METTL3 promotes immature dental pulp stem cells-induced angiogenesis by regulating ETS1 mRNA stability in an m6A-HuR-dependent manner.

Angiogenesis serves as the determinate element of pulp regeneration. Dental pulp stem cell (DPSC) implantation can promote the regeneration of dental pulp tissue. Herein, the role of m6A methyltransferase methyltransferase-like 3 (METTL3) in regulating DPSCs-induced angiogenesis during pulp regeneration therapy was investigated. Cell DPSC viability, HUVEC migration, and angiogenesis ability were analyzed by CCK-8 assay, wound healing, Transwell assay, and tube formation assay. The global and EST1 mRNA m6A levels were detected by m6A dot blot and Me-RIP. The interactions between E26 transformation-specific proto-oncogene 1(ETS1), human antigen R(HuR), and METTL3 were analyzed by RIP assay. The relationship between METTL3 and the m6A site of ETS1 was performed by dual-luciferase reporter assay. ETS1 mRNA stability was examined with actinomycin D. Herein, our results revealed that human immature DPSCs (hIDPSCs) showed stronger ability to induce angiogenesis than human mature DPSCs (hMDPSCs), which might be related to ETS1 upregulation. ETS1 knockdown inhibited DPSCs-induced angiogenesis. Our mechanistic experiments demonstrated that METTL3 increased ETS1 mRNA stability and expression level on DPSCs in an m6A-HuR-dependent manner. ETS1 upregulation abolished sh-METTL3's inhibition on DPSCs-induced angiogenesis. METTL3 upregulation promoted DPSCs-induced angiogenesis by enhancing ETS1 mRNA stability in an m6A-HuR-dependent manner. This study reveals a new mechanism by which m6A methylation regulates angiogenesis in DPSCs, providing new insights for stem cell-based tissue engineering.

Klotho inhibits H2 O2 -induced oxidative stress and apoptosis in periodontal ligament stem cells by regulating UCP2 expression.

Periodontitis, a human chronic inflammatory disease, has affected the lives of millions of individuals. Periodontal ligament stem cells (PDLSCs), derived from the periodontal ligament, exhibit tissue specificity and impaired differentiation ability and are closely associated with tissue regeneration in periodontitis. Klotho, a single-pass transmembrane protein, has been reported to positively affect H2 O2 -induced oxidative stress and inflammation in PDLSCs. The ultimate damage of oxidative stress stimulation in PDLSCs was cell apoptosis, which was also the major lesion in periodontitis. Thus, the present study aimed to figure out the effect of klotho on H2 O2 -injured PDLSCs and its underlying mechanism to provide new therapeutic targets in periodontitis. The expression of klotho and uncoupling protein 2 (UCP2) was investigated in the gingival tissues, gingival crevicular fluid (GCF), and periodontal ligament stem cells (PDLSCs) in patients with chronic periodontitis. Then, under klotho treatment, oxidative stress was evaluated by measuring SOD and GSH-PX levels. Cell apoptosis and cell necrosis were also detected by measuring the cell death-relevant proteins, including Caspase-3, BAX, Bcl, MLKL, RIP1, and RIP3. Finally, a rescue assay was performed by inhibiting the expression of UCP2. The results showed that klotho and UCP2 were downregulated in patients with chronic periodontitis. In addition, klotho upregulated the production of UCP2 in H2 O2 -treated PDLSCs. Klotho inhibited H2 O2 -induced oxidative stress and cellular loss in PDLSCs, moreover, the rescue assay suggested that UCP2 knockdown suppressed the effects of klotho on PDLSCs. In conclusion, this study showed that klotho inhibits H2 O2 -induced oxidative stress and apoptosis in PDLSCs by regulating UCP2 expression. This novel discovery might provide a potential target for chronic periodontitis treatment.

Role of hormone replacement therapy in relieving oral dryness symptoms in postmenopausal women: a case control study.

BACKGROUND: To evaluate the efficacy of hormone replacement therapy in relieving oral symptoms in postmenopausal women presenting with genitourinary symptoms along with oral dryness. METHODS: A case-control study was conducted after selecting 60 postmenopausal women. Oral dryness status of all the patients was evaluated with the help of questionnaire related to oral dryness. These subjects were divided into case group and control group on the basis of response to questionnaire of oral dryness. Unstimulated saliva samples were obtained and analyzed for estimation of salivary estradiol levels by enzyme linked immune sorbent assay technique. After analyzing the result of salivary estradiol levels, case group was subjected to hormone replacement therapy (HRT). The patients were followed up for their response towards oral dryness as well as salivary estradiol levels after the therapy. RESULTS: The mean salivary estradiol level before HRT was significantly more among control group as compared to case group (p value < 0.001). Most of the patients complained of dry mouth (26 out of 30); reduced amount of saliva in the mouth (25 out of 30); dry mouth at night (28 out of 30); dry mouth during the day (25 out of 30) before HRT. These complains were significantly reduced after the therapy. The mean salivary estradiol in the case group levels increased significantly after HRT (p value < 0.001). CONCLUSION: The salivary estradiol levels were reduced in post menopausal women with the complain of xerostomia as compared to those without the complain of xerostomia. Further these levels can be recovered with the help of hormone replacement therapy.

Expression of ß-catenin and MMP-8 in gingival crevicular fluid and gingival tissue indicates the disease severity of patients with chronic periodontitis.

The aim of the present study was to investigate the interaction among ß-catenin, matrix metalloproteinase-8 (MMP-8) and severity in patients with chronic periodontitis. Both gingival crevicular fluid (GCF) and gingival tissue was collected from 21 healthy control individuals, 21 patients with moderate chronic periodontitis (mCP) and 23 patients with severe chronic periodontitis (sCP). The concentration of MMP-8 in GCF was detected via ELISA and the mRNA levels of ß-catenin and MMP-8 in GCF and gingival tissue was detected via reverse transcription-quantitative PCR. The protein levels of ß-catenin and MMP-8 in gingival tissue was detected using western blotting and the interaction between ß-catenin and MMP-8 in gingival tissue was detected by co-immunoprecipitation. The expression of ß-catenin and MMP-8 was significantly higher in the GCF and gingival tissue of patients with chronic periodontitis (mCP and sCP) compared with the control patients. Furthermore, the expression of ß-catenin and MMP-8 in GCF and gingival tissue was positively correlated with the clinical attachment level. In addition, a positive interaction was identified between ß-catenin and MMP-8, and the expression of ß-catenin was positively correlated with the expression of MMP-8 in GCF and gingival tissue. The CGF and gingival tissue expression of ß-catenin and MMP-8 may indicate disease severity in patients with chronic periodontitis.

Distribution of 8 periodontal microorganisms in family members of Chinese patients with aggressive periodontitis.

OBJECTIVE: To date, no information on the distribution of periodontal microorganisms among family members of Chinese patients with aggressive peridontitis (AgP) is available. The aim of the present study was to investigate the probability of transmission of eight periodontal microorganisms between patients with aggressive periodontitis and their family members. DESIGN: Saliva and pooled subgingival plaque samples were collected from 103 participants from 41 nuclear families (including 41 AgP probands, 19 mothers, 22 fathers, 21 siblings). Eight periodontal microorganisms, including Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia, Treponema denticola, Campylobacter rectus, Prevotella intermedia, Prevotella nigrescens and Fusobacterium nucleatum were detected in these samples by the polymerase chain reaction (PCR). In addition, the distribution of fimA genotypes was assessed in P. gingivalis-positive individuals by PCR. RESULTS: P. gingivalis, T. forsythia, T. denticola, C. rectus and F. nucleatum were the most frequently detected species both in AgP probands and in their relatives. Kappa statistical analysis revealed that the detection of A. actinomycetemcomitans (Kappa = 0.503) and F. nucleatum (Kappa = 0.565) in probands was highly consistent with that in their relatives. Most probands shared the identical fimA genotype of P. gingivalis with their relatives. CONCLUSIONS: Our results suggested that the intrafamilial transmission of periodontal microorganisms may occur between Chinese patients with aggressive periodontitis and their relatives.