positive predictive value and negative predictive value of adverse events of prognosis were calculated, and ROC curve was drawn to analyze the values in different grouping methods for the prognostic evaluation.Results Thirteen adverse events occurred in all patients. The single test showed that sensitivity, specificity, positive predictive value, and negative predictive value of hs-TnT for predicting adverse events were respectively 84.6%,55.2%,22%,and 96%. When sPESI was used alone, sensitivity, specificity, positive predictive value and negative predictive value were 92.3%, 48.8%, 21.1% , and 97.7% . Results of combined testing showed 100% sensitivity, 29% specificity, 17.6% positive predictive value, and 100% negative predictive value. The ROC curve area of the sPESI, hs-TnT, sPESI and hs-TnT are 0.832 (95% CI, 0.705-0.958), 0.825 (95%CI, 0.694-0.957),0.872 (95%CI, 0.773-0.971) . Conclusions PESI and hs-TnT have clinical value in evaluating the short-term prognosis of hemodynamically stable pulmonary embolism. sPESI combined with hs-TnT has higher significance, especially in patients with low-risk PE.

The role of Fas mutation on non-alcoholic steatohepatitis in mice / 中华肝脏病杂志

<p><b>OBJECTIVE</b>Our previous study indicated that the death receptor Fas played a key role on hepatocyte apoptosis in nutritional steatohepatitis in mice. This study aimed to explore whether Fas mutation accelerated hepatic steatosis and inflammatory infiltration in methionine-choline deficient (MCD) diet feeding mice.</p><p><b>METHODS</b>Mice homozygous for the lymphoproliferation spontaneous mutation (C57BL/6J-Faslpr) and wild type C57BL/6J mice were fed with MCD diet for three weeks to induce non-alcoholic steatohepatitis (NASH). Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), triglyceride (TG) and total cholesterol (TC) levels were detected by an Olympus AU5400 automatic chemical analyzer. The role of Fas gene mutation on NASH was assessed by comparing the severity of hepatic steatosis and inflammation in the liver sections, the mRNA and protein expressions of hepatic inflammatory and fibrogenesis related factors, proliferating cell nuclear antigen (PCNA) and transforming growth factor beta 1 (TGFb1).</p><p><b>RESULTS</b>The serum ALT levels of the wild type and Faslpr mice fed with MCD were significant higher than that of the control mice (126.33+/-10.50 U/L vs (25.00+/-10.14) U/L, (160.33+/-48.29) U/L vs (18.33+/-9.08) U/L, with the LSD-t value 12.02, 5.08 respectively, the P value<0.001, 0.007 respectively. The serum ALT levels showed no significant difference between the Faslpr and wild type mice fed with MCD, with the LSD-t value 1.19, the P value 0.229. The serum AST, TG and TC levels showed neithere significant difference among the four groups. MCD diet induced hepatic steatosis and inflammatory infiltration in both of the wild type and Faslpr mice. Especially, severer hepatic injury was observed in Faslpr mice as compared with wild type mice. The mRNA expression levels of cell proliferation factor PCNA and fibrogenesis growth factor TGF b1 in wild type mice fed with MCD were significantly higher than that of the control mice (2.84+/-0.73, 2.77+/-0.54 vs 1.31+/-0.18, 0.89+/-0.18), with the LSD-t value 4.99, 8.08 respectively, the P value 0.001, <0.001 respectively. The mRNA expression levels of PCNA and TGFb1 in Faslpr mice fed with MCD were significantly higher than that of the Faslpr control mice and the wild type mice fed with MCD (5.57+/-1.13, 5.73+/-0.89 vs 1.04+/-0.16, 0.85+/-0.11 and 2.84+/-0.73, 2.77+/-0.54), with the LSD-t value 10.15, 13.19 and 5.33, 6.91 respectively, the P value<0.001. The protein expressions levels of PCNA and TGFb1 were concordant with the mRNA.</p><p><b>CONCLUSIONS</b>Faslpr promoted hepatic steatosis and inflammatory infiltration in mice fed with MCD diet, which might associated with excessive release of cell proliferative, inflammatory and fibrogenesis factors.</p>

The role of heme oxygenase-1 in non-alcoholic steatohepatitis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To investigate the potential role of heme oxygenase-1 on preventing non-alcoholic steatohepatitis (NASH) in mice.</p><p><b>METHODS</b>Experimental models of NASH were established by feeding male C57BL/6J mice with choline-methionine deficient diet (MCD) for four weeks. Control animals were fed with choline-methionine supplemented diet. The treatment groups were fed with MCD diet combined with HO-1 inducer hemin or inhibitor zinc protoporphyrin IX (ZnPP-IX). Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were tested by enzymic method with automatic biochemistry analyzer. The degree of hepatic steatosis, inflammation and fibrosis were examined under HE staining. The hepatic mRNA and protein expressions of HO-1, TNFalpha and IL-6 were analyzed by RT-PCR and Western blot respectively. MCD fed mice showed increased serum ALT and AST levels and moderate to severe hepatic steatosis with inflammatory infiltration, hepatic spot or focal necrosis, light portal and sinus hepaticus fibrosis in the liver sections, which associated with enhanced expression of HO-1, TNFalpha and IL-6 mRNA and protein (1.13+/-0.11, 1.74+/-0.05; 0.20+/-0.01, 1.92+/-0.10; 0.58+/-0.02, 2.06+/-0.05 vs 0.43+/-0.02, 0.75+/-0.05; 0.08+/-0.00, 0.59+/-0.02; 0.22+/-0.01, 0.91+/-0.02). Administration of hemin significantly decreased serum ALT and AST levels and attenuated hepatic steatosis and necroinflammation which associated with up-regulation of antioxidative gene HO-1 and down-regulation of pro-inflammatory cytokines TNFalpha and IL-6 (P < 0.01). A contrary effect on serum aminotransferase levels and liver histopathology was observed in mice injected with ZnPP-IX (P < 0.01).</p><p><b>CONCLUSIONS</b>The effect was associated with suppressed HO-1 expression and increased TNFaLPHA and IL-6 expression. The data provided a biochemical, morphological and molecular biological evidence for the protective role of HO-1 in ameliorating hepatic steatosis, necroinflammation in experimental nutritional steatohepatitis.</p>

Phylogenetic Analysis of Homologous Proteins Encoded by UL2 and UL23 genes of Herpesviridae / 中国病毒学

The proteins encoded by the Herpesviridae β-gene play a critical role in the replication stage of the virus. In this paper, phylogenetic analyses provided evidence that someβ-gene products, such as UL2 and UL23 from HSV1, have their homologous genes in its family, and also exist in prokaryotic organisms, indicating that these viruses appear to have been assembled over evolutionary time by numerous independent events of horizontal gene transfer.

The role of apoptosis and the related genes in non-alcoholic steatohepatitis / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To study the role of apoptosis and the expression of apoptosis-related genes Fas ligand (FasL), Fas, caspase-3 and caspase-8 in an animal model of non-alcoholic steatohepatitis (NASH).</p><p><b>METHODS</b>An experimental progressive NASH model was established by feeding male C57BL6/J mice with a high fat, methionine-choline deficient (MCD-) diet for two days, five days, ten days, three weeks and eight weeks. Control mice were fed methionine-choline supplemented (MCD+) diet. Hepatic steatosis, inflammation and fibrosis were graded by examining their H and E stained liver sections. Hepatocyte apoptosis was detected by TUNEL assay. Expressions of mRNA and protein of FasL, Fas and caspase-8 were performed by quantitative real time RT-PCR and Western blot. Caspase-3 activity assay was conducted using ApoAlert caspase-3 assay kit.</p><p><b>RESULTS</b>In MCD- mice, minimal hepatic steatosis was observed at day 5, and by day 10, mild steatosis with inflammatory infiltration was found. Severe steatohepatitis was noted at week 3, and fibrosis at week 8. TUNEL assay showed that apoptotic index in MCD- group was higher than that in MCD+ group at week 3 (15.59%+/-4.87% vs 5.17%+/-3.19%, P less than 0.05) and at week 8 (11.29%+/-3.22% vs 5.41%+/-1.54%, P less than 0.05). Compared to MCD+ group, the expression of FasL was dramatically increased on day 10 and in week 3 in MCD- mice both at the mRNA and protein levels (P less than 0.05 and P less than 0.01). Expression of Fas mRNA was up-regulated in weeks 3 and 8 (P less than 0.01), and expression of Fas in protein level was higher at week 8 (P less than 0.01) in MCD- group. Expression of caspase-8 significantly increased at the mRNA level at week 3 and week 8 (P less than 0.01 and P less than 0.05 respectively) and at the protein level at week 8 (P less than 0.05) in MCD- group. In all of the time points except for day 5, caspase-3 activities were significantly more enhanced in MCD- group than that in MCD+ group (P less than 0.05).</p><p><b>CONCLUSIONS</b>In our experimental NASH model, hepatic apoptosis was frequently detected. Increased apoptosis was probably attributable to up-regulation of apoptosis-related genes, such as FasL/Fas system, and activation of the caspase pathway. These changes may provoke hepatic apoptosis and the development of inflammation and fibrosis.</p>

Expression and purification of ATP sulfurylase from Saccharomyces cerevisias in Escherichia coli and its application in pyrosequencing / 生物工程学报

ATP sulfurylase (ATPS,EC 2.7.7.4) reversibly catalyzes the reaction between ATP and sulfate to produce APS and pyrophosphate (PPi), and has been used in pyrosequencing. The gene coding ATP sulfurylase was amplified from the genomic DNA of Saccharomyces cerevisias (CICC 1202), and cloned into prokaryotic expression plasmid pET28a( + ) to provide a recombinant expression plasmid pET28a( + )-ATPS. Upon IPTG induction, ATP sulfurylase was produced by E. coli BL21 (DE3) harboring the recombinant expression plasmid pET28a( + )-ATPS. The relative molecular weight of recombinant ATP sulfurylase with His tag was about 60 kD. The recombinant ATP sulfurylase with electrophoretic pure grade was obtained only by two purification steps: His * Bind Resin affinity chromatography and ultrafiltration. The specific activity of the purified recombinant ATP sulfurylase was as high as 5.1 x 10(4) u/mg. The successful application of the enzyme in pyrosequencing was also demostrated.

The intervention-therapeutic effect of lipiodol-arsenic trioxide emulsion for primary hepatic carcinoma / 介入放射学杂志

Objective To discuss the short-term clinical effect including functional change of lipiodol- arsenic trioxide emulsion on the primary hepatic carcinoma.Methods Fifty-two patients undergone arterial chemoemblization were selected and then randomly divided into two groups:treatment group(n=27)and control group(n=25).Patients in treatment group were treated with lipiodol-arsenic trioxide,while those in control group treated with mitomycin,epirubicin,cisplatin or lipiodol.Clinical symptoms and six liver function parameters were observed and analized.Results The clinical symptoms of patients in treatment group improved much better than those in control group,and the liver function impairment of patients in treatment group also decreased more than those in control group.Conclusions Lipiodol-arsenic trioxide is an effective and safe intervention-therapeutic embolization material for primary hepatic carcinoma.

Experimental study on metastasis and correlation of metastasis with angiogenesis of hepatic tumor in rabbits after transcatheter arterial chemoembolization using As_2O_3 and lipiodol / 中华放射学杂志

Objective To investigate the effect of transcatheter arterial chemoembolization(TACE) using As_2O_3 and Lipiodol on the growth and metastasis of the implanted hepatic tumor in rabbits and the correlation of metastasis with angiogenesis of the residual tumor.Methods Forty-eight New Zealand white rabbits were randomly divided into four groups and VX_2 carcinoma was implanted in the left lobes of the livers.Two weeks later,a catheter was inserted into the hepatic artery and infusion was performed via the hepatic artery using physiological saline(group A),Lipiodol(group B),ADM-Lipiodol(group C),and As_2O_3-Lipiodol(group D),respectively.One week after the treatment,the value of microvessel density (MVD)of tumors(samples got by biopsy)was examined by immunohistochemistry.Three weeks after the treatment,the volume and necrotic area of the implanted tumor were measured.The metastases in the liver, lungs and other organs were recorded.Results One week after the treatment,the value of MVD of the tumorswas(21.8?5.3),(23.4?3.9),(22.4?4.5),and(14.3?3.4)/400 power LM(F= 11.246,P=0.000).Three weeks after the treatment,the mean volume of the implanted tumor was (35.5?7.1),(21.2?8.3),(20.7?9.1),and(11.8?3.7)cm~3(F=21.203,P=0.0000)in groups A,B,C and D,respectively.There was significant difference between group D and group B(q= 4.398,P

The X-ray findings of spondylometaphyseal dysplasia / 中华放射学杂志

Objective To explore the clinical,radiological findings and pathogenic factor of spondylometaphyseal dysplasia.Methods Five cases were reported and the relevant documents were studied retrospectively.Plain X-ray film was performed in all patients.Results There were 4 male and 1 female,age ranged from 4 to 15 years with average of 9 years.The main features of 5 cases included delayed bone age,stature short,short trunk,waddling gait noted,scoliosis in 2 cases,kyphoscoliosis in 1 case,severe genu valgum in 2 cases.The main X-ray appearance of 5 cases is multiple irregularities of long bones metaphyses associated with platyspondylia,epiphysis is normal.Type Ⅰ SMD in 2 cases,Type Ⅱ SMD in 1 case,Type Ⅲ SMD in 2 cases.Conclusion When we meet children with delayed bone age and stature short,and muhiple irregularities of long bones metaphyses associated with platyspondylia were seen in X-ray plain film.we should think about spondylometaphyseal dysplasia.