复发性阿弗他溃疡患者唾液中表皮生长因子和病损区表皮生长因子受体的定量研究

目的探讨复发性阿弗他溃疡（RAU）患者溃疡期和间隙期唾液表皮生长因子（EGF）质量浓度的变化以及病损区表皮生长因子受体（EGFR）的表达有无缺陷。方法选择27例在溃疡期和间隙期均成功获取唾液样本的轻型RAU患者作为RAU1组,采集33例正常人的唾液样本作为对照1组,采用酶联免疫吸附法检测两组唾液样本中EGF质量浓度。另外选择31例轻型RAU溃疡期患者在溃疡基底部剪取小块组织样本作为RAU2组,采集35例无RAU者的正常黏膜组织作为对照2组,采用荧光定量逆转录聚合酶链反应检测两组组织样本内EGFR的RNA表达情况。结果RAU1组溃疡期患者唾液EGF质量浓度高于RAU1组间隙期和对照1组,而间隙期EGF质量浓度低于对照1组（P<0.05）。RAU1组和对照1组唾液EGF质量浓度在不同性别间差别不大,与年龄也没有相关性（P>0.05）。RAU2组EGFR的RNA表达强度高于对照2组,两组间有统计学差异（P<0.05）。结论RAU患者口腔溃疡的复发性可能与间隙期唾液EGF质量浓度减少有关;口腔溃疡的自愈性可能与溃疡期唾液EGF质量浓度增加和溃疡区EGFR表达增加有关。     

Secretion of epidermal growth factor in parotid saliva in diabetic patients: role of autonomic innervation

Parotid saliva was collected over a 12-min period from 24 insulin dependent diabetic patients with varying degrees of autonomic neuropathy and 12 age and sex matched non-diabetic controls. Epidermal growth factor (EGF) concentrations in saliva were measured by radio-immunoassay. The EGF concentrations in diabetics with no autonomic neuropathy or with combined autonomic neuropathy were equivalent but secretion of EGF was significantly elevated at the 6- and 12-min periods of collection in diabetic patients with early or established autonomic neuropathy. It is postulated that when parasympathetic autonomic neuropathy is present a relative "over-activity" of the sympathetic innervation promotes release of salivary EGF. This sympathetic predominance may maintain salivary EGF concentration despite the elevated salivary flow and volume which is associated with parasympathetic autonomic neuropathy.     

Increased adhesion of peripheral blood neutrophils from patients with localized juvenile periodontitis

Adhesion of peripheral blood neutrophils from 5 patients with localized juvenile periodontitis (LJP) and age- and gender-matched healthy controls was measured using a semi-automated 96-well microtiter plate assay method. Both unstimulated and formyl-methionyl-leucyl-phenylalanine (FMLP, 10-1000 nM)-stimulated neutrophils from LJP patients showed in general higher adhesion than did their controls. After 15-60 min incubation with 100 and 1000 nM FMLP the numbers of adherent cells were significantly (p<0.05), 2.1-2.6-fold higher in LJP patients than in controls. Neutrophils from these LJP patients showed also enhanced respiratory burst activity in response to unopsonized zymosan stimulation. To test whether a decrease in intracellular diacylglycerol (DAG) kinase activity could account for the increased neutrophil adhesion of LJP patients normal neutrophils were treated with R59949 (10 μm), a DAG-kinase inhibitor. Both unstimulated and FMLP-stimulated normal neutrophils showed significantly (p<0.05) enhanced adhesion after R59949-treatment. Taken together, our data indicate that neutrophils from the 5 LJP patients investigated here exhibit 2 parallel hyperactivities, namely increased adhesion and enhanced production of reactive oxygen species. Furthermore, our present and previous (Hurttia et al., J Periodont Res 1997; 32 : 401-407) results suggest that the observed neutrophil functional abnormalities in some LJP patients may be associated with decreased cellular DAG-kinase activity. It is proposed that the hyperadherent and -active neutrophils may promote the development of LJP by causing tissue damage in the periodontium.     

Increased adhesion of peripheral blood neutrophils from patients with localized juvenile periodontitis

Adhesion of peripheral blood neutrophils from 5 patients with localized juvenile periodontitis (LJP) and age- and gender-matched healthy controls was measured using a semi-automated 96-well microtiter plate assay method. Both unstimulated and formyl-methionyl-leucyl-phenylalanine (FMLP, 10–1000 nM)-stimulated neutrophils from LJP patients showed in general higher adhesion than did their controls. After 15–60 min incubation with 100 and 1000 nM FMLP the numbers of adherent cells were significantly (p<0.05), 2.1–2.6-fold higher in LJP patients than in controls. Neutrophils from these LJP patients showed also enhanced respiratory burst activity in response to unopsonized zymosan stimulation. To test whether a decrease in intracellular diacylglycerol (DAG) kinase activity could account for the increased neutrophil adhesion of LJP patients normal neutrophils were treated with R59949 (10 μM), a DAG-kinase inhibitor. Both unstimulated and FMLP-stimulated normal neutrophiis showed significantly (p<0.05) enhanced adhesion after R59949-treatment. Taken together, our data indicate that neutrophils from the 5 LJP patients investigated here exhibit 2 parallel hyperactivities, namely increased adhesion and enhanced production of reactive oxygen species. Furthermore, our present and previous (Hurttia et al, J Periodont Res 1997; 32: 401-407) results suggest that the observed neutrophil functional abnormalities in some LJP patients may be associated with decreased cellular DAG-kinase activity. It is proposed that the hyperadherent and -active neutrophils may promote the development of LJP by causing tissue damage in the periodontium.     

口腔扁平苔藓组织中表皮生长因子及其受体表达变化的研究

目的探讨表皮生长因子(EGF)、表皮生长因子受体(EGFR)及两种原癌基因c-fos、c-jun,在口腔扁平苔藓中的基因表达、蛋白含量变化及其生物学意义。方法选择新鲜组织标本31例,包括糜烂型扁平苔藓10例、网状型扁平苔藓12例和正常口腔黏膜9例,使用逆转录聚合酶链反应技术(RT-PCR)检测EGF、EGFR、c-fos和c-jun基因在不同组织中的表达;同时使用免疫组化方法比较这4种蛋白在不同组织中的定位和表达量的变化。结果糜烂型扁平苔藓EGFR基因的mRNA和蛋白含量分别为(55.9±23.1)%、(71.1±10.1)%,均明显高于网状型扁平苔藓及正常口腔黏膜组织(P<0.05)。糜烂型扁平苔藓中的c-fos和c-jun基因表达呈相同的变化规律,两种基因的mRNA含量均显著高于正常口腔黏膜。糜烂型扁平苔藓的C-fos、C-jun蛋白的阳性细胞率分别是正常黏膜的1.3和1.5倍,蛋白含量明显高于正常口腔黏膜(P<0.01)。结论糜烂型扁平苔藓的癌变率可能大于网状型扁平苔藓,其中EGF及其受体引导的信号传导通路可能起着十分重要的作用。     

Diacylglycerol in peripheral blood neutrophils from patients with localized juvenile periodontitis

Neutrophils from patients with localized juvenile periodontitis (LJP) show several functional abnormalities. Recently, it has become increasingly apparent that the reason for these changes lies in part at the post receptor level of cellular metabolism. In this study we have analyzed intracellular diacylglycerol (DAG), a second messenger and an endogenous activator of protein kinase C, in un-stimulated and agonist-stimulated neutrophils. from five LJP patients showing a chemotaxis defect and matched normal individuals. No difference was observed in the basal cellular DAG between the two groups. In neutrophils from LJP patients the DAG levels increased by 67% and 111% from the basal level following stimulation with N-formyl-methionyl-leucyl-phenylalanine (FMLP) and unopsonized zymosan particles, respectively, while in control cells the mean increases were 36% and 65%, respectively. Incubation with serum-opsonized zymosan particles produced an identical rise in DAG in both groups. These data indicate that the stimulation of receptors for FMLP and unopsonized zymosan may produce an enhanced accumulation of DAG in neutrophils from LJP patients. In addition to DAG mass analysis, we determined the effect of R59022, a DAG-kinase inhibitor, on zymosan-stimulated luminol-amplified chemiluminescence (CL) of neutrophils. In control cells R59022 significantly enhanced unopsonized zymosan induced CL, but it had no effect on cells from LJP patients, suggesting a possible change in the regulation of DAG-kinase in LJP.     

Detection of epidermal growth factor receptor in inflammatory periapical lesions

Epithelial cell proliferation is often observed in periapical lesions of endodontic origin. The mechanisms which stimulate the epithelial cell rests of Malassez to proliferate are not understood fully. Fifteen inflammatory periapical lesions (10 granulomas and five cysts) obtained from periapical surgery and six additional periapical lesions (four granulomas and two cysts) collected from extracted teeth were examined using immunohistochemical staining and ¹²⁵I-EGF (epidermal growth factor) binding assay to detect the presence of epidermal growth factor receptor. The results indicated that the periapical lesions without epithelial cell proliferation had a weak immunoperoxidase staining or low specific binding of i²⁵I-EGF. In contrast, the periapical lesions with epithelial cell proliferation and cyst formation exhibited a strong immunoperoxidase staining in the epithelial cells or high specific binding of ¹²⁵I-EGF.     

Prevalence of juvenile periodontitis in schoolchildren in Lagos, Nigeria

The aim of this survey was to study the prevalence of juvenile periodontitis in schoolchildren aged 12-19 yr in Lagos, Nigeria. 1001 children, 565 boys and 436 girls, took part in the study. Initially all the children were screened clinically at school according to the recommendations of CPITN. A mirror and WHO 621 pattern probe were used for this examination. Any child with two or more sextants or teeth with CPITN code 3 or one sextant code 4 was taken for a radiographic and full clinical examination. Only 19 children, 12 boys and seven girls, fulfilled these requirements. Of these 19 children, five boys and three girls showed radiographic evidence of bone loss as well as increased probing depths, and were diagnosed as having juvenile periodontitis. The other children showed no evidence of bone loss radiographically, despite maximum probing depths of 4-5 mm. The results indicate a prevalence of juvenile periodontitis of 0.8% in this Negro population.     

Prevalence of juvenile periodontitis in Chile

The aim of this survey was to study the prevalence of juvenile periodontitis in schoolchildren aged 15-19 years in Santiago, Chile. A random sample of 2500 schoolchildren (1318 male, 1182 female) that represented the full range of different socio-economic strata and ethnic groups seen in the population of Santiago, was used. Initially, the children were screened clinically at school by assessment of probing depths around the incisors and first molars with a WHO 621 pattern probe. Children with 2 or more teeth with 5.5 mm or deeper pockets were invited for a radiographic examination comprising bitewing radiographs of molars, and periapical radiographs of incisors. Any subject with 2 mm or more alveolar bone loss was invited for a full clinical and radiographic examination. After screening, 27 subjects had a tentative diagnosis of juvenile periodontitis. 4 of these refused radiographic examination and only accepted a thorough clinical examination. 23 subjects presented themselves for the radiographic and complete clinical examination. Of the 27 subjects selected for detailed examination, 8 subjects (7 female, and 1 male) were diagnosed as having juvenile periodontitis. There was an overall prevalence of juvenile periodontitis of 0.32% with 95% confidence, which gives a range of +/- 0.10%. When prevalence was assessed by socio-economic status, juvenile periodontitis was found more commonly in low socio-economic group. The results of the current study suggest that in Chile, there might be a relationship between socio-economic status and prevalence of juvenile periodontitis, and that this disease is more frequent in women.     

The predominant cultivable organisms in juvenile periodontitis

abstract — Quantitative and qualitative examinations of the predominant cultivable microflora of the advanced tissue destruction (pocket) associated with juvenile periodontitis and of control pockets without loss of attachment were carried out by using anaerobic tube culture technique. The microflora from the control sites consisted primarily of streptococci, Gram-negative rods, and Gram-positive rods. In contrast, the test pockets had a relatively stable and characteristic microflora, dominated by various Gram-negative, anaerobic rods. The Gram-negative rods were divided into main groups, but a satisfactory classification of many of these microorganisms could not be performed with the methods used and the data available. The predominating Gram-negative rods differed from individual to individual. The role of the Gram-negative rods in the etiology of juvenile periodontitis is unknown. Some potential pathogenic mechanisms of these microorganisms are discussed.     

Differential in vitro response to epidermal growth factor by prenatal murine cranial-base chondrocytes

The retrognathic Brachyrrhine (Br) heterozygote mouse mutant has a very localized morphological deficiency in the sphenoethmoidal region of the anterior cranial base. The purpose of this study was to test the hypothesis that a primary growth defect occurs in that region of Br mice. Primary cell cultures were derived from presumptive nasal septal and sphenoethmoidal regions of Br and wild-type littermates. Cultures were stimulated with 1.0 ng/ml epidermal growth factor (EGF), and [3H]thymidine and [35S] incorporation was measured. Growth of the nasal septal chondrocytes did not differ significantly between groups. In the cultures derived from the sphenoethmoidal region [35S] incorporation was greater, but not significantly so, in the normal group. However, EGF did significantly stimulate proliferation of the sphenoethmoidal chondrocytes in wild-type cultures above that measured in Br cultures. Therefore, the Br genetic aberration is associated with a primary growth defect in the sphenoethmoidal region of the cranial base. These results suggest that growth of the anterior cranial base occurs differentially and that the defect in Br mice results in reduced sphenoidal but not nasal septal growth.     

Cell-mediated immunity in juvenile periodontitis and levamisole treatment

abstract — Immunostimulation with levamisole was attempted in eight patients with juvenile periodontitis and six reference patients with gingivitis but without loss of periodontal attachment. The following parameters were studied before and after levamisole treatment: gingival status, the concentration of serum immunoglobulins and complement, T and B lymphocyte ratio, leukocyte migration inhibition and lymphocyte transformation responses by dental plaque bacteria, PPD or PHA, and lymphocyte ATP-ase activity. In juvenile periodontitis cell-mediated immunity to dental plaque antigens seemed to be impaired, but the response was not restored by treatment with levamisole. There was no evidence of a broader suppression of cell-mediated immunity in juvenile periodontitis and there was no significant clinical effect of levamisole treatment. It is suggested that the apparent suppression of in vitro cell-mediated immunity to dental plaque bacteria in juvenile periodontitis is a secondary change, caused by a long-standing chronic infection.     

Increased luminol enhanced chemi-luminescence from peripheral granulocytes in juvenile periodontitis

Abstract – The granulocyte function in patients with juvenile periodontitis (JP) (eight men and six women, aged 13–33 yr) and sex and age matched controls was determined by the luminol enhanced chemiluminescence (CL) induced by opsonized and unopsonized bacteria or latex beads. Maximal CL induced by latex beads with or without autologus serum did not differ between the two groups. However, bacteria with or without autologous serum seemed to induce a higher maximal CL in the JP group. The difference was statistically significant for the results obtained with opsonized bacteria. Serum opsonized bacteria induced a much higher maximal CL than the unopsonized. The opsonin dependent reaction indicates a close association with the process of phagocytosis. The increased CL of granulocytes in JP could indicate the formation of extracellularly free oxygen radicals with the potential to damage tissue. These observations support a possible involvement of the granulocytes in the pathogenesis of JP.     

Determination of salivary levels of mucin and amylase in chronic periodontitis patients

Sánchez GA, Miozza V, Delgado A, Busch L. Determination of salivary levels of mucin and amylase in chronic periodontitis patients. J Periodont Res 2011; 46: 221–227. © 2010 John Wiley & Sons A/S Background and Objective: Patients with periodontal disease show differences in the profile of proteins in whole saliva. This profile reflects the nature and amplitude of the host response to a periodontal microbial challenge. Since periodontitis is a chronic inflammatory disease with different progression stages, the aim of the study was to evaluate the host response in these different clinical stages by assessing salivary flow rate, the concentrations of proteins and mucin and the amylase activity. Material and Methods: Sixty adult subjects were clinically examined and distributed into four groups (n = 15) according to the periodontal status, namely, healthy, mild, moderate and severe periodontitis. Whole saliva was collected for 5 min, followed by a second 5 min sampling period with stimulation by chewing a paraffin block, and flow rate was determined. Salivary proteins, amylase and mucin were determined by colorimetric methods. Results: The concentrations of proteins, amylase and mucin increased in subjects with moderate and severe periodontal disease in unstimulated saliva, while flow rate decreased. A positive correlation was found between proteins and amylase or mucin concentrations among the different groups, indicating that the concentrations changed in the same way, being the response of salivary glands to the disease, possibly to enhance the protective potential of saliva. Mucin concentration was lower in the mild periodontitis group. Mechanical stimulation induced an increase in flow rate and output of proteins, amylase and mucin. Conclusion: Periodontitis induces an increase in the output of proteins, including mucin and amylase, thereby enhancing the protective potential of saliva, but this is accompanied by a decrease in flow rate.     

碱性成纤维细胞生长因子对牙周膜细胞表皮生长因子受体基因表达的影响

目的研究碱性成纤维细胞生长因子（bFGF）对人牙周膜细胞（PDLC）表达表皮生长因子受体（EGFR）的影响,探讨bFGF在牙周组织分化再生中的意义。方法体外原代培养人PDLC,有限稀释法形成单细胞克隆,用外源性bFGF刺激单细胞克隆,采用逆转录聚合酶链反应（RT-PCR）检测克隆细胞内EGFR基因表达的变化。结果bFGF促进人PDLC内EGFR mRNA的合成,并且随着质量浓度的增加促进作用增强。结论bFGF对EGFR的促进作用很可能是牙周炎损伤修复过程中一个重要的调节因素,为牙周组织分化再生提供部分理论基础。