| Backgrounds and Objectives:Natural environment in high altitude region, including low atmospheric pressure andpartial pressure of oxygen, cold, strong winds and intense radiation, can affect human’s oraland systemic health seriously. A large number of domestic studies show that the incidencesand process speed of periodontitis in plateau region are much higher than that in plainregion. However, current much of research focus on epidemiological investigation, thereare little known about the relation between pathophysiology and pathogenesis ofperiodontitis and environment in high altitude region.The dental plaque was an initiating factor of periodontitis. The mainly periodontalpathogenic bacteria were anaerobe. The occurrence and development of periodontitis werecorrelated with the defense mechanisms of host, the interaction among bacteria and thenumber of bacteria planting in periodontal pockets and so on. Due to complexity of thelocal ecological environment and relationship among bacteria as well as technical obstacles,many problems were still not fully elucidated. In our preliminary studies, it was found thatthe periodontal pathogenic bacteria of Sprague-Dawley rats under the stimulated hypoxiaenvironment were various and the species of bacteria were significantly different from thatunder the normoxia environment by polymerase chain reaction-denaturing gradient gelelectrophoresis (PCR-DGGE). There was little known about the effects of the number andendotoxin levels of anaerobe on pathophysiology and pathogenesis of periodontitis.In this study periodontitis rabbit models under stimulated hypoxia environment wereestablished, then the bacterial types, quantity and endotoxin levels in subgingival plaques inrabbit models were investigated by isolating, culturing, microscoping of anaerobe andendotoxin detection etc, and the correlations between the pH value of GCF, viable count, endotoxin levels and clinical index were analyzed. Moreover, the detection rate and theendotoxin level of Porphyromonas gingivalis(Pg), one of the chief periodontal pathogenicbacteria, in subgingival plaques were detected, in order to enrich the fundament ofperiodontal microbiologic under hypoxia environment.Materials and Methods:1. Forty male rabbits were divided randomly into four groups：the normoxia controlgroup, the normoxia ligated group, the hypoxia control group and the hypoxia ligated group.Each group included ten rabbits. The ligated groups were combined with the methods of bligating both of the central incisors of mandible by steel ligature and being raised usingperiodontitis diets to set up periodontitis models. The hypoxia groups were raised underhypoxia environment (simulating the altitude5000meters,23h per day), while thenormoxia groups were raised under the normoxia environment (Chongqing, the averagealtitude400meters).2. After8weeks, the clinical indexes were detected, including attachment loss (AL),plaque index (PLI) and gingival index (GI). The pH value of gingival crevicular fluid (GCF)was tested by pH test strips. The rabbit periodontitis models were evaluated throughobserving radiographic manifestations and histopathologic slides under a light microscope.3. Subgingival plaques of both of the central incisors of mandible were collected forisolating, culturing, microscoping of anaerobe and endotoxin detection etc. and thecorrelations between the pH value of GCF, viable count, endotoxin level and clinical indexwere analyzed.4. The periodontal anaerobe was collected for isolating, culturing, identifying anddetecting endotoxin of Pg.Results:Ⅰ. Experimental results of animal models1. Histopathological observationIn the normoxia control group and the hypoxia control group, there were nopathological change of epithelia of gingival crevice, the periodontal ligament fibers andcells arranged in order, no alveolar bone destruction and resorption. But in the normoxialigated group, there was erosion of epithelia of gingival crevice, infiltrated manyinflammatory cells, the periodontal membrane space widened and alveolar bone destructed. Meanwhile, in the hypoxia ligated group, it was observed that the erosion of epithelia ofgingival crevice, infiltrated a large number of inflammatory cells, such as neutrophils andlymphocytes, the periodontal membrane space widened, the periodontal ligament fibersdestroyed and the alveolar ridge reduced.2. X-ray examinationIn the normoxia control group and the hypoxia control group, there were no apparentchange of the periodontal membrane space and alveolar ridge crest, while in the normoxialigated group and the hypoxia ligated group, there were widened periodontal membranespace and serious bone resorption, especially in the latter group.3. Clinical parametersAL, PLI and GI of the hypoxia ligated group were higher than other groups,significantly different from the normoxia ligated group（P<0.01）.Ⅱ. Experimental results of isolating and culturing of periodontal anaerobe1. After Gram staining, it was observed microscopically that there were single bacteriaspecies in the periodontal anaerobe of rabbit models in the normoxia groups, mainlyincluding Gram-negative bacilli and cocci, while there were abundant bacteria species inthat of the hypoxia groups, mainly including Gram-negative （G-） bacilli, cocci,Gram-positive（G+）bacilli and cocci.2. The pH value of GCF, viable count and endotoxin level in the hypoxia ligated groupwere higher than other groups, significantly different from the normoxia ligated group（P<0.05）.3. The pH value of GCF and viable count were positively correlated with AL, PLI andGI（P<0.01）. Endotoxin level was positively correlated with AL（P<0.01）and have nocorrelation with PLI and GI（P>0.05）.Ⅲ. Experimental results of isolating and culturing Pg1. By being purified and cultured on CDC blood plate after five days, there wasformed diameter of1-2mm bacteria colonies, which were brown or black, convex shape,smooth surface and edge, metallic luster and complete hemolysis. Microscopic morphologyof pure culture of Pg was G-bacilli or coccibacilli, uneven dyeing.2. Pg was successfully isolated in each group except for the normoxia control group.The detection rate and the endotoxin levels of Pg in the hypoxia ligated group were significantly different from the normoxia ligated group（P<0.05）.Conclusion:1. The periodontitis rabbit models exposed to hypoxia and normoxia environment wereestablished successfully, which was confirmed by that the hypoxia ligated group wassignificantly different from the normoxia ligated group in the periodontal clinicalmanifestation, clinical indexes and periodontal flora. Moreover, the results suggest that theprocess speed and damage degree of periodontitis under hypoxia environment are muchhigher than those changes under normoxia environment.2. The hypoxia environment was more preferred to the settlement of G-anaerobe,which led to the increasing of oxygen consumption and the decreasing of redox potential,thus the pH value was increased in periodontal pockets.3. It was more suitable for Pg colonizing in subgingival plaques under hypoxiaenvironment, and maybe under hypoxia environment more endotoxin were released by Pg,representing more powerful virulence. |
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