A single application of hydrogen sulphide induces a transient osteoclast differentiation with RANKL expression in the rat model

Objective

Oral malodor is mainly attributed to volatile sulphur compounds (VSCs) such as hydrogen sulphide (H₂S), methyl mercaptan and dimethyl sulphide. VSC accelerate periodontal soft tissue destruction. However, there is little information about the potential role of H₂S in alveolar bone loss. The purpose of this animal study was to examine the effects of sodium hydrogen sulphide (NaHS), H₂S donor drug, on osteoclast differentiation in rat periodontal tissue.

Design

Twenty-four male Wistar rats (8 weeks old) were divided into four groups: a control group and three experimental groups, which were examined at 3 h, 1 day, and 3 days after topical application of 3 μl NaHS (l M in physiological saline) into the gingival sulcus of rat first molar. Expression of tumour necrosis factor (TNF)-α, RANKL, NF-κB and tartrate-resistant acid phosphatase (TRAP) was evaluated in the periodontal tissue.

Results

Three hours after NaHS application, TNF-α expression increased in the periodontal ligament. The numbers of RANKL-positive osteoblasts and TRAP-positive osteoclasts significantly increased progressively with time and reached a maximum level after 1 day. Significant up-regulation of RANKL and NF-κB mRNA was observed at 3 h after NaHS application.

Conclusions

H₂S application caused a transient increase of osteoclast differentiation with up-regulation of RANKL expression in osteoblasts. H₂S, which is primarily responsible for halitosis, may also contribute to alveolar bone resorption through RANKL expression.     

Oral administration of vitamin C prevents alveolar bone resorption induced by high dietary cholesterol in rats

BACKGROUND: A high-cholesterol diet stimulates alveolar bone resorption, which may be induced via tissue oxidative damage. Vitamin C reduces tissue oxidative damage by neutralizing free radicals and scavenging hydroxyl radicals, and its antioxidant effect may offer the clinical benefit of preventing alveolar bone resorption in cases of hyperlipidemia. We examined whether vitamin C could suppress alveolar bone resorption in rats fed a high-cholesterol diet. METHODS: In this 12-week study, rats were divided into four groups: a control group (fed a regular diet) and three experimental groups (fed a high-cholesterol diet supplemented with 0, 1, or 2 g/l vitamin C). Vitamin C was provided by adding it to the drinking water. The bone mineral density of the alveolar bone was analyzed by microcomputerized tomography. As an index of tissue oxidative damage, the 8-hydroxydeoxyguanosine level in the periodontal tissue was determined using a competitive enzyme-linked immunosorbent assay. RESULTS: Hyperlipidemia, induced by a high-cholesterol diet, decreased rat alveolar bone density and increased the number of tartrate-resistant acid phosphatase-positive osteoclasts. The expression of 8-hydroxydeoxyguanosine was upregulated in the periodontal tissues. Intake of vitamin C reduced the effect of a high-cholesterol diet on alveolar bone density and osteoclast differentiation and decreased periodontal 8-hydroxydeoxyguanosine expression. CONCLUSION: In the rat model, vitamin C suppressed alveolar bone resorption, induced by high dietary cholesterol, by decreasing the oxidative damage of periodontal tissue.     

Vitamin C intake attenuates the degree of experimental atherosclerosis induced by periodontitis in the rat by decreasing oxidative stress

Objective

Periodontitis has been causally linked to cardiovascular disease, which is mediated through the oxidative stress induced by periodontitis. Since vitamin C has been suggested to limit oxidative damage, we hypothesized that vitamin C intake may reduce endothelial oxidative stress induced by periodontitis in the aorta. The aim of this study was to investigate the effects of vitamin C intake on the initiation of atherosclerosis in a ligature-induced rat periodontitis model.

Design

Eighteen 8-week-old-male Wistar rats were divided into three groups of six rats and all rats received daily fresh water and powdered food through out the 6-week study. In the vitamin C and periodontitis groups, periodontitis was ligature-induced for the first 4 weeks. In the vitamin C group, rats were given distilled water containing 1 g/L vitamin C for the 2 weeks after removing the ligature.

Results

In the periodontitis group, there was lipid deposition in the descending aorta and significant increases of serum level of hexanoyl-lysine (HEL), and aortic levels of nitrotyrosine expression, HEL expression and 8-hydroxydeoxyguanosine (8-OHdG) compared to the control group. Vitamin C intake significantly increased plasma vitamin C level and GSH:GSSG ratio (178% and 123%, respectively), and decreased level of serum HEL and aortic levels of nitrotyrosine, HEL and 8-OHdG (23%, 87%, 84%, and 38%, respectively).

Conclusions

These results suggest that vitamin C intake attenuates the degree of experimental atherosclerosis induced by periodontitis in the rat by decreasing oxidative stress.     

牙龈蛋白及其对破骨和成骨细胞功能的影响

牙龈卟啉单胞菌分泌的牙龈蛋白作为牙周病的一种重要的毒力因子,一方面通过降解骨保护蛋白（OPG）及促进核因子-κB（NF-κB）受体活化因子配体（RANKL）的释放激活OPG/RANKL/NF-κB受体活化因子信号转导通路,进而诱导破骨细胞前体细胞向破骨细胞分化,增强破骨细胞功能;另一方面,牙龈蛋白可通过线粒体依赖性内在程序性细胞死亡通路和肿瘤坏死因子受体家族介导的外在程序性细胞死亡通路诱导成骨细胞程序性细胞死亡,抑制成骨细胞功能,最终导致牙槽骨丧失。本文就近年来牙龈蛋白及其对破骨和成骨细胞功能的影响相关研究进展作一综述。     

HDAC5-Mediated Acetylation of p100 Suppresses Its Processing

IntroductionPeriodontitis is a condition involving chronic inflammation in the gums, periodontal ligaments, cementum, and alveolar bone. Nuclear factor-κB (NF-κB) activation is the prominent mediator of inflammation and osteoclast differentiation. The role of histone deacetylase 5 (HDAC5) in periodontitis development and NF-κB regulation is not fully understood.MethodsWe used primary mouse bone marrow–derived osteoclast cultures in vitro and a mouse model of chronic periodontists (CPD) treated with the HDAC4/5 inhibitor LMK-235. Real-time polymerase chain reaction, micro computed tomography, flow cytometry, western blot, and immunoprecipitation were used to study proinflammatory cytokines, NF-κB activation, HDAC5 activity, and the interaction of HDAC5 with NF-κB p100.ResultsLMK-235, a selective inhibitor of HDAC4 and HDAC5, reduced osteoclast marker gene expression (Cstk, Acp5, and Calcr) and tartrate-resistant acid phosphatase activity in primary osteoclast cultures. LMK-235 reduced the increase in cementoenamel junction–alveolar bone crest distance, inflammatory cell infiltration of gingival tissues, and expression levels of interleukin (IL)-1β, tumor necrosis factor alpha, IL-6, and IL-23a, indicating an ameliorative effect on CPD. Immunoprecipitation experiments have further confirmed p100–HDAC5 interaction, acetylation levels of p100, and NF-κB activation.ConclusionsThese results indicate that HDAC5 binds and deacetylates p100, leading to its activation, increased proinflammatory cytokine production, gingival infiltration, and osteoclast differentiation, thus promoting alveolar bone resorption. HDAC5 inhibition is therefore a potentially promising therapeutic strategy for the treatment of periodontitis.     

In vivo host interactions with mineral trioxide aggregate and calcium hydroxide: inflammatory molecular signaling assessment

Introduction

Mineral trioxide aggregate (MTA) and calcium hydroxide [Ca(OH)₂] are promising biomaterials for stimulating dentinogenesis and cementogenesis. This research was undertaken to understand how MTA and CA(OH)₂ participate in the inflammatory, healing, and biomineralization processes. In this part of the study, we evaluated inflammatory signaling molecules promoted by in vivo host interaction with MTA and Ca(OH)₂.

Methods

Human dentin tubes were filled with ProRoot MTA (Dentsply Tulsa Dental, Tulsa, OK), Ca(OH)₂, or kept empty. After 12 hours and 1, 3, 7, 15, 30, and 60 days of implantation in subcutaneous tissues in the backs of mice, the tubes and surrounding tissues were retrieved for cytokine level quantification and histological and immunohistochemical analysis.

Results

MTA and Ca(OH)₂ induced proinflammatory cytokine up-regulation for up to 3 days. Moreover, interleukin-10 overexpression was noted on the tissue in contact with the biomaterials during the acute phase of the inflammatory reaction. Immunohistochemical analyses showed an increased expression of myeloperoxidase, nuclear factor kappa B (NF-κB), cyclooxygenase-2, inducible nitric oxide synthase enzymes, and vascular endothelial growth factor on day 1 for all groups.

Conclusions

MTA and Ca(OH)₂ increased the activation of the NF-κB signaling system on day 1 for all groups. This finding can be associated with a proinflammatory and pro-wound healing environment, which was promoted earlier by MTA.     

Histomorphometric analysis and immunolocalization of RANKL and OPG during the alveolar healing process in female ovariectomized rats treated with oestrogen or raloxifene

Objective

To investigate the effects of bone-resorption inhibitors (oestrogen and raloxifene) on the RANKL/OPG balance during the chronology of the alveolar healing process in ovariectomized (OVX) rats by means of immunocolocalization and histomorphometric analysis.

Materials and methods

One hundred sixty female Wistar rats at 70 days of age were either OVX or sham-operated and divided into four groups: sham, OVX/Oil, OVX with E₂ replacement (17β-estradiol, 400 μg/month), OVX with RLX treatment (1 mg/kg bw/day). The 60-day treatment started 8 days after ovariectomy. The incisors were extracted to allow analysis of 7, 14, 21, 28 and 42 days of wound healing. After obtaining the histological samples, slides were stained with hematoxylin and eosin or subjected to immunocolocalization reaction for RANKL and OPG. Results were quantitatively evaluated.

Results

Histomorphometric analysis showed that the sham group presented the highest and OVX/Oil group the lowest mean bone formation value in the post-extraction period. The immunocolocalization analysis showed a larger increase in bone turnover at 7 postoperative days in OVX/Oil and sham groups and decreasing bone turnover in the other periods. The OVX/Oil group showed a large decrease in bone turnover at 14 postoperative days, a period demonstrated by mild cellular activity. OVX/E₂ and sham groups showed a decreased bone turnover at 28 postoperative days while OVX/RLX group showed a decreased bone turnover at 21 postoperative days. On the 42nd postoperative day, sham and OVX/RLX groups showed an established alveolar bone healing process.

Conclusions

Ovariectomy delays the alveolar healing process and interferes with bone turnover through the balance between RANKL and OPG. Oestrogen replacement or raloxifene treatment did not totally recover the oestrogen-deficient state. However raloxifene treatment showed more satisfactory results than oestrogen replacement.     

Reactive oxygen species removal activity of davallialactone reduces lipopolysaccharide-induced pulpal inflammation through inhibition of the extracellular signal-regulated kinase 1/2 and nuclear factor kappa b pathway

Introduction

Davallialactone, hispidin analogues derived from the mushroom Inonotus xeranticus, has antioxidant properties. This study examined whether the reactive oxygen species (ROS) removal activity of davallialactone affects the lipopolysaccharide (LPS)-induced anti-inflammatory activity in human dental pulp cells.

Methods

The LPS-induced formation of ROS was analyzed by using dichlorofluorescein diacetate with fluorescence-activated cell sorter, and the expression of inflammatory molecules in primary cultured human dental pulp cells was determined by immunoblotting. The inflammatory mechanism of the davallialactone-involved signal pathway was examined by immunoblotting.

Results

Davallialactone acted as an antioxidant to confirm the elimination of ROS formation and elevation of Cu/Zn superoxide dismutase and Mn superoxide dismutase expression in LPS-induced pulp cells. The antioxidant activity of davallialactone leads to inhibition of LPS-induced inflammation by blocking the extracellular signal-regulated kinase (ERK1/2) and nuclear factor kappa B (NF-κB) pathway, which decreases the expression of inflammatory molecules such as intercellular adhesion molecule-1, vascular cell adhesion molecule-1, matrix metalloproteinase-2, matrix metalloproteinase-9, inducible nitric oxide synthase, and cyclooxygenase-2. The character of davallialactone was more effective in comparison with N-acetylcysteine as the control antioxidant in this study.

Conclusions

Davallialactone has antioxidant activity and anti-inflammatory effects in LPS-induced human dental pulp cells through the suppression of ERK1/2 activation followed by blockage of NF-κB translocation from cytosol into nuclear. Therefore, the good anti-inflammatory capacity of davallialactone might be used for oral diseases such as pulpitis and periodontitis.     

牙槽裂两侧正畸牙移动后RANKL和OPG表达的研究

目的：研究牙移动后牙槽裂隙两侧破骨细胞核因子κB受体活化因子配体（RANKL）和骨保护素（OPG）的表达和作用.方法：选用4只4月龄雄性Beagle犬建立牙槽裂模型,正畸牵引裂隙两侧牙齿向裂隙区移动,实时荧光定量聚合酶链反应（real-time PCR）检测裂隙两侧牙槽骨中RANKL和OPG mRNA的表达水平.结果：正畸牵引16～20周后,牙槽裂间隙缩小以至关闭,原裂隙处有新骨生成.裂隙两侧牙槽骨中RANKL和OPG mRNA表达增加.结论：牙槽裂两侧牙齿受正畸牵引后,RANKL与OPG参与骨重建,裂隙处骨改建活跃.此方法为修复牙槽裂提供新的思路.     

Osteogenic induction and 1,25-dihydroxyvitamin D3 oppositely regulate the proliferation and expression of RANKL and the vitamin D receptor of human periodontal ligament cells

Objective

Human periodontal ligament cells (hPDLCs) may play an important role in osteoclastogenesis in alveolar bone by expressing the receptor activator of NF-KappaB ligand (RANKL) and osteoprotegerin (OPG). The present study aimed to investigate the differences between the effects of osteogenic induction and 1,25-dihydroxyvitamin D₃ (VD₃) on hPDLC proliferation and the expression of RANKL, osteoprotegerin, and the vitamin D receptor (VDR) in hPDLCs.

Methods

Primary cultures of 11 hPDLC populations from 11 donors were obtained. Three samples of each hPDLC population from passage 3 were, respectively, treated with osteogenic induction medium, 10⁻⁸ M VD₃, or vehicle as a control. Cell proliferation at days 0, 1, 3, 5, and 7 was estimated with the MTT method. At day 6, the mRNA levels of RANKL, OPG and VDR were determined with real-time RT-PCR.

Results

Osteogenic induction significantly promoted hPDLC proliferation, while VD₃ inhibited proliferation. Osteogenic induction significantly down-regulated the mRNA level of RANKL by 1.61-fold (P = 0.033) and decreased the level of VDR by 2.13-fold (P = 0.003), while there was no change in the level of OPG and OPG/RANKL ratio with osteogenic induction. On the contrary, VD₃ significantly up-regulated the level of RANKL by 9.58-fold (P = 0.001) and increased the level of VDR by 3.15-fold (P = 0.004), while down-regulating the OPG/RANKL ratio by 7.14-fold (P = 0.004).

Conclusion

Osteogenic induction and VD₃ exert opposite effects in regulating hPDLC proliferation and mRNA expression of RANKL and VDR. This may induce hPDLCs to play different roles in alveolar bone metabolism.     

Blockade of sympathetic β-receptors inhibits Porphyromonas gingivalis-induced alveolar bone loss in an experimental rat periodontitis model

Introduction

Periodontal disease is characterised by alveolar bone loss. Some studies have suggested the involvement of sympathetic nervous system in the deterioration of periodontal disease. Noradrenaline, released from sympathetic nerve terminals due to various stimuli, binds to specific adrenergic receptors on immune cells. Recently, we reported that restraint stress augmented the alveolar bone loss induced by Porphyromonas gingivalis infection. In this study, we investigated the effects of the β-blocker (propranolol) on alveolar bone loss induced by P. gingivalis infection to examine the involvement of sympathetic nerves in periodontal breakdown.

Methods

Sprague-Dawley rats were treated as follows: saline injection (Group A), propranolol injection (Group B), saline injection and oral challenge with P. gingivalis (Group C), and propranolol injection and oral challenge with P. gingivalis (Group D). Horizontal alveolar bone loss was evaluated by measuring the distance between the cemento-enamel junction and the alveolar bone crest. Specimens from periodontal tissue were evaluated by staining with hematoxylin-eosin and tartrate-resistant acid phosphatase.

Results

Blockade of β-receptors in periodontal tissue by propranolol inhibited osteoclast differentiation and prevented alveolar bone loss induced by P. gingivalis infection. Histological study revealed that the number of osteoclasts detected was proportional to the level of bone loss.

Conclusions

These results indicate that the sympathetic nervous system is involved in the development of periodontitis and suggest that sympathetic signal modulation with β-blockers enables the control of alveolar bone mass metabolism.     

Comparative immunohistochemical expression of RANK, RANKL and OPG in radicular and dentigerous cysts

Objective

Receptor activator of nuclear factor-κB (RANK), RANK ligand (RANKL) and osteoprotegerin (OPG) are members of the superfamily of ligands and receptors of tumour necrosis factor family involved in bone metabolism. The formation, differentiation and activity of osteoclasts are regulated by these proteins. To clarify the roles of osteoclast regulatory factors in cystic expansion of odontogenic cysts, expression of these proteins were analysed in radicular and dentigerous cysts.

Design

The immunohistochemistry expression of these biomarkers were evaluated and measured in lining epithelium and fibrous capsule of the radicular (n = 20) and dentigerous cysts (n = 20).

Results

A similar expression in lining epithelium was observed in the lesions. The fibrous capsule of dentigerous cyst showed a higher content of RANK-positive and RANKL-positive cells than fibrous capsule of radicular cyst. In the lining epithelium the RANKL/OPG ratio showed higher numbers of OPG-positive than RANKL-positive cells, whereas fibrous capsule of the cysts had a tendency to present a similar expression (OPG = RANKL).

Conclusion

Ours findings indicate the presence of RANK, RANKL and OPG in cysts. Moreover, increased expression of OPG compared to RANKL in the lining epithelium could contribute to the differential bone resorption activity in theses lesions.     

中间普氏菌培养上清对牙龈成纤维细胞RANKL／OPG表达的影响

目的：探讨中间普氏菌培养上清对牙龈成纤维细胞核因子-κB受体活化因子配体（receptor activator for nuclear factor-κB ligand,RANKL）、骨保护素（osteoprotegerin,OPG）表达的影响。方法：将系列浓度的中间普氏菌培养上清作用于牙龈成纤维细胞,显微镜下观察细胞的变化。实时定量 RT-PCR法和 Western blot法分别检测 RANKL、OPG mRNA及蛋白水平的表达,并计算 RANKL/OPG的比值。结果：牙龈成纤维细胞的数量随中间普氏菌培养上清浓度的增加而减少,50μg/mL 时,显微镜下没有活细胞存在。在mRNA水平上和蛋白水平上,RANKL/OPG的比值在12．5μg/mL处理组均高于对照组,差异具有统计学意义（P＜0．01）。结论：中间普氏菌培养上清可诱导人牙龈成纤维细胞表达 RANKL 和 OPG,破坏 RANKL/OPG比例的平衡,影响破骨细胞分化的细胞因子微环境,在破骨细胞分化和牙周炎骨吸收中起着一定的调节作用。     

Porphyromonas endodontalis lipopolysaccharides induce RANKL by mouse osteoblast in a way different from that of Escherichia coli lipopolysaccharide

Introduction

Porphyromonas endodontalis lipopolysaccharide (LPS) has been shown to have a high positive rate in infected root canals and symptomatic apical periodontitis. It may play an integral role as a potent stimulator of inflammatory cytokines involved in apical lesions. The receptor activator of nuclear factor-κB ligand (RANKL) has been proven to be the key regulator of bone remodeling. This study investigated P. endodontalis LPS-induced RANKL production and LPS signaling in mouse osteoblasts.

Methods

LPS-induced RANKL production in mouse osteoblast MC3T3-E1 cells was measured by Western blot and real-time polymerase chain reaction, and the Toll-like receptors (TLRs) were determined by the blocking test using anti-TLRs antibodies. In addition, specific inhibitors were used to analyze the intracellular signaling pathways. Escherichia coli LPS was used as the control.

Results

Both of the anti-TLR2 and anti-TLR4 antibodies significantly (P < .05) inhibited the expression of RANKL from osteoblasts stimulated with P. endodontalis LPS; only anti-TLR2 antibody had a significant (P < .05) inhibitory effect on E. coli LPS signaling. SP600125 (c-Jun N-terminal kinase [JNK] inhibitor) prevented the up-regulation of RANKL expression in P. endodontalis LPS-infected osteoblasts (P < .05). The inhibitory effect of wortmannin (phosphatidylinositol 3-kinase inhibitor) and PD98059 (mitogen-activated protein kinase [MAPK]/extracellular signal-regulated kinase [ERK] kinase-1/2 [MEK 1/2] inhibitor) were observed in E. coli LPS-treated mouse osteoblasts (P < .05).

Conclusions

Results from this study showed that P. endodontalis LPS has the ability to promote the expression of RANKL in mouse osteoblasts, and this induction was mainly through the TLR2/4-JNK signaling pathway, a situation quite different from that of typical bacterial endotoxin (E. coli LPS).     

铜掺入型钛合金对破骨细胞分化影响的体外研究

目的:研究铜掺入型钛合金对破骨细胞分化的影响.方法:通过激光熔化技术制备掺铜的钛合金.使用RANKL诱导小鼠单核细胞分化,制备破骨细胞.将掺铜的钛合金与破骨细胞共培养,通过观察细胞形态、记录破骨细胞数目以及破骨相关基因的表达,分析掺铜的钛合金对破骨细胞分化相关基因表达的影响.结果:小鼠单核细胞与RANKL共培养后可分化...     

Dexamethasone promotes regeneration of crushed inferior alveolar nerve by inhibiting NF-κB activation in adult rats

PurposeNuclear factor kappa B (NF-κB), which is closely related to inflammation, has become a topic of interest for research. The aim of this study is to investigate the effects of dexamethasone (Dex), an inhibitor of NF-κB, on inferior alveolar nerve injury in adult rats.Materials and methodsThe crushed inferior alveolar model is established in Wistar rats and they are randomly divided into three groups according to treatment: pyrrolidine dithiocarbamate (PDTC), dexamethasone (Dex), and saline (physiological saline). After treatment, the rats are respectively sacrificed at 3, 7, and 14 d, and inferior alveolar nerves are extracted for histochemical and western blot analysis.ResultCompared with the PDTC and saline groups, nerve fibers in the Dex group are regularly arranged with few vacuoles, which is similar to normal inferior alveolar nerves. Immunofluorescent results show significantly decreased NF-κB expression in the Dex group. Western bolt shows higher expression of GAP-43 and lower expression of NF-κB.ConclusionTaken together, all results show that dexamethasone significantly improved the regeneration of crushed inferior alveolar nerves by inhibiting NF-κB activation in adult rats.     

间歇性低氧对牙周炎大鼠牙周组织中 NF-κB、IL-6及 PGE2含量的影响

目的：建立慢性间歇性低氧（CIH）及牙周炎大鼠模型,研究 NF-κB、IL-6及 PGE2水平的变化。方法：将32只普通级6周龄雄性 SD 大鼠随机分为4组（n ＝8）：A：常氧空白组、B：常氧牙周炎组、C：CIH 组、D：CIH 合并牙周炎组。B、D 组大鼠上颌第二磨牙进行结扎处理,辅以高糖饮食;A、C 组正常饮食。C、D 组置于低氧舱8 h／d。8周后处死,HE 染色,免疫组化检测牙周组织 NF-κB 含量,ELISA 检测牙龈组织 IL-6、PGE2。结果：HE 染色：8周后 B 组、D 组牙周炎症表现明显。免疫组化：B、C、D 组 NF-κB 表达均高于 A 组（P ＜0．05）;ELISA 检测：B、C、D 组 IL-6、PGE2含量高于 A 组（P ＜0．05）,且 D 组 IL-6、PGE2和NF-κB 表达较其他各组明显升高。结论：CIH 导致炎症因子升高,加重牙周组织破坏,使牙周炎症加剧。     

Hydrogen sulfide synergistically upregulates Porphyromonas gingivalis lipopolysaccharide-induced expression of IL-6 and IL-8 via NF-κB signalling in periodontal fibroblasts

Objectives

The periodontal pathogen Porphyromonas gingivalis produces hydrogen sulfide (H₂S). H₂S in the oral cavity is positively correlated with periodontitis but the mechanism by which H₂S contributes to periodontal diseases is obscure. We investigated the effect of H₂S in combination with P. gingivalis lipopolysaccharide (LPS) on expression of the pro-inflammatory cytokines interleukin (IL)-6 and IL-8 in periodontal fibroblasts and the underlying mechanism of action.

Material and methods

Gingival fibroblasts (GFs) and periodontal ligament cells (PDLCs) were treated with different concentrations of the H₂S donor NaHS in the presence/absence of P. gingivalis LPS for different time periods. Expression of IL-6 and IL-8 was detected by real-time PCR and ELISA. The activity of nuclear factor-kappa B (NF-κB) signalling was investigated using western blotting, EMSA and pathway blockade assays.

Results

Real-time PCR and ELISA results showed that H₂S not only upregulated expression of IL-6 and IL-8 at mRNA and protein levels in a dose- and time-dependent manner, but also aggravated P. gingivalis LPS-induced expression of IL-6 and IL-8 in GFs and PDLCs. Western blotting and EMSA showed that NF-κB signalling was activated by NaHS, P. gingivalis LPS, and both, which was in accordance with the expression levels of IL-6 and IL-8 in GFs and PDLCs. These results were confirmed using a NF-κB pathway blockade assay.

Conclusions

H₂S synergistically upregulated P. gingivalis LPS-induced expression of IL-6 and IL-8 in GFs and PDLCs via activation of NF-κB signalling, which could promote the development of periodontitis.