Measuring methane emissions during the installation of residential and commercial natural gas meters in China.

Methane is a potent greenhouse gas that accounts for one-quarter of the world's radiative forcing. Methane emissions from the natural gas sector are prevalent throughout the natural gas (NG) chain. Studies have shown that methane emissions from post-meter uses of natural gas are vastly understated. A surge in the number of natural gas users, for example, would amplify the climate impact of methane emissions during the installation of natural gas meters. Thus, quantifying methane emissions during the installation of natural gas meters is critical in light of severe global climate change and urgent reduction targets. In this study, we used a mass balance approach to calculate methane emissions during the separate installation of 1444 residential natural gas meters and 51 commercial natural gas meters. Our results revealed the methane emission had a fat tail distribution. Specifically, the estimated mean methane emissions for household users were 0.008 (0.001-0.022) kg per household and 0.192 (0.013-0.816) kg per commercial user. Extrapolating these statistics to the whole of China, total emissions from 2007 to 2021 were 3.80 million metric tons (MMt) CH4, with an annual average of 0.25 MMt. Notably, in terms of economic development and population size, the provinces with the highest methane emissions were concentrated in the southeast. Our findings close a gap in measuring CH4 emissions in China across the natural gas chain and provide data to support the reduction targets set and the development of reduction technologies.

Occlusal trauma inhibits osteoblast differentiation and bone formation through IKK-NF-κB signaling.

BACKGROUND: Occlusal trauma is an important factor promoting bone loss caused by periodontal diseases. Although there are reports of traumatic force promoting bone resorption in periodontal diseases, no studies examining the inhibition of bone formation by traumatic force and the underlying mechanism have been reported. The aim of this study was to investigate the mechanism whereby traumatic force inhibits bone formation. METHODS: MC3T3-E1 cells were induced to undergo osteogenic differentiation and subjected to cyclic uniaxial compressive stress with or without stimulation with Pg. LPS. The expression of osteoblast markers and the activation of IKK-NF-κB signaling were evaluated in vitro. Then, MC3T3-E1 cells were induced to undergo osteogenic differentiation and subjected to cyclic uniaxial compressive stress with or without IKK-2 Inhibitor VI. The expression of osteoblast markers was determined. Then, the classic Wnt signaling pathway (ß-catenin, Gsk3ß, p-Gsk3ß, and Dkk1) was further evaluated in vitro. Finally, occlusal trauma was induced in Wistar rats with or without the injection of IKK-2 Inhibitor VI, to evaluate changes in bone mass and IKK-NF-κB and Wnt/ß-catenin signaling in vivo. RESULTS: After stimulation with Pg. LPS and traumatic force, IKK-NF-κB signaling was significantly activated in vitro. The expression of osteoblast markers and the activity of alkaline phosphatase in MC3T3-E1 cells declined after traumatic force loading and were rescued when IKK-NF-κB signaling was blocked. Wnt/ß-catenin signaling was accordingly inhibited upon force loading, but this inhibition was reversed when IKK-NF-κB was antagonized in vitro. X-ray and Micro-CT analysis of the mandibles of the rats as well as HE and TRAP staining showed that bone loss induced by occlusal trauma declined after IKK-NF-κB was inhibited. The expression of p65 and IκBα was increased when occlusal trauma was induced in Wistar rats, whereas ß-catenin, OCN, and Runx2 levels were decreased. After blocking IKK-NF-κB, significant upregulation of ß-catenin, OCN, and Runx2 was observed in rats suffering from occlusal trauma. CONCLUSIONS: IKK-NF-κB signaling could be activated by traumatic force or occlusal trauma. Its activation promoted the degradation of ß-catenin, ultimately inhibiting osteogenic differentiation in vitro and bone formation in vivo.

The effect of NLRP inflammasome on the regulation of AGEs-induced inflammatory response in human periodontal ligament cells.

BACKGROUND AND OBJECTIVE: Diabetes influences the frequency and development of periodontitis. Inflammation of human periodontal ligament cells (HPDLCs) participates in this pathologic process. Hence, this study aims to explore whether advanced glycation end products (AGEs), by-products of diabetes, could exaggerate inflammation induced by muramyl dipeptide (MDP) in HPDLCs, and whether nucleotide-binding oligomerization domain-like receptors (NLRs) signaling pathway was involved. MATERIAL AND METHODS: Human periodontal ligament cells were pre-treated with 100 µg/mL AGEs for 24 hours and stimulated with 10 µg/mL MDP for 24 hours. IL-6, IL-1ß, and RAGE were detected, and the activation of NF-κB signaling pathway was observed. The expression of NLRs was evaluated with or without silencing RAGE or blocking NF-κB pathway under AGEs stimulation. Statistical analyses were performed by using independent sample t test. RESULTS: Advanced glycation end products induced significant increase of inflammatory cytokines in HPDLCs (P < 0.05). Results of western blot (WB) showed that after 45 minutes stimulation of AGEs, p-p65/p65 ratio peaked; AGEs promoted the expression of NLRP1, NLRP3, and apoptosis-associated speck-like protein containing a CARD (ASC). After silencing RAGE or blocking NF-κB pathway, the up-regulation of NLRs protein caused by AGEs was attenuated. Additionally, AGEs pre-treatment could enhance the inflammatory response of MDP and the expression of NLRs, which were demonstrated by more expression of IL-6, IL-1ß, NOD2, NLRP1, NLRP3, and ASC. CONCLUSION: Advanced glycation end products induced inflammatory response in HPDLCs via NLRP1-inflammasome and NLRP3-inflammasome activation in which NF-κB signal pathway was involved. Besides, AGEs promoted the inflammatory response of MDP via NOD2, NLRP1-inflammasome, and NLRP3-inflammasome.

Dickkopf-1 may regulate bone coupling by attenuating wnt/ß-catenin signaling in chronic apical periodontitis.

OBJECTIVE: Alveolar bone loss is a common outcome of chronic apical periodontitis. In this study, we investigated the involvement of the Dickkopf-1-Wnt/ß-catenin signaling pathway in the attenuation of osteogenic differentiation induced by Escherichia coli lipopolysaccharide, and we evaluated the use of Dickkopf-1 inhibitor and Dickkopf-1 recombinant protein to reverse bone loss in different phases of osteogenic differentiation. METHODS: MC3T3-E1 cells grown in osteogenic medium were treated with Escherichia coli lipopolysaccharide for 24h during osteogenic induction on days 0, 1, 7, 14 and 21. Dickkopf-1 siRNA was added on days 0 and 1, and Dickkopf-1 recombinant was added on days 7, 14, and 21. Quantitative real-time PCR, Western blotting and alkaline phosphatase activity assays were performed to measure osteogenic marker expression and Wnt/ß-catenin signaling. A rat apical periodontitis model was used to further evaluate the function of Dickkopf-1 in relation to bone loss. RESULTS: MC3T3-E1 cells treated with Escherichia coli lipopolysaccharide showed decreased mRNA expression of osteogenic markers. Wnt/ß-catenin signaling was also inhibited, and Dickkopf-1 showed corresponding variations as quantified by Western blotting. Using Dickkopf-1 inhibitor or Dickkopf-1 recombinant protein at different phases of osteogenic differentiation in vitro partially reversed the decrease in osteogenic marker expression. The rat apical periodontitis model indicated that the Dickkopf-1 inhibitor could restore bone loss in the periapical area in vivo. CONCLUSIONS: Dickkopf-1 may play a key regulatory role in determining the outcome for bone in inflammatory environments, and modulating the Wnt/ß-catenin signaling pathway via Dickkopf-1 inhibitor or recombinant protein may provide a potential therapeutic option to prevent bone destruction in endodontic disease.

Expression Profiles of Inflammation-associated microRNAs in Periapical Lesions and Human Periodontal Ligament Fibroblasts Inflammation.

INTRODUCTION: microRNAs (miRNAs) have been identified to be closely related to inflammatory diseases. The aim of our study was to identify expression profiles of miRNAs associated with inflammation in apical periodontitis (AP) lesions and human periodontal ligament fibroblasts (HPDLFs) inflammation. METHODS: Total RNAs were extracted from 10 AP lesions, 6 control tissues, and HPDLFs using lysis buffer. Expressions of miRNAs (miR-29b, 106b, 125b, 143, 155, and 198) were detected by real-time polymerase chain reaction. The dual luciferase assay was used to test miR-155 directly targeted semaphorein3a (SEMA3A). Western Blot and the enzyme-linked immunosorbent assay were used to detect the protein expressions of SEMA3A and proinflammatory cytokines, respectively. All experiments were repeated at least 3 times. Data were analyzed using the independent sample t test. RESULTS: The previously mentioned miRNAs were all significantly up-regulated in AP lesions (P < .05), whereas they were not in HPDLFs inflammation, in which miR-29b, 106b, 125b, and 198 were down-regulated (P < .05) and miR-143 and 155 were unchanged (P > .05). Overexpression of miR-155 induced proinflammatory phenotype, and down-regulation reduced the other miRNAs in HPDLFs (P < .05). Moreover, miR-155 directly targeted SEMA3A, which was significantly up-regulated (increased) in acute HPDLFs inflammation and down-regulated (decreased) in AP lesions (P < .05). Knockdown of SEMA3A led to induction of the proinflammatory phenotype; down-regulation of miR-29b, 106b, 125b, 143, and 198 (P < .05); and, more importantly, up-regulation of miR-155 (P < .01). CONCLUSIONS: Our study showed that the expression profiles of inflammation-associated miRNAs associated with inflammation in AP lesions and HPDLFs inflammation were different. miR-155 may play a crucial role in apical periodontitis progression by directly inhibiting SEMA3A.

Expression of programmed death 1 ligand 1 on periodontal tissue cells as a possible protective feedback mechanism against periodontal tissue destruction.

Programmed death 1 ligand 1 (PD­L1) is a negative co­stimulatory molecule in immune responses. Previous reports have indicated that inflammatory cytokines can upregulate the expression of PD­L1 in tumor cells, which in turn suppresses host immune responses. Periodontitis is characterized by persistent inflammation of the periodontium, which is initiated by infection with oral bacteria and results in damage to cells and the matrices of the periodontal connective tissues. In the present study, the expression and function of PD­L1 in periodontal tissue destruction were examined. Periodontal ligament cells (PDLCs) were stimulated by inflammatory cytokines and periodontal pathogens. The expression and function of PD­L1 on the surface of PDLCs was investigated using flow cytometry in vitro. Periodontal disease was induced by the injection of Porphyromonas gingivalis in mouse models. The expression levels of PD­L1 in the periodontal tissues of the mice were analyzed using flow cytometry and immunohistochemistry. PD­L1 was inducibly expressed on the PDLCs by the inflammatory cytokines and periodontal pathogens. The inflammation­induced expression of PD­L1 was shown to cause the apoptosis of activated T lymphocytes and improve the survival of PDLCs. Furthermore, in the mouse model of experimental periodontitis, the expression of PD­L1 in severe cases of periodontitis was significantly lower, compared with that in mild cases. By contrast, no significant differences were observed between the healthy control and severe periodontitis groups. The results of the present study showed that the expression of PD­L1 may inhibit the destruction of periodontal tissues, indicating the involvement of a possible protective feedback mechanism against periodontal infection.

Compressive mechanical stress may activate IKK-NF-κB through proinflammatory cytokines in MC3T3-E1 cells.

OBJECTIVE: To determine whether IKK-NF-κB is activated either directly by compressive mechanical stress or by proinflammatory cytokines produced by MC3T3-E1 cells under compressive stress loading. RESULTS: MC3T3-E1 cells subjected to cyclic uniaxial compressive stress showed increased expression of proinflammatory cytokines and activation of the IKK-NF-κB signaling pathway with nuclear translocation of p65. Following treatment with antibodies to neutralize the action of the proinflammatory cytokines, IL-1ß and IL-6, the activation of IKK-NF-κB signaling was notably inhibited in MC3T3-E1 cells subjected to force loading. CONCLUSION: IKK-NF-κB signaling in MC3T3-E1 cells may be activated by proinflammatory cytokines that are produced as a consequence of mechanical stress loading and not by direct compressive mechanical stress.

Role of saliva proteinase 3 in dental caries.

Salivary analysis can be used to assess the severity of caries. Of the known salivary proteins, a paucity of information exists concerning the role of proteinase 3 (PR3), a serine protease of the chymotrypsin family, in dental caries. Whole, unstimulated saliva was collected from children with varying degrees of active caries and tested using a Human Protease Array Kit and an enzyme-linked immunosorbent assay. A significantly decreased concentration of salivary PR3 was noted with increasing severity of dental caries (P<0.01); a positive correlation (r=0.87; P<0.01; Pearson's correlation analysis) was also observed between salivary pH and PR3 concentration. In an antibacterial test, a PR3 concentration of 250 ng·mL⁻¹ or higher significantly inhibited Streptococcus mutans UA159 growth after 12 h of incubation (P<0.05). These studies indicate that PR3 is a salivary factor associated with the severity of dental caries, as suggested by the negative relationship between salivary PR3 concentration and the severity of caries as well as the susceptibility of S. mutans to PR3.

Inflammation promotes oral squamous carcinoma immune evasion via induced programmed death ligand-1 surface expression.

The association between inflammation and cancer provides a new target for tumor biotherapy. The inflammatory cells and molecules within the tumor microenvironment have decisive dual roles in antitumor immunity and immune evasion. In the present study, phytohemagglutinin (PHA) was used to stimulate peripheral blood mononuclear cells (PBMCs) to simulate the tumor inflammatory microenvironment. The effect of immune cells and inflammatory cytokines on the surface expression of programmed cell death-1 ligand 1 (PD-L1) and tumor immune evasion was investigated using flow cytometry (FCM) and an in vivo xenotransplantation model. Based on the data, PHA-activated, but not resting, immune cells were able to promote the surface expression of PD-L1 in Tca8113 oral squamous carcinoma cells via the secretion of inflammatory cytokines, but not by cell-cell contact. The majority of the inflammatory cytokines had no significant effect on the proliferation, cell cycle progression and apoptosis of the Tca8113 cells, although they each induced the expression of PD-L1 in a dose-dependent manner. In total, 99% of the Tca8113 cells expressed PD-L1 following treatment with the supernatant of PHA-stimulated PBMCs. The PHA-supernatant pretreated Tca8113 cells unusually induced Tca8113 antigen-specific CD8+ T cell apoptosis in vitro and the evasion of antigen-specific T cell attraction in a nude mouse tumor-bearing model. These results indicate a new mechanism for the promotion of tumor immune evasion by the tumor inflammatory microenvironment.