[Interaction between glycogen synthase kinase-3ß and endoplasmic reticulum stress is involved in high glucose-induced injury in human umbilical vein endothelial cells].

OBJECTIVE: To explore the role of the interaction between glycogen synthase kinase-3ß (GSK-3ß) and endoplasmic reticulum stress (ERS) in the high glucose (HG)-induced injury in human umbilical vein endothelial cells (HUVECs). METHODS: HUVECs treated with 40 mmol/L glucose for 24 h were examined for expression levels of GSK-3ß, GRP78, CHOP and cleaved caspase-3 protein using Western blotting. The cell viability was examined using CCK-8 assay and cell apoptosis was detected with Hoechst 33258 nuclear staining and photofluorography. The intracellular level of reactive oxygen species (ROS) was measured with dichlorfluoresein staining and photofluorography. Mitochondrial membrane potential (MMP) was tested by rhodamine 123 (Rh123) staining and photofluorography. RESULTS: Treatment of HUVECs with 40 µmol/L glucose for 3-24 h activated GSK-3ß in a time-dependent manner, leading to significantly down-regulated expression of phosphorylated (p)-GSK-3ß (P<0.05). HG exposure of the cells for 1-24 h induced ERS, evidenced by time-dependently up-regulated expression of GRP78 and CHOP (P<0.05). LiCl, an inhibitor of GSK-3ß, attenuated HG-induced ERS and significantly lowered the expression levels of GRP78 and CHOP (P<0.01). 4-PBA, an inhibitor of ERS, obviously ameliorated the activation of GSK-3ß by HG as shown by the increase in p-GSK-3ß expression level (P<0.01). HG exposure for 24 h induced obvious injuries in HUVECs, which exhibited decreased cell viability, increased cell apoptosis, increased expression of cleaved caspase-3 and ROS generation, and loss of MMP. Pretreatment of the cells with LiCl or 4-PBA for 60 min before HG exposure significantly lessened the cell injuries (P<0.01). CONCLUSION: Interactions between GSK-3ß and ERS occur in HUVECs exposed to HG and participate in HG-induced cell injuries.

[Angiotensin-(1-7) protects cardiac myocytes against high glucose-induced injury by inhibiting ClC-3 chloride channels].

OBJECTIVE: To explore whether angiotensin-(1-7) [Ang-(1-7)] protects cardiac myocytes against high glucose (HG)-induced injury by inhibiting ClC-3 chloride channels. METHOD: H9c2 cardiac cells were exposed to 35 mmol/L glucose for 24 h to establish a cell injury model. The cells were treated with Ang-(1-7) or the inhibitor of chloride channel (NPPB) in the presence of HG for 24 h to observe the changes in HG-induced cell injury. Cell counter kit 8 (CCK-8) assay was used to test the cell viability, and the morphological changes of the apoptotic cells were detected using Hoechst 33258 staining and fluorescent microscopy. The intracellular level of reactive oxygen species (ROS) was examined by DCFH-DA staining, SOD activity in the culture medium was measured using commercial kits, and the mitochondrial membrane potential (MMP) of the cells was tested with rodamine 123 staining. The expression level of cardiac ClC-3 chloride channels was detected with Western blotting. RESULTS: Exposure of H9c2 cardiac cells to 35 mmol/L glucose for 24 h markedly enhanced the expressions of cardiac ClC-3 channel protein (P<0.01). Co-treatment of the cells with 1 µmol/L Ang-(1-7) and HG for 24 h significantly attenuated HG- induced upregulation of ClC-3 channel protein expression (P<0.01). Co-treatment of the cells exposed to HG with 1 µmol/L Ang-(1-7) or 100 µmol/L NPPB for 24 h obviously ameliorated HG-induced injuries as shown by increased cell viability, enhanced SOD activity, decreased number of apoptotic cells, and reduced intracellular ROS generation and loss of MMP (P<0.01). CONCLUSION: ClC-3 channels are involved in HG-induced injury in cardiac cells. Ang-(1-7) protects cardiac cells against HG-induced injury by inhibiting ClC-3 channels.

Case-Control Study on Impact of the Telomerase Reverse Transcriptase Gene Polymorphism and Additional Single Nucleotide Polymorphism (SNP)- SNP Interaction on Non-Small Cell Lung Cancers Risk in Chinese Han Population.

AIMS: To investigate the impact of telomerase reverse transcriptase (TERT) gene polymorphism and additional SNP-SNP interaction on non-small cell lung cancer (NSCLC) risk in Chinese population. METHODS: A total of 828 participants (526 males, 302 females), with a mean age of 71.3 ± 15.7 years old, were selected, including 410 NSCLC patients and 418 normal participants. Logistic regression was performed to investigate association between single nucleotide polymorphism (SNP) and NSCLC risk. Generalized multifactor dimensionality reduction (GMDR) was used to analysis the interaction among four SNPs. RESULTS: Non-small cell lung cancer risk was significantly higher in carriers of G allele of the rs2736100 polymorphism than those with TT (TG + GG vs. TT, adjusted OR (95%CI = 1.68 (1.28-2.07). In addition, we also found that NSCLC risk was also significantly higher in carriers of A allele of the rs2736098 polymorphism than those with GG (GA + AA vs. GG, adjusted OR (95%CI) = 1.52 (1.19-1.93). GMDR analysis indicated that there was a significant two-locus model (P = 0.0100) involving rs2736098 and rs2736100, indicating a potential gene-gene interaction between rs2736098 and rs2736100. Overall, the two-locus models had a cross-validation consistency of 10 of 10, and had the testing accuracy of 62.17%. We found that patients with GA or AA of rs2736098 and TG or GG of rs2736100 genotype have the highest NSCLC risk, compared to patients with GG of rs2736098 and TT of rs2736100 genotype, OR (95%CI) was 2.52 (1.68-3.68), after covariates adjustment. CONCLUSIONS: Minor allele of rs2736098 and rs2736100 in TERT gene and interaction between the two SNP were associated with increased risk of NSCLC risk.

The serum glucan level and pathological changes of antifungal treatment for lower respiratory tract infection of Candida albicans.

Due to the fact that Candida albicans colonizes in the upper respiratory tracts of healthy people, whether or not its isolation from airway secretions is sufficient to warrant treatment remains controversial. The animal models of immunosuppressive rats with pulmonary candidiasis were established by the intratracheal inoculating suspensions of C. albicans, and the animals were divided into the following three groups: (1) antifungal treatment group, (2) saline control group, and (3) blank control group. We noted the following in our studies: (1) The fungal load of the saline control group gradually increased such that it was higher than those of the antifungal treated group and was significant from the fourth day of treatment (P < 0.01). (2) The serum (1,3)-ß-D-glucan (BG) in the saline control group also gradually increased so that it was significantly higher than found with the treated group by the sixth day of treatment (P < 0.05), and in fact, the rank of pulmonary colony count and BG in the two groups at different time points showed an almost perfect linear correlation. (3) The median survival period of the rats in the antifungal treated group and saline control group was 15 and 8 days respectively, no rats died in the blank control group. (4) The lung lesions from the saline control group gradually became more aggravated than those in the antifungal treated group; no significant pathological changes were found in the blank control group. Antifungal treatment (micafungin) is capable of efficaciously decreasing the lung fungal burden, and continuous monitoring of BG is useful for the evaluation of therapeutic effect of antifungals. Infection of C. albicans with associated pathological damage implies the need for antifungal therapy.

Evaluation of galactomannan enzyme immunoassay and quantitative real-time PCR for the diagnosis of invasive pulmonary aspergillosis in a rat model.

Since there is no consensus about the most reliable assays to detect invasive aspergillosis from samples obtained by minimally invasive or noninvasive methods, we compared the efficacy of an enzyme-linked immunosorbent assay (ELISA) for galactomannan (GM) detection and quantitative real-time PCR assay (qRT-PCR) for the diagnosis of invasive pulmonary aspergillosis. Neutropenic, male Sprague-Dawley rats (specific pathogen free; 8 weeks old; weight, 200 ± 20 g) were immunosuppressed with cyclophosphamide and infected with Aspergillus fumigatus intratracheally. Tissue and whole blood samples were harvested on days 1, 3, 5, and 7 post-infection and examined with GM ELISA and qRT-PCR. The A. fumigatus DNA detection sequence was detected in the following number of samples from 12 immunosuppressed, infected rats examined on the scheduled days: day 1 (0/12), day 3 (0/12), day 5 (6/12), and day 7 (8/12) post-infection. The sensitivity and specificity of the qRT-PCR assay was 29.2% and 100%, respectively. Receiver operating characteristic curve (ROC) analysis indicated a Ct (cycle threshold) cut-off value of 15.35, and the area under the curve (AUC) was 0.627. The GM assay detected antigen in sera obtained on day 1 (5/12), day 3 (9/ 12), day 5 (12/12), and day 7 (12/12) post-infection, and thus had a sensitivity of 79.2% and a specificity of 100%. The ROC of the GM assay indicated that the optimal Ct cut-off value was 1.40 (AUC, 0.919). The GM assay was more sensitive than the qRT-PCR assay in diagnosing invasive pulmonary aspergillosis in rats.

Activation of the p38 MAPK/NF-κB pathway contributes to doxorubicin-induced inflammation and cytotoxicity in H9c2 cardiac cells.

A number of studies have demonstrated that inflammation plays a role in doxorubicin (DOX)-induced cardiotoxicity. However, the molecular mechanism by which DOX induces cardiac inflammation has yet to be fully elucidated. The present study aimed to investigate the role of the p38 mitogen-activated protein kinase (MAPK)/nuclear factor-κB (NF-κB) pathway in DOX-induced inflammation and cytotoxicity. The results of our study demonstrated that the exposure of H9c2 cardiac cells to DOX reduced cell viability and stimulated an inflammatory response, as demonstrated by an increase in the levels of interleukin-1ß (IL-1ß) and IL-6, as well as tumor necrosis factor-α (TNF-α) production. Notably, DOX exposure induced the overexpression of phosphorylated p38 MAPK and phosphorylation of the NF-κB p65 subunit, which was markedly inhibited by SB203580, a specific inhibitor of p38 MAPK. The inhibition of NF-κB by pyrrolidine dithiocarbamate (PDTC), a selective inhibitor of NF-κB, significantly ameliorated DOX-induced inflammation, leading to a decrease in the levels of IL-1ß and IL-6, as well as TNF-α production in H9c2 cells. The pretreatment of H9c2 cells with either SB203580 or PDTC before exposure to DOX significantly attenuated DOX-induced cytotoxicity. In conclusion, our study provides novel data demonstrating that the p38 MAPK/NF-κB pathway is important in the induction of DOX-induced inflammation and cytotoxicity in H9c2 cardiac myocytes.

Interleukin-12 and interleukin-2 alone or in combination against the infection in invasive pulmonary aspergillosis mouse model.

Aspergillus fumigatus is an intracellular opportunistic fungus causing invasive pulmonary mycosis, characterised by hyphal invasion and destruction of pulmonary tissue. Th1 cytokines could enhance fungicidal activity. The effects from the combination of interleukin-12 (IL-12) and IL-2 are rarely known in invasive pulmonary aspergillosis infection. To assess the cleaning of A. fumigatus infection in the pulmonary tissues by IL-12 and IL-2, interferon-γ (IFN-γ) was detected in the sera using ELISA, quantification of IFN-γ mRNA using real-time RT-PCR and lung Colony-forming unit was assayed by cultivation. Morphology was analysed by histopathological examination. Our results showed that IL-12 and/or IL-2 could enhance the IFN-γ expression in the pulmonary tissue, reduce the colony load in the pulmonary tissue and increase the survival rate of mouse. The combination of IL-12 and IL-2 could assist in increasing the IFN-γ expression in the pulmonary tissue, but neither reduce colony load in the pulmonary tissue nor increase the survival rate of mouse significantly. It was demonstrated that IL-12 and IL-2 were strong immunomodulatory cytokines as a prerequisite for protecting the host from infectious agents.

Retrospective analysis on the impact of tuberculosis on patients with systemic lupus erythematosus (SLE).

Up to now, there have been few reports concerning changes in lupus activity and immune indices of tuberculosis in patients with systemic lupus erythematosis (SLE). A retrospective investigation was given to survey the case data of SLE patients companied with tuberculosis that were treated in our hospital from 2001 to 2010 and compared with that of sex- and age-matched patients with single SLE. Changes in autoantibodies, lupus activity, inflammatory indices, positive rates of tuberculin (PPD) test and tuberculosis antibody (TB-Ab) of both groups were observed. It was indicated by results that ANA antibody level and positive rates of anti-Sm, anti-SSA and anti-SSB antibodies were significantly lower in the TB group than those in the control group (P < 0.05); C3 and C4 levels were significantly higher in the TB group than those in the control group; damage of hematological system (predominantly platelet) was less severe in the TB group than that in the control group (P < 0.05); no significant differences in IgG, IgM and IgA were noted between two groups (P > 0.05); ESR, C-reactive protein and LDH levels were significantly higher in the TB group than those in the control group (P < 0.05); PPD-IgG were significantly higher in the TB group than those in the control group (P < 0.05). These results suggested that after SLE patients were infected with tuberculosis, immune function was altered and lupus activity was inhibited as well.

[Analysis of the risk factors of pulmonary fungal infections related to mechanical ventilation and prognosis of the patients].

OBJECTIVE: To investigate the risk factors of pulmonary fungal infections related to mechanical ventilation and the prognosis of patients. METHODS: A retrospective case-controlled study was conducted to analyze the culture results of the pulmonary secretions in patients with pulmonary fungal and nonfungal infections in association with mechanical ventilations. The risk factors of pulmonary fungal infections related to mechanical ventilation were identified and their impact on the clinical outcome of the patients was evaluated. RESULTS: Of the 127 patients included in this study, 81 (63.78%) were positive and 46 (36.22%) negative for pulmonary fungal infections according to the diagnostic criteria of ventilator-associated pneumonia (VAP). The mortality of the patients with fungal infection was 82.7%, significantly higher than that of patients with non-fungal infection (67.39%, chi2=3.910, P<0.05). Univariate analysis and multivariate logistic regression showed that such factors as old age, duration of mechanical ventilation, tracheal intubation or incision for over 7 days, diabetes, blood glucose over 6.1 mmol/L, multi-organ dysfunction, combined use of antibiotics, at least 3-time changes antibiotics, administration of glucocorticosteroid for over 7 days, and immunodepressant use were all the independence risk factors of pulmonary fungal infection related to mechanical ventilation. Old age, multi-organ dysfunction, blood glucose over 6.1 mmol/L, glucocorticosteroid use for over 7 days, anesthetic use for over 3 days and high APACHE III scores were the risk factors for mortality in patients with the infections. CONCLUSIONS: Pulmonary fungal infection associated to mechanical ventilation is often the results of presence of multiple risk factors, and early identification of these factors for timely antifungal treatment may improve the prognostics of the patients and help reduce the mortality rate.