Effect of oral bacteria on peripheral blood leukocyte interleukin-6 and soluble interleukin-6 receptor production

To determine the effect of pathogenic oral bacteria on interleukin 6 (IL-6) and soluble IL-6 receptor production, we measured their release by human peripheral blood mononuclear cells in vitro. Unseparated peripheral blood mononuclear cells, peripheral blood lymphocytes (monocyte depleted), pure T cells, or monocytes were cultured with Actinobacillus actinomycetemcomitans, Capnocytophaga gingivalis, Capnocytophaga ochracea, Fusobacterium nucleatum or Porphyromonas gingivalis for 24 h. Supernatants were tested for IL-6 and soluble IL-6 receptor by enzyme-linked immunosorbent assay. Only monocytes and peripheral blood mononuclear cells responded with significant IL-6 release in the presence of all bacteria tested. However, peripheral blood lymphocytes were capable of producing IL-6 when activated by phytohemagglutinin or IL-2 followed by bacteria, though substantially less than cultures containing monocytes. No bacteria tested increased soluble IL-6 receptor release over spontaneous soluble IL-6 receptor release. We conclude that monocytes release IL-6 after contact with oral pathogens; however, soluble IL-6 receptor from T cells and monocytes is constitutively produced and may modulate EL-6 actions.     

IL—1ra对IL—1β诱导人牙龈成纤维细胞分泌IL—6的拮抗作用

目的：研究白细胞介素－1受体拮抗剂对IL－1β所介导生物学功能的影响，为进一步临床应用提供理论依据。方法：采用细胞培养技术和ELISA夹心法。结果：经IL－1β单独作用，牙龈成纤维细胞培养上清液中IL－6含量明显增高，当一定浓度的IL－1ra与1μg/L IL-1β共同作用人牙龈成纤维细胞后，培养上清液中IL－6的含量比单独IL－1β作用组低。结论：IL－1ra能够抑制IL－1诱导的人牙龈成纤维细胞分泌IL－6。     

IL-lra对IL-1β诱导人牙龈成纤维细胞分泌IL-6的拮抗作用

目的:研究白细胞介素-1受体拮抗剂对IL-1β所介导生物学功能的影响,为进一步临床应用提供理论依据.方法:采用细胞培养技术和ELISA夹心法.结果:经IL-1β单独作用,牙龈成纤维细胞培养上清液中IL-6含量明显增高,当一定浓度的IL-lra与1ug/L IL-1β共同作用人牙龈成纤维细胞后,培养上清液中IL-6的含量比单独IL-1β作用组低.结论:IL-lra能够抑制IL-l诱导的人牙龈成纤维细胞分泌IL-6.     

Interleukin-2 stimulates osteoclastic activity; Increased acid production and radioactive calcium release

Recombinant human interleukin-2 (IL-2) was studied to determine effects on acid production by individual osteoclasts in situ on mouse calvarial bones. This analysis was performed using a microspectrofluorimetric technique to quantify acid production in individual cells. Radioactive calcium release was determined using calvarial bones in a standard tissue culture system. This allowed us to correlate changes in acid production with a measure of bone resorption. IL-2 stimulated acid production and bone resorbing activity. Both effects were inhibited by calcitonin. No stimulation of bone resorption occurred when IL-2-containing test media was incubated with a specific anti-IL-2 antibody and ultrafiltered. Our data demonstrated a correlation between acid production and bone resorbing activity in mouse calvaria exposed to parathyroid hormone (PTH). The data obtained from cultured mouse calvaria exposed to IL-2 demonstrated similar stimulatory effects to those seen during PTH exposure. These data suggest that calvaria exposed to IL-2 in vitro have increased osteoclastic acid production corresponding with increased bone resorption.     

Role of soluble interleukin-6 receptor in inflamed gingiva for binding of interleukin-6 to gingival fibroblasts

Interleukin-6 (IL-6), frequently detected in periodontitis, is known to mediate important signals in the inflammatory cytokine network. Gingival fibroblasts (GF) secrete cytokines upon stimulation with inflammatory mediators. However, it is not clear if GF respond to IL-6. We examined the IL-6 receptor gene expression in GF. Furthermore, we tested whether GF are target cells for IL-6 by examination of binding of IL-6. GF were found to contain trace amounts of mRNA for IL-6 receptor (IL-6R), but had high levels of mRNA for 130-kDa glycoprotein (gp130), which is a signal transducer for IL-6/IL-6R complex. Based on this observation, we hypothesized that IL-6 could bind GF if exogenous soluble forms of IL-6R (sIL-6R) existed in the gingiva or culture condition. Thus, we investigated the existence of sIL-6R in gingiva using enzyme-linked immunosorbent assay and whether sIL-6R influenced the binding of IL-6 to GF in vitro. In inflamed gingiva, sIL-6R was detected and its concentration ranged from 150 to 700 pg/microgram protein. The sIL-6R enhanced the binding of IL-6 to GF in a dose-dependent manner. This enhancement was inhibited by an antibody against gp130, suggesting that the IL-6/sIL-6R complex bound to the fibroblasts via gp130. These data demonstrated that gingival fibroblasts can be target cells for IL-6 in the presence of appropriate amounts of sIL-6R. This situation may exist during inflammation in periodontal tissue.     

Analysis of tumor necrosis factor-alpha, interleukin-6, interleukin-1beta, soluble tumor necrosis factor receptors I and II, interleukin-6 soluble receptor, interleukin-1 soluble receptor type II, interleukin-1 receptor antagonist, and protein in the synovial fluid of patients with temporomandibular joint disorders

OBJECTIVE: To measure the levels of various cytokines, cytokine receptors, and cytokine antagonists in the synovial fluid of patients with temporomandibular disorders (TMD) and to determine the correlations among these expression levels. STUDY DESIGN: Synovial fluid was obtained from 55 patients with TMD and from 5 asymptomatic healthy volunteers as controls. The concentrations of tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, IL-1beta, soluble tumor necrosis factor receptors I and II (sTNFR-I and sTNFR-II), IL-6 soluble receptor (IL-6sR), IL-1 soluble receptor type II, and IL-1 receptor antagonist were determined by enzyme-linked immunosorbent assay. RESULTS: The concentrations of TNF-alpha, IL-6, IL-1beta, sTNFR-I, and sTNFR-II were significantly higher in the synovial fluid of patients than in controls (P < .05). TNF-alpha level was positively correlated with those of IL-6, sTNFR-I, and sTNFR-II. In particular, there was a highly significant positive correlation between sTNFR-I and sTNFR-II. CONCLUSION: TNF and sTNFRs in the synovial fluid of patients with TMD may be important in the pathogenesis of these disorders.     

Clodronate inhibits PGE(2) production in compressed periodontal ligament cells

Periodontal ligament (PDL) cells play an essential role in orthodontic tooth movement. We recently reported that clodronate, a non-N-containing bisphosphonate, strongly inhibited tooth movement in rats, and thus could be a useful adjunct for orthodontic treatment. However, it is not clear how clodronate affects the responses of PDL cells to orthodontic force. In this study, we hypothesized that clodronate prevents the mechanical stress-induced production of prostaglandin E(2) (PGE(2)), interleukin-1beta (IL-1beta), and nitric oxide (NO) in human PDL cells. A compressive stimulus caused a striking increase in PGE(2) production, while the responses of IL-1beta and NO were less marked. Clodronate concentration-dependently inhibited the stress-induced production of PGE(2). Clodronate also strongly inhibited stress-induced gene expression for COX-2 and RANKL. These results suggest that the inhibitory effects of clodronate on tooth movement and osteoclasts may be due, at least in part, to the inhibition of COX-2-dependent PGE(2) production and RANKL expression in PDL cells.     

Interleukin-1beta, interleukin-1 receptor antagonist, and interleukin-1 soluble receptor II in temporomandibular joint synovial fluid from patients with chronic polyarthritides.

PURPOSE: The aim of this study was to investigate whether interleukin-1beta (IL-1beta), interleukin-1 receptor antagonist (IL-1ra), or soluble IL-1 receptor II (sIL-1RII) in synovial fluid or plasma is associated with joint pain or signs of tissue destruction in patients with temporomandibular joint (TMJ) involvement of polyarthritides. PATIENTS AND METHODS: Forty-three patients with TMJ involvement of polyarthritides were included. TMJ resting pain, tenderness to palpation, pressure pain threshold, pain on mandibular movement, and anterior open bite were assessed. TMJ synovial fluid samples and plasma were obtained for analysis of IL-1beta, IL-1ra, and sIL-1RII. RESULTS: IL-1beta was detected in 18% of the synovial fluid samples and in 44% of the plasma samples. The concentrations of IL-1ra in plasma were lower than in the synovial fluid, whereas the opposite condition was found for sIL-1-RII. IL-1ra in synovial fluid and plasma was associated with low intensity of TMJ pain. sIL-1RII in synovial fluid was associated with low degree of anterior open bite, whereas sIL-1RII in plasma was associated with widespread musculoskeletal pain, TMJ pain and tenderness, and decreased pressure pain threshold over the TMJ. CONCLUSION: IL-1ra and sIL-1RII are present in different proportions in TMJ synovial fluid and blood plasma from patients with TMJ involvement of polyarthritis. Both of these molecules seem to influence the clinical features of these forms of TMJ inflammation.     

Interleukin-2, interleukin-2 receptor and interleukin-4 levels are elevated in the sera of patients with periodontal disease

Three serological markers of immune cell activation, interleukin-2 (IL-2), soluble interleukin-2 receptor (sIL-2R), and interleukin-4 (IL-4), were measured by enzyme-linked immunosorbent assays in 20 control subjects and 26 periodontitis patients. The experimental group comprised 19 juvenile/post-juvenile and 7 severe generalized periodontitis patients with radiographic evidence of alveolar bone loss. Although some control sera contained immunoreactive IL-2 (2 of 20) and IL-4 (3 of 20), all contained sIL-2R, the levels of which correlated well with age (r = 0.644). Mean levels of all three markers were significantly elevated in the sera of patients with periodontal disease compared to control values. However, there was a wide variation in the amounts detected; IL-2 (0.21-173.33 ng/ml); sIL-2R (217.95-1177.27 units/ml); IL-4 (3.17-16.35 pg/ml), which did not correlate with either the degree of bone loss or pocket formation observed clinically. Moreover, there was no correlation between the levels of IL-2, sIL-2R or IL-4 for any given individual in the experimental group. The finding that only 2 of the control sera contained IL-2 (10%) compared to 23 of the periodontitis patients (88.5%) suggests that, of the three parameters investigated, the measurement of IL-2 could provide a sensitive laboratory test for assessing periodontal disease activity. Nevertheless, a definitive study to determine the relationship of serum IL-2 levels to clinical parameters of disease activity will be necessary to confirm this observation.(ABSTRACT TRUNCATED AT 250 WORDS)     

头颈部恶性肿瘤患者血清可溶性白介素2受体水平

目的:研究头颈部恶性肿瘤患者血清可溶性白介素2受体(sIL-2R)水平的变化。方法:用双抗体夹心法对60例头颈部恶性肿瘤患者,60例良性肿瘤患者以及50例健康对照者的血清sIL-2R水平进行检测。结果:头颈部恶性肿瘤患者组血清sIL-2R水平显著高于健康对照组(P<0.01),而良性肿瘤患者组血清sIL-2R水平与健康对照组无统计学差异(P>0.05)。结论:血清中高sIL-2R水平可能与头颈部恶性肿瘤的发生有相关性。     

Antigenicity of primary endodontic infection against macrophages by the levels of PGE(2) production

Introduction

Root canal contents are potent stimuli for proinflammatory cytokines involved in apical periodontitis. This study investigated target gram-negative bacterial species and endotoxins in primary endodontic infection with apical periodontitis, determined their antigenicity against macrophages through the levels of PGE₂, and evaluated their relationship with clinical findings.

Methods

Samples were taken from 21 root canals with primary infection and apical periodontitis by using paper points. Polymerase chain reaction (16S rDNA) was used for bacterial detection and limulus amebocyte lysate assay for endotoxin measurement. Levels of prostaglandin E2 (PGE₂) were measured by enzyme-linked immunosorbent assay (Duoset Kit; R&D, Minneapolis, MN).

Results

Prevotella nigrescens (13/21), Fusobacterium nucleatum (6/21), and Porphyromonas endodontalis (6/21) were the most frequently observed species. A positive association was found between F. nucleatum and P. endodontalis (P < .05). A correlation was found between the number of gram-negative bacterial species and the levels of endotoxins, as well as PGE₂ (P < .05). Higher levels of endotoxin were detected in teeth with exudation, whereas elevated levels of PGE₂ were found in teeth with tenderness to percussion and pain on palpation.

Conclusions

Our findings imply an additive effect between the number of gram-negative bacterial species involved in endodontic infection regarding the induction of proinflammatory cytokine by macrophage cells. Moreover, teeth with clinical symptomatology were related to higher levels of endotoxins and PGE₂ secretion.     

Mangiferin inhibits lipopolysaccharide-induced production of interleukin-6 in human oral epithelial cells by suppressing toll-like receptor signaling

ObjectiveOral epithelial cells have currently been found to play an important role in inflammatory modulation in periodontitis. Mangiferin is a natural glucosylxanthone with anti-inflammatory activity. The aim of this study was to investigate the regulatory effect of mangiferin on lipopolysaccharide (LPS)-induced production of proinflammatory cytokine interleukin-6 (IL-6) in oral epithelial cells and the underlying mechanisms.DesignThe levels of LPS-induced IL-6 production in OKF6/TERT-2 oral keratinocytes were detected using enzyme-linked immunosorbent assay (ELISA). The expression of Toll-like receptor (TLR) 2 and TLR4 was determined using western blot analysis. And the phosphorylation of TLR downstream nuclear factor-κB (NF-κB), p38 mitogen-activated protein kinase (p38 MAPK) and c-Jun N-terminal kinase (JNK) was examined using cell-based protein phosphorylation ELISA kits.ResultsWe found that mangiferin reduced LPS-upregulated IL-6 production in OKF6/TERT-2 cells. Additionally, mangiferin inhibited LPS-induced TLR2 and TLR4 overexpression, and suppressed the phosphorylation of NF-κB, p38 MAPK and JNK. Moreover, mangiferin repressed IL-6 production and TLR signaling activation in a dose-dependent manner after 24 h treatment.ConclusionsMangiferin decreases LPS-induced production of IL-6 in human oral epithelial cells by suppressing TLR signaling, and this glucosylxanthone may have potential for the treatment of periodontitis.     

压力对髁突软骨细胞白介素4、6分泌的影响

目的:探讨下颌骨髁突软骨细胞白细胞介素4和白细胞介素6的表达随机械力的变化过程。方法:体外培养下颌骨髁突软骨细胞,采用可控液压细胞加载装置90kPa压强下分别加载60、360min,免疫组化染色观察加压前后白细胞介素4和白细胞介素6的表达、分布变化。结果:正常对照的MCC中IL-4和IL-6只有很弱的表达,90kPa加压60min后IL-6在胞浆、胞核中表达强阳性,IL-4也在胞浆、胞核中阳性表达;加压时间延长至360min后IL-6表达较60min时有所减弱,而IL-4的表达比60min时持续增强。结论:适宜的压力刺激可引起髁突软骨细胞中白介素4、6表达的增强。     

Interleukin-1 and interleukin-1 inhibitor production by human adherent cells stimulated with periodontopathic bacteria

This study examined the effect of the putative periodontopathic bacteria Bacteroides gingivalis and Fusobacterium nucleatum on the production of interleukin-1 (IL-1) and IL-1 inhibitors by human plastic-adherent mononuclear cells from normal donors. Fusobacterium mortiferum was used as a non-oral, non-pathogenic control organism. Unstimulated adherent cells spontaneously secreted an IL-1 inhibitor, whereas stimulation with B. gingivalis induced the synthesis and secretion of IL-1. With both fusobacteria IL-1 was present in the intracellular environment, whereas the predominant secretory product was either IL-1 or an IL-1 inhibitor. These results suggest that bacteria are capable of modulating cytokine production by monocytes and may thereby alter the local immune response.     

Prostaglandin F2alpha upregulates interleukin-6 production in human gingival fibroblasts

Prostaglandin F2alpha (PGF2alpha) is a bioactive lipid mediator which has been suggested to be involved in the pathogenesis of periodontal disease. However, the roles of PGF2alpha in periodontal lesions are poorly understood. In the present study, we investigated the effect of PGF2alpha on interleukin (IL)-6 production in human gingival fibroblasts (HGF). PGF2alpha stimulated IL-6 production in a time- and concentration-dependent fashion. IL-1beta and tumor necrosis factor alpha (TNFalpha), proinflammatory cytokines, induced IL-6 production in a time-dependent manner, and PGF2alpha synergistically enhanced IL-6 production induced by IL-1beta and TNFalpha. IL-6 mRNA was expressed in PGF2alpha-stimulated HGF, and PGF2alpha increased IL-6 mRNA levels induced by IL-1beta and TNFalpha. Fluprostenol, a selective FP receptor agonist, could mimic PGF2alpha-induced IL-6 production. Since FP receptors are coupled to elevation of intracellular calcium and activation of protein kinase C (PKC), the mechanism of IL-6 production by PGF2alpha was investigated using TMB-8, an inhibitor of Ca2+ mobilization from intracellular stores, and calphostin C, an inhibitor of PKC. TMB-8 significantly suppressed PGF2alpha-induced IL-6 production, whereas calphostin C showed a stimulatory effect on PGF2alpha-induced IL-6 production. From these data, we suggest that PGF2alpha upregulates IL-6 production through FP receptors in HGF, that PGF2alpha synergistically enhances IL-6 production in IL-1beta- and TNFalpha-stimulated HGF, and that PGF2alpha-induced IL-6 production may be dependent on intracellular Ca2+ mobilization and be downregulated by PKC activation. PGF2alpha may be involved in the pathogenesis of periodontal disease by enhancing IL-6 levels in periodontal lesions.