Exosomal Long Noncoding RNAs are Differentially Expressed in the Cervicovaginal Lavage Samples of Cervical Cancer Patients.

BACKGROUND: As the second leading cause of cancer morbidity and death in women, cervical cancer remains an important public health problem worldwide. Novel biomarkers with high sensitivity and specificity for the early detection and diagnosis of cervical cancer are urgently needed. Increasing evidence shows that long noncoding RNAs (lncRNAs) are differentially expressed in cancer tissues and may serve as diagnostic markers. In multiple tumor types, exosomes harboring lncRNAs are actively released from tumor cells. In this study, we investigate the potential association of exosomal lncRNA expression with cervical cancer. METHODS: Cervicovaginal lavage specimens were collected from patients with cervical cancer and cancer-free volunteers who are HPV-positive or HPV-negative. Exosomes in these specimens were isolated by ultracentrifugation and confirmed by transmission electron microscopy. The exosomal lncRNAs HOTAIR, MALAT1, and MEG3 were quantified by qRT-PCR. RESULTS: Expression of HOTAIR, MALAT1 and MEG3 was predominantly observed in cervical cancer-derived exosomes in cervicovaginal lavage samples. The expression levels of lncRNAs were significantly different in exosomes isolated from cervical cancer patients compared to normal controls. CONCLUSIONS: Our data suggest that lncRNAs in exosomes isolated from cervicovaginal lavage are differentially expressed in cervical cancer patients and cancer-free volunteers. Exosomal lncRNAs may have great potential to be used for the early detection and diagnosis of cervical cancer, and serve as convenient and noninvasive biomarkers.

Silent information regulator sirtuin 1 ameliorates acute liver failure via the p53/glutathione peroxidase 4/gasdermin D axis.

BACKGROUND: Acute liver failure (ALF) has a high mortality with widespread hepatocyte death involving ferroptosis and pyroptosis. The silent information regulator sirtuin 1 (SIRT1)-mediated deacetylation affects multiple biological processes, including cellular senescence, apoptosis, sugar and lipid metabolism, oxidative stress, and inflammation. AIM: To investigate the association between ferroptosis and pyroptosis and the upstream regulatory mechanisms. METHODS: This study included 30 patients with ALF and 30 healthy individuals who underwent serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) testing. C57BL/6 mice were also intraperitoneally pretreated with SIRT1, p53, or glutathione peroxidase 4 (GPX4) inducers and inhibitors and injected with lipopolysaccharide (LPS)/D-galactosamine (D-GalN) to induce ALF. Gasdermin D (GSDMD)-/- mice were used as an experimental group. Histological changes in liver tissue were monitored by hematoxylin and eosin staining. ALT, AST, glutathione, reactive oxygen species, and iron levels were measured using commercial kits. Ferroptosis- and pyroptosis-related protein and mRNA expression was detected by western blot and quantitative real-time polymerase chain reaction. SIRT1, p53, and GSDMD were assessed by immunofluorescence analysis. RESULTS: Serum AST and ALT levels were elevated in patients with ALF. SIRT1, solute carrier family 7a member 11 (SLC7A11), and GPX4 protein expression was decreased and acetylated p5, p53, GSDMD, and acyl-CoA synthetase long-chain family member 4 (ACSL4) protein levels were elevated in human ALF liver tissue. In the p53 and ferroptosis inhibitor-treated and GSDMD-/- groups, serum interleukin (IL)-1ß, tumour necrosis factor alpha, IL-6, IL-2 and C-C motif ligand 2 levels were decreased and hepatic impairment was mitigated. In mice with GSDMD knockout, p53 was reduced, GPX4 was increased, and ferroptotic events (depletion of SLC7A11, elevation of ACSL4, and iron accumulation) were detected. In vitro, knockdown of p53 and overexpression of GPX4 reduced AST and ALT levels, the cytostatic rate, and GSDMD expression, restoring SLC7A11 depletion. Moreover, SIRT1 agonist and overexpression of SIRT1 alleviated acute liver injury and decreased iron deposition compared with results in the model group, accompanied by reduced p53, GSDMD, and ACSL4, and increased SLC7A11 and GPX4. Inactivation of SIRT1 exacerbated ferroptotic and pyroptotic cell death and aggravated liver injury in LPS/D-GalN-induced in vitro and in vivo models. CONCLUSION: SIRT1 activation attenuates LPS/D-GalN-induced ferroptosis and pyroptosis by inhibiting the p53/GPX4/GSDMD signaling pathway in ALF.

Assessing recent recurrence after hepatectomy for hepatitis B-related hepatocellular carcinoma by a predictive model based on sarcopenia.

BACKGROUND: Sarcopenia may be associated with hepatocellular carcinoma (HCC) following hepatectomy. But traditional single clinical variables are still insufficient to predict recurrence. We still lack effective prediction models for recent recurrence (time to recurrence < 2 years) after hepatectomy for HCC. AIM: To establish an interventable prediction model to estimate recurrence-free survival (RFS) after hepatectomy for HCC based on sarcopenia. METHODS: We retrospectively analyzed 283 hepatitis B-related HCC patients who underwent curative hepatectomy for the first time, and the skeletal muscle index at the third lumbar spine was measured by preoperative computed tomography. 94 of these patients were enrolled for external validation. Cox multivariate analysis was per-formed to identify the risk factors of postoperative recurrence in training cohort. A nomogram model was developed to predict the RFS of HCC patients, and its predictive performance was validated. The predictive efficacy of this model was evaluated using the receiver operating characteristic curve. RESULTS: Multivariate analysis showed that sarcopenia [Hazard ratio(HR) = 1.767, 95%CI: 1.166-2.678, P < 0.05], alpha-fetoprotein ≥ 40 ng/mL (HR = 1.984, 95%CI: 1.307-3.011, P < 0.05), the maximum diameter of tumor > 5 cm (HR = 2.222, 95%CI: 1.285-3.842, P < 0.05), and hepatitis B virus DNA level ≥ 2000 IU/mL (HR = 2.1, 95%CI: 1.407-3.135, P < 0.05) were independent risk factors associated with postoperative recurrence of HCC. Based on the sarcopenia to assess the RFS model of hepatectomy with hepatitis B-related liver cancer disease (SAMD) was established combined with other the above risk factors. The area under the curve of the SAMD model was 0.782 (95%CI: 0.705-0.858) in the training cohort (sensitivity 81%, specificity 63%) and 0.773 (95%CI: 0.707-0.838) in the validation cohort. Besides, a SAMD score ≥ 110 was better to distinguish the high-risk group of postoperative recurrence of HCC. CONCLUSION: Sarcopenia is associated with recent recurrence after hepatectomy for hepatitis B-related HCC. A nutritional status-based prediction model is first established for postoperative recurrence of hepatitis B-related HCC, which is superior to other models and contributes to prognosis prediction.

Predicting disease progression in cirrhotic patients after transjugular intrahepatic portosystemic shunt implantation: A sex-stratified analysis.

BACKGROUND: Transjugular intrahepatic portosystemic shunt (TIPS) has been extensively used to treat portal hypertension-associated complications, including cirrhosis. The prediction of post-TIPS prognosis is important for cirrhotic patients, as more aggressive liver transplantation is needed when the post-TIPS prognosis is poor. AIM: To construct a nutrition-based model that could predict the disease progression of cirrhotic patients after TIPS implantation in a sex-dependent manner. METHODS: This study retrospectively recruited cirrhotic patients undergoing TIPS implantation for analysis. Muscle quality was assessed by measuring the skeletal muscle index (SMI) by computed tomography. Multivariate Cox proportional hazard models were utilized to determine the association between SMI and disease progression in cirrhotic patients after TIPS implantation. RESULTS: This study eventually included 186 cirrhotic patients receiving TIPS who were followed up for 30.5 ± 18.8 mo. For male patients, the 30-mo survival rate was significantly lower and the probability of progressive events was higher (3.257-fold) in the low-level SMI group than in the high-level SMI group. According to the multivariate Cox analysis of male patients, SMI < 32.8 was an independent risk factor for long-term adverse outcomes after TIPS implantation. A model was constructed, which involved creatinine, plasma ammonia, SMI, and acute-on-chronic liver failure and hepatic encephalopathy occurring within half a year after surgery. This model had an area under the receiver operating characteristic curve of 0.852, sensitivity of 0.926, and specificity of 0.652. According to the results of the DeLong test, this model outperformed other models (Child-Turcotte-Pugh, Model for End-Stage Liver Disease, and Freiburg index of post-TIPS survival) (P < 0.05). CONCLUSION: SMI is strongly associated with poor long-term outcomes in male patients with cirrhosis who underwent TIPS implantation.

Identification and validation of a T-cell-related MIR600HG/hsa-mir-21-5p competing endogenous RNA network in tuberculosis activation based on integrated bioinformatics approaches.

Background: T cells play critical roles in the progression of tuberculosis (TB); however, knowledge regarding these molecular mechanisms remains inadequate. This study constructed a critical ceRNA network was constructed to identify the potentially important role of TB activation via T-cell regulation. Methods: We performed integrated bioinformatics analysis in a randomly selected training set from the GSE37250 dataset. After estimating the abundance of 18 types of T cells using ImmuCellAI, critical T-cell subsets were determined by their diagnostic accuracy in distinguishing active from latent TB. We then identified the critical genes associated with T-cell subsets in TB activation through co-expression analysis and PPI network prediction. Then, the ceRNA network was constructed based on RNA complementarity detection on the DIANA-LncBase and mirDIP platform. The gene biomarkers included in the ceRNA network were lncRNA, miRNA, and targeting mRNA. We then applied an elastic net regression model to develop a diagnostic classifier to assess the significance of the gene biomarkers in clinical applications. Internal and external validations were performed to assess the repeatability and generalizability. Results: We identified CD4+ T, Tr1, nTreg, iTreg, and Tfh as T cells critical for TB activation. A ceRNA network mediated by the MIR600HG/hsa-mir-21-5p axis was constructed, in which the significant gene cluster regulated the critical T subsets in TB activation. MIR600HG, hsa-mir-21-5p, and five targeting mRNAs (BCL11B, ETS1, EPHA4, KLF12, and KMT2A) were identified as gene biomarkers. The elastic net diagnostic classifier accurately distinguished active TB from latent. The validation analysis confirmed that our findings had high generalizability in different host background cases. Conclusion: The findings of this study provided novel insight into the underlying mechanisms of TB activation and identifying prospective biomarkers for clinical applications.

Homology analysis of 51 penicillin-intermediate Streptococcus pneumoniae isolates from Wenzhou City, China.

OBJECTIVE: To investigate drug resistance features and homology among penicillin-intermediate Streptococcus pneumoniae isolates from Wenzhou City, China. METHODS: Fifty-one penicillin-intermediate S. pneumoniae isolates were obtained from respiratory samples of infants and children hospitalized with lung infections. An antimicrobial susceptibility test was used to assess drug resistance. Polymerase chain reaction and agarose gel electrophoresis were used to identify S. pneumoniae isolates and pulsed-field gel electrophoresis (PFGE) was used to analyze molecular subtypes. Hierarchical cluster analysis of PFGE fingerprints was used to compare genetic diversity and relatedness of S. pneumoniae isolates. The Quellung test was used for serotyping. RESULTS: Fifty-one penicillin-intermediate S. pneumoniae isolates showed evidence of multi-drug resistance and polyclonal origins. The isolates were classified into 25 subtypes through hierarchical cluster analysis of PFGE fingerprints. Three of these subtypes formed a supertype (15/51, 16/51 and 8/51 isolates), while the remaining subtypes occurred sporadically (12/51 isolates). CONCLUSIONS: Transmission of penicillin-intermediate S. pneumoniae is mostly vertical and to a lesser extent horizontal. Effective prevention strategies, including respiratory tract management and contact isolation, are essential to control nosocomial S. pneumoniae infection. Once susceptibility is confirmed, vancomycin, high-dose penicillin or third-generation cephalosporins (cefotaxime and ceftriaxone) may be used to treat penicillin-intermediate S. pneumoniae.

[Identification of differential proteins of serum in the patients suffering from coal-burning arsenism].

OBJECTIVE: To compare and analyze the differential expression of proteins between coal-burning arsenism serum and normal human serum and identify the proteins related with arseniasis caused by coal-burning. METHODS: Serum samples were collected from 6 normal subjects and 6 patients suffering from coal-burning arsenism. 2-DE was performed to separate serum proteins. After silver staining, the differential expression of proteins was analyzed and then identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). RESULTS: There were an average of 779 +/- 35 spots and 865 +/- 30 spots on 2-DE matching of two groups and the matching rate was 90.1% between two groups. From these two groups, 60 different protein spots were identified. Up-regulated expression was observed in 25 proteins and down-regulated expression in 35 proteins in the patient serum group. Among which 35 with differential expression above three times were singled out and MALDI-TOF-MS analysis was carried out on them. Thirteen proteins were identified, including keratin 10, apolipoprotein A-V, transferrin, alpha-1-antitrypsin, human zinc-alpha-2-glycoprotein, mitogen-activated protein kinase 3, vacuolar protein sorting 33A, O-linked GlcNAc transferase and etc. Up-regulated expression was observed in 5 proteins and down-regulated expression in 8 proteins in the patient serum group. CONCLUSION: The well-resolved and reproducible 2-DE serum patterns of patients suffering from coal-burning arsenism were established and some differentially expressed proteins characterized. These data will be used to screen the biomarker and to further study arseniasis caused by coal-burning.

Expression of PERK-eIF2α-ATF4 pathway signaling protein in the progression of hepatic fibrosis in rats.

OBJECTIVE: This study aims to observe changes in the expression of the hepatic endoplasmic reticulum stress PERK-eIF2α-ATF4 signaling pathway protein in the progression of CCl4-induced hepatic fibrosis in rats. METHODS: Male Wistar rats were sacrificed at the end of the 2nd, 4th, 8th and 12th weeks, respectively. A specific test was performed to compare the pathological changes of hepatic tissues in the model and normal groups. Immunohistochemical staining and Western blot were carried out to detect the expression of p-PERK, p-eIF2α and ATF4 proteins in the hepatic tissue group. Furthermore, real-time PCR was used to detect changes in the expression of ATF4 mRNA in hepatic tissues. RESULTS: In the eight-week and twelve-week hepatic fibrosis group, significant fibrosis hyperplasia was identified in the livers of rats, and pseudo-lobules were also formed in the livers of rats in the twelve-week hepatic fibrosis group. Immunohistochemical staining and Western blot results indicated that the expression levels of p-PERK, p-eIF2α and the ATF4 protein in the livers of rats were significantly increased from the 8th week compared with the normal group (P<0.05). Real-time PCR results revealed that the expression of ATF4 mRNA was significantly increased in hepatic tissues in the hepatic fibrosis group compared with the normal group (P<0.05), and this was gradually enhanced as hepatic fibrosis progressed. CONCLUSIONS: CCl4 can induce an increase in the expression of the PERK-eIF2α-ATF4 signaling protein in the development of hepatic fibrosis along with phosphorylation-mediated activation, indicating that the activation of the PERK-eIF2α-ATF4 signaling pathway may contribute to the onset and development of hepatic fibrosis by regulating downstream target genes.

[Differential analysis of associated proteome in hepatic fibrosis tissue].

OBJECTIVE: To develop the 2-DE profiles of proteome from hepatic fibrosis tissue and preliminarily analyze the differential expression of proteome. METHODS: The differential expression of proteins were analyzed by imaging analysis and MALDI-TOF-MS after the total protein was extracted from the hepatic fibrosis tissue and the normal liver tissue by 2-DE. 3 proteins were verified by in Western-blotting. RESULTS: Fifty-nine differentially expressed protein were found in the proteome profile analysis of these two types of tissue, among which 30 protein spots were up-regulated and 29 protein spots were down-regulated in these hepatic fibrosis tissues. Fifteen protein spots with the differential expressions more than 3 times were analyzed by peptide mass fingerprint (PMF) and 10 differentially expressed proteins were found to be related with cell signal transduction, cell proliferation, and oxidative stress. Western-blotting results of 14-3-3beta, DJ-1, and PEBP were consistent with those by the 2-DE examination. CONCLUSION: Some differentially expressed proteins have been found in the hepatic fibrosis tissue and the normal liver tissue. These data provided a fundamental basis for further study of the mechanism of hepatic fibrogenesis. Proteome;

Focal Adhesion Kinase Regulates Hepatic Stellate Cell Activation and Liver Fibrosis.

Understanding the underlying molecular mechanisms of liver fibrosis is important to develop effective therapy. Herein, we show that focal-adhesion-kinse (FAK) plays a key role in promoting hepatic stellate cells (HSCs) activation in vitro and liver fibrosis progression in vivo. FAK activation is associated with increased expression of α-smooth muscle actin (α-SMA) and collagen in fibrotic live tissues. Transforming growth factor beta-1 (TGF-ß1) induces FAK activation in a time and dose dependent manner. FAK activation precedes the α-SMA expression in HSCs. Inhibition of FAK activation blocks the α-SMA and collagen expression, and inhibits the formation of stress fibers in TGF-ß1 treated HSCs. Furthermore, inhibition of FAK activation significantly reduces HSC migration and small GTPase activation, and induces apoptotic signaling in TGF-ß1 treated HSCs. Importantly, FAK inhibitor attenuates liver fibrosis in vivo and significantly reduces collagen and α-SMA expression in an animal model of liver fibrosis. These data demonstrate that FAK plays an essential role in HSC activation and liver fibrosis progression, and FAK signaling pathway could be a potential target for liver fibrosis.

Tristetraprolin Down-Regulation Contributes to Persistent TNF-Alpha Expression Induced by Cigarette Smoke Extract through a Post-Transcriptional Mechanism.

RATIONALE: Tumor necrosis factor-alpha (TNF-α) is a potent pro-inflammatory mediator and its expression is up-regulated in chronic obstructive pulmonary disease (COPD). Tristetraprolin (TTP) is implicated in regulation of TNF-α expression; however, whether TTP is involved in cigarette smoke-induced TNF-α expression has not been determined. METHODS: TTP expression was examined by western blot analysis in murine alveolar macrophages and alveolar epithelial cells challenged without or with cigarette smoke extract (CSE). TNF-α mRNA stability, and the decay of TNF-α mRNA, were determined by real-time quantitative RT-PCR. TNF-α protein levels were examined at the same time in these cells. To identify the molecular mechanism involved, a construct expressing the human beta-globin reporter mRNA containing the TNF-α 3'-untranslated region was generated to characterize the TTP targeted site within TNF-α mRNA. RESULTS: CSE induced TTP down-regulation in alveolar macrophages and alveolar epithelial cells. Reduced TTP expression resulted in significantly increased TNF-α mRNA stability. Importantly, increased TNF-α mRNA stability due to impaired TTP function resulted in significantly increased TNF-α levels in these cells. Forced TTP expression abrogated the increased TNF-α mRNA stability and expression induced by CSE. By using the globin reporter construct containing TNF-α mRNA 3'-untranslated region, the data indicate that TTP directly targets the adenine- and uridine-rich region (ARE) of TNF-α mRNA and negatively regulates TNF-α expression at the post-transcriptional level. CONCLUSION: The data demonstrate that cigarette smoke exposure reduces TTP expression and impairs TTP function, resulting in significantly increased TNF-α mRNA stability and excessive TNF-α expression in alveolar macrophages and epithelial cells. The data suggest that TTP is a novel post-transcriptional regulator and limits excessive TNF-α expression and inflammatory response induced by cigarette smoke.

Effect of Danshao Huaxian capsule on expression of matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 in fibrotic liver of rats.

AIM: To investigate the effects of Danshao Huaxian (DSHX) capsules, a preparation of traditional Chinese medicine, on the expression of matrix metalloproteinase-1 (MMP-1), and tissue inhibitor of metalloproteinase-1 (TIMP-1) in the fibrous livers of rats. METHODS: Eighty male Wistar rats were randomly divided into normal control group (group A), CCl(4)-induced hepatic fibrosis group (group B), non-DSHX-treated group (group C), low dose-treated group (group D), and high dose-treated group (group E). Fibrous liver models in rats were induced by subcutaneous injection of CCl(4), oral administration of alcohol and high-lipid/low-protein diet for 8 wk. After the models were established, the rats in groups D and E were orally given a low dose (0.5 g/kg) and a high dose (1.0 g/kg) of DSHX daily for 8 wk, respectively. Then, the liver indexes, serum hyaluronic acid (HA) and alanine aminotransferase (ALT) were examined. The degree of hepatic fibrosis was evaluated by optical microscopy. Hydroxyproline (Hyp) in the urine was determined, and the expression of MMP-1 and TIMP-1 was detected by immunohistochemical techniques. RESULTS: In groups D and E, the liver indexes, levels of serum HA and ALT reduced and development of hepatic fibrosis weakened significantly. The urinary Hyp and expression of MMP-1 in the liver tissues elevated, but the expression of TIMP-1 decreased obviously, as compared to groups B and C. CONCLUSION: DSHX enhances the expression of MMP-1 but decreases that of TIMP-1 in liver tissues of CCl(4)-induced hepatic fibrotic rats, which may result in its elevated activity that contributes to fighting against hepatic fibrosis.

In vivo effects of Chinese herbal recipe, Danshaohuaxian, on apoptosis and proliferation of hepatic stellate cells in hepatic fibrotic rats.

AIM: To investigate the effects of Danshaohuaxian (DSHX), a Chinese herbal recipe, on the apoptosis and cell cycles of hepatic stellate cells (HSCs) in rat hepatic fibrosis and its possible mechanisms. METHODS: Seventy-six male Wistar rats were randomly divided into normal control group, hepatic fibrosis group, non-DSHX-treated group and DSHX-treated group. Except for the normal control group, rat hepatic fibrotic models were induced by subcutaneous injection of carbon tetrachloride (CCl4), drinking alcohol, giving diet of hyperlipid and hypoprotein for 8 wk. When the hepatic fibrotic models were produced, 12 rats of hepatic fibrosis group (15 rats survived, others died during the 8 wk) were sacrificed to collect blood and livers. HSCs were isolated from the other 3 rats to detect the apoptotic index (AI) and cell cycles by flow cytometry. DSHX was then given to the DSHX-treated group (1.0 g/kg, PO, daily) for 8 wk. At the same time, normal control group and non-DSHX-treated group were given normal saline for 8 wk. At end of the experiment, some rats in these three groups were sacrificed to collect blood and livers, the other rats were used for HSC isolation to detect the apoptotic index (AI) and cell cycles. Then the liver index, serum hyaluronic acid (HA) and alanine aminotransferase (ALT), degree of hepatic fibrosis, urinary excretion of hydroxyproline (Hyp) and expression of collagen types I and III (COL I and III) in these four groups were detected respectively. RESULTS: Compared with the indexes of the hepatic fibrosis group and non-DSHX-treated group, the DSHX-treated group revealed a liver index of (0.0267+/-0.0017 vs 0.0423+/-0.0044, 0.0295+/-0.0019, P<0.05), levels of serum HA (200.78+/-31.71 vs 316.17+/-78.48, 300.86+/-72.73, P<0.05) and ALT (93.13+/-5.79 vs 174.5+/-6.02, 104.75+/-6.54, P<0.01), and stage of hepatic fibrosis (1.30 vs 4.25, 2.60, P<0.01) all reduced. The urinary excretion of Hyp increased (541.09+/-73.39 vs 62.00+/-6.40, 182.44+/-30.83, P<0.01), the COL I and III expression decreased (COL I: 1.07+/-0.96 vs 4.18+/-2.26, 3.22+/-1.44, P<0.01; COL III: 1.09+/-0.58 vs 3.04+/-0.62, 2.23+/-0.58, P<0.01), the HSCs apoptotic index of HSCs (7.81+/-0.47 vs 1.63+/-0.25, 1.78+/-0.4, P<0.05) and the ratio of G0-G1 phase cells increased (94.30+/-1.33 vs 62.27+/-17.96, 50.53+/-2.25, P<0.05). The ratios of S-phase cells (3.11+/-1.27 vs 9.83+/-1.81, 11.87+/-1.9, P<0.05) and G2-M phase cells (2.58+/-0.73 vs 23.26+/-10.95, 13.60+/-1.15, P<0.01) declined. CONCLUSION: DSHX capsule shows certain therapeutic effects on hepatic fibrosis in rats and inhibits abnormal deposition of COL I and III in rat livers by promoting the apoptosis of HSCs and preventing their proliferation.

Effects of Dan-Shao-Hua-Xian on the expression of collagen type I and III in rats with hepatic fibrosis.

BACKGROUND: Hepatic fibrosis is the common pathological change in various chronic liver diseases, and its major cause is the imbalance between the production and degradation of the extracellular matrix, which is mainly composed of collagens. Dan-Shao-Hua-Xian (DSHX) capsule, a traditional Chinese herbal compound, has shown marked preventive effects on hepatic fibrosis in rats in our previous studies. The present study was designed to further investigate its therapeutic actions on hepatic fibrosis in rats and its possible mechanisms. METHODS: Eighty male Wistar rats were randomly divided into normal control group, hepatic fibrosis group, non-DSHX-treated group, low-dose-treated group, and high-dose-treated group. The rat models of hepatic fibrosis were established by subcutaneous injecton of CCl4, drinking alcohol, giving diet of hyperliprosis and hypoprotein for 8 weeks. The two DSHX-treated groups were treated respectively with low dose (0.5 g/kg) and high dose (1.0 g/kg) of DSHX capsule p.o. everyday for 8 weeks. At the end of the experiment, liver indexes were calculated in each group in addition to the levels of the serum hyaluronic acid and alanine aminotransferase. Their degree of hepatic fibrosis and urinary excretion of hydroxyproline and expression of collagen I, III were detected. RESULTS: Comparison of the indexes of the hepatic fibrosis group and non-DSHX-treated group revealed that the liver indexes, levels of serum hyaluronic acid and alanine aminotransferase, and stage of hepatic fibrosis were all significantly reduced in the two DSHX treated groups. The urinary excretion of hydroxyproline was increased and the expression of collagen I and III in liver tissue was lessened. These alterations were more obviously observed in the high-dose-treated group. CONCLUSION: DSHX capsule has certain therapeutic effect on hepatic fibrosis in rats.

Roles of virulence genes (PsaA and CpsA) on the invasion of Streptococcus pneumoniae into blood system.

BACKGROUND: Streptococcus pneumoniae (SP) is the major cause of childhood mortality worldwide, we need to understand virulence genes of SP so can better target the treatment.We investigated the expression of virulence genes PsaA and CpsA in different strains of SP interacting with monocyte cell line (THP-1) or pneumocyte cell line (A549) and the possible mechanism of SP invasion of the blood system. METHODS: A total of 23 strains of SP were collected from hospitalized patients (blood-derived and sputum-derived) in the Second Affiliated Hospital of Wenzhou Medical College. The strains and ATCC 49619 were cultured, and RNAs were extracted. THP-1 and A549 cells were stimulated by different SP and ATCC 49619 for 4 h and 8 h, respectively. Quantitative real-time PCR was used to analyze the mRNA expression of PsaA and CpsA. The data were analyzed by SPSS 17.0. RESULTS: The mRNA level of PsaA and CpsA were all significantly increased in clinical SP strains when compared to ATCC49619 after tedTHP-1 and A549 cells were stimulated. Clinical SPs showed higher virulence compared with ATCC49619. CONCLUSIONS: The expression of CpsA is the basis of the pathogenicity of SP. The expression of virulence gene PsaA may be helpful to the invasion of SP to the blood system.

Effects of clinical isolates of Streptococcus pneumoniae on THP-1 human monocytic cells.

Twenty­three clinical Streptococcus pneumoniae (SP) strains were isolated from blood and sputum specimens from the Second Affiliated Hospital of Wenzhou Medical College in 2009. These strains and the ATCC 49619 standard strain were cultured and suspended in normal saline (at a turbidity of 1.0 McFarland). The production of interleukin (IL)­8, intracellular adhesion molecule­1 (ICAM­1) and IL­10 in THP­1 cells following stimulation with the SP suspension was analyzed by an enzyme-linked immunosorbent assay. The concentrations of IL­8, ICAM­1 and IL­10 from the THP­1 monocytes were greater than those of the blank control following stimulation with the SP suspension. No significant difference was identified in the levels of IL­8, ICAM­1 and IL­10 secretion between THP­1 monocytes stimulated by blood­borne SP (bb­SP) and sputum­borne SP (sb­SP).

Prevalence of plasmid-mediated quinolone resistance in Escherichia coli isolates in Wenzhou, Southern China, 2002-2008.

A total of 514 consecutive clinical Escherichia coli isolates, irrespective of resistance background, were collected in the period 2002-2008 in Wenzhou, southern China, to investigate the prevalence of plasmid-mediated quinolone resistance (PMQR). The dominant PMQR gene was aac(6')-Ib-cr, followed by qnr, whereas qepA was absent. A total of 253 (49.2%) of these isolates were aac(6')-Ib-positive. Subsequently, 134 of these isolates were sequenced and 42 (31.3%) found to harbor aac(6')-Ib-cr, 18 to harbor new aac(6')-Ib mutants, and 74 to harbor wild-type aac(6')-Ib. The genes qnrA, qnrB, and qnrS were found in 2 (0.4%), 6 (1.2%), and 14 (2.7%) of 514 isolates, respectively, with 2 isolates co-harboring qnrB and qnrS genes. Sequencing allowed us to identify qnrA1, qnrB4, qnrB6, and qnrS1 in 20 qnr-positive isolates, with qnrS1 being the most prevalent allele. The genes qnrC and qnrD were not found in any isolates. Interestingly, 35% of qnr-positive isolates and 16.7% of aac(6')-Ib-cr-positive isolates were susceptible to ciprofloxacin. PMQR genes are therefore present in both quinolone-resistant and -susceptible isolates and can also be transferred by conjugation experiments, thus suggesting a likely future increase in quinolone resistance.