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Glucose fluctuations may contribute to large conductance calcium activated potassium (BK) channel dysfunction. However, the underlying mechanisms remain elusive. The aim of this study was to investigate the molecular mechanisms involved in BK channel dysfunction as a result of glucose fluctuations. A rat diabetic model was established through the injection of streptozotocin. Glucose fluctuations in diabetic rats were induced via consumption and starvation. Rat coronary arteries were isolated and coronary vascular tensions were measured after three weeks. Rat coronary artery smooth muscle cells were isolated and whole-cell BK channel currents were recorded using a patch clamp technique. Human coronary artery smooth muscle cells in vitro were used to explore the underlying mechanisms. After incubation with iberiotoxin (IBTX), the Δ tensions (% Max) of rat coronary arteries in the controlled diabetes mellitus (C-DM), the uncontrolled DM (U-DM) and the DM with glucose fluctuation (GF-DM) groups were found to be 84.46 ± 5.75, 61.89 ± 10.20 and 14.77 ± 5.90, respectively (P .05), while the current densities of the BK channels in the three groups were 43.09 ± 4.35 pA/pF, 34.23 ± 6.07 pA/pF and 17.87 ± 4.33 pA/pF, respectively (P .05). The Δ tensions (% Max) of rat coronary arteries after applying IBTX in the GF-DM rats injected with 0.9% sodium chloride (NaCl) (GF-DM + NaCl) and the GF-DM rats injected with N-acetyl-L-cysteine (NAC) (GF-DM + NAC) groups were found to be 8.86 ± 1.09 and 48.90 ± 10.85, respectively (P .05). Excessive oxidative stress and the activation of protein kinase C (PKC) α and nuclear factor (NF)-κB induced by glucose fluctuations promoted the decrease of BK-β1 expression, while the inhibition of reactive oxygen species (ROS), PKCα, NF-κB and muscle ring finger protein 1 (MuRF1) reversed this effect. Glucose fluctuations aggravate BK channel dysfunction via the ROS overproduction and the PKCα/NF-κB/MuRF1 signaling pathway.

Diabetes mellitus (DM) is an independent risk factor for atrial fibrillation, but the underlying ionic mechanism for this association remains unclear. We recently reported that expression of the small-conductance calcium-activated potassium channel 2 (SK2, encoded by KCCN2) in atria from diabetic mice is significantly down-regulated, resulting in reduced SK currents in atrial myocytes from these mice. We also reported that the level of SK2 mRNA expression is not reduced in DM atria but that the ubiquitin-proteasome system (UPS), a major mechanism of intracellular protein degradation, is activated in vascular smooth muscle cells in DM. This suggests a possible role of the UPS in reduced SK currents. To test this possibility, we examined the role of the UPS in atrial SK2 down-regulation in DM. We found that a muscle-specific E3 ligase, F-box protein 32 (FBXO-32, also called atrogin-1), was significantly up-regulated in diabetic mouse atria. Enhanced FBXO-32 expression in atrial cells significantly reduced SK2 protein expression, and siRNA-mediated FBXO-32 knockdown increased SK2 protein expression. Furthermore, co-transfection of SK2 with FBXO-32 complementary DNA in HEK293 cells significantly reduced SK2 expression, whereas co-transfection with atrogin-1ΔF complementary DNA (a nonfunctional FBXO-32 variant in which the F-box domain is deleted) did not have any effects on SK2. These results indicate that FBXO-32 contributes to SK2 down-regulation and that the F-box domain is essential for FBXO-32 function. In conclusion, DM-induced SK2 channel down-regulation appears to be due to an FBXO-32-dependent increase in UPS-mediated SK2 protein degradation.

Objectives We assessed the effectiveness and safety of cryoballoon ablation (CBA) in the Chinese population with paroxysmal atrial fibrillation (AF) with a 1-year follow-up and determined the association of early recurrence of atrial tachyarrhythmias (ERAT) with late recurrence (LR). Methods A total of 114 patients (age 61 ± 10 years, 78 males) with paroxysmal AF who underwent CBA were consecutively enrolled. After procedures, patients were observed for 3 days with continuous electrocardiogram monitoring in the hospital with routine follow-up visits at 3 months, 6 months, and 1 year. Documented atrial tachyarrhythmia >30 seconds was defined as recurrence. ERAT was defined as any recurrence during the first 3 months, and LR was recurrence between 3 and 12 months. Results With the first 3 months as blanking period, 76% of patients were free of LR at 12 months. Five patients (4%) experienced complications, including phrenic nerve palsy, stroke, and groin complications. Forty-five percent of patients had ERAT in the first 3 months and 31% of patients had ERAT in the first 3 days. Patients with ERAT had higher LR rate (LRR) than those without ERAT (43% vs 8%, P

The L-type calcium channel (LTCC) is one of the major ion channels that are known to be associated with the electrical remodeling of atrial fibrillation (AF). In AF, there is significant downregulation of the LTCC, but the underlying mechanism for such downregulation is not clear. We have previously reported that microRNA-499 (miR-499) is significantly upregulated in patients with permanent AF and that KCNN3, the gene that encodes the small-conductance calcium-activated potassium channel 3 (SK3), is a target of miR-499. We found that CACNB2, an important subunit of the LTCC, is also a target of miR-499. We hypothesize that miR-499 plays an important role in AF electrical remodeling by regulating the expression of CACNB2 and the LTCC. In atrial tissue from patients with permanent AF, CACNB2 was significantly downregulated by 67% (n = 4, p, Highlights • LTCC is downregulated with electrical remodeling of atrial fibrillation. • MiR-499 is increased and CACNB2 is reduced in atria from patients with atrial fibrillation. • MiR-499 binds to the 3′UTR of CACNB2 and inhibits its protein expression. • Downregulation of CACNB2 results in the downregulation of LTCC pore-forming subunit. • MiR-499 contributes to the electrical remodeling of AF through regulation of CACNB2.

Objective The aim of this study was to induce a rat model of oral submucous fibrosis (OSF) using bleomycin. Study Design A randomized trial was performed. Bleomycin dilution or phosphate-buffered saline was injected daily into the buccal mucosa of the test and control rats, respectively, for 2, 4, 6, or 8 weeks. Changes in histopathologic features, myofibroblasts, ultrastructure, and the levels of collagen type I and III, transforming growth factor–beta 1, and interferon-γ were measured. Results Compared with controls, the buccal mucosa of the test rats appeared pale and stiff. There was collagen deposition in the laminae propria. Type I and type III collagen increased steadily, transforming growth factor–beta 1 increased and then decreased, and interferon-γ was gradually reduced. The ultrastructure was also similar to that of OSF. Conclusions Using bleomycin, we induced a rat model of OSF that is very similar to human OSF in clinical manifestations, pathology, and ultrastructural changes.

To forecast the future trend of betel nut-associated oral cancer and the resulting burden on health based on historical oral cancer patient data in Hunan province, China.Oral cancer patient data in five hospitals in Changsha (the capital city of Hunan province) were collected for the past 12 years. Three methods were used to analyse the data; Microsoft Excel Forecast Sheet, Excel Trendline, and the Logistic growth model. A combination of these three methods was used to forecast the future trend of betel nut-associated oral cancer and the resulting burden on health.Betel nut-associated oral cancer cases have been increasing rapidly in the past 12 years in Changsha. As of 2016, betel nuts had caused 8,222 cases of oral cancer in Changsha and close to 25,000 cases in Hunan, resulting in about ¥5 billion in accumulated financial loss. The combined trend analysis predicts that by 2030, betel nuts will cause more than 100,000 cases of oral cancer in Changsha and more than 300,000 cases in Hunan, and more than ¥64 billion in accumulated financial loss in medical expenses.The trend analysis of oral cancer patient data predicts that the growing betel nut industry in Hunan province will cause a humanitarian catastrophe with massive loss of human life and national resources. To prevent this catastrophe, China should ban betel nuts and provide early oral cancer screening for betel nut consumers as soon as possible.

Aims Betel-quid (BQ) contains biologically psychoactive ingredients; however, data are limited concerning the symptoms and syndrome of BQ dependence among chewers. The aims of this study were to evaluate the ingredients-associated BQ dependence syndrome and country-specific chewing features and behaviour for BQ dependence among chewers from six Asian communities. Design An intercountry Asian Betel-quid Consortium study. Setting Six Asian general communities in Taiwan, Mainland China, Indonesia, Malaysia, Sri Lanka and Nepal. Participants Six multi-stage random samples of BQ chewers in the Asian Betel-quid Consortium study (n = 2078). Measurements All chewers were evaluated for BQ dependence using the DSM-IV and ICD-10 criteria. Findings The 12-month BQ dependence rate was 12.5–92.6% and 47.9–99.3% (P = 0.023) among tobacco-free and tobacco-added BQ chewers across the six Asian communities, with a higher dependence rate in chewers who used tobacco-free BQ with lime added than without (23.3–95.6% versus 4.0%, P ≤ 0.001). Taiwanese and Hunanese BQ chewers both notably endorsed the dependency domain of ‘time spent chewing’. ‘Tolerance’ and ‘withdrawal’ were the major dependence domains associated with the Nepalese and Indonesian chewers, with high BQ dependence rates. Malaysian and Sri Lankan chewers formed a BQ dependence cluster linked closely to ‘craving’. In Sri Lanka, the quantity consumed explained 90.5% (P

Tongue squamous cell carcinoma (TSCC) is one of the most common types of oral cancer; however, its molecular mechanisms remain unclear. In this study, methylated DNA immunoprecipitation (MeDIP) coupled with methylation microarray analysis was performed to screen for aberrantly methylated genes in adjacent normal control and TSCC tissues from 9 patients. Roche NimbleGen Human DNA Methylation 385K Promoter Plus CpG Island Arrays were used to detect 28,226 CpG sites. A total of 1,269 hypermethylated CpG sites covering 330 genes and 1,385 hypomethylated CpG sites covering 321 genes were found in TSCC tissue, compared to the adjacent normal tissue. Furthermore, we chose three candidate genes (FBLN1, ITIH5 and RUNX3) and validated the DNA methylation status by methylation-specific PCR (MS-PCR) and the mRNA expression levels by reverse transcription PCR (RT-PCR). In TSCC tissue, FBLN1 and ITIH5 were shown to be hypermethylated and their expression was found to be decreased, and RUNX3 was shown to be hypomethylated, however, its mRNA expression was found to be increased. In addition, another three genes (BCL2L14, CDCP1 and DIRAS3) were tested by RT-PCR. In TSCC tissue, BCL2L14 and CDCP1 expressions were markedly upregulated, and DIRAS3 expression was significantly downregulated. Our data demonstrated that aberrant DNA methylation is observed in TSCC tissue and plays an important role in the tumorigenesis, development and progression of TSCC.

J Oral Pathol Med (2012) 41: 748–754 Background: The development of betel-quid (BQ) processing industry in Hunan has increased the usage of this substance and may produce significant health consequences. However, the update data for the prevalence of BQ chewing and its effect on oral submucous fibrosis (OSF) are limited. Methods: A population-based survey was conducted to investigate the prevalence of BQ chewing and OSF in Hunan. A questionnaire interview and an oral examination were conducted. A multivariate logistic regression model was used to study the effect of socio-demographic characteristics on area-specific behaviors of BQ chewing. Results: A total of 2356 subjects participated in this survey, with a 78.5% of response rate. The lifetime prevalence of BQ chewing was 16.2% (2.9% for ex-chewers and 13.3% for current-chewers). The findings from multivariate logistic regression model showed that a higher prevalence of this habit was among individuals who were males, urban residents, 15–49 years old, educated for 9–12 years, smokers, and drinkers. Male chewers occupied a fairly higher percentage of the combined use of tobacco and/or alcohol (86.1–87.5%) than the non-chewers (50.6–56.7%). The overall prevalence of OSF was 1.0%. Compared with the subjects without OSF (1.7–23.2%), a significantly higher proportion of OSF patients were a current-chewer (62.3% in men and 100.0% in women). Conclusions: BQ chewing is a vital risk factor for OSF in Hunan. A high prevalence of BQ chewing found among the younger cohort (15–49 years old) is an urgent warning for this oral premalignant disorder.

Background: Myofibroblasts play an important role in the development of oral submucous fibrosis (OSF). In the current study, we investigate the effect of curcumin on growth and apoptosis of myofibroblasts derived from human oral mucosa. Methods: Myofibroblasts were generated by incubating fibroblasts, obtained from human oral mucosa, with transforming growth factor-β1 (TGF-β1). MTT, PI staining, and FACS assays were used to investigate curcumin’s effect on proliferation and cell cycle of fibroblasts and myofibroblasts. Annexin V/PI binding and FACS assays were used to examine apoptosis of myofibroblasts, Western blotting to determine the levels of Bcl-2 and Bax, and enzyme-linked immunosorbant assay was employed to examine the levels of collagen type I and III in the supernatants of myofibroblasts. Results: Curcumin inhibits proliferation of fibroblasts and myofibroblasts; it also disturbs the cell cycle, induces apoptosis and decreases the generation of collagen type I and III in myofibroblasts, which are more sensitive to its effects than fibroblasts. Curcumin induces apoptosis in myofibroblasts by down-regulating the Bcl-2/ Bax ratio. Conclusion: Our results demonstrate the antifibrotic effect of curcumin in vitro. It may therefore be a candidate for the treatment of OSF.

The small conductance Ca(2+)-activated K(+) (SK) channels have recently been found to be expressed in the heart, and genome-wide association studies have shown that they are implicated in atrial fibrillation. Diabetes mellitus is an independent risk factor of atrial fibrillation, but the ionic mechanism underlying this relationship remains unclear. We hypothesized that SK channel function is abnormal in diabetes mellitus, leading to altered cardiac electrophysiology. We found that in streptozotocin-induced diabetic mice, the expression of SK2 and SK3 isoforms was down-regulated by 85 and 92%, respectively, whereas that of SK1 was not changed. SK currents from isolated diabetic mouse atrial myocytes were significantly reduced compared with controls. The resting potentials of isolated atrial preparations were similar between control and diabetic mice, but action potential durations were significantly prolonged in the diabetic atria. Exposure to apamin significantly prolonged action potential durations in control but not in diabetic atria. Production of reactive oxygen species was significantly increased in diabetic atria and in high glucose-cultured HL-1 cells, whereas exposure of HL-1 cells in normal glucose culture to H2O2 reduced the expression of SK2 and SK3. Tyrosine nitration in SK2 and SK3 was significantly increased by high glucose culture, leading to accelerated channel turnover. Treatment with Tiron prevented these changes. Our results suggest that increased oxidative stress in diabetes results in SK channel-associated electrical remodeling in diabetic atria and may promote arrhythmogenesis.

Background Lactotransferrin (LTF) has been confirmed to act as a tumor suppressor in multiple cancers; however, its roles in oral squamous cell carcinoma (OSCC), one of malignant head and neck carcinomas, has not been explored. Methods Here, the expression of LTF in OSCC tissues and TCA8113 cells was detected with RT-PCR, qPCR, and IHC. And the correlation between LTF expression and OSCC metastasis was assessed. MS-PCR was performed to reveal the methylation status in promoter regions of LTF both in OSCC tissue samples and cells. The influences of 5-Aza-Cdc treatment to the methylation status and expression levels of LTF were also analyzed. At last, the functions of LTF in OSCC progression were demonstrated by MTT analysis, clone formation assay, and cell cycle analysis in TCA8113 cells with forced ectopic expression of LTF. Results LTF showed a low or null expression pattern in OSCC tissues and cells, at least partially, due to the hypermethylated status in promoter regions for 5-Aza-Cdc, a methyltransferase inhibitor, could restore the expression of LTF in TCA8113 cells. And the expression level of LTF exhibited a negative correlation with OSCC metastasis. Conclusions Re-expression of LTF inhibited the growth, proliferation, as well as cell cycle progression of TCA8113 cells. In conclusion, hypermethylation contributes much to LTF inactivation in OSCC. And LTF can partially reverse the malignant phenotypes of OSCC cells and may be served as a potential target for diagnosis and therapy of OSCC in future.

To study the effect of Erigeron Breviscapus (EB) at different concentrations and different intervention time points on the mRNA and protein expression of OPG/RANKL/RANK in MG63 osteoblast-like cells and RAW264. 7 pre-osteoclast cells cultured in vitro, thus exploring roles EB played in bone rebuilding and its mechanisms.MG63 osteoblast-like cells and RAW264.7 pre-osteoclast cells were cultured in vitro. The 3rd passage cells were divided into the control group and different experimental groups. Total RNA and protein were respectively isolated from cells treated with different concentrations of EB (0, 0.001, 0.01, 0.1, and 1.0 mg/mL) for 48 h. Meanwhile, the protein was extracted from 0 and 1 mg/mL EB groups at 12, 24, and 48 h respectively. Expression of OPG mRNA and RANKL mRNA in MG63 osteoblast-like cells, and expression of RANK mRNA in RAW264.7 pre-osteoclast cells were detected by semi-quantitative RT-PCR. Expression of OPG protein and RANKL protein in MG63 osteoblast-like cells, and expression of RANK protein in RAW264. 7 pre-osteoclast cells were detected by Western blot.Along with increased EB concentration, expression of OPG mRNA and protein in MG63 osteoblast-like cells was gradually lowered (P0.05) after 48-h intervention of EB, the expression of RANKL mRNA and protein in MG63 osteoblast-like gradually increased (P0.05); the expression of RANK mRNA in RAW264.7 pre-osteoclast cells increased (P0.05). But the expression of RANK mRNA was slightly lower in the 0.1 mg/mL EB group than in the 0.01 mg/mL EB group, and the expression of RANK protein in RAW264.7 pre-osteoclast cells gradually increased (P0.05). After treatment with 1 mg/mL EB for 12, 24, 48 h, the expression of OPG protein in MG63 osteoblast-like cells gradually decreased as time went by (P0.05), and the expression of RANKL protein in MG63 osteoblast-like and RANK protein in RAW264.7 pre-osteoclast cells gradually increased (P0.05). The expression of RANKL protein in RAW264.7 pre-osteoclast cells increased as time went by (P0.05).EB could inhibit the expression of OPG in osteoblasts in a dose- and time-dependent manner, promote the expression of RANKL in osteoblasts and the secretion of RANK in pre-osteoclast, indicating EB might play roles in promoting bone resorption.

Oral submucous fibrosis is a high risk precancerous condition and is suggested to be caused by areca nut chewing. Areca nut chewing is popular in Hunan Province of China, and is more concentrated in Xiangtan City. Two and nine cases of oral submucous fibrosis (OSF) were first noticed in 1984 and 1985 respectively, and an epidemiologic survey was subsequently performed in 1986. The epidermiologic method of cluster sampling was used. The Yuhu District, one of the five urban districts of the Xiangtan City with a population of 100,000 was selected as a whole body in the survey, 57 independent units of various professions were randomly selected as group samples and more than 70% of subjects in each unit were examined. Definite fibrous band on palpation was used as a main diagnostic criterion for OSF. A total of 11046 individuals were examined; among them were 3907 areca nut chewers (35.37%) and 7139 non-chewers (64.63%). 335 cases of OSF were found, comprising a prevalence rate of 3.03%. The disease involved mainly the middle third of the oral cavity. All of the OSF cases were areca nut chewers. No case had been found in non-chewers. Four cases of oral carcinoma were found on the basis of OSF, the malignant transformation rate was 1.19%. The high prevalence of OSF may be due to areca nut chewing plus extensive and heavy use of hot pepper in Xiangtan people. The result supports the role of the areca nut as an etiologic factor in the development of OSF. The low malignant transformation rate of 1.19% compared with the 7.6% in an Indian report may be because Xiangtan people chew areca nut without tobacco.

Betel-quid (BQ) contains biologically psychoactive ingredients; however, data are limited concerning the symptoms and syndrome of BQ dependence among chewers. The aims of this study were to evaluate the ingredients-associated BQ dependence syndrome and country-specific chewing features and behaviour for BQ dependence among chewers from six Asian communities.An intercountry Asian Betel-quid Consortium study.Six Asian general communities in Taiwan, Mainland China, Indonesia, Malaysia, Sri Lanka and Nepal.Six multi-stage random samples of BQ chewers in the Asian Betel-quid Consortium study (n = 2078).All chewers were evaluated for BQ dependence using the DSM-IV and ICD-10 criteria.The 12-month BQ dependence rate was 12.5-92.6% and 47.9-99.3% (P = 0.023) among tobacco-free and tobacco-added BQ chewers across the six Asian communities, with a higher dependence rate in chewers who used tobacco-free BQ with lime added than without (23.3-95.6% versus 4.0%, P ≤ 0.001). Taiwanese and Hunanese BQ chewers both notably endorsed the dependency domain of 'time spent chewing'. 'Tolerance' and 'withdrawal' were the major dependence domains associated with the Nepalese and Indonesian chewers, with high BQ dependence rates. Malaysian and Sri Lankan chewers formed a BQ dependence cluster linked closely to 'craving'. In Sri Lanka, the quantity consumed explained 90.5% (P 0.001) of the excess dependence risk for tobacco-added use, and could be a mediator between tobacco-derived psychoactive effect and BQ dependence development.DSM-IV criteria for dependence apply to a significant proportion of betel quid users in Asian communities, more so if they use it with tobacco or lime.

To examine the expression of cytochrome P450 related genes in oral submucous fibrosis tissue and to investigate the possible role of the genes in pathogenesis of oral submucous fibrosis (OSF).Buccul mucosa tissues were obtained from OSF patients in early, medium and advanced stages, with each stage including 10 patients. Normal buccul mucosa tissues were collected from 10 patients undergoing oral and maxillofacial surgery as control. Oral submucous fibrosis-related genes were analysed by cDNA chips, and the results were submitted to the gene network database. Differentially expressed genes related to the pathway of CYP metabolism were indentifyed by the database analysis. Reverse transcription-polymerase chain reaction (RT-PCR) was used to verify the results from cDNA chips by increasing sample volume.There were eight genes [CYP2B6, CYP2C18, CYP2F1, CYP3A5, microsomal glutathione S-transferase 2 (MGST2), alcohol dehydrogenase (ADH), UDP glucuronosyl transferase 2B15 (UGT2B15), ADH1C] which were related to the pathway of CYP metabolism. These genes were low expressed in all stages of OSF (P0.001).There were no differences in genes expression among the three stages of OSF (P0.05).There were down-regulated genes related to the pathway of CYP metabolism in oral submucous fibrosis tissue. The ability of the pathway of CYP to metabolize and clear betel nut ingredients was reduced in OSF patients, which may play a role in the pathogenesis of OSF.

MicroRNAs are important regulators of gene expression, including those involving electrical remodeling in atrial fibrillation (AF). Recently, KCNN3, the gene that encodes the small-conductance calcium-activated potassium channel 3 (SK3), was found to be strongly associated with AF.To evaluate the changes in atrial myocardial microRNAs in patients with permanent AF and to determine the role of microRNA on the regulation of cardiac SK3 expression.Atrial tissue obtained during cardiac surgery from patients (4 sinus rhythm and 4 permanent AF) was analyzed by using microRNA arrays. Potential targets of microRNAs were predicted by using software programs. The effects of specific microRNAs on target gene expression were evaluated in HL-1 cells from a continuously proliferating mouse hyperplastic atrial cardiomyocyte cell line. Interactions between microRNAs and targets were further evaluated by using luciferase reporter assay and by using Argonaute pull-down assay.Twenty-one microRNAs showed significant (2-fold) changes in AF. MicroRNA 499 (miR-499) was upregulated by 2.33-fold (P.01) in AF atria, whereas SK3 protein expression was downregulated by 46% (P.05). Transfection of miR-499 mimic in HL-1 cells resulted in the downregulation of SK3 protein expression, while that of miR-499 inhibitor upregulated SK3 expression. Binding of miR-499 to the 3' untranslated region of KCNN3 was confirmed by luciferase reporter assay and by the increased presence of SK3 mRNA in Argonaute pulled-down microRNA-induced silencing complexes after transfection with miR-499.Atrial miR-499 is significantly upregulated in AF, leading to SK3 downregulation and possibly contributing to the electrical remodeling in AF.

To explore the effect of DNAX-associated protein 12 (DAP12) pathway on the transformation from mouse monocytes RAW264.7 to osteoclasts induced by tensile strain.DAP12shRNA plasmid was constructed and introduced to RAW264.7 cells. Then we supplied tensile strain to RAW264.7 cells by four-point bending system. The mRNA or protein expression of DAP12, tartrate-resistant acid phosphatase (TRAP), tyrosine kinases Btk and Tec and nuclear facior of activated T cells 1 (NFATc1) was measured by reverse transcription PCR (RT-PCR) and Western blotting respectively.The expression of DAP12 mRNA (0.112 ± 0.025) and protein (0.193 ± 0.015) both declined sharply after plasmid being introduced into monocytes RAW264.7 (P0.05). After silencing DAP12 expression in RAW264.7 cells by RNA interference, tensile strain-induced TRAP mRNA expression of RAW264.7 cells increased at 6 h (0.671 ± 0.031) and 12 h (0.800 ± 0.043) (P0.05), but it was weaker than non-RNA-interference-groups at each time point (P0.05). After silencing DAP12 expression in RAW264.7 cells by RNA interference, the expressions of Btk, Tec, NFATc1 increased as time passed (6, 12 h) (P0.05), but the expressions on corresponding time decreased sharply compared with those in control groups (P0.05).DAP12 pathway play an important role in regulating osteoclast differentiation induced by tensile strain.

Oral squamous cell carcinoma (OSCC) is the most common type of head and neck malignant tumor. however, its pathological mechanisms have not yet been elucidated. In the present study, we screened for candidate tumor-suppressor genes (TSGs) related to OSCC among 10 candidate genes located in 3p21.3, a region abundant with TSGs based on previous studies, using semi-quantitative reverse transcription PCR (RT-PCR). Three genes, GNAT1, SEMA3B and AXUD1, with low or no expression in OSCC tissues and the cell line TCA8113 were selected, and the promoter methylation status was further analyzed by methylation-specific PCR (MS-PCR). Hypermethylation in the promoter regions of SEMA3B was found in OSCC tissues, and a significant difference in the frequency of methylation of SEMA3B was observed between OSCC and non-cancerous tissues. Furthermore, TCA8113 cells treated with 5-Aza-Cdc started to re-express SEMA3B at a concentration of 5 µM or higher. Our study confirmed that three candidate TSGs with low expression may be involved in OSCC and that hypermethylation in promoter regions may contribute to the low expression of SEMA3B. These findings offer novel insights for clarifying the molecular mechanisms of tumorigenesis of OSCC as well as for aiding in its clinical diagnosis and therapeutic strategy.

The development of betel-quid (BQ) processing industry in Hunan has increased the usage of this substance and may produce significant health consequences. However, the update data for the prevalence of BQ chewing and its effect on oral submucous fibrosis (OSF) are limited.A population-based survey was conducted to investigate the prevalence of BQ chewing and OSF in Hunan. A questionnaire interview and an oral examination were conducted. A multivariate logistic regression model was used to study the effect of socio-demographic characteristics on area-specific behaviors of BQ chewing.A total of 2356 subjects participated in this survey, with a 78.5% of response rate. The lifetime prevalence of BQ chewing was 16.2% (2.9% for ex-chewers and 13.3% for current-chewers). The findings from multivariate logistic regression model showed that a higher prevalence of this habit was among individuals who were males, urban residents, 15-49 years old, educated for 9-12 years, smokers, and drinkers. Male chewers occupied a fairly higher percentage of the combined use of tobacco and/or alcohol (86.1-87.5%) than the non-chewers (50.6-56.7%). The overall prevalence of OSF was 1.0%. Compared with the subjects without OSF (1.7-23.2%), a significantly higher proportion of OSF patients were a current-chewer (62.3% in men and 100.0% in women).BQ chewing is a vital risk factor for OSF in Hunan. A high prevalence of BQ chewing found among the younger cohort (15-49 years old) is an urgent warning for this oral premalignant disorder.

To compare the shaping ability and the influence on apical foramen among hand ProTaper, stainless steel K-files and rotary ProTaper in preparing different curved root canal.Forty simulated resin root canal blocks were randomly divided into four groups and prepared by hand ProTaper, stainless steel K-file and rotary ProTaper, respectively. Of them, 12 blocks in group A, B, C consist of six 200 curved root canals and six 30 degrees curved root canals each group. The curvature of the other 4 blocks in group D was less than 5 degrees. Taking photos of the models to the root canal orthotopically and apical foramen using digital camera before and after instrumentation. Finally, the transportation of root canal and the size of apical foramen were analyzed using special image software Auto-CAD.The transportation of center in group B was the highest than that in group A and group C (P0.05). In some portions of root canal, the transportation of center in group C was higher than that in group A. The size of the apical foramen in group B was significantly bigger than the other groups and the size of the apical foramen in 30 degrees root canal was significantly bigger than that in 20 degrees root canal after instrumentation (P0.05). There was no significantly different between group A and group C, though the size of apical foramen in group C was bigger than that in group A at the same curvature, and that in 30 degrees root canal was bigger than in 20 degrees root canal (P0.05).Both of the two instruments engender root canal transportation, and curvature is the main reason of transportation. Comparing with stainless steel K-files, NiTi files can maintain the shape of the root canal and apical foramen well.

To investigate the effects of arecoline and nicotine on the expression of human telomerase reverse transcriptase (hTERT) mRNA and protein in cultured normal human oral keratinocytes (KC).The experiments were divided into arecoline group, arecoline/nicotine group and control group. The hTERT mRNA and protein expression of KC was examined by reverse transcription polymerase chain reaction (RT-PCR) and Western blot.Arecoline could induce the hTERT mRNA and protein expression of KC in a dose dependent manner, the hTERT mRNA and protein expression of KC was higher in 0.030, 0.060, 0.090 g/L arecoline group than control group (P0.001). Nicotine (0.025 g/L) increased hTERT mRNA and protein expression of KC induced by arecoline.Arecoline could increase the expression of hTERT mRNA and protein in oral keratinocytes. Nicotine had a synergistic effect on arecoline. hTERT over-expression induced by arecoline and nicotine may play an important role in the malignant transformation of oral submucous fibrosis.

To detect the effect of erigeron breviscapus on the expression of vascular endothelial growth factor (VEGF) in the periodontal tissues during orthodontic tooth movement.45 rabbits were divided into 3 groups (groups A, B and C). Groups A and B included experimental group of 1, 3, 7 and 14 days respectively. The mandibular first molar of each experimental rabbit was observed. The rabbits of group A and group B received iontophoresis with erigeron breviscapus into the right (group A-R and group B-R) and with normal sodium into the left as the control (group A-L and group B-L). Additionally, the rabbits of group B were designed orthodontic appliance, by which 0.78 N mesial force was applied to pull the mandibular first molars. Group C, group of 0 day, was no iontophoresis and orthodontic appliance as the control. After killed on schedule, the amount of experimental tooth movement was measured and the expression of VEGF was examined by immunohistochemical method.The amount of experimental tooth movement increased successively from 1 to 14 days. The differences among days 3, 7 and 14 were significant in the comparison between group B-R and group B -L (P0.01). The expression of VEGF in groups A-R and B-L enhanced apparently compared with that in groups C and A-L (P0.01), but that in group B-R was the most apparent (P0.01). The expression of VEGF reached the peak level on day 3 in groups A-R and B-R (P0.01), but it reached the peak level on day 7 in group B-L (P0.01).Erigeron breviscapus by iontophoresis can accelerate orthodontic tooth movement, and can meanwhile up-regulate the expression of VEGF in periodontium in the earlier period of orthodontic tooth movement. Thus it can be presumed that one of its mechanisms for erigeron breviscapus to accelerate orthodontic tooth movement is erigeron breviscapus effects the metabolism and differentiation of osteoblast and osteoclast through up-regulating the expression of VEGF in periodontium.

To prepare the magnetic pingyangmycine-bearing microspheres (MPMs), test its characteristics, and evaluate the target ability of MPMs.MPMs were prepared by a cold-condensation method at first. Then the scan electron microscope, high performance liquid chromatography (HPLC) and atomic absorption spectrometry were used to measure the diameter of MPMs and the contents of PYM and Fe3O4 in MPMs. Drug-releasing instrument was used to test the quantity of PYM released from MPMs. The distribution of intravenously injected MPMs in SD rat liver was investigated under an externally magnetic field.MPMs are spherical with the mean diameter of 32.23 microns. The content of pingyangmycine in MPMs was 3.8% and Fe3O4 was 11.7% (w/w). The quantity of PYM released from MPMs in 3 hours was 63.1%. The content of Fe3O4 in the targeting sites of rats was obviously higher than that in the non-targeting sites (P0.01).MPMs have good properties of drug-targeting and drug sustain releasing. MPMs may be ideal for the use of the treatment of cavernous hemangioma.

To investigate the probability and mechanism magnetic pingyangmycine-bearing microspheres(MPMs) in the treatment of cavernous hemangioma.Twenty rabbits were divided into four groups. MPMs, pingyangmycine and magnetic nonpingyangmycine-bearing microspheres were injected into marginal ear veins respectively. All animals were killed after 21 days and histological varations of the veins were observed under the light microscope and transmission electron microscope.The occlusion of the veins in the group of MPMs with the external magnetic field was more prominent than that in the other groups.MPMs can occlude veins by means of its embolized effect and PYM drug effect. MPMs may be better used in the therapy of cavernous hemangioma.