Transfer of Invitro CD4 + T Cells with Hypomethylation of Perforin Promoter into Rats' Abdomens Causes Autoimmune Emphysema.

Our previous study suggested that hypomethylation of perforin promoter of CD4 + T cells might be involved in the pathogenesis of autoimmune emphysema of rats. Whether transfer of this kind of cells hypomethylated in vitro into naive immunocompetent rats also results in emphysema is unknown yet. To test the hypothesis above, thirty Sprague Dawley (SD) rats were randomly divided into three groups: a model group (n = 10), a normal control group (n = 10) and a sham operation group (n = 10). In the model group, spleen-derived CD4 + T cells of normal rats were treated with 5-azacytidine (5-Aza), complete Freund's adjuvant and Phosphate Buffered Saline (PBS), then transferred into naive immunocompetent rats. The normal control group was injected with CD4 + T lymphocytes from spleens of normal rats and the same amount of adjuvant and PBS as above. In sham operation group, normal rats were injected intraperitoneally with complete Freund's adjuvant and PBS. Histopathological evaluations (mean linear Intercept (MLI) and mean alveolar numbers (MAN)), anti-endothelial cell antibodies (AECA) in serum and bronchoalveolar lavage fluid (BALF), lung vascular endothelial growth factor (VEGF)), the apoptotic index (AI) of alveolar septal cells and the methylation levels of perforin promoter of CD4 + T cells were investigated. The levels of the methylation above and MAN were lower in the model group than in the control and the sham operation group, while the AECA in serum and BALF, VEGF, MLI and the AI were greater (all p < 0.05). The methylation levels of perforin promoter were positively correlated with the MAN (r = 0.747, p < 0.05) and negatively correlated with AI, AECA, MLI, and VEGF (r was -0.789, -0.746, -0.743, -0.660, respectively, all p < 0.05). This study suggests that transfer of invitro CD4 + T cells with hypomethylation of perforin promoter into rats causes autoimmune emphysema, possibly by increasing expression of VEGF and promoting alveolar septal cell apoptosis.

Hypomethylation of perforin regulatory elements in CD4+ T cells from rat spleens contributes to the development of autoimmune emphysema.

BACKGROUND AND OBJECTIVE: It is widely accepted that perforin regulatory elements are hypomethylated in CD4+ T cells from patients with active lupus, but whether this is the case in autoimmune emphysema is not known. METHODS: Twenty rats were randomly divided into a normal control group and an emphysema group. Rat models of emphysema were established by intraperitoneal injection with xenogeneic endothelial cells. The levels of tumour necrosis factor-α, interleukin-8 and matrix metalloproteinase (MMP)-9 in bronchoalveolar lavage fluid (BALF) were measured, lung mean linear intercept and destructive index measured. Mean methylation of perforin gene promoter in CD4+ T cells and the expression of perforin were investigated. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling methods were used to examine the percentage of apoptotic cells in the alveolar septa. RESULTS: The levels of MMP-9 in BALF were higher in emphysema group than in control group (P < 0.05). The mean linear intercept and destructive index were higher in emphysema group than in control group (P < 0.05). The mean perforin gene promotor methylation of emphysema group was significantly decreased as compared with control group, while the expression levels of perforin gene were relatively higher (P < 0.05). There were more terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling-positive cells in the alveolar septa in control group than in emphysema group. CONCLUSIONS: Hypomethylation of perforin regulatory elements in CD4+ T cells may result in the lung septal cell apoptosis associated with the development of experimental autoimmune emphysema. MMP-9 may play an important role in the pathogenesis of this kind of disease.

Design, synthesis and antibacterial evaluation of novel AHL analogues.

Two series of novel AHL analogues were designed, synthesized and evaluated for antibacterial activity under cell membrane conditions in vitro. Analogues 4a-c and 4g-m presented potent activity against Gram-positive bacteria. Especially the analogue 4l exerted the most potent inhibition against Bacillus subtilis with MIC50 value of 1.443µg/ml. To our surprise, analogues 6a-c and 6g showed weak inhibition against Gram-negative bacteria with MIC50 values ranging from 17.589 to 67.840µg/ml. This was the first report about synthesis and antibacterial evaluation in vitro of AHL analogues containing dithioester linkage.

SAM protects against alveolar septal cell apoptosis in autoimmune emphysema rats.

BACKGROUND: Hypomethylation of the perforin gene promoter in CD4 + T cells, inflammation and oxidative stress, might be involved in alveolar septal cell apoptosis associated with emphysema in rats. This study aimed to investigate the effects of S-adenosylmethionine (SAM) on this kind of apoptosis in rats with autoimmune emphysema. METHODS: Twenty-four rats were randomly divided into three groups: a normal control group, a model group, and a SAM group. Pathological changes in lung tissues were observed, and the mean linear intercept (MLI) and mean alveolar number (MAN) were measured. The levels of anti-endothelial cell antibodies (AECA) in serum, alveolar septal cell apoptosis, perforin gene promotor methylation in CD4 + T cells in the spleen, and the levels of cytokines, malondialdehyde (MDA), and glutathione (GSH) and the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in bronchoalveolar lavage fluid (BALF) were investigated. RESULTS: The MLI, apoptosis index (AI) of alveolar septal cells, levels of AECA in serum, and levels of tumour necrosis factor-α (TNF-α), matrix metalloproteinase-9 (MMP-9) and MDA in BALF were increased, while the MAN, methylation levels, and the activities of GSH, SOD and GSH-Px in BALF were decreased in the model group compared with those in the normal control group and the SAM group (all P < 0.05). The levels of interleukin-8 (IL-8) in BALF were greater in the model group than in the normal control group (P < 0.05). CONCLUSIONS: SAM protects against alveolar septal cell apoptosis, airway inflammation and oxidative stress in rats with autoimmune emphysema possibly by partly reversing the hypomethylation of the perforin gene promoter in CD4 + T cells.

Mitochondrial separation protein inhibitor inhibits cell apoptosis in rat lungs during intermittent hypoxia.

Obstructive sleep apnoea (OSA) is a very common sleep and breathing disorder that occurs in worldwide. It is important to develop a more effective treatment for OSA to overcome lung cell apoptosis during intermittent hypoxia (IH). A mitochondrial separation protein inhibitor (Mdivi-1) has been demonstrated to be a powerful tool for inhibiting apoptosis. In the present study, the protective effect and possible mechanism of apoptosis in lung cells during IH was investigated using in vivo and in vitro experiments. Following IH exposure for 4 weeks, the lung tissues of Sprague Dawley rats exhibited interstitial lesions, while Mdivi-1 reduced these pulmonary interstitial lesions. B-cell lymphoma (Bcl)-2 mRNA and protein expression levels were decreased however caspase-3, caspase-9 and dynamin-related protein 1 (Drp-1) mRNA and protein expression levels were increased. Following Mdivi-1 intervention, Bcl-2 mRNA and protein expression levels were increased while caspase-3, caspase-9 and Drp-1 mRNA and protein expression levels were decreased (P<0.05). After exposure to IH for 12 h, the apoptosis rate of WTRL1 cells in rats increased gradually with the IH time (P<0.05). Bcl-2 mRNA and protein expression levels were decreased, whereas caspase-3, caspase-9, cytochrome C (Cyt-C) and Drp-1 mRNA levels were increased, and caspase-3, caspase-9 and Drp-1 protein expression levels were increased. After Mdivi-1 intervention, Bcl-2 mRNA and protein expression levels were increased but caspase-3, caspase-9, Cyt-C and Drp-1 mRNA levels were decreased along with caspase-9, Cyt-C and Drp-1 protein expression levels which were decreased (P<0.05). The results of the present study suggest that Mdivi-1 may be a potential agent for treating OSA because it inhibits the mitochondrial pathway and reduces apoptosis.

miR-138 inhibits proliferation by targeting 3-phosphoinositide-dependent protein kinase-1 in non-small cell lung cancer cells.

OBJECTIVE: Underlying mechanisms of non-small cell lung cancer (NSCLC) development remain poorly understood. miR-138 and 3-phosphoinositide-dependent protein kinase-1 (PDK1) have been reported to be involved in the genesis of NSCLC. The aim of this study was to investigate the role and mechanisms of miR-138 and PDK1 in human NSCLC cells. METHODS: The effect of miR-138 on proliferation of A549 lung cancer cells was first examined using 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide assay. The expression of PDK1 in A549 lung cancer cells was assessed by real-time polymerase chain reaction further. A luciferase reporter activity assay was conducted to confirm target association between miR-138 and 3' untranslated region (3'-UTR) of PDK1. Finally, the role of PDK1 on proliferation of A549 cells was evaluated by transefection of PDK1 small interfering RNA (siRNA). RESULTS: Proliferation of A549 lung cancer cells was suppressed by miR-138 in a concentration-dependent manner. Furthermore, miR-138 can bind to the 3'-UTR of PDK1 and downregulate expression of PDK1 at both mRNA and protein levels. Knockdown of PDK1 by siRNA significantly inhibits the proliferation of A549 lung cancer cells. CONCLUSIONS: These findings suggest that miR-138 as a potential tumor suppressor could inhibit cell proliferation by targeting PDK1 in NSCLC cells, which could be employed as a potential therapeutic target for miRNA-based NSCLC therapy.

Design and synthesis of novel 1,2,3-triazole-dithiocarbamate hybrids as potential anticancer agents.

A series of novel 1,2,3-triazole-dithiocarbamate hybrids were designed, synthesized and evaluated for anticancer activity against four selected human tumor cell lines (MGC-803, MCF-7, PC-3, EC-109). Majority of the synthesized compounds exhibited moderate to potent activity against MGC-803 and MCF-7. Among them, compounds 3a and 3c showed excellent broad spectrum anticancer activity with IC50 values ranging from 0.73 to 11.61 µM and 0.49-12.45 µM, respectively. Particularly, compound 3a was more potent than 5-fluorouracil against all tested human cancer cell lines. Flow cytometry analysis demonstrated that treatment of MGC-803 with 3c led to cell cycle arrest at G2/M phase accompanied by an increase in apoptotic cell death after 12 h.

Design, synthesis and antiproliferative activity studies of novel 1,2,3-triazole-dithiocarbamate-urea hybrids.

A series of novel 1,2,3-triazole-dithiocarbamate-urea hybrids were designed, synthesized and their antiproliferative activities against four selected human cancer cell lines were evaluated. The results showed that a number of the hybrids exhibited potent activity in selected human cancer cell lines. Among them, compounds 27 and 34 showed broad spectrum anticancer activity with IC50 values ranging from 1.62 to 20.84 µM and 0.76 to 13.55 µM, respectively. Interestingly, compounds 27 and 34, being very potent against MGC-803 cells, exhibited no significant cytotoxicity against normal human embryonic kidney cells at up to 55 µM and 70 µM, respectively. Evidences of cell cycle arrest and apoptosis induction were obtained for the most effective compounds 27 and 34 by means of flow cytometry and microscopic techniques.

Efficient construction of novel D-ring modified steroidal dienamides and their cytotoxic activities.

Two series of steroidal dienamides 4a-q and 5a-f were designed, synthesized and evaluated for cytotoxic activities against five human cancer cell lines (MGC-803, EC109, PC-3, SMMC-7721 and MCF-7). The protocol developed efficiently achieved the construction of carbon-carbon double bond and selective conversion of nitrile group into carboxamide in one-pot procedure. Besides, compounds 4a-q and 5a-f showed moderate to excellent cytotoxic activities with the IC50 values ranging from 0.1 to 40 µM and most of them were more potent than 5-fluorouracil. Particularly, four compounds 4d, 4e, 4q and 5a showed excellent selectivity against MGC-803 with the IC50 values less than 1 µM. Flow cytometry analysis demonstrated that compound 4c caused the cellular early apoptosis and cell cycle arrest in G2/M phase in a concentration-independent manner.

Triazole-dithiocarbamate based selective lysine specific demethylase 1 (LSD1) inactivators inhibit gastric cancer cell growth, invasion, and migration.

Lysine specific demethylase 1 (LSD1), the first identified histone demethylase, plays an important role in epigenetic regulation of gene activation and repression. The up-regulated LSD1's expression has been reported in several malignant tumors. In the current study, we designed and synthesized five series of 1,2,3-triazole-dithiocarbamate hybrids and screened their inhibitory activity toward LSD1. We found that some of these compounds, especially compound 26, exhibited the most specific and robust inhibition of LSD1. Interestingly, compound 26 also showed potent and selective cytotoxicity against LSD1 overexpressing gastric cancer cell lines MGC-803 and HGC-27, as well as marked inhibition of cell migration and invasion, compared to 2-PCPA. Furthermore, compound 26 effectively reduced the tumor growth bared by human gastric cancer cells in vivo with no signs of adverse side effects. These findings suggested that compound 26 deserves further investigation as a lead compound in the treatment of LSD1 overexpressing gastric cancer.

Hemophagocytic Lymphohistiocytosis Induced by Severe Pandemic Influenza A (H1N1) 2009 Virus Infection: A Case Report.

After early outbreaks in North America in April 2009, the pandemic influenza A (H1N1) virus spread rapidly around the world, and even some patients developed certain severe complications. We reported one case of hemophagocytic lymphohistiocytosis (HLH) induced by severe pandemic influenza A (H1N1) virus infection. A 17-year-old girl had acute onset of fever, dry cough, rhinorrhea, and sore throat Her family members and close friends also had the similar symptoms. Anti-infection treatment with penicillin was given after 8 days of the onset of symptoms in the local hospital, and her chest radiograph showed consolidation of the left lung. Then, she was sent to the People's Hospital of Guizhou Province in China and endotracheal intubation were underwent on the ninth day for acute hypoxic respiratory failure. She was diagnosed with HLH induced by severe pandemic influenza A (H1N1) 2009 virus. Oseltamivir, steroids, immunoglobulin, and plasmapheresis were given immediately after admission. After being treated in the People's Hospital of Guizhou Province for 16 days, she was discharged. This experience shows that HLH may be a life-threatening complication for severe pandemic influenza A (H1N1) 2009 virus infection and responds well to therapy.

Infliximab protects against pulmonary emphysema in smoking rats.

BACKGROUND: It is widely accepted that tumor necrosis factor-α (TNF-α) plays an important role in the pathogenesis of emphysema. This study aimed at investigating the protective effects of anti-TNF-α antibody, infliximab, in the development of emphysema induced by passive smoking in rats. METHODS: Thirty-nine rats were randomly divided into a normal control group (group 1), an emphysema group (group 2), and an infliximab-intervention group (group 3). Rat models of emphysema were established by exposure to cigarette smoking daily for 74 days. After 1 month, the infliximab intervention group was treated with infliximab via subcutaneous injection. The levels of TNF-α, IL-8 and vascular endothelial growth factor (VEGF) in bronchoalveolar lavage fluid (BALF) were measured with enzyme linked immunosorbent assay (ELISA). The number and classification of cells in the BALF were measured. Lung tissue sections stained by hematoxylin and eosin (HE) were observed, and mean linear intercept (MLI) and mean alveolar numbers (MAN) were measured. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) methods were used to examine the percentage of positive cells and distribution of apoptotic cells. RESULTS: The levels of TNF-α and IL-8 in BALF were higher in group 2 than in group 1 and group 3. The MLI was greater in group 2 than that in group 1 and group 3 while MAN was decreased. The concentration of VEGF in BALF of group 2 was significantly decreased as compared with group 1. The total cells and neutrophils number was significantly increased in group 2 as compared with group 1 and group 3, so was the percentage of neutrophils. The number of TUNEL positive cells in the alveolar septa was significantly increased in group 2 as compared with group 1 and group 3. CONCLUSION: Infliximab protects against cigarette smoking-induced emphysema by reducing airway inflammation, attenuating alveolar septa cell apoptosis and improving pathological changes.

Helicobacter pylori seroprevalence in patients with obstructive sleep apnea syndrome among a Chinese population.

OBJECTIVE: To investigate the seroprevalence of Helicobacter pylori (H. pylori) in patients with obstructive sleep apnea syndrome (OSAS), and to determine any association between H. pylori infection and severity of OSAS. METHODS: Two hundred and forty-three subjects were recruited in this cross-sectional study at the Department of Respiratory Medicine in the West China Hospital, Sichuan, P. R. China, from October 2006 to April 2008. Polysomnography (PSG) was used to determine the apnea-hypopnea index (AHI), and enzyme-linked immunosorbent assay was used to test H. pylori IgG. According to the AHI, subjects were divided into 4 groups: the control group (AHI < 5/hours), patients with mild OSAS group (AHI: 5-14/hours), moderate OSAS group (AHI: 15-29/hours), and severe OSAS group (AHI: > or = 30/hours). RESULTS: The prevalence of H. pylori infection in patients with OSAS was 75.5%, and in the controls it was 53.4% (p=0.000). The prevalence of H. pylori infection in patients with mild OSAS was 57.1%, with moderate OSAS was 76.5%, and with severe OSAS was 90.9%. There were significant differences between patients with moderate and severe OSAS and the controls, as well as among the mild, moderate, and severe OSAS groups. CONCLUSION: Helicobacter pylori infection may be associated with OSAS. In addition, increased severity of OSAS might be associated with higher seroprevalence of H. pylori.