Quantification of tryptase and T-cell immunoglobulin mucin 1 double-positive mast cells in different degrees of human periodontitis

AIM：To examine the expression of T-cell immunoglobulin mucin 1 (TIM-1) on tryptase-positive mast cells (MCs) in different severities of human chronic periodontitis. METHODS：Human gingival specimens (n=92) were involved in this study, including healthy control (n=27), mild chronic periodontitis (n=34) and severe chronic periodontitis (n=31). The gingival specimens were fixed in 4% formaldehyde. Paraffin embedding and serial sectioning with hematoxylin and eosin staining were performed for histopathological examination, and double-immunofluorescence staining was conducted for identification of tryptase-TIM-1 double-positive MCs in the gingival tissues. RESULTS：Compared with the healthy controls, the densities (cells/mm2) of tryptase-TIM-1 double-positive MCs were significantly increased in both mild chronic periodontitis group (P<0.05) and severe chronic periodontitis group (P<0.01). However, compared with mild chronic periodontitis group, both the score of gingival tissue inflammation and the density of tryptase-TIM-1 double-positive MCs in the gingival tissues were significantly increased in severe periodontitis group (P<0.05). CONCLUSION：Significantly increased number of tryptase-TIM-1 double-positive MCs has the similar tendency as the severity of periodontitis inflammation in human chronic periodontitis, suggesting that tryptase-TIM-1 double-positive MCs may play an important role in human chronic periodotitis.     

Expression of hypoxia-inducible factor 1α in human gingival tissues with chronic periodontitis

AIM：To observe the expression of hypoxia-inducible factor 1α (HIF-1α) in human gingival tissues with chronic periodontitis. METHODS：A total of 55 volunteers, including 15 healthy controls, 20 cases of moderate chronic periodontitis and 20 cases of severe chronic periodontitis, were involved in this study, and their gingival specimens were taken and fixed in 4% neutral formalin. The histological changes of gingival tissues were observed by HE staining, and the expression of HIF-1α in gingival tissues was detected by immunohistochemical staining. RESULTS： The proportion of HIF-1α positive cells in gingival tissues was significantly higher in chronic periodontitis groups than that in healthy control group (P<0.01), and that in severe chronic periodontitis group was significantly higher than that in moderate chronic periodontitis group (P<0.05). There was a significantly positive correlation between the severity of chronic periodontitis and the proportion of HIF-1α positive cells in gingival tissues. CONCLUSION：The expression of HIF-1α in human gingival tissues is increased with the severity of chronic periodontitis, suggesting that hypoxia may play an important role in chronic periodontitis.     

Expression of TLR2 and TLR4 on mast cells in human chronic periapical diseases

AIM: To observe the expression of TLR2 and TLR4 on mast cells(MCs) in the periapical tissues from different types of human chronic periapical diseases, and to analyze the role of TLR2 and TLR4 on tryptase-positive MCs in the immunopathogenesis of human chronic periapical diseases. METHODS: A total of 60 donors, including healthy control group, periapical granuloma group and periapical cyst group, were enrolled in the study. The periapical tissue specimens were fixed in 10% buffered formalin and stained with hematoxylin and eosin for histopathology, or stained with double-immunofluorescence for identification of TLR2-tryptase and TLR4-tryptase double-positive MCs in the periapical tissues. RESULTS: Compared with the healthy control, the densities of TLR2-tryptase and TLR4-tryptase double-positive MCs in periapical tissues were significantly increased in human chronic periapical diseases(P<0.01). The densities of TLR2-tryptase and TLR4-tryptase double-positive MCs in periapical cyst group were significantly higher than those in periapical granuloma group(P<0.01). CONCLUSION: TLR2 and TLR4 were expressed on the MCs in the periapical tissues of human chronic periapical diseases. TLR2-tryptase and TLR4-tryptase double-positive MCs may participate in the pathogenesis of chronic periapical diseases.     

Expression of GATA3 in human breast cancer tissues and its clinical significance

AIM: To investigate the expression of GATA3 in human breast carcinoma and its clinical significance. METHODS: The expression level of GATA3 in breast cancer tissues from 124 patients was detected by the method of immunohistochemistry and the relationships between GATA3 expression and other clinicopathological factors were analyzed. RESULTS: Low expression of GATA3 in breast cancer tissues was associated with estrogen receptor (ER)/progesterone receptor (PR) negative, high histological tumor grade, p53 mutation and vascular invasion (P<005), but not with age, tumor size,human epidermal growth factor receptor 2 (HER-2) expression and lymph node metastasis (P>005). In all breast cancer tissues, the positive expression rate of GATA3 was 56.4%. The positive expression rate of GATA3 in luminal breast cancer is 684%, higher than that in non-luminal breast cancer (326%, P<005). In all breast cancer tissues, the expression of GATA3 in middle recurrence risk group was higher than that in high recurrence risk group (P<005). CONCLUSION: GATA3 expression in breast cancer is related to differentiation and biological characteristics of the tumor, which can be a factor for evaluation of the treatment and prognosis.     

Effects of antihistamine treatment on mast cell infiltration and c-Kit and SCF expression in liver tissues of rats with experimental hepatitis

AIM：To study the infiltration of mast cells and the expression of c-Kit and stem cell factor (SCF) in liver tissues of rats with experimental hepatitis and their changes after antihistamine (AH) treatment. METHODS：Thirty Wistar rats were divided into 3 groups at random: normal control (NC) group, chronic hepatitis (CH) group and AH group. The rat model of CH was established by composite factors (subcutaneous injection of carbon tetrachloride, accompanied by a diet containing high cholesterol, high alcohol, low protein and low choline). The rats in AH group were treated with ketotifen based on CH. At the end of the 4th week, blood samples were taken to determine plasma tryptase (TS) and histamine (HA) levels. Liver tissues were taken to detect HA content, observe the histological changes with HE staining and count the number of mast cells with toluidine blue (TB) staining. The mRNA and protein expression of c-Kit and SCF in liver tissues was detected by RT-PCR and immunohistochemistry. RESULTS：(1) The plasma TS and HA levels and liver HA content in CH group were significantly increased compared with NC group (P<0.05), while those in AH group were obviously decreased compared with CH group (P<0.05). (2) Fatty degeneration and fibrosis were observed in CH group under light microscope, but the hepatic injury was obviously attenuated in AH group. TB staining showed there were many degranulating and degranulated mast cells filled with purple granules around liver blood vessels and in fiber interval in CH group, and there were few purple granules in the cytoplasm of mast cells in AH group. The number of mast cells in CH group was increased compared with NC group (P<0.05), and that in AH group was reduced compared with CH group (P<0.05). (3) The results of RT-PCR showed that AH down-regulated the expression of c-Kit and SCF mRNA (P<0.05). The expression of c-Kit and SCF proteins in liver tissues increased in CH rats (P<0.05 vs NC group), decreased after AH treatment (P<0.05 vs CH group) and was positively correlated with liver HA content (P<0.05). CONCLUSION：These data suggest that an inflammatory pathway mediated by mast cell activation is involved in experimental hepatitis. Ketotifen can reduce mast cell degranulation by down-regulating the expression of mast cell membrane receptor c-Kit and its ligand SCF, thereby attenuating the liver inflammation.     

Correlation between Helicobacter pylori infection and apoptosis in gingival tissues

AIM:To observe the effects of Helicobacter pylori(Hp) on the apoptosis of human gingival tissue.METHODS:Gingival tissue samples were taken from 30 patients without chronic periodontitis,and Hp was detected by conventional PCR.The apoptosis of the gingivival cells was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) to analyze the correlation between Hp infection and apoptosis of the gingival tissues.RESULTS:The Hp positive detections were 12 in the 30 patients without periodontitis,so the positive rate of Hp in the gingival tissue samples was 40%.The gingival tissue showed a large number of apoptotic cells in Hp positive group,and less apoptotic cells in Hp negative group.The apoptotic index in Hp positive group (0.498±0.092) was significantly higher than that in normal group (0.207±0.053)(P<0.05).CONCLUSION:Hp might play a role in the apoptosis of gingival tissues.     

Effects of metoprolol on phosphorylated connexin 43 expression in myocardial tissues and apoptosis of myocardial cells in heart failure rats

AIM：To explore the in vivo effects of metoprolol on the expression of phosphorylated connexin 43 (p-Cx43) in myocardial tissues and the apoptosis of myocardial cells in rats with heart failure (HF).METHODS：One hundred Sprague-Dawley rats were randomly divided into 5 groups (each n=20): sham group, HF group, low-dose (1.25 mg·kg-1·d-1) metoprolol treatment (MetoA) group, middle-dose (5 mg·kg-1·d-1) metoprolol treatment (MetoB) group and high-dose (20 mg·kg-1·d-1) metoprolol treatment (MetoC) group.The rats in HF group and metoprolol treatment groups were subject to abdominal aortic ligation, and different doses of metoprolol were given 4 weeks later till 8 weeks after operation.Echocardiography was conducted to monitor the hemodynamic parameters at the 4th and 8th weeks, and the rat hearts were taken at the 8th week after operation.The morphological changes and the proliferation of collagen fibers in myocardial tissues were observed by HE and Masson staining, respectively.The expression level of p-Cx43 was detected by Western blotting and the apoptosis of myocardial cells was assessed by TUNEL method.The relationship between p-Cx43 expression level and apoptotic index was analyzed by Pearson’s correlation.RESULTS：(1) Echocardiography showed that metoprolol could effectively improved cardiac hemodynamics in HF rats, and pathological findings suggested that metoprolol could effectively reverse HF-induced cardiac remodeling in a dose-dependent manner within the therapeutic dose range.(2) Western blotting showed that p-Cx43 expression in HF group was significantly higher than that in sham group (P<001), and that in all metoprolol treatment groups was significantly decreased compared with HF group (P<005 or P<001), among which pairwise comparisons also showed significant differences (P<001).(3) The myocardial apoptotic index in HF group ［(51.17±6.94)%］ was significantly increased compared with sham group ［(4.62±160)%, P<001］.Compared with HF group, myocardial apoptotic indexes in MetoA group ［(40.60±4.15)%］, MetoB group ［(30.66±4.00)%］ and MetoC group ［(22.24±5.69)%］ were significantly decreased (P<001), among which pairwise comparisons also showed significant differences (P<001).(4) The expression level of p-Cx43 was positively correlated with the apoptotic index (r=0.905, P<001).CONCLUSION: The mechanism of metoprolol against HF-induced myocardial apoptosis may be related to inhibition of p-Cx43 expression.     

Correlation between Toll-like receptor 4 on peripheral blood monocytes and serum IL-6 in patients with chronic severe hepatitis B

AIM:To study the change of Toll like receptor 4(TLR4) on peripheral blood monocytes (PBMCs) and its role in the pathogenesis of chronic severe hepatitis B.METHODS:The expression of TLR4 on CD14+ PBMCs was determined by flow cytometry in 30 healthy control,31 patients with chronic hepatitis B and 30 patients with chronic severe hepatitis B. The level of serum interleukin-6 (IL-6) was detected by ELISA. RESULTS:The expressions of TLR4 on PBMCs and serum IL-6 in the groups of healthy control,patients with chronic hepatitis B and patients with chronic severe hepatitis B were 2.3±1.1,3.7±2.3,6.9±4.1 mean fluorescence intensity (MFI) and （11.5±7.2) ng/L,(40.8±31.2) ng/L,(77.6±33.3） ng/L. The TLR4 value in the group of patients with chronic severe hepatitis B was significant higher than that in the group of healthy control and the group of patients with chronic hepatitis B (P<0.05). However,there was no significant difference between the group of patients with chronic hepatitis B and the group of healthy control (P>0.05). Serum IL-6 increased gradually and significantly between the group of healthy control and the groups of patients with chronic hepatitis B and patients with chronic severe hepatitis B. There was a significant positive correlation between the expression of TLR4 and the content of serum IL-6 in the group of chronic severe hepatitis B. CONCLUSION:TLR4 may play a role in the pathogenesis of chronic severe hepatitis B.     

Effect of hyperbaric oxygen on periodontitis with psychological stress in rats

AIM: To investigate the role of chronic psychological stress on periodontitis and the effects of hyperbaric oxygen (HBO) on periodontitis with psychological stress in rats. METHODS: Male special pathogen-free Wistar rats (n=80) were randomly divided into 4 groups: (1)normal control group; (2)experimental periodontitis group: the periodontitis model was induced by wrapping 3/0 silk ligature inoculated with Porphyromonas gingivalis around the left maxillary second molar of the rats; (3)psychological stress stimulation group; (4)periodontitis model with stress stimulation group. Psychological stress was removed at the 9th week after ligature, and 4 rats from each experimental group were randomly chosen for HBO treatment. The rats were sacrificed at the 2nd, 4th, 8th and 10th weeks after ligature. The levels of blood glucose, adrenocorticotropic hormone (ACTH), corticosterone and adrenaline were measured as the stress markers. The histological changes of periodontal tissues were observed under microscope with HE staining. RESULTS: The levels of blood glucose, ACTH, corticosterone and adrenaline in psychological stress stimulation group and periodontitis with stress group were significantly higher than those in control group and experimental periodontitis group at the 2nd and 4th weeks after ligature (P<0.05). The levels of the stress markers were significantly lower than those in untreated groups in the 10th week after HBO (P<0.01). The sites of gingival attachment were normal in control group and psychological stress stimulation group. Periodontal pocket, and periodontal attachment loss (AL) were observed in experimental periodontitis group. The tissue damage was much heavier in periodontitis model with stress stimulation group as the furcation of tooth was exposed and the tissue damage was observed on both sides of the adjacent teeth. No significant difference of AL between psychological stress stimulation group and normal control group during the experiment was observed. The AL in periodontal model with stress stimulating group was significantly higher than that in experimental periodontitis group at the 2nd, 4th and 8th weeks (P<0.01). The level of AL was attenuated at the 10th week after HBO (P<0.01). No difference of histological change in periodontal tissues was observed between control group and psychological stress stimulation group. Severer inflammatory changes and alveolar bone destruction were observed in periodontitis with stress group than those in experimental periodontitis group. The levels of inflammation reduced at the 10th week after HBO. CONCLUSION: Stress stimulation is one of the inducing factors of periodontitis in rats, which aggravates periodontitis. HBO may represent a useful way in treating psychological stress periodontitis.     

Effects of transplantation of adipose stromal vascular fraction on cardiac function in adriamycin-induced heart failure rats

AIM: To investigate the effects of transplantation of adipose stromal vascular fraction (SVF) on the cardiac function in adriamycin-induced heart failure rats. METHODS: SVF was isolated from adipose tissue of a Sprague-Dawley (SD) rat by collagenase digestion and marked with green fluorescent protein (GFP) in vitro. Twenty-eight SD rats were randomized into normal control group (n=8), adriamycin control group (n=10) and SVF treatment group (n=10). Adriamycin at dose of 15 mg/kg was intraperitoneally injected into the rats twice a week for 4 weeks to induce heart failure. SVF cells (05 mL, 1×107/L) were injected via penis vein, and PBS vehicle was injected into the control animals in the same way. Four weeks later, the cardiac function was determined by multichannel physiologic recorder via cardiac catheterization. SVF was demonstrated in the myocardium by frozen section fluorescence microscopy. The CD31 expression was determined by an immunohistochemical test. RESULTS: Compared with adriamycin control group, SVF transplantation increased left ventricular peak systolic pressure ［LVSP, （13565±21.58) mmHg vs (10558±2262) mmHg, P<005］, left ventricular pressure maximal rise rate ［+dp/dtmax, (4 81565±56624) mmHg/s vs (3 53550±46528) mmHg/s, P<005］, and left ventricular pressure maximal decline rate ［-dp/dtmax, (3 67756±46775) mmHg/s vs (2 73865±51251) mmHg/s, P<005］. The results of the CD31 immunohistochemical test showed that the nuclear staining and granule distribution were more uniform, and the number of blood vessels per visual field increased in SVF treatment group as compared with adriamycin control group (P<005). CONCLUSION: SVF transplantation improves the cardiac function in the rat model of heart failure, possibly and partly through the promotion of myocardial neovascularization.     

The expression of receptor of epidermal growth factor in gingiva of patient with periodontitis

AIM:To observe the expression of epidermal growth factor receptors(EGFR) of gingival tissue in periodontitis. METHODS: The expression of EGFR was determined by using immunohistochemical techniques in gingival tissue of 15 healthy individual, 32 cases with adult periodontitis (AP) and 12 cases with juvenile periodontitis (JP). RESULTS: Expression of EGFR was mainly located on basal cell membranes in healthy gingiva, and the staining intensity was faint. In AP cases, expression of high level EGFR was mostly observed on the membranes of epithelial cell in the periodontal endopocket or junctional epithelium, intensity of staining appeared to decrease gradually with the differentiation of keratinocytes, and the horny cell layer was not stained by the antibody. In JP cases, strong positive staining was present on membrane of epithelial cells in the germinative layer of gingival tissue. There was an apparent difference between healthy gingiva and AP or JP (P＜ 0.01 ), or AP and JP (P＜ 0.01). CONCLUSION: The distribution and expression of EGFR in gingival epithelium of periodontitis showed obvious differ- euce. The data indicated that EGFR may affect apical migration of junctional epithelium, and may play a role in development of AP and JP.     

Hypoxia-inducible factor-1α attenuates myocardial inflammatory injury induced by ischemia and reperfusion in rats

AIM:To investigate the role of hypoxia-inducible factor-1α (HIF-1α) stable expression in myocardial inflammatory injury induced by ischemia and reperfusion (I/R) in rats. METHODS:Male Wistar rats were randomly divided into 4 groups:sham operation (sham) group, I/R group, HIF-1α stabilizer dimethyloxalyl glycine (DMOG)+I/R group and HIF-1α inhibitor YC-1+I/R group. The protein expression of myocardial Toll-like receptor 4 (TLR4) and nuclear factor-κB (NF-κB) was determined by Western blot. The mRNA levels of interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), IL-6, TLR4 and NF-κB were detected by real-time PCR. The myeloperoxidase (MPO) activity in the myocardial tissues was measured. HE staining was used to observe the infiltration of inflammatory cells. RESULTS:HIF-1α decreased the infiltration of inflammatory cells, the MPO activity, and the mRNA levels of inflammatory factors IL-1β, IL-6 and TNF-α in the myocardial tissues. HIF-1α also reduced the expression of TLR4 and NF-κB at mRNA and protein levels (P<0.05). CONCLUSION:The stable expression of HIF-1α has an anti-inflammatory effect on the myocardial tissues after I/R injury in rats. The mechanism may be related to the inhibition of TLR4/NF-κB signaling pathway.     

Effect of simvastatin on function of mouse pancreatic beta cells and its mechanisms

AIM: To observe the influence of simvastatin on insulin secretion function of mouse pancreatic beta cell line MIN6 and to explore its possible mechanisms.
METHODS: MIN6 cells were randomly divided into normal control group and low-, middle-and high-concentration simvastatin treatment groups, which were cultured for 48 h with high-glucose DMEM containing 15% fetal bovine serum plus 0, 2, 5 and 10 μmol/L simvastatin, respectively. The insulin secretion of MIN6 cells was measured by radioimmunoassay. The content of ATP in MIN6 cells was measured by biochemiluminescence method. The mRNA and protein expression levels of inwardly rectifying potassium channel 6.2 (Kir6.2), voltage-dependent calcium channel 1.2 (CaV1.2) and glucose transporter 2 (GLUT2) were detected by real-time fluorescence quantitative PCR and Western blotting, respectively.
RESULTS: Compared with normal control group, middle-and high-concentration simvastatin treatment markedly decreased the synthesis and secretion of insulin in MIN6 cells (P<005). The content of ATP in MIN6 cells was markedly decreased in simvastatin treatment groups (P<005). The mRNA expression level of Kir6.2 in MIN6 cells was significantly up-regulated in simvastatin treatment groups (P<001), while the mRNA expression levels of CaV1.2 and GLUT2 were significantly down-regulated in middle-and high-concentration simvastatin treatment groups (P<001). The protein expression of Kir6.2 was significantly increased but that of CaV1.2 was significantly decreased in middle-and high-concentration simvastatin treatment groups (P<001), and the protein expression level of GLUT2 was markedly decreased in high-concentration simvastatin treatment group (P<001).
CONCLUSION: Simvastatin inhibits insulin synthesis and secretion in mouse pancreatic beta cell line MIN6 via suppressing ATP production, up-regulating the expression of Kir6.2 and down-regulating the expression of CaV1.2 and GLUT2.     

Role of endoplasmic reticulum stress in Bim-mediated cardiomyocyte apoptosis induced by hypoxia

AIM:To investigate the role of endoplasmic reticulum stress (ERS) in the process of Bim-mediated cardiomyocyte apoptosis induced by hypoxia. METHODS:Cardiomyocytes were isolated from neonatal Sprague-Dawley rats aged 1~3 days, and primarily cultured in vitro. The antibody targeting α-striated muscle actin was used to identify the cardiomyocytes. The siRNAs targeting bim were transfected into cardiomyocytes with liposome, followed by detecting the expression of Bim by Western blotting. Cardiomyocytes were divided into five groups: blank control group, hypoxia group, hypoxia+liposome group, hypoxia+negative control siRNA group and hypoxia+Bim-siRNA group. The cell viability was determined by MTT assay, and the cell apoptotic rate and the intracellular calcium concentration were measured by flow cytometry. The protein expression of caspase-12 and inositol 1,4,5-triphosphate (IP3) was detected by Western blotting. RESULTS:Immunohistochemical identification confirmed that rat cardiomyocytes were successfully cultured. Green fluorescence was observed in the cells transfected with negative control siRNA under fluorescence microscope. The expression of Bim was obviously inhibited after transfected with Bim-siRNAs and the silencing efficiency of Bim-siRNA-2 was the highest (86.73%). Compared with blank control group, the viability of cardiomyocytes in hypoxia group was significantly reduced (P<005). Compared with hypoxia+negative control siRNA group, the viability of cardiomyocytes in hypoxia+Bim-siRNA group was significantly increased (P<005). The apoptotic rate and the intracellular calcium concentration of cardiomyocytes were obviously increased in hypoxia group (P<0.01), and were both decreased after bim silencing (P<005 or P<0.01). The expression of caspase-12 and IP3 was up-regulated in hypoxia group (P<005), and was down-regulated after bim silencing (P<005 or P<0.01). CONCLUSION: Cardiomyocyte apoptosis induced by hypoxia can be inhibited by silencing the expression of bim gene. Caspase-12 and IP3, as markers of ERS, may participate in the process of Bim-mediated cardiomyocyte apoptosis induced by hypoxia.     

Influence of amylin on apoptosis of human pancreatic islet β-cells and its molecular mechanism

AIM: To investigate the molecular mechanism of amylin in inducing apoptosis of human pancreatic islet β-cells. METHODS: Human pancreatic islet cells were isolated and cultured. The cells were treated with amylin or amylin and aminoguanidine (AG group) for 24 h, respectively. Apoptosis of pancreatic islet β-cells was studied by in situ TUNEL method combined with double staining for insulin and ELISA. The levels of insulin, NO2-/NO3- and glutathione (GSH), p53 mRNA and bcl-2 mRNA were also detected. RESULTS: (1) The enrichment factor and the apoptosis rate of pancreatic islet β-cells in amylin group were markedly higher than that in control group and AG group (P<001). (2) The insulin level in amylin group was significantly lower than that in control group and AG group (P<005). (3) The levels of NO2-/NO3- and p53 mRNA in amylin group were significantly higher than that in control group and AG group (P<001), while the levels of GSH and bcl-2 mRNA were markedly decreased as compared with control group and AG group. CONCLUSIONS: Amylin increases apoptosis of human pancreatic islets β cells, resulting in decrease in insulin secretion. This may be due to increased expression of p53 and decreased expression of bcl-2 during the of oxidative stress induced by amylin.     

Effect of hyperbaric oxygen on HIF-1α expression in rat experimental periodontitis with psychological stress

AIM: To investigate the effect of hyperbaric oxygen (HBO) on hypoxia-inducible factor-1α (HIF-1α) expression in rat experimental periodontitis with psychological stress. METHODS: Male special pathogen-free Wistar rats (n=120) were randomly divided into 4 groups: normal control group; psychological stress stimulation group; experimental periodontitis group: the periodontitis model was induced by wrapping 3/0 silk ligature inoculated with Porphyromonas gingivalis around the left maxillary second molar of the rats; periodontitis model with stress stimulation group. Psychological stress was removed at the 9th weeks after ligature, 6 rats from each experiment group were randomly chosen to HBO treatment. The rats were sacrificed at the 2nd, 4th, 8th and 10th weeks after ligature. Gingival index (GI) and attachment loss (AL) were measured before sacrifice. The histological changes of periodontal tissues were observed under microscope with HE staining. The expression of HIF-1α was observed by the method of immunohistochemistry. RESULTS: The sites of gingival attachment were normal in control group and psychological stress stimulation group. Periodontal pocket, and periodontal attachment loss were observed in experimental periodontitis group. The tissue damage was much serious in periodontitis model with stress stimulation group. No significant difference of GI and AL among psychological stress stimulation group and normal control group during the experiment was observed. GI and AL in periodonitis model with stress stimulating group were significantly higher than those in experimental periodontitis group at the 4th and 8th weeks (P < 0.01). The levels of GI and AL were significantly lower at the 10th weeks after HBO treatmnt than those in untreated groups (P < 0.05). No significant difference of HIF-1α expression scores among psychological stress stimulation group and normal control group was found. HIF-1α expression scores in periodonitis model with stress stimulating group was significantly higher than that in experimental periodontitis group at the 4th and 8th weeks (P < 0.01). At the 10th weeks after HBO treatment the levels of HIF-1α were significantly lower than that in untreated groups (P < 0.01). CONCLUSION: Stress stimulation may aggravate periodontitis by decreasing tissue oxygenation in rats. HBO may represent a useful way in psychological stress periodontitis therapy.     

Effect of azithromycin on airway inflammation and airway mucus hypersecretion in rats with chronic obstructive pulmonary disease

AIM: To observe the effect of azithromycin on the rats with chronic obstructive pulmonary disease (COPD), and to explore the underlying mechanism about the airway inflammation and mucus hypersecretion. METHODS: Male SD rats were randomly divided into normal control group, COPD model group, azithromycin treatment group. The COPD model was established by the method of cigarette smoking combined with intratracheal injection of LPS. Pathological changes of the bronchi and lung tissues of the rats were observed with HE staining. Pulmonary ventilation function in the rats was detected with pulmonary function instrument. The levels of IL-8, IL-17 and TNF-α in bronchoalveolar lavage fluid (BALF) were measured by ELISA. The expression of MUC5ac and TLR4 at mRNA and protein levels in bronchi and lung tissues was determined by real-time PCR and Western blot.RESULTS: HE staining showed that the changes of bronchi and lung tissues in model group were consistent with typical pathological manifestations of COPD. Compared with model group, these changes were alleviated in treatment group. The pulmonary functions in model group were significantly decreased compared with control group. The levels of IL-8, IL-17 and TNF-α in the BALF in model group were significantly increased compared with control group (P <0.05). The expression of MUC5ac and TLR4 at mRNA and protein levels in model group was significantly higher than that in control group (P <0.05). Compared with model group, the degree of the descent in pulmonary function in treatment group was significantly lessened. Compared with model group, the levels of IL-8, IL-17 and TNF-α in treatment group were significantly inhibited (P <0.05). Furthermore, the expression of MUC5ac and TLR4 at mRNA and protein levels in treatment group was significantly lower than that in model group (P <0.05). CONCLUSION: Azithromycin decreases the levels of IL-8, IL-17 and TNF-α in the BALF of COPD model rats, inhibits the protein expression of MUC5ac and TLR4 in the lung tissues, thus playing a preventive and therapeutic role to reduce airway inflammation and airway mucus hypersecretion.     

Effects of baicalin on serum levels of IL-6 and IL-4 in mouse periodontitis

AIM: To investigate the effect of baicalin on experimental periodontitis in mouse model by comparing the histological changes in periodontal tissues and serum levels of inter leukin(IL)-6/IL-4 in mice, and to analyze the role of baicalin in immune regulation and anti-inflammatory mechanisms. METHODS: Twenty-seven male Kunming mice (SPF grade, 12-week-old) were randomly divided into 3 groups. The naive mice were used in normal control group. In experimental periodontitis group, the periodontitis model was produced by ligature of braided silk around the first maxillary molar and inoculation with putative periodontopathic bacteria. Five weeks after the ligature, the mice were fed with 10% glucose, and gavaged with distilled water. In baicalin treatment+periodontitis group, the periodontitis model was induced as above, then gavaged with baicalin at the beginning of the fifth week after the ligature. The mice were sacrificed at week 4, 6 and 8. The histological changes of the periodontal tissues were observed under microscope with hematoxylin and eosin (HE) staining. The serum level of IL-6 and IL-4 in the mice were determined by ELISA. RESULTS: The periodontal tissues showed moderate inflammatory damages in experimental periodontitis group. The periodontal destruction was significantly reduced in baicalin treatment+periodontitis group. The serum level of IL-6 in experimental periodontitis group was significantly higher than that in control group and baicalin treatment+periodontitis group (P<0.01), and the serum level of IL-6 in baicalin treatment+periodontitis group was significantly lower than that in periodontitis group at week 6 and 8 (P<0.01). The serum level of IL-4 in periodontitis group was significantly lower than that in control and baicalin treatment+periodontitis group (P<0.01). The serum level of IL-4 in baicalin treatment+periodontitis group was significantly higher than that in periodontitis group at weeks 6 and 8 (P<0.01).CONCLUSION: The pathogenesis of periodontitis is closely related to the imbalance of Th1/Th2 cytokines, characterized by increased serum level of IL-6 and the decreased serum level of IL-4. Baicalin plays a significant role in treating mouse periodontitis by decreasing the serum level of IL-6 and increasing the serum level of IL-4.     

Effect of Toll-like receptor 4 signal transduction on airway inflammation in infant asthmatic rat

AIM: To study the change and regulatory mechanism of Toll-like receptor 4 (TLR4) on eosinophil (EOS) apoptosis. METHODS: Twenty-seven SD rats were randomly divided into control group (A), asthma group (B) and dexamethasone group (D). Asthmatic model rats were sensitized and repeatedly exposed to aerosolized ovalbumin. Pulmonary tissues were observed under light microscope (LM). The inflammatory cells in BALF were counted. The levels of IL-10 in serum were measured by ELISA. Expressions of TLR4 mRNA were tested by hybridization. The apoptotic EOS was detected by TUNEL.RESULTS: (1) LM showed that inflammatory cells infiltrated around the bronchus, airway mucous plug in group B, obviously lightened in group D. (2) Inflammatory cells count in BALF: the total cellular score, EOS absolute count and EOS% in group B were significantly increased (P<0.01). Compared to group B, a significant decrease in group D was observed (P<0.01). (3) The level of IL-10 in group B was significantly higher than that in group A and in group D (P<0.01). (4) No significant difference (P>0.05) of TLR4 mRNA expression was observed between group A and group B. However, that in group D were significantly increased (P< 0.01). (5) Percentages of apoptotic EOS in group B were significantly lower than those in group A (P<0.01), those in group D were significantly increased (P<0.01). A significant correlation between TLR4 mRNA and apoptotic EOS (r=0.612, P<0.01) was observed. CONCLUSION: Dexamethasone can increase IL-10 secretion, induce EOS apoptosis, which may correlate with TLR4 signal transduction.     

Effect of nimesulide on ligature-induced periodontitis in rats

AIM: The purpose of this study was to observe the effect of nimesulide on periodontitis. METHODS: The gingival index (GI) was measured before the rats were sacrificed at the ends of week 4, 5 and 8. The periodontal tissues were stained with hematoxylin and eosin. Histological changes were observed by microscope. The periodontal attachment loss (AL) was measured by Tiger cell image analyzer. RESULTS: (1) Experimental periodontitis was successfully induced in rats by placing a piece of 3/0 braided silk around the cervix of the lower incisors at week 4 after the ligature. (2) In ligature-induced periodontitis group, at week 4 after the ligature, the GI and AL were significantly higher than those in control group (P<0.01). The histopathologic changes of periodontium in periodontitis group showed obvious inflammation, and the severity of destruction for periodontium was increased as time passed. (3) In the nimesulide prevention group, the GI and AL were significantly lower than those in periodontitis group (P<0.01). The histopathologic examination showed less inflammatory responses, and no obvious alveolar bone resorption was observed. (4) In the nimesulide treatment group, the GI and AL were significantly lower than those in periodontitis group at the end of week 5 and 8 after the ligature (P<0.01).CONCLUSION: (1) In ligature-induced periodontitis, nimesulide inhibits effectively its progress. (2) In the developing periodontitis, a significant improvement is observed in GI and AL following the treatment with nimesulide.