Single-cell transcriptomic analysis of radiation-induced lung injury in rat.

Radiation-induced lung injury (RILI) frequently occurs as a complication following radiotherapy for chest tumors like lung and breast cancers. However, the precise underlying mechanisms of RILI remain unclear. In this study, we generated RILI models in rats treated with a single dose of 20 Gy and examined lung tissues by single-cell RNA sequencing (scRNA-seq) 2 weeks post-radiation. Analysis of lung tissues revealed 18 major cell populations, indicating an increase in cell-cell communication following radiation exposure. Neutrophils, macrophages, and monocytes displayed distinct subpopulations and uncovered potential for pro-inflammatory effects. Additionally, endothelial cells exhibited a highly inflammatory profile and the potential for reactive oxygen species (ROS) production. Furthermore, smooth muscle cells (SMC) showed a high propensity for extracellular matrix (ECM) deposition. Our findings broaden the current understanding of RILI and highlight potential avenues for further investigation and clinical applications.

Foxp3 expression in CD4+CD25+Foxp3+ regulatory T cells promotes development of colorectal cancer by inhibiting tumor immunity.

The mechanism underlying CD4+CD25+Foxp3+ regulatory T cells (Tregs) promoting the development of colorectal cancer (CRC) was elucidated in the present study. Forty-eight cases of colorectal carcinomas, 22 cases of colon polyps and 21 cases of normal colorectal tissues were collected. The correlation among Foxp3, IL-10 and Stat3, and the clinical relevance of these three indexes were analyzed. The results showed that the levels of Foxp3 expressed in infiltrating CD4+CD25+Foxp3+Tregs, and IL-10 and Stat3 in CRC tissues were all significantly higher than those in polypus tissues and normal colon tissues (P< 0.01). Pearson correlation analysis indicated that the expression level of Foxp3 was positively correlated with Stat3 at mRNA level (r=0.526, P=0.036), and was positively correlated with IL-10 at protein level (r=0.314, P=0.030). The Foxp3 expressed in CD4+CD25+Foxp3+Tregs was correlated with the histological grade, lymph node metastasis and TNM stage of CRC (P<0.05 for all). The IL-10 expression was correlated with the histological grade and TNM stage (both P<0.05). The Stat3 expression was correlated with the lymph node metastasis and TNM stage (both P<0.05). It was concluded that CD4+CD25+Foxp3+Tregs can inhibit tumor immunity in combination with some other related inhibitory cytokines and that Foxp3 expression in CD4+CD25+Foxp3+Tregs correlates with CRC progression.

Altered expression profile of micrornas in gastric stromal tumor.

MicroRNAs (miRNAs) play important roles in carcinogenesis, but the global miRNA expression profile in gastric stromal tumor tissues remains unclear. This study was to examine the miRNA expression profile in gastric stromal tumor tissues and explore the function of dysregulated miRNAs by performing gene ontology (GO) and pathway enrichment analysis. Total RNA was extracted and purified from 3 pairs of frozen gastric stromal tumor tissues and the adjacent non-tumor tissues by using mirVana™ miRNA isolation kit. The miRNA expression was analyzed with Affymetrix microarrays (version 4.0) containing 2578 human mature microRNA probes. The dysregulated microRNAs were validated by quantitative RT-PCR in 30 pairs of gastric stromal tumor tissues. The target gene of the dysregulated microRNAs was predicted by miRanda, TargetScan and PicTar. GO and pathway enrichment analysis was conducted to examine the potential function of miR-3178 and miR-193a-5p. The results showed that there were 12 differently expressed microRNAs in gastric stromal tumor tissues, among which 10 miRNAs were down-regulated, and 2 were up-regulated (P<0.05). The validation results by RT-PCR were in accordance with those by microRNA microarry. GO analysis found that the target genes of miR-3178 were involved in 5 GO terms and those of miR-193a-5p in 7 GO terms in level 2. Pathway enrichment analysis suggested that miR-3178 and miR-193a-5p were related to 57 and 122 signaling pathways, respectively. It was concluded that gastric stromal tumor displays a unique miRNA signature. This specific expression may become a new diagnostic and prognostic biomarker for gastric stromal tumor. miR-3178 and miR-193a-5p function as suppressive microRNAs, and they may also become diagnosis and treatment targets for gastric stromal tumor.

EUS-Guided FNA for Diagnosis of Pancreatic Cystic Lesions: a Meta-Analysis.

BACKGROUND: Preoperative diagnosis of pancreatic cystic lesions (PCLs) must be reliable as the current standard treatment, major or total pancreatectomy, dramatically affects quality of life. Additionally, early diagnosis of malignancy is essential to an improved prognosis. The diagnostic accuracy of fluid analysis using endoscopic ultrasonography-guided fine-needle aspiration (EUS-FNA) has been demonstrated in pancreatic solid lesions. The utility of this technique in the diagnosis of PCLs is still unknown. METHODS: A comprehensive search was performed in multiple databases. Studies differentiating benign and malignant PCLs via EUS-FNA were included in this meta-analysis. The quality of diagnostic accuracy studies (QUADAS) was adopted to evaluate the selected studies. Pooled sensitivity, specificity, likelihood ratio, diagnostic odds ratio, and summary receiver operating characteristic (sROC) curve analyses were conducted. Two main classification types of malignancy were characterized and analyzed. We also generated a subgroup analysis of available clinical factors. Publication bias was evaluated by Begg's and Egger's tests. RESULTS: Sixteen studies containing 1024 subjects have been published. The pooled sensitivity for malignant cytology according to classification 1 was 0.51 (95% CI, 0.45-0.58), and pooled specificity was 0.94 (95% CI, 0.92-0.96). When the detected PCLs were identified as classification 2, suspicious malignancy or potential malignancy, sensitivity and specificity were similar, 0.52 (95% CI, 0.46-0.57) and 0.97 (95% CI, 0.95-0.98) respectively. CONCLUSION: This meta-analysis demonstrates that EUS-FNA is a reliable clinical tool for the diagnosis of PCLs. However, a more accurate algorithm is needed to reduce various biases and to improve the sensitivity of EUS-FNA in the detection of malignant PCLs.

Altered MiRNA expression in gastric cancer: a systematic review and meta-analysis.

BACKGROUND/AIMS: Aberrant microRNA expression has the potential to be used for early diagnosis of gastric cancer or to predict survival and treatment response. This study performed a systematic review and meta-analysis of altered miRNAs in gastric cancer in order to assess the use of miRNAs as novel biomarkers for early detection and prognosis prediction of gastric cancer. METHODS: We retrieved published articles from the PubMed online database and obtained different sets of data on miRNAs expression profiling in gastric cancer and highlighted the most frequently dysregulated miRNAs in gastric cancer. We then extracted studies that used quantitative RT-PCR and then pooled them together by using meta-disc software (version 1.4). RESULTS: We found that there were 47 aberrantly expressed miRNAs in gastric cancer (29 up-regulated and 18 down-regulated) that were most frequently reported in the literature. In publications that provided information on specific miRNA expression vs. diagnostic value, the pooled data showed good sensitivity and specificity as well as high levels of overall accuracy. However, specimen types could be a factor that introduces substantial heterogeneity. Published studies also showed association of altered miRNA expression with clinicopathological data from gastric cancer patients. CONCLUSION: Thus, various miRNAs are differentially expressed in gastric cancer and some of them could be further evaluated as biomarkers for early diagnosis of gastric cancer and prediction of prognosis or treatment response.

Accuracy of early detection of colorectal tumours by stool methylation markers: a meta-analysis.

AIM: To evaluate the accuracy of methylation of genes in stool samples for diagnosing colorectal tumours. METHODS: Electronic databases including PubMed, Web of Science, Chinese Journals Full-Text Database and Wanfang Journals Full-Text Database were searched to find relevant original articles about methylated genes to be used in diagnosing colorectal tumours. A quality assessment of diagnostic accuracy studies tool (QADAS) was used to evaluate the quality of the included articles, and the Meta-disc 1.4 and SPSS 13.0 software programs were used for data analysis. RESULTS: Thirty-seven articles met the inclusion criteria, and 4484 patients were included. The sensitivity and specificity for the detection of colorectal cancer (CRC) were 73% (95%CI: 71%-75%) and 92% (95%CI: 90%-93%), respectively. For adenoma, the sensitivity and specificity were 51% (95%CI: 47%-54%) and 92% (95%CI: 90%-93%), respectively. Pooled diagnostic performance of SFRP2 methylation for CRC provided the following results: the sensitivity was 79% (95%CI: 75%-82%), the specificity was 93% (95%CI: 90%-96%), the diagnostic OR was 47.57 (95%CI: 20.08-112.72), the area under the curve was 0.9565. Additionally, the results of accuracy of SFRP2 methylation for detecting colorectal adenomas were as follows: sensitivity was 43% (95%CI: 38%-49%), specificity was 94% (95%CI: 91%-97%), the diagnostic OR was 11.06 (95%CI: 5.77-21.18), and the area under the curve was 0.9563. CONCLUSION: Stool-based DNA testing may be useful for noninvasively diagnosing colorectal tumours and SFRP2 methylation is a promising marker that has great potential in early CRC diagnosis.

Detection of promoter hypermethylation of Wnt antagonist genes in fecal samples for diagnosis of early colorectal cancer.

AIM: To investigate the feasibility of detecting aberrantly hypermethylated Wnt-antagonist gene promoters (SFRP2 and WIF-1) in fecal DNA as non-invasive biomarkers for early colorectal cancer (CRC). METHODS: The methylation-specific polymerase chain reaction assay was performed to blindly analyze the methylation status of SFRP2 and WIF-1 gene promoters in fecal samples from 48 subjects with CRC, 35 with adenomas, 32 with hyperplastic polyps and 30 endoscopically normal subjects. Additionally, we compared the diagnostic efficiency of measuring the hypermethylated SFRP2 and WIF-1 genes in the feces to the fecal occult blood test (FOBT) for the early detection of CRC. RESULTS: Hypermethylated SFRP2 was detected in the feces of 56.3% (27/48) of CRC cases, 51.4% (18/35) of adenoma cases and 12.5% (4/32) of patients with hyperplastic polyps. The hypermethylation of WIF-1 was detected in 60.4% (29/48), 45.7% (16/35) and 18.7% (6/32) of fecal samples from CRC, adenoma and hyperplastic polyp patients, respectively. At least one hypermethylated gene was detected in 81.3% (39/48) of CRC and 65.7% (23/35) of adenoma samples. In contrast, only a hypermethylated WIF-1 gene was detected in one case of normal fecal samples. Moreover, no significant associations were observed between SFPR2 and WIF-1 hypermethylation and clinicopathological features. Additionally, 81.8% of CRC cases diagnosed as Dukes A stage or advanced adenomas had at least one hypermethylated gene detected, while the detection rate with the FOBT was only 31.8% (P < 0.001). CONCLUSION: Hypermethylated SFRP2 and WIF-1 genes in fecal DNA are novel and promising molecular biomarkers that have great diagnostic potential for early CRC.

[Therapeutic effects of muscovite on ulcerative colitis in rats].

OBJECTIVE: To explore the efficacy of muscovite on iodoacetamide -induced ulcerative colitis in rats and elucidate its possible mechanism. METHODS: Ulcerative colitis was induced in female Sprague-Dawley (SD) rats by an intracolonic injection of iodoacetamide. A total of 48 rats were divided randomly(by the method of random digits table) into 6 groups: control group, model group, low-dose muscovite group (360 mg/kg), high-dose muscovite group (720 mg/kg), 5-aminosalicylie acid (5-ASA) group and muscovite plus 5-ASA group (combined treatment), and each group had 8 rats. The body weight, disease activity index (DAI), macroscopic damage and microscopic score of rats in each group were subsequently evaluated after dosing for 7 days. The protein levels of tumor necrosis factor-α (TNF-α), interleukin-8 (IL-8) and myeloperoxidase (MPO) activity were detected by enzyme-linked immunosorbent assay(ELISA) while the activity of nuclear facor(NF)-κB was determined by immunohistochemistry.One way ANOVA and rank-sum test were used. RESULTS: After doing, body weight macroscopic damage, microscopic score, TNF-α concentration, MPO and NF-κB activity of rats in each group were all significantly correlated with the dose of muscovite (r = 0.573, -0.647, -0.569, -0.681, -0.811, -0.842, all P < 0.05). High-dose muscovite group had no significant difference with 5-ASA group in body weightt, DAI, macroscopic damage, microscopic score, IL-8 concentration, TNF-α concentration, MPO and NF-κB activities((166 ± 5) vs (167 ± 5) g, 0.33 (0.00, 1.17) vs 0.17 (0.00, 0.83), 2.50 (2.00, 4.00) vs 3.00 (2.00, 3.00), 3.00 (2.00, 3.00) vs 2.50 (2.00, 3.00), (109 ± 17) vs (111 ± 15) pg/ml, (166 ± 38) vs (155 ± 45) pg/ml, (52 ± 6) vs (49 ± 4) U/g, 7.39 ± 0.42 vs 7.41 ± 0.34, all P > 0.05). The MPO and NF-κB activities of combined treatment group were lower than those of 5-ASA group((40 ± 4) vs (49 ± 4) U/g, 4.67 ± 0.72 vs 7.41 ± 0.34, all P < 0.05). However, other indices showed no significant difference with 5-ASA group (all P > 0.05). CONCLUSIONS: Rectal administration of muscovite ameliorates colonic inflammation of iodoacetamide-induced colitis. Its underlying mechanism is probably due to the regulation of inflammatory response. Muscovite may be a potential therapeutic agent for treatment of ulcerative colitis.

Effects of matrine on the proliferation of HT29 human colon cancer cells and its antitumor mechanism.

Matrine is one of the main active components that is extracted from the dry roots of Sophora flavescens. The compound has potent antitumor activity in various cancer cell lines. However, the anticancer activity of matrine in colon cancer cells remains unclear. The purpose of the present study was to investigate the effects of matrine on the growth of human colon cancer cells and the expression of the associated proteins. Cancer cell proliferation was measured by 3-(4,5-dimethylthiazolyl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. The cell cycle distribution and apoptosis were analyzed by flow cytometry (FCM). The activation of the caspases and the expression of pro-apoptotic and anti-apoptotic factors were examined using western blot analysis. Matrine was shown to significantly inhibit the proliferation of HT29 cells in a dose- and time-dependent manner, and also to reduce the percentage of cells in the G2/M phase, which was most frequently associated with an increase of cells arrested in the G0/G1 phase of the cell cycle. Western blot analysis revealed that matrine induced the activation of caspase-3 and -9 and the release of cytochrome C (Cyto C) from the mitochondria to the cytosol. Furthermore, the pro-apoptotic factor, Bax, was upregulated and the anti-apoptotic factor, Bcl-2, was downregulated, eventually leading to a reduction in the ratio of Bcl-2/Bax proteins. The results demonstrated that matrine inhibits proliferation and induces apoptosis of HT29 human cells in vitro. The induction of apoptosis appears to occur through the upregulation of Bax, the downregulation of Bcl-2, the release of Cyto C from the mitochondria to the cytosol and the activation of caspase-3 and caspase-9, which subsequently trigger major apoptotic cascades. Matrine has potent antitumor activity in HT29 cells and may be used as a novel effective reagent in the treatment of colon cancer.

Roles of lipoxin A4 in preventing paracetamol-induced acute hepatic injury in a rabbit model.

The objective of this research is to investigate the potential role of lipoxin A4 in preventing paracetamol (PCM)-induced hepatic injury. One hundred male New Zealand white rabbits were randomly divided into control group, PCM group, N-acetylcysteine (NAC) group, lipoxin A4 (LXA4) group, and LXA4 + NAC group. The rabbits were assigned to receive 300 mg/kg weight PCM in 0.9 % saline or equivalent volume of saline via gastric lavage. LXA4 (1.5 µg/kg) and equivalent volume of 2 % ethanol were separately given to the rabbits in LXA4-treated and PCM groups 24 h after PCM administration. Meanwhile, the rabbits in the NAC-treated groups received a loading dose of 140 mg/kg of N-acetylcysteine. The blood samples and liver tissue were collected for biochemical and histological evaluation 36 h after paracetamol administration. The administration of LXA4 24 h after paracetamol poisoning resulted in significant improvement in hepatic injury as represented by decrease of hepatocellular enzyme release and attenuation of hepatocyte apoptosis and necrosis. In LXA4-treated groups, the expression of TNF-α was significantly lower than those in PCM and NAC groups (p < 0.05). In contrast, the level of IL-10 was significantly higher than PCM and NAC groups (p < 0.05). Moreover, the expressions of NF-κB p65 in PCM and NAC groups were significantly increased compared with those of LXA4-treated groups and control group (respectively, p < 0.05 and p < 0.01). LXA4-treated groups also showed significantly higher survival rates. Lipoxin A4 significantly mitigates paracetamol-induced hepatic injury, in which anti-inflammation effect may play an important role, leading to hepatic apoptosis and necrosis.

Treating TNBS-induced colitis in rats with probiotics.

BACKGROUND/AIMS: We aimed to investigate the therapeutic effects of Peifeikang, a probiotics compound, on colitis in rats induced by trinitrobenzene sulfonic acid and to elucidate its potential mechanism. MATERIAL AND METHODS: We evaluated the therapeutic effects of Peifeikang by analysis of the disease activity index, colonic mucosa damage index, and histopathological score of the inflamed colons, by measurement of colonic myeloperoxidase activity through spectrophotometric assay, by determination of colonic positivities of tumor necrosis factor-alpha and interleukin-10 via immunochemical staining, and by detection of serum levels of tumor necrosis factor-alpha and interleukin-10 with enzyme-linked immunosorbent assay. RESULTS: Intrarectal administration of trinitrobenzene sulfonic acid caused colonic inflammation similar to human ulcerative colitis with significantly increased disease activity index, colonic mucosa damage index, histopathological score, and colonic myeloperoxidase activities (p<0.05). After treatment with Peifeikang or olsalazine alone, or both, disease activity index, colonic mucosa damage index, histopathological score, and colonic myeloperoxidase activities decreased significantly (p<0.05). The relief of trinitrobenzene sulfonic acid-induced colitis was accompanied by significantly decreased production of tumor necrosis factor-alpha in both serum and intra-colon (p<0.05) and by significantly increased production of interleukin-10 in both serum and intra-colon (p<0.05). CONCLUSIONS: Peifeikang can effectively ameliorate trinitrobenzene sulfonic acid-induced colitis in rats, the underlying mechanism of which may be attributed to the modulatory effects of Peifeikang on the production of tumor necrosis factor-alpha and interleukin-10.

Involvement of mitochondrial pathway in NCTD-induced cytotoxicity in human hepG2 cells.

BACKGROUND: Norcantharidin, the demethylated analog of cantharidin derived from a traditional Chinese medicine, Mylabris, has been used in the treatment of anti-cancer effects. However, the detailed mechanisms underlying this process are generally unclear. The aim of this study was to investigate the mechanism of NCTD-induced apoptosis in HepG2 cells. METHODS: The cytotoxicity was measured by MTT assay for cellular viability and by flow cytometry. The mitochondrial membrane potential and reactive oxygen species production was evaluated by flow cytometry analysis. The role of caspase activities were assayed using caspase apoptosis detection kit . Western blot analysis was used to evaluate the level of Cyto-C, Bcl-2, Bax, Bid, caspase 3, -9, -8 and PARP expression RESULTS: After treatment with NCTD, a decrease in the viability of HepG2 cells and increase in apoptosis were observed. NCTD-induced apoptosis was accompanied by an increase in ROS production, loss of mitochondrial membrane potential and release of cytochrome c(cyto-c) from the mitochondria to the cytosol and down-regulation of anti-apoptotic protein Bcl-2 levels with concurrent up-regulation in pro-apoptotic protein Bax levels. However, another pro-apoptotic molecule, Bid, showed no change in such same treatment. NCTD-increased activity of caspase 9,caspase 3 and the subsequent cleavage caspase substrate PARP were also observed. The expression levels of pro-caspase-8 were not changed after NCTD treatment. CONCLUSION: These results indicate that NCTD induced cytotoxicity in HepG2 cells by apoptosis, which is mediated through ROS generation and mitochondrial pathway.

[Probiotic therapy using live combined bifidobacterium, lactobacillus and enterococcus for experimental colitis in rats model].

OBJECTIVE: To evaluate the effect of live combined bifidobacterium, lactobacillus and enterococcus capsules for colitis in rats induced by trinitrobenzenesulfonic acid (TNBS), so as to explore a new therapy for inflammatory bowel diseases (IBD). METHODS: 50 female adult Sprague-Dawley rats were randomly divided into 5 groups i.e. normal control group (G1), untreated TNBS-induced colitis (G2), TNBS-induced colitis treated with live combined bifidobacterium, lactobacillus and enterococcus (G3), TNBS-induced colitis treated with olsalazine (G4) and TNBS-induced colitis treated with both live combined bifidobacterium, lactobacillus and enterococcus and olsalazine at the same dose and duration (G5). Each group received its respective treatment. Serum levels of C-reactive protein (CRP), TNFα and IL-10 were measured with ELISA, colonic myeloperoxidase (MPO) activity was determined with spectrophotometric method, histopathologic picture of the colon of each rat was studied with microscope and colonic mucosa damage index (CMDI) was recorded. RESULTS: Serum CRP, TNFα, IL-10, CMDI and colonic MPO in G1 were significantly lower than those in G2 (P < 0.001) with normal colonic architecture.G2 exhibited the most severe colonic inflammation and the highest levels of CRP, TNFα, IL-10, CMDI and colonic MPO with staszica significance. Treatment groups G3, G4 and G5 showed more obvious colonic inflammatory remission and lower levels of serum CRP, TNFα, IL-10 and colonic MPO, G5 being most notable when compared to G2 with staszica significance. In G2, serum levels of CRP, TNFα, IL-10 and colonic MPO activity each correlated positively with CMDI (P < 0.001). CONCLUSIONS: Live combined bifidobacterium, lactobacillus and enterococcus can effectively ameliorate colitis in rats induced by TNBS; the underlying mechanism may possibly be associated with the serum levels of cytokines.

Expression and functional research of TLR4 in human colon carcinoma.

INTRODUCTION: To verify whether human colon carcinoma cells can express Toll-like receptor 4 (TLR4) and investigate the biological function of TLR4 on human colon carcinoma cells. METHODS: Human colon carcinoma cells SW480 was cultured with RPMI 1640 medium. The expression of TLR4 was analyzed by reverse transcription polymerase chain reaction and quantified by flow cytometry. Cell proliferation was evaluated by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay. The production of transforming growth factor-beta, vascular endothelial growth factor, and interleukin-8 induced by lipopolysaccharide (LPS)-the TLR4 ligand was tested by enzyme-linked immunosorbent assay. Whether mitogen-associated protein kinases and nuclear factor-kappaB (NF-kappaB) proteins were activated was analyzed by Western blot. Apoptosis was analyzed by Annenxin V/PI staining. RESULTS: TLR4 is expressed on SW480 cells. LPS could not affect TLR4 expression and the proliferation of SW480 cells. LPS increased the phosphorylation of ERK1/2 and P38 and activated NF-kappaB. LPS promoted the production of vascular endothelial growth factor, transforming growth factor-beta, and interleukin-8. In addition, LPS induced resistance of SW480 cells to tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis. Furthermore, NF-kappaB activation was necessary for apoptosis resistance of SW480 cells induced by LPS. CONCLUSION: TLR4 is expressed on human colon carcinoma cells and functionally active. It may play important roles in promoting immune escape of human colon carcinoma cells by inducing immunosuppressive factors and apoptosis resistance.

[Association of hemorrhagic fever with renal syndrome and HLA-DRB allele polymorphisms in Han nationality, Hubei province].

OBJECTIVE: To understand the association of human leukocyte antigen (HLA)-DRB polymorphism and patients diagnosed as hemorrhagic fever with renal syndrome (HFRS). METHODS: HLA-DR allele polymorphism was detected by PCR-sequence specific primers (PCR-SSP). Hantavirus (HV) typed as Hantaan virus (HTNV) and Seoul virus (SEOV) in patients were detected by RT-heminested PCR. RESULTS: The gene frequency of DRB1*0401-0411, *1001 and *1101-1105 in HFRS case group were 3.1%, 2.2% and 15.7% respectively. Compared with control group, it was significant higher in HFRS case group (RR = 13.87, 9.72 and 2.00 respectively with Chi-square value as 10.006, 6.324 and 6.472 respectively, P < 0.05). When compared with HFRS case group, the gene frequency of DRB1*1501-1502, DRB4 and DRB5 in control group were 11.0%, 19.0% and 16.9% respectively, markedly lower than in patients (RR = 0.45, 0.58 and 0.23 respectively. Chi-square values were 6.138, 4.583 and 21.076 respectively, P < 0.05). There was no significant difference in other HLA-DR gene frequencies. Mixed infection was found in Hubei, with HTNV slightly more than SEOV. Distinct hantaviruses could coexist in either different or the same geographic or ecological zores in Hubei province. Patients with HLA-DRB1*1101-1105 alleles were 81.8% (27/33) infected by HTNV and only 18.2% infected by SEOV, which had significant difference (P < 0.05). CONCLUSION: DRB1*0401-0411, *1001 and *1101-1105 were possibly associated with increased susceptibility to HV infection. On the other hand there was an inverse correlation among HFRS, DRB1*1501-1502, DRB4 and DRB5.

Targeting the most upstream site of Wnt signaling pathway provides a strategic advantage for therapy in colorectal cancer.

The Wnt signaling pathway has important functions in development, tissue homeostasis, and regeneration. Deregulation of canonical Wnt/beta-catenin signaling is frequently found in various human cancers, particularly in colorectal cancer, and non-canonical Wnt signaling pathways also have been implicated in neoplasia. Colorectal cancer is a multi-pathway disease. Activation of Wnt signaling by both genetic and epigenetic alterations has been found to be important for both, initiation and progression of colorectal cancer. In addition, since Wnt signaling results in diverse downstream intracellular events, targeted inhibition of Wnt/beta-catenin signaling at the most upstream site of this pathway is a rational and an advantageous new approach for the therapy of colorectal cancer.

Epigallocatechin-3-gallate suppressed the over-expression of HSP 70 and MDR1 induced by heat shock in SGC 7901.

This study investigated the effects of epigallocatechin-3-gallate (EGCG) on the expression of HSP 70 and MDR 1. SGC-7901 cells were cultured with RPMI 1640 medium. The single or combined effects of EGCG (0.1, 1, 10, 20, and 40 micromol/L) and heat shock were examined by MTT assay. The expression of HSP 70 and MDR 1 was semi-quantified by the reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry method (SP staining). EGCG suppressed cell proliferation at a time- and dose-dependent manner. The effects of combined treatment with EGCG and heat shock on the growth of SGC-7901 cells were stronger than single effects of EGCG. After using EGCG for 24 h, 48 h and 72 h, the IC50s were 112.5 micromol/l, 21.41 micromol/l and 5.24 micromol/l, respectively. Heat shock stimulated the over-expression of HSP 70, especially after heat shock for 8 h, as well as MDR1 after heat shock for 24 h. But EGCG suppressed the over-expression induced by heat shock. The authors conclude that EGCG inhibited the proliferation of SGC-7901, and EGCG combined with heat shock strengthened the effects. Heat shock weakened the over-expression of HSP 70 and MDR1; however, EGCG suppressed the over-expression of HSP 70 and MDR1 induced by heat shock. EGCG combined with heat shock may enhance the sensitivity of drugs to tumors.

[The role of secreted Wnt-antagonist genes hypermethylation in early detection of colorectal tumor].

OBJECTIVE: To investigate the functions of promoter hypermethylation of secreted Wnt-antagonist genes in colorectal tumorigenesis and progression. METHODS: Two colorectal cancer cell lines, HCT116 and SW480, were treated by 5-aza-2'-deoxycytidine (DAC) and trichostatin A (TSA) for demethylation. The promoter hypermethylation and expression of sFRP and WIF-1 genes in different stages of colorectal tumor and colorectal cancer cell lines were detected by methylation-specific PCR and reverse transcription PCR, respectively. RESULTS: None of the normal colorectal mucosa samples showed methylated bands of any sFRP and WIF-1genes. Hypermethylation of sFRP1, 2, 4, 5 and WIF-1 was detected in 93.1% (67/72), 83.3% (60/72), 36.1% (26/72), 52.8% (38/72) and 84.7% (61/72) of adenocarcinomas, 87.9% (29/33), 81.8% (27/33), 24.2% (8/33), 57.6% (19/33) and 72.7% (24/33) of adenomas, 52.6%, 28.9%, 2.6%, 18.4%, 23.7% of the adjacent normal mucosa. Methylation was more frequently found in colorectal tumors than in normal mucosa and adjacent normal mucosa from patients with tumor (P < 0.05). No significant association between Wnt-antagonist genes hypermethylation and clinicopathological characteristics was found (P > 0.05). SFRP1, 2, 4, 5 and WIF-1 genes were methylated in HCT116 cell line. SFRP1, 2 and WIF-1 were methylated in SW480 cell line. The mRNA expression of sFRPs and WIF-1 genes was absent or significantly downregulated (P < 0.01) when they were methylated in two colorectal cancer cell lines. SFRP3 was expressed in two colorectal carcinoma cell lines. DAC/TSA combination treatment re-expressed the silenced sFRPs and WIF-1 genes mRNA expressions effectively. A single application of TSA could not re-express sFRPs and WIF-1 genes mRNA expressions. The influence of demethylation treatment on sFRP3 expression was minimal. CONCLUSION: Hypermethylation of Wnt-antagonist genes is a common early event in the evolution of colorectal tumor. Methylation of sFRP1, 2, 5 and WIF-1 genes might serve as biomarkers for the early detection of colorectal tumor.

Synergistic effect of triptolide combined with 5-fluorouracil on colon carcinoma.

AIM: To investigate the inhibitive effects of triptolide (TPL) combined with 5-fluorouracil (5-FU) on colon carcinoma HT-29 cells in vitro and in vivo and their side effects. METHODS: HT-29 cells were cultured with RPMI 1640 medium. The single or combined effects of TPL and 5-FU on HT-29 cells were examined by MTT assay, flow cytometry. The combined effects were evaluated by the median-effect principle. The model of tumour xenografts was established in nude mice. TPL 0.25 mg/kg/day and 5-FU 12 mg/kg/day, either in combination or on their own, were injected into mice and the inhibitive effects and side effects were observed. RESULTS: TPL and 5-FU either combined or alone inhibited significantly the proliferation of HT-29 cells and induced obvious apoptosis. Mean (SD) growth inhibition rate reached 94.92 (2.76)% and the apoptic rate at 48 h reached 41.71 (1.38)%. The combined effects were synergistic (CI<1) at lower concentrations. TPL or 5-FU alone inhibited significantly the growth of tumour xenografts and the inhibition rates were 78.53% and 84.16%; the drugs combined had more significant effect, the tumour inhibition rate reaching 96.78%. During the course of chemotherapy, no obvious side effect was observed. CONCLUSION: The combined effects of TPL and 5-FU on the growth of colon carcinoma in vitro and in vivo were superior to the effects when the agents were used individually. TPL combined with 5-FU had synergistic effects at lower concentrations and promoted apoptosis, but did not increase the side effects of chemotherapy.

[Hypermethylation and regulation of expression of secreted frizzled-related protein genes in colorectal tumor].

OBJECTIVE: To investigate the functions of promoter hypermethylation of secreted frizzled-related protein (sFRP) genes in colorectal tumorigenesis and progression. METHODS: Three colorectal cancer cell lines, RKO, HCTll6 and SW480, were treated hy 5-aza-2'-deoxycytidine and trichostatin A for demethylation. The promoter hypermethylation and expression of sFRP genes in colorectal tumor tissue and colorectal cancer cell lines were detected hy methylation-specific PCR and reverse transcription PCR, respectively. RESULTS: None of the normal colorectal mucosa tissues showed methylation of sFRP genes. sFRP1, 2, 4 and 5 were frequently methylated in colorectal adenocarcinoma, adenoma and aberrant crypt foci (ACF) (sFRP1 > 85%, sFRP2 > 75%, sFRP5 > 50%), the differences between any two of them were not significant (P >0.05). Methylation was more frequent in colorectal tumors than in normal mucosa and adjacent normal mucosa from patients with tumor. Hypermethylation of sFRP genes was present in three colorectal cancer cell lines. When sFRP genes were methylated, their corresponding mRNA expression was absent. After cells were treated by DAC/TSA combination, the silenced sFRP expression could be effectively re-expressed. CONCLUSION: Hypermethylation of sFRP genes is a common early event in the evolution of colorectal tumors that occurs frequently in ACF. Methylation of sFRP1, 2 and 5 genes might serve as biomarkers for the early detection of colorectal tumors. Demethylation can effectively reverse gene expression that appears possibly to be an effective way for tumor therapy.