Diabetes modulates gene expression in the gingival tissues of patients with chronic periodontitis

AIM: This study evaluated whether diabetes modulates gene expression [interleukin (IL)-1beta, IL-1ra, IL-6, IL-8, IL-10; tumor necrosis factor (TNF)-alpha; interferon (IFN)-gamma, receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG)] in sites with periodontitis. MATERIALS AND METHODS: Gingival biopsies were harvested and divided into three groups--Control group: systemically and periodontally healthy subjects (n = 10); Periodontitis group: systemically healthy subjects diagnosed with chronic periodontitis (n = 20); Diabetes group: type 1 diabetic subjects, diagnosed with chronic periodontitis (n = 20). Total RNA was obtained and analyzed by quantitative polymerase chain reaction. RESULTS: Data analysis demonstrated that, except for OPG, mRNA levels for all factors were increased by inflammation (P < 0.001). Interleukin-1beta, IL-1ra, IL-6, IL-8, IFN-gamma, and RANKL mRNA levels were higher in the diabetic group when compared with the control non-periodontitis group (P < 0.05), whereas IL-10 and OPG were lower (P < 0.05). No difference was observed for TNF-alpha between diabetic and control groups (P > 0.05). Diabetes lowered IL-1beta, IL-8, IL-10, TNF-alpha, RANKL, and OPG mRNA levels in sites with comparable type of periodontitis (P < 0.001). Moreover, increased RANKL:OPG and IL-6:IL-10 ratios were found. CONCLUSION AND CLINICAL RELEVANCE: Taken together, these data suggest that decreased levels of IL-10 and OPG may play an important role in the periodontal breakdown in diabetic patients.     

Comprehensive gene expression analysis in human periodontal ligaments of the mandibular third molars performing vertical movement and the maxillary second premolars with occlusal contact

OBJECTIVES: The periodontal ligament (PDL) is thought to be an important tissue in vertical movement during tooth eruption, but the precise molecular mechanism is not known. Thereto, comprehensive gene expression was analyzed in human PDL of mandibular third molars performing vertical movement and maxillary second premolars with occlusal contact. DESIGN: The expression profile of 9,243 genes in the PDL of one subject was compared between vertically moving third molars and second premolars with occlusal contact by DNA microarray. RESULTS: The expression of 27 genes showed more than a 10-fold difference between third molars and second premolars. The expression of CALB1 (encoding calbindin 1), CYP26A1 (encoding cytochrome P450, family 26, subfamily A, polypeptide 1), SPOCK3 (encoding testican-3), CCK (encoding cholecystokinin) and SCRG1 (encoding scrapie responsive protein 1) was more than 30-fold higher in PDLs of the third molars than the second premolars. CALB1 is reported to increase at the pressure side of PDL during experimental orthodontic tooth movement in rats. Interestingly, in this study, CALB1 expression showed the largest difference. In contrast, CRCT1 (encoding cysteine-rich C-terminal 1), SPRP3 (encoding small proline-rich protein 3), IL8 (encoding interleukin 8) and MMP12 (encoding matrix metalloproteinase 12) showed more than 100-fold higher expression in PDLs of the second premolars than the third molars. CONCLUSION: The present comprehensive gene expression in PDLs provides new insights into the molecular mechanism during the vertical tooth movement.     

Gene expression signatures in chronic and aggressive periodontitis: a pilot study

This pilot study examined gene expression signatures in pathological gingival tissues of subjects with chronic or aggressive periodontitis, and explored whether new subclasses of periodontitis can be identified based on gene expression profiles. A total of 14 patients, seven with chronic and seven with aggressive periodontitis, were examined with respect to clinical periodontal status, composition of subgingival bacterial plaque assessed by checkerboard hybridizations, and levels of serum IgG antibodies to periodontal bacteria assayed by checkerboard immunoblotting. In addition, at least two pathological pockets/patient were biopsied, processed for RNA extraction, amplification and labeling, and used to study gene expression using Affymetrix U-133 A arrays. Based on a total of 35 microarrays, no significantly different gene expression profiles appeared to emerge between chronic and aggressive periodontitis. However, a de novo grouping of the 14 subjects into two fairly robust clusters was possible based on similarities in gene expression. These two groups had similar clinical periodontal status and subgingival bacterial profiles, but differed significantly with respect to serum IgG levels against the important periodontal pathogens Porphyromonas gingivalis, Tannerella forsythensis and Campylobacter rectus. These early data point to the usefulness of gene expression profiling techniques in the identification of subclasses of periodontitis with common pathobiology.     

Influence of gingival crevicular washing on the expression of polymorphonuclear leukocyte membrane receptors before and after periodontal therapy

Abstract. Extensive data demonstrate that polymorphonuclear leukocytes (PMN) are the predominant cell type involved in periodontal disease and that gingival crevicular fluid constituents are influenced by the inflamed gingiva. The aim of the present study was to evaluate the ability of gingival crevicular washing (GCW) (a dilution of gingival crevicular fluid) from periodontal sites in different clinical conditions of modulating the PMN membrane receptors involved in motility, adhesion and phagocytosis before and after periodontal treatment. 10 patients affected by adult periodontitis (AP) were selected. From each patient. 2 test sites (TS) were chosen on the basis of a probing depth > 5 mm and attachment loss, and 2 control sites (CS) with probing depth < 3 mm without attachment loss. Modifications of membrane receptor density of PMN from healthy donors incubated with GCW harvested from TS and CS was evaluated using fluorescent probes and flow cytometry. Compared to CS-GCW. TS-GCW before therapy increased the expression of the β2 integrin CD 11b and the chemotactic receptor for the oligopeptide N-formyl methionyl leucyl phenylalanine (FMLP-R) while it reduced the expression of L-selection. GCW collected from the same TS after the successful completion of periodontal treatment did not influence PMN receptors, indicating that the clinical improvement paralleled the disappearance of the PMN modulating capability contained in TS-GCW before therapy. In conclusion, the present data illustrate the relevant modifications occurring at PMN membrane in chronic adult periodontitis exerted by GCW obtained by a simple fluid collection technique. Thus, monitoring gingival crevicular fluid PMN activating capability may help disclose the presence of chronic periodontitis and may be useful in assessing successful treatment.     

Microarray analysis of nicotine-induced changes in gene expression in a macrophage-like human cell line

BACKGROUND AND OBJECTIVE: Cigarette smoking has been suggested as a risk factor for periodontitis. Thousands of components are present in cigarette smoke, including nicotine, which may play an important role in the observed effects of smoking on cell metabolism. However, the mechanisms underlying these effects are unclear. Using DNA microarrays, we monitored differentially expressed genes, responsive to nicotine, in a macrophage-like human cell line. MATERIAL AND METHODS: Human U937 cells were treated for 1 h, with or without 1.0 microg/ml of nicotine. For differentiation, cultures were incubated with 10 nm phorbol myristate acetate for 48 h. Analysis of gene expression was performed using a DNA microarray of 8500 genes. RESULTS: The expression of 4914 genes was detected. Screening was carried out on those genes whose expression in three separate experiments showed an average change of twofold or greater, and 118 up-regulated genes and 97 down-regulated genes were identified. Among these were genes related to inflammation and other immune responses, such as phospholipase A2 and interferon. Consistent with the array findings, we found similar changes in mRNA expression after analysis using the real-time polymerase chain reaction. CONCLUSION: The results suggest that nicotine causes excess inflammation and disturbs host defense mechanisms against pathogens.     

TLR2 Arg753Gly, TLR4 Asp299Gly and Thr399Ile gene polymorphisms are not associated with chronic periodontitis in a Turkish population

AIM: Toll-like receptor (TLR) gene polymorphisms could affect the host's ability to respond to microbial pathogens. In this case-control study, the association of TLR2 and TLR4 gene polymorphisms with chronic periodontitis (CP) was investigated. MATERIALS AND METHODS: Genomic DNA was obtained from the peripheral blood of 83 patients with CP and 106 periodontally healthy subjects. The TLR2 Arg753Gly, Arg677Trp and TLR4 Asp299Gly, Thr399Ile gene polymorphisms were genotyped by the polymerase chain reaction-restriction fragment length polymorphism method. The data were analysed by a chi2 test, logistic regression analysis and the Mann-Whitney U test. RESULTS: The 753Gln allele was found in 6.1% of the CP patients as compared with 6.6% in the control group (p>0.05). The frequency of the 299Gly and 399Ile allele was 2.4% and 1.8% in CP patients. For the healthy subjects, the frequency was 2.8% for the 299Gly and 2.5% for the 399Ile allele (p>0.05). None of the CP patients or healthy subjects showed homozygosity for the TLR2 and TLR4 alleles. Percentage of sites with bleeding on probing and plaque were significantly higher in 299Gly-positive patients compared with 299Gly-negative patients (p<0.05). CONCLUSION: These results showed that the TLR2 and TLR4 gene polymorphisms studied are not associated with susceptibility to CP in Turkish patients.     

口腔癌细胞诱导的淋巴管内皮细胞基因表达谱的变化

目的探讨在口腔癌细胞影响下淋巴管内皮细胞分子表型的变化。方法利用体外共培养模型,将淋巴管内皮细胞与口腔癌细胞进行共培养,模拟肿瘤中淋巴管内皮细胞的变化。应用Affymetrix U133 Plus 2.0基因芯片对肿瘤组织中淋巴管内皮细胞（TLEC）与淋巴管内皮细胞（LEC）基因表达的差异进行了检测和比较。并对功能相似的表达差异基因进行分类。结果差异基因表达谱共发现了677个基因表达差异在1倍以上,其中在TLEC中表达上调的基因有384条,下调的基因有293条。这些基因与细胞黏附、凋亡、运动、发育及血管生成有关。同时这些基因还参与细胞的信号传导、免疫应答、细胞代谢等过程。结论LEC与TLEC在分子水平是有差别的。以此为基础,可以针对淋巴管内皮细胞进行靶向阻断,达到治疗口腔癌淋巴道转移的目的。     

Longitudinal analysis of metalloproteinases, tissue inhibitors of metalloproteinases and clinical parameters in gingival crevicular fluid from periodontitis-affected patients

BACKGROUND: The aim of this work was to improve the assessment of the periodontal disease status through measurements of extracellular matrix metalloproteinases (MMPs) and their tissular inhibitors (TIMPs) in the gingival crevicular fluid from patients diagnosed with chronic periodontitis. METHODS: Gingival crevicular fluid samples from patients (n = 13) were taken from 60 sites initially, and from 51 and 41 sites, respectively, 3 and 6 months after scaling and root planing. Gingival crevicular fluid samples were also taken from healthy subjects (n = 11, 24 sites). The presence of MMP-9 and MMP-8 was assessed by zymography and immunowestern blotting, respectively. The actual MMP activity (gelatinase and collagenase) was measured using the fluorogenic substrate assay. TIMP-1 and -2 levels were measured by immunodot blot. RESULTS: The fluorogenic substrate assay determinations showed higher MMP activity in sites with probing depth > or = 4 mm, with significant reduction post-treatment. Gelatinase activity followed by zymography consisted mainly of MMP-9. A different pattern of MMP-8 in control and patient sites was found. Controls only showed species of a partially active form (69 kDa), whereas patient sites showed a high frequency of the active form (56 kDa), and in some cases the latent form (85 kDa) was also observed. The active form reduced its frequency in sites with probing depth > or = 4 mm. TIMP-1 and -2 levels in patients were significantly lower than in controls, and after treatment the recovery of TIMP-1 level similar to control was observed. CONCLUSION: Significant correlations between the severity of the periodontal disease and the actual MMP activity, the active form of MMP-8 and the low level of both TIMP-1 and TIMP-2 were found.