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.     

Change of nitric oxide concentration in ligature-induced periodontitis in rats

AIM: To study the possible role of nitric oxide (NO) in the development of periodontitis and the relationship between the NO concentration and the attachment loss. METHODS: Seventy-two Sprague-Dawley rats were randomly assigned to two groups, the control group and periodontitis group. Experimental periodontitis in rats was produced by a ligature of braided silk. The nitric oxide concentration was indirectly ascertained by the concentration of nitrite (NO2-) and nitrate (NO3-) in the gingival tissue, which was assayed by spectrophotometry. The attachment loss (AL) was measured by the technology of the cellular graphics engineering research. The histopathologic change in periodontium was observed under a light microscope by using the histotomy. RESULTS: Compared to control group, the NO2-/NO3- concentration in gingival tissue was significantly higher in periodontitis group at four weeks and eight weeks following ligation (P<0.01). In periodontitis group, the NO2-/NO3- concentration in gingival tissue was higher at eight weeks than that at four weeks following ligation (P<0.01). At four weeks and eight weeks, the AL in experimental periodontitis in rats was significantly increased than that at one week after ligation (P<0.01); and the AL was also much higher at eight weeks than that at four weeks (P<0.01). CONCLUSIONS: The NO2-/NO3- concentration in the gingival tissue in periodontitis group was significantly higher than that in control group. These results demonstrate that the NO2-/NO3- concentration is related to the severity of AL, and NO synthesis is very important to the process of inflammation and lesion in periodontium. Reducing NO production may be of great therapeutic value in the treatment of periodontitis.     

The inhibitory effect of aminoguanidine on periodontitis in rats

AIM: To study the inhibitory effect of aminoguanidine(AG) on periodontitis. METHODS: Experimental periodontitis was produced in rats by a ligature of braided silk, the NO concentration was assayed by spectrophototometry, the attachment loss of periodontium was measured by using technology for image and graphics engineering research and the histopathologic changes of periodontium were also examined under a light microscope. RESULTS: The NO concentration significantly decreased after AG treatment for 4 weeks, and the inflammation and the damages of periodontium was also reduced significantly. CONCLUSION: The inhibition of iNOS by AG may be a potential therapeutic strategy for the treatment of periodontitis.     

Effect of Zhiqi oral solution on adult periodontitis

AIM:To evaluate therapeutic effect of Zhiqi oral solution on adult periodontitis.METHODS:Supragingival scaling and root planning were carried out in 100 patients with adult periodontitis. Then the patients with periodontitis were divided into two groups randomly. 32 patients in the control group took orally Metronidazole and 68 patients in the experimental group took orally Zhiqi oral solution. The pain reaction, bleeding index, probing depth, plaque index, changes of microorganisms in periodontal pocket and side effect before and after the treatment were observed at a 30 days observation period.RESULTS:Zhiqi oral solution had a therapeutic effect on periodontitis, compared with the control group (P＜0.05). The bleeding index and plaque index in patients of two groups were improvedobviously compared with before treatment (P＜0.05). However Zhiqi oral solution had an obvious effect on decreasing the probing depth(P＜0.05).CONCLUSION:Zhiqi oral solution improves effectively the condition of periodontium, and its supplementary treatment effect on adult periodontitis is obvious.     

Effect of psychological stress on development of periodontitis and expression of periodontal hypoxia-inducible factor 1α in rats

AIM: To investigate the effect of psychological stress on the development of periodontitis and the expression of periodontal hypoxia-inducible factor 1α(HIF-1α) in rats.METHODS: Forty-eight male Wistar rats(SPF grade) were randomly divided into 4 groups:(1) normal control group, i.e. naive rats;(2) experimental periodontitis group: the periodontitis model was induced by wrapping 3-0 silk ligature inoculated with putative periodontopathic bacteria around the left maxillary second molar of the rats;(3) stress group: the rats were treated with stress alone;(4) periodontits with stress group, the periodontitis model was induced as above,and the rats were treated with stress. The rats were sacrificed at week 1, 4, 6 and 8 after the ligature. The attachment losses(AL) were measured by home-made probe. The histological changes of periodontal tissues stained with hematoxylin and eosin(HE) were observed under microscope. The HIF-1α expression level in the periodontal epithelium was determined by immunohistochemistry that was used to evaluate the severity of hypoxia by measuring the average rate of HIF-1α-positive cells.RESULTS: No significant difference of AL between stress group and normal control group was observed(P>0.05).The AL and the average rate of HIF-1α-positive cells in periodontitis with stress group were significantly higher than those in experimental periodontitis group at time points of week 4,6 and 8 after ligature(both P<0.01).CONCLUSION: Psychological stress is one of the periodontitis inducing factors in the animal model. Psychological stress may aggravate periodontitis by decreasing tissue oxygenation in rats.     

Influence of chronic psychological stress on periodontitis and serum isoenzyme LDH1 in rats

AIM: To investigate the role of chronic psychological stress on periodontitis and serum level of isoenzyme LDH1 in rats. METHODS: Eighty female special pathogen-free Wistar rats 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 putative periodontopathic bacteria around the left maxillary second molar of the rats; (3) psychological stress stimulation group: the rats were treated with stress stimulation alone; (4) periodontitis model with stress stimulation group: the periodontitis models were induced by wrapping 3/0 silk ligature inoculated with putative periodontopathic bacteria around the left maxillary second molar of the rats, and then treated with stress stimulation. The rats were sacrificed at 2nd, 4th, 6th and 8th weeks after ligature. The levels of blood glucose and adrenocorticotropic hormone (ACTH) were measured as the stress markers, and the serum isoenzyme lactate dehydrogenase 1(LDH1) was used to evaluate the severity of hypoxia. The histological changes of periodontal tissues were observed under microscope with HE staining. RESULTS: The levels of blood glucose and ACTH in stress 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<0.01). The level of serum LDH in periodontitis with stress group was significantly higher than that in control group. The level of serum LDH1 in periodontitis with stress group was significantly lower than that in control group (P<0.01). No difference of histological change in periodontal tissues was observed in control group and stress group. Severer inflammatory changes and alveolar bone destruction were observed in periodontitis with stress group than those in experimental periodontitis group. CONCLUSION: Stress stimulation is one of the inducing factors of periodontitis in rats that aggravates periodontitis by decreasing the tissue oxygenation.     

Study on COX activity and its mRNA expression,and PGE2 release from cerebral microvascular endothelial cells stimulated by IL-1β

AIM: To study the cyclooxygenase(COX) activity and its mRNA expression, and PGE₂ release from rats cerebral microvascular endothelial cells (rCEMC) stimulated by IL-1β(30 μg/L) at different times. METHODS: rCMEC were cultured, and identified by immunohistochemistry for von Willebrand factor (Ⅷ factor, a marker for all endothelial cells) in cytoplasm of the cells. After rCEMC grew to confluency, they were stimulated with IL-1β for 0.5, 1, 2, 4, 8, 12 and 24 h, respectively. Activity of COX-1 and COX-2 in rCEMC and production of PGE₂ in the conditioned media were detected by ELISA. COX-1 and COX-2 mRNA expressions were measured by real-time quantity PCR. The amplification product was tested by melting curve and identified by electrophoretic gel. RESULTS: ① Positive immunostaining for Ⅷ factor was present diffusely in the cytoplasm in more than 90% rCMEC. ② Compared to the cells without IL-1β stimulation, the production of PGE₂ increased significantly (P<0.05) at 4 h after rCEMC were incubated with IL-1β and reached the top level at 12 h (P<0.01), then declined thereafter at 24 h (P<0.05). ③ There was no significant difference on COX-1 activity between IL-1β group and non-IL-1β group. COX-2 activity increased significantly compared with those in non-IL-1β (P<0.05) at 8 h after rCEMC were incubated with IL-1β and reached the top level at 12 h (P<0.01), then declined thereafter at 24 h (P<0.05). ④ There was no significant difference on COX-1 mRNA expression between IL-1β group and non-IL-1β group. COX-2 mRNA was induced and became detectable at 1 h, and reached the top level at 4 h, then declined thereafter at 8 h and became undetectable by 12 h and 24 h after incubation with IL-1β. The melting curve showed there was no nonspecific amplification and electrophoretic gel showed the lengths of amplification products accorded with the predicted lengths. CONCLUSION: While rCEMC are stimulated by IL-1β, the excretion of PGE₂ increases and reaches the top level at 12 h, which is related with its induction on COX-2 mRNA expression and COX-2 activity.     

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 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.     

Toll-like receptor 4 expression on mast cells in human gingival tissues with chronic periodontitis

AIM: To observe the expression of Toll-like receptor 4 (TLR4) on mast cells in human gingival tissues with chronic periodontitis. METHODS: A total of 68 volunteers, including 23 cases of mild chronic periodontitis, 25 cases of severe chronic periodontitis and 20 healthy controls, 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. The expression of TLR4 in gingival tissues was detected by immunohistochemical staining, and TLR4 expression on mast cells was detected by immunofluorescence double staining. RESULTS: The expression of TLR4 in gingival tissues and on mast cells in chronic periodontitis groups was significantly higher than that in normal control group (P<005), and that in severe chronic periodontitis group was significantly higher than that in mild chronic periodontitis group (P<005). CONCLUSION: The expression of TLR4 in gingival tissues and on mast cells is increased with the severity of chronic periodontitis, suggesting that TLR4, especially TLR4 on mast cells, may play an important role in human chronic periodontitis.     

Relationship between TNF-α gene polymorphism-308 and type 2 diabetes with periodontitis

AIM: To investigate the gene polymorphism-308 of tumor necrosis factor alpha (TNF-α) in the relation with the susceptibility to periodontitis combined with type 2 diabetes mellitus (DM) and its severity.METHODS: Human DNA samples were obtained from 240 DM patients with periodontitis (periodontitis group, n=120) and without periodontitis (control group, n=120). All patients were genotyped by PCR-RFLP analysis. The frequencies of genotypes and alleles were analyzed. Sulcus bleeding index (SBI) and probing depth (PD) in all patients were measured. The polymorphism-308 of TNF-α gene in the relation with the susceptibility to periodontitis combined with DM and its severity was analyzed.RESULTS: No significant difference in the frequency of genotype and allele was found between DM patients with mild periodontitis and DM patients without periodontitis (P>0.05). However, the frequencies of these genotypes and alleles in DM patients with moderate and severe periodontitis were significantly higher than those in DM patients without periodontitis (P<0.01). The findings showed that the level of TNF-α was associated with SBI and PD (P<0.01).CONCLUSION: TNF-α -308 G/A polymorphism is not associated with DM patients with mild periodontitis, whereas it may have a role in pathogenesis and prognosis of moderate and severe periodontitis combined with DM through TNF-α level.     

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.     

Changes of PGE2, PGI2 and TXA2 concentration in rat traumatic pulpitis

AIM: To study the relationship between prostaglandins and acute pulpitis. METHODS: Rat traumatic pulpitis model was established by pulp exposure. The kinetic pathological changes in dental pulpal tissues and changes of PGE₂,6-Keto-PGF_1α and TXB₂ concentration in dental pulp were observed. RESULTS: After pulpal trauma, the dental pulp showed inflammatory changes and the concentrations of PGE₂,6-Keto-PGF_1α and TXB₂ were increased, which peaked at 6 hour post-trauma. CONCLUSION: Prostaglandins play a significant role in the pathogenesis of pulpitis.     

Effect of Ginkgo biloba extract on retinopathy in diabetic rats

AIM: To investigate the effect of Ginkgo biloba extract(GBE) on diabetic retinopathy(DR) and its possible mechanism in rats. METHODS: Goto-Kakizaki(GK) rats were used as a DR model, and were treated with different doses of GBE. Normal Wistar rats were used as the control. Blood glucose and retina barrier injury were analyzed, respectively. Ganglion cell apoptosis was detected by TUNEL staining. Moreover, the protein expression of nuclear factor E2-related factor 2(Nrf2), ERK, Bcl-2 and P53, and ERK phosphorylation were examined by Western blot.RESULTS: GBE reduced blood glucose in the DR rats, attenuated retina barrier injury, and decreased the apoptosis of ganglion cells. Furthermore, the expression of Nrf2 and Bcl-2, and phosphorylation of ERK were increased after GBE treatment, whereas P53 expression was decreased. CONCLUSION: GBE protects ganglion cells against apoptosis in DR rats, which may be through activation of Nrf2/ERK pathway and regulating Bcl-2 and P53 expression.