[A preliminary in vivo and in vitro study of endothelial cell pyroptosis in the periodontal inflammatory environment].

Objective: To observe whether endothelial cells undergo pyroptosis in the inflammatory periodontal environment by using a model in vivo and in vitro, providing an experimental basis for indepth understanding of the underlying pathogenesis of periodontitis. Methods: According to the classification of periodontal diseases of 2018, gingival tissues were collected from periodontally healthy subjects and patients with stage Ⅲ-Ⅳ, grade C periodontitis, who presented Department of Oral and Maxillofacial Surgery and Department of Periodontology, School of Stomatology, The Fourth Military Medical University from April to May 2022. Immunohistochemical staining was performed to detect the expression level and distribution of gasdermin D (GSDMD), a hallmark protein of cell pyroptosis, in gingival tissues. Periodontitis models were established in each group by ligating the maxillary second molar teeth of three mice for 2 weeks (ligation group). The alveolar bone resorption was determined by micro-CT (mice without ligation treatment were used as the control group), and the colocalization of GSDMD and CD31 were quantitatively analyzed by immunofluorescence staining in gingival tissues of healthy and inflammatory mice. Human umbilical vein endothelial cells (HUVECs) were cultured in vitro and treated with lipopolysaccharide (LPS) of Porphyromonas gingivalis (Pg) combined with adenosine triphosphate (ATP) at various concentrations of 0.5, 1.0, 2.5, 5.0, and 10.0 mg/L, respectively, and the 0 mg/L group was set as the control group at the same time. Scanning electron microscopy was used to observe the morphology of HUVECs. Western blotting was used to detect the expression of gasdermin D-N terminal domains (GSDMD-N) protein and immunofluorescence cell staining was used to detect the expression and distribution of GSDMD. Cell counting kit-8 (CCK-8) was used to detect the proliferative ability of HUVECs, and propidium iodide (PI) staining was used to detect the integrity of cell membrane of HUVECs. Results: Immunohistochemistry showed that GSDMD in gingival tissues of periodontitis was mainly distributed around blood vessels and its expression level was higher than that in healthy tissues. Micro-CT showed that alveolar bone resorption around the maxillary second molar significantly increased in ligation group mice compared with control subjects (t=8.88, P<0.001). Immunofluorescence staining showed significant colocalization of GSDMD with CD31 in the gingival vascular endothelial cells in mice of ligation group. The results of scanning electron microscopy showed that there were pores of different sizes, the typical morphology of pyroptosis, on HUVECs cell membranes in the inflammatory environment simulated by ATP combined with different concentrations of LPS, and 2.5 mg/L group showed the most dilated and fused pores on cell membranes, with the cells tended to lyse and die. Western blotting showed that the expression of GSDMD-N, the hallmark protein of cell pyroptosis, was significantly higher in 2.5 and 5.0 mg/L groups than that in the control group (F=3.86, P<0.01). Immunofluorescence cell staining showed that the average fluorescence intensity of GSDMD in 2.5 mg/L group elevated the most significantly in comparison with that in the control group (F=35.25, P<0.001). The CCK-8 proliferation assay showed that compared to the control group (1.00±0.02), 0.5 mg/L (0.52±0.07), 1.0 mg/L (0.57±0.10), 2.5 mg/L (0.58±0.04), 5.0 mg/L (0.55±0.04), 10.0 mg/L (0.61±0.03) groups inhibited cell proliferation (F=39.95, P<0.001). PI staining showed that the proportion of positive stained cells was highest [(56.07±3.22)%] in 2.5 mg/L group (F=88.24, P<0.001). Conclusions: Endothelial cells undergo significant pyroptosis in both in vivo and in vitro periodontal inflammatory environments, suggesting that endothelial cell pyroptosis may be an important pathogenic factor contributing to the pathogenesis of periodontitis.

[Trend of hypertension death and its impact on life expectancy in 6 northwestern provinces of China, 2013-2021].

Objective: To analyze the mortality trend of hypertension in Xinjiang Uygur Autonomous Region, Qinghai Province, Shaanxi Province, Gansu Province, Ningxia Hui Autonomous Region and Inner Mongolia Autonomous Region (6 provinces) in northwestern China, from 2013 to 2021, and evaluate the influence of hypertension on people's life expectancy. Methods: Based on national death surveillance data and demographic data in the 6 provinces from 2013 to 2021, the mortality rate, standardized mortality rate, life expectancy, cause eliminated life expectancy (CELE), potential gains in life expectancy (PGLEs) and life loss rate of hypertension were calculated. Software Joinpoint was used to analyze the mortality trends and calculate average annual percentage change (AAPC) and annual percentage change (APC) in hypertension deaths. Results: From 2013 to 2021, the overall standardized mortality rate of hypertension in the 6 provinces showed a downward trend (AAPC=-1.82%, P=0.050). The mortality rate in rural area was always higher than that in urban area, and showed an increasing trend after 2016 (APC=4.74%, P=0.003), and the mortality rate in men was always higher than that in women. The incidence trend of deaths of different types of hypertension were different, and the deaths caused by hypertensive heart disease accounted for the highest proportion (72.69%). In 2021, the life expectancy of the population in the 6 provinces increased by 1.01 years, the CELE increased by 0.93 years, the PGLEs decreased by 0.08 years, and the life loss rate decreased by 0.11% compared with 2013. Conclusions: The overall standardized mortality rate of hypertension showed a decreasing trend in the 6 northwestern provinces from 2013 to 2021, but it showed an increasing trend in rural area after 2016. Prevention of hypertension should be further strengthened in rural area, men and elderly population.

[Depression status of elderly patients with metabolic syndrome in three provinces of China].

Objective: To understand the depression status and its influencing factors in elderly patients with MS in China and to explore the correlation between various components of elderly MS and depression. Methods: This study is based on the "Prevention and Intervention of Key Diseases in Elderly" project. We used a multi-stage stratified cluster random sampling method to complete 16 199 elderly aged 60 years and above in 16 counties (districts) in Liaoning, Henan, and Guangdong Provinces in 2019, excluding 1 001 missing variables. Finally, 15 198 valid samples were included for analysis. The respondents' MS disease was obtained through questionnaires and physical examinations, and the respondents' depression status within the past half month was assessed using the PHQ-9 Depression Screening Scale. The correlation between elderly MS and its components and depression and its influencing factors were analyzed by logistic regression. Results: A total of 15 198 elderly aged 60 years and above were included in this study, with the prevalence of MS at 10.84% and the detection rate of depressive symptoms in MS patients at 25.49%. The detection rates of depressive symptoms in patients with 0, 1, 2, 3, and 4 MS abnormal group scores were 14.56%, 15.17%, 18.01%, 25.21%, and 26.65%, respectively. The number of abnormal components of MS was positively correlated with the detection rate of depressive symptoms, and the difference between groups was statistically significant (P<0.05). The risk of depression symptoms in patients with MS, overweight/obesity, hypertension, diabetes, and dyslipidemia was 1.73 times (OR=1.73, 95%CI:1.51-1.97), 1.13 times (OR=1.13, 95%CI:1.03-1.24), 1.25 times (OR=1.25, 95%CI:1.14-1.38), 1.41 times (OR=1.41, 95%CI:1.24-1.60), 1.81 times (OR=1.81,95%CI:1.61-2.04), respectively, more than those without the disease. Multivariate logistic regression analysis showed that the detection rate of depressive symptoms in patients with sleep disorders was higher than that with normal sleep (OR=4.89, 95%CI: 3.79-6.32). The detection rate of depressive symptoms in patients with cognitive dysfunction was 2.12 times higher than that in the average population (OR=2.12, 95%CI: 1.56-2.89). The detection rate of depressive symptoms in patients with impaired instrumental activities of daily living (IADL) was 2.31 times (OR=2.31, 95%CI: 1.64-3.26) higher than that in the average population. Tea drinking (OR=0.73, 95%CI: 0.54-0.98) and physical exercise (OR=0.67, 95%CI: 0.49-0.90) seemed to be protective factors for depression in elderly MS patients (P<0.05). Conclusions: Older patients with MS and its component abnormalities have a higher risk of depression than the average population. Sleep disorders, cognitive impairment, and IADL impairment are important influencing factors for depression in elderly MS patients, while tea drinking and physical exercise may help to reduce the risk of the disease.

[Analysis on liver cancer mortality and cause eliminated life expectancy in key areas of 4 provinces, China, 2008-2018].

Objective: To explore the changes of liver cancer mortality and the effect of liver cancer on life expectancy in key areas of four provinces in China from 2008 to 2018 and provide the basis for the evaluation of comprehensive prevention and control of cancer and promotion of the rational allocation of health resources. Methods: Based on the national cause-of-death surveillance in key areas of the 4 provinces from 2008 to 2018, we analyzed the mortality of liver cancer, cause eliminated life expectancy (CELE) and potential gains in life expectancy (PGLEs). Software Joinpoint 4.9.0.0 was used to calculate the average annual percentage change (AAPC). Arriaga's decomposition method was used to estimate the contribution of the changes of liver cancer mortality in each age group to life expectancy. Results: The standardized mortality of liver cancer in key areas of the 4 provinces showed a downward trend from 2008 to 2018 (AAPC=-4.37%, P<0.001). The changes of liver cancer mortality had a positive effect on the increase of life expectancy, with a contribution value of 0.240 years and a contribution degree of 5.62%. The positive effect was greatest in age group 45-49 years (0.041 years, 0.96%), and the negative effect was greatest in age group 50-54 years (-0.015 years, -0.35%). Compared with 2008, the life expectancy increased by 4.27 years (AAPC=0.59%, P<0.001), the liver cancer CELE increased by 4.20 years (AAPC=0.58%, P<0.001), the PGLEs decreased by 0.07 years (AAPC=-0.62%,P<0.001), and life loss rate decreased by 0.13% (AAPC=-1.18%, P=0.001). The liver cancer PGLEs increased in Yongqiao district, Anhui province (0.09 years), and decreased in other districts (counties), with the largest decline was in Fugou county, Henan province (-0.21 years). Conclusions: From 2008 to 2018, the standardized mortality rate of liver cancer in key areas of the 4 provinces decreased gradually, contributing to the growth of life expectancy. The life loss caused by liver cancer decreased gradually, but the PGLEs varied with districts (counties).

[Distribution and risk factors of self-reported cancer cases in adults in China, 2015].

Objective: To analyze the prevalence and risk factors of self-reported cancer in adults in China in 2015. Methods: The data used in this study were from China Chronic Disease and Risk Factors Surveillance in 2015. The frequency and proportion of the classified variables were analyzed by descriptive statistics, the disordered classified variables were compared by χ2 test, and the possible risk factors of cancer patients were screened by univariate and multivariate logistic regression analyses. Results: In 2015, there were 1 809 self-reported tumors patients in China, including 689 males (0.63%), 1 120 females (1.03%), 769 (0.71%) in the eastern region, 465 (0.43%) in the central region and 575 (0.53%) in the western region. The patients were mainly distributed in people aged 45- and 55- years old, being overweight or obese, living in eastern urban area, having low education level, being married, having low annual household income and being occupational population. The results of multivariate logistic regression showed that compared with the western region, the prevalence rate of cancer was higher in the eastern region (OR=1.05, 95%CI: 1.04-1.06), while lower in the central region (OR=0.94, 95%CI: 0.93-0.95); the risk for cancer in people with family history of malignancy was higher than that in people without family history of malignancy (OR=1.95, 95%CI:1.94-1.96) the risk for cancer in people with an annual household income of less than 10 000 yuan or between 10 000 and 50 000 yuan was higher than that in people with an annual household income of more than 50 000 yuan (<10 000 yuan: OR=1.59, 95%CI: 1.58-1.60; between 10 000 and 50 000 yuan: OR=1.27, 95%CI: 1.26-1.28); and the risk for cancer in people living urban areas was lower than that in people living in rural areas (OR=0.98, 95%CI: 0.97-0.99). In terms of personal behavior and diet, the risk for cancer in smokers was 1.25 times higher than that in non-smokers (OR=1.25, 95%CI: 1.24-1.26), and the risk for cancer in alcoholics was 1.16 times higher than that in non-alcoholics (OR=1.16, 95%CI: 1.15-1.17), the risk for cancer in people with insufficient vegetable and fruit intakes was 1.29 times and 1.03 times higher than those in people with sufficient intakes of vegetables and fruits, respectively (OR=1.29, 95%CI: 1.28-1.30;OR=1.03,95%CI: 1.02-1.04). People with low frequency of high-intensity exercise had a higher risk for cancer compared with those with high frequency of high-intensity exercise (OR=1.32, 95%CI: 1.31-1.33), the risk for cancer was higher in people with low frequency of moderate exercise than in people with high frequency of moderate exercise (OR=1.08, 95%CI: 1.07-1.09). The risk for cancer in people with sedentary time less than 2 hours was higher than that in those with sedentary time more than 2 hours (OR=1.69, 95%CI: 1.68-1.70), and the risk for cancer in people who ate moderate amount of red meat was lower than that in people who ate excessive amount of red meat (OR=0.86, 95%CI: 0.85-0.87). Conclusions: The number of female self-reported cancer was more than that in males, and the number of self-reported cancer in the eastern region was higher than that in the central and western regions. Living in eastern region, with family history of malignancy, having low annual household income, smoking, drinking, insufficient vegetable intake, insufficient fruit intake and low frequency of high-intensity exercise and low frequency of moderate intensity exercise were the main risk factors for cancer, while living in central region, living in urban area and low red meat intake were protective factors.

[Mesenchymal stem cells derived apoptotic extracellular vesicles attenuate pro-inflammatory macrophages induced by Porphyromonas gingivalis lipopolysaccharide].

Objective: To investigate whether bone marrow mesenchymal stem cells (BMMSCs) derived apoptotic extracellular vesicles (ApoEVs) could regulate the polarization of mouse macrophage cell line RAW264.7 and whether BMMSCs derived ApoEVs could attenuate pro-inflammatory condition of RAW264.7 induced by Porphyromonas gingivalis lipopolysaccharide (Pg-LPS), so as to provide experimental evidence and theoretical basis for using BMMSCs derived ApoEVs as a method to treat periodontitis. Methods: The Operetta CLS high-content analysis system was used to observe the time-dependent apoptosis process of BMMSCs. Besides, field emission scanning electron microscopy (FESEM), dynamic light scattering technology and streaming potential method were used to measure the surface characteristics of BMMSCs derived ApoEVs. The Operetta CLS high-content analysis system was used to observe the process of RAW264.7 phagocyting 5-carboxy-tetramethylrhodamine, succinimidyl ester (5-TAMRA-SE) labeled ApoEVs. Real-time quantitative PCR was used to detect the mRNA expression of arginase-1 (Arg-1). Cell immunofluorescence and Western blotting were used to detect the number of inducible nitric oxide synthase (iNOS)(+) macrophages and iNOS protein expression level in each experiment group. Enzyme linked immunosorbent assay was used to detect tumor necrosis factro-α (TNF-α) level in the Pg-LPS induced pro-inflammatory macrophage culture supernatant in each experiment group. Results: After treating with 0.5 µmol/L staurosporine for 12 hours, mouse BMMSCs underwent shrinking with obvious vesicles structure around. The FESEM showed the ApoEVs were in spherical shapes. The size range of ApoEVs was about 100-1 000 nm and the average Zeta potential was -16.6 mV. The Operetta CLS high-content analysis system showed RAW264.7 could phagocytose 5-TAMRA-SE labeled ApoEVs by pseudopodia. The relative mRNA expression of Arg-1 was significantly increased in RAW 264.7 after being treated with interleukin 4 (IL-4) and ApoEVs (261.97±15.91) compared to that with IL-4 alone (115.29±15.42) (P<0.01). Cell immunofluorescence showed that ApoEVs could reduce the number of iNOS(+) macrophages induced by Pg-LPS (39.33±4.70) comparing to those without ApoEVs (95.33±4.70) (P=0.007). In the meanwhile, ApoEVs could also down-regulate the iNOS protein level of macrophages induced by Pg-LPS (5.84±1.05) comparing to those without ApoEVs (14.91±3.87) (P<0.01). Besides, ApoEVs could also reduce the TNF-α secretion in the culture supernatant of pro-inflammatory macrophages induced by Pg-LPS [(21 899.71±409.73) ng/L] comparing to those without ApoEVs [(71 296.50±2 344.22) ng/L] (P=0.003). Conclusions: BMMSCs derived ApoEVs could regulate the polarization of macrophages and could also attenuate the pro-inflammatory condition of macrophages induced by Pg-LPS.

[Role and mechanism of low-dose lipopolysaccharide-treated human periodontal ligament stem cells on the expression of macrophage pro-inflammatory factors].

Objective: To investigate the effect of low dose lipopolysaccharide (LPS)-treated human periodontal ligament stem cells (HPDLSC) on the expression of macrophage pro-inflammatory factors and the mechanism involved. Methods: The primary HPDLSCs were obtained from healthy third molar periodontal ligament tissue. Phosphate buffer saline (PBS), 100 µg/L or 10 mg/L of LPS were used to treat HPDLSCs for 48 h, and their conditioned media were respectively co-cultured with THP-1-derived macrophages for 48 h. The corresponding experimental groups were PBS-treated HPDLSC-derived conditioned medium (CM-C) group, low dose LPS-treated HPDLSC-derived conditioned medium (CM-L) group, and high dose LPS-treated HPDLSC-derived conditioned medium (CM-H) group. Quantitative real-time PCR was performed to explore the mRNA expressions of macrophage interleukin (IL)-6, IL-8, IL-12, tumor necrosis factor-α (TNF-α) in the CM-C, CM-L and CM-H groups, and the expressions of nuclear factor (erythroid-derived 2)-like 2 (NRF2), glutamate-cysteine ligase catalytic subunit (GCLC), NAD(P)H quinone dehydrogenase 1 (NQO1) and heme oxygenase 1 (HO-1) in the CM-C and CM-L groups. Meanwhile, Western blotting was used to detect the change of nuclear and cytoplasmic NRF2 and the levels of GCLC and HO-1 in the CM-C and CM-L groups. The 2', 7'-dichlorofluorescein probe was adopted to detect the reactive oxygen species (ROS) levels of macrophages in the CM-C and CM-L groups and the data were characterized by the mean fluorescent intensity (MFI). Results: The mRNA expressions of macrophage pro-inflammatory factors IL-6, IL-8, IL-12 and TNF-α in the CM-H group (2.332±0.594, 3.601±0.639, 2.120±0.677 and 2.468±0.236) were significantly upregulated compared with those in the CM-C group (1.000±0.321, 1.000±0.151, 1.000±0.059 and 1.000±0.095) (P<0.05); while the relative mRNA levels of IL-6, IL-12 and TNF-α in the CM-L group (0.056±0.002, 0.215±0.024 and 0.567±0.071) were much lower than those in the CM-C group (1.000±0.209, 1.000±0.220 and 1.000±0.220) (P<0.05). At the mRNA level, the expression of NRF2 was significantly increased in the CM-L group (1.864±0.198) compared with that in the CM-C group (1.000±0.094) (P<0.05). At the protein level, the cytoplasmic NRF2 and nuclear NRF2 were increased in CM-L group (1.175±0.104 and 1.308±0.082) compared with those in the CM-C group (1.000±0.025 and 1.000±0.049) (P<0.05). Furthermore, the antioxidative genes, i.e. GCLC and NQO1, localized in NRF2 downstream, were significantly upregulated in the CM-L group (1.786±0.278 and 1.444±0.078) compared with the CM-C group (1.000±0.139 and 1.000±0.226) (P<0.05). The protein levels of GCLC and HO-1 were augmented in the CM-L group (1.159±0.036 and 1.412±0.075) in contrast with those in the CM-C group (1.000±0.050 and 1.000±0.013) (P<0.05). In addition, the MFI in the CM-L group (123 419±1 302) was significantly lower than that in the CM-C group (139 193±1 241) (P<0.05). Conclusions: Low-dose LPS-treated HPDLSCs could regulate oxidative stress response through activating the NRF2 signaling pathway of macrophages and further downregulating the expressions of macrophage pro-inflammatory factors.

Protective effect of oral contraceptive against Helicobacter pylori infection in US adult females: NHANES 1999-2000.

Recently, the antibacterial properties of oestrogen and progestogen were discovered. The aim of this study was to find the cross-sectional association between oral contraceptive use and Helicobacter pylori seroprevalence. Data were obtained from the US National Health and Nutrition Examination Survey (NHANES). The H. pylori immunoglobulin G (IgG) enzyme-linked immunosorbent assays were used to categorise participants as seropositive or seronegative. The study population included 799 female participants who had information on H. pylori seroprevalence and all other covariates and had not been taking any medications (except oral contraceptives). The bivariate Rao-Scott chi-square test indicated a significant association between H. pylori seroprevalence and contraceptive use (P < 0.01). The variables of race, education, poverty income ratio, smoking, and blood lead and cadmium levels were also significantly associated with H. pylori seroprevalence (P < 0.01). Multiple logistic regression analysis of the age-adjusted model revealed that contraceptive users are 65% less likely of being H. pylori seropositive as compared to non-contraceptive users (odds ratio (OR): 0.35, 95% confidence interval (CI): 0.18-0.68). This association is stronger with the final multivariate model (OR: 0.46, 95% CI: 0.23-0.89). Conclusions: This finding reveals the potential protective effect of oral contraceptives against H. pylori infection and serves as a foundation study for further investigations.

[Inflammatory periodontal stem cells mediate interleukin-1ß secretion of macrophage by regulating macrophage endoplasmic reticulum stress].

Objective: To investigate the effect and mechanism of periodontal ligament stem cell (PDLSC) from inflammatory environment on the secretion of interleukin-1ß (IL-1ß) by macrophages. Methods: PDLSCs were pretreated with lipopolysaccharide (LPS) in order to simulate the inflammatory environment. Human monocyte cell line (THP-1) cells were treated with conditioned media collected from healthy and inflammatory PDLSCs respectively and divided into conditioned medium of health PDLSC (CM-H) group and conditioned medium of LPS-PDLSC (CM-LPS) group. After 24 h of co-culture, the condition media were abandoned and THP-1 cells were then cultured for another 24 h. The expression of IL-1ß in THP-1 cells supernatant was detected by enzyme-linked immunosorbent assay (ELISA). Quantitative real time-PCR (qRT-PCR) was used to detect the expression of glucose regulated protein 78 (GRP78), activating transcription factor-6 (ATF6), inositol requiring enzyme 1 (IRE1), protein kinase R-like endoplasmic reticulum kinase (PERK), CCAAT enhancer binding protein homologous protein (CHOP), activating transcription factor-4 (ATF4) and X box binding protein 1 spliced (XBP1s), which were all related with endoplasmic reticulum stress (ERS), in THP-1 cells. The expressions of proteins GRP78 and CHOP were detected by Western blotting. Furthermore, THP-1 cells, which pretreated with ER inhibitor 4-phenylbutyrate (4-PBA) for intervention experiments were grouped by various concentrations of 4-PBA including groups 0 (control group), 1, 10 and 20 mmol/L and treated with condition medium of inflammatory PDLSC. ELISA was used to detect IL-1ß expression and qRT-PCR to detect expression of ERS related genes. Results: ELISA results showed that the expression of IL-1ß in THP-1 cells of group CM-LPS [(31.35±2.11) ng/L] was significantly higher than group CM-H [(8.19±1.51) ng/L] (t=12.60, P<0.01). qRT-PCR results showed that the relative expressions of GRP78, ATF6, IRE1, PERK, CHOP, ATF4 and XBP1s genes in THP-1 cells of group CM-LPS (1.782±0.070, 1.387±0.204, 1.404±0.119, 1.777±0.187, 1.325±0.156, 1.295±0.066 and 1.137±0.149, respectively) were significantly higher than those in group CM-H (P<0.05). In the 4-PBA intervention experiment, compared with group 0 mmol/L, the expressions of GRP78, IRE-1, ATF-6, PERK and CHOP were significantly lower in group 1, 10 and 20 mmol/L (P<0.05). Moreover, compared with control group [(31.23±1.98) ng/L], the expression of IL-1ß in THP-1 cells were significantly lower in group 10 mmol/L [(21.20±0.37) ng/L] and group 20 mmol/L [(23.85±1.80) ng/L] (P<0.05) with ERS inhibited. Conclusions: PDLSC from inflammatory environment could promote IL-1ß secretion of macrophages through upregulating macrophages ERS.

[Research study on nucleic acid testing of hemostatic stickers used and the surface of the recipients' hands after the inoculation of inactivated SARS-CoV-2 vaccine].

Among the staff of Beijing Chao-Yang Hospital, Capital Medical University, who received the inactivated SARS-CoV-2 vaccine on January 30 in 2021, 28 recipients were selected for this research. Samples for nucleic acid tests were collected from the surface of the recipients' both hands before and after vaccination. The hemostatic stickers used after the inoculation were also collected for nucleic acid tests. The nucleic acid tests of the samples collected from the surface of both hands of the 28 recipients before vaccination were all negative. After vaccination, the nucleic acid tests of the samples collected from the surface of both hands of recipients were positive in 3 cases, and suspicious in 8 cases, with a positive rate of 10.7%. A total of 25 hemostatic stickers used were collected, 24 of them had positive nucleic acid tests, and the rest one had suspicious nucleic acid test result, with a positive rate of 96%. The hemostatic stickers used after the inoculation have the risk of nucleic acid contamination.

[Evaluation and prevention of periodontal risk related to orthodontic therapy].

The importance for evaluating the periodontal risk related to orthodontic therapy and ensuring the treatment safety was reviewed and emphasized in the present article according to the characteristics of periodontal tissues and common clinical problems of various orthodontic patients. The proposals of periodontal management during orthodontic therapy were given in outline for risk prevention and multidisciplinary cooperation.

[A pilot study on the consistency of biological widths measured by periodontal probe and cone-beam CT].

Objective: To compare the consistency of the biological widths measured by using cone-beam CT (CBCT) and periodontal probe in patients with two different gingival biotypes. Methods: Totally 27 patients [13 males, 14 females, (37.6±13.7) years old], who planned to receive the crown lengthening surgery, were recruited under the inclusion and exclusion criteria in Department of Periodontology, School of Stomatology, The Fourth Military Medical University during November 2017 to June 2018. A total of 40 teeth (14 front teeth, 26 posterior teeth) were involved in this study. The patients were divided into two groups according to their gingival biotypes: thin gingival biotype [5 males, 8 females, (40.2±15.0) years old, 21 teeth] and thick gingival biotype [8 males, 6 females, (35.1±11.9) years old, 19 teeth]. All the teeth were checked before crown lengthening procedures by using CBCT, and the biological widths and sulcus depths were measured during the surgery by using periodontal probes (Hu-Friedy, U S A). The data were recorded and statistically analyzed. Results: There were no significant differences of the biological widths between the two measuring methods amongst all of the 40 teeth [periodonial probe: (1.64±0.26) mm; CBCT: (1.69±0.20) mm], amongst 21 thin gingival biotype teeth [periodontal probe: (1.49±0.19) mm; CBCT: (1.57±0.12) mm] and amongst 19 thick gingival biotype teeth [periodontal probe: (1.80±0.21) mm; CBCT: (1.87±0.18) mm] (P>0.05). There were no significant differences of the biological widths [anterior teeth: (1.59±0.15) mm, posterior teeth: (1.67±0.29) mm, P=0.42] and of the sulcus depths [anterior teeth: (2.00±0.28) mm, posterior teeth: (2.11±0.43) mm, P=0.44] between anterior teeth and posterior teeth. The difference of biological widths, measured by two methods respectively, between thin and thick gingival biotype groups was statistically significant (P<0.01). There were significant differences of the sulcus depths, measured by the periodontal probes, between the thin [(1.93±0.28) mm] and thick [(2.24±0.41) mm] gingival biotype groups (P<0.01). Conclusions: The biological widths measured by CBCT is consistent with those measured by using periodontal probes. The biological widths and the depths of the sulcus of thin and thick gingival biotypes are different.

Autophagy promotes MSC-mediated vascularization in cutaneous wound healing via regulation of VEGF secretion.

Vascularization deficiency caused a lot of diseases, such as diabetes ulcer and myocardial infarction. Mesenchymal stem cells (MSCs), with the self-renewal and multipotent differentiation capacities, have been used for many diseases treatment through regulation microenvironment. Numerous studies reported that MSCs transplantation could largely improve cutaneous wound healing via paracrine secretion of growth factors. However, whether MSCs take part in the angiogenesis process directly remains elusive. Previous study proved that autophagy inhibited immunosuppressive function of MSCs and prevented the degradation of MSCs function in inflammatory and senescent microenvironment. Here, we proved that autophagy determines the therapeutic effect of MSCs in cutaneous wound healing through promoting endothelial cells angiogenesis and demonstrated that the paracrine of vascular endothelial growth factor (VEGF) in MSCs was required in wound site. We further revealed that autophagy enhanced the VEGF secretion from MSCs through ERK phosphorylation directly. Collectively, we put forward that autophagy mediated paracrine of VEGF plays a central role in MSCs cured cutaneous wound healing and may provide a new therapeutic method for angiogenesis-related diseases.

[Effects of cell-to-cell communication and histone acetyltransferase on the change of osteogenic differentiation ability among single-cell clones from healthy periodontium with heterogeneity of osteogenic differentiation abilities].

Objective: To investigate the effect of cell-to-cell communication amongst single-cell clones from healthy periodontium with different osteogenic differentiation potentials on change of osteogenic differentiation capabilities and the role histone acetyltransferase partaken in this process. Methods: In order to research the change of osteogenic differentiation ability via cell-to-cell communication, indirect co-culture method was used by placing two single-cell clones with different osteogenesis potentials in each of the 6-well plates. Blank control, weak and strong osteogenic groups were set up, corresponding to Transwell chambers with blank, cells of weak osteogenesis ability and cells of strong osteogenesis ability, respectively. Each group was made in triplicate. After co-culture for four days, Transwell chamber was removed. Quantitative real-time PCR (qPCR) and alizarin red staining were employed to detect the change of osteogenic differentiation ability. The acetylation level of H3 was measured by using Western blotting. Histone acetyltransferases were detected by qPCR. Results: Single-cell clones were ensured from mesenchymal stem cells by flow cytometer, the positive expression of CD29, CD90, CD105, CD146 was (99.80±0.02)%, (99.36±0.18)%, (99.41±0.05)% and (95.10±2.11)%, respectively. And CD31 and CD34 expression were (0.29±0.11)% and (0.22±0.13)%, respectively. Alizarin red and oil red O staining confirmed that single-cell clones had the abilities of adipogenesis and osteogenesis. Alkaline phosphatase (ALP) and alizarin red staining indicated that different single-cell clones were heterogeneity in osteogenesis differentiation. Indirect co-culture indicated that the mRNA expression of osteocalcin (OCN) were 14.24±5.60 and 4.78±2.90, respectively and Runt-related transcription factor 2 (RUNX2) were 2.75±1.44 and 1.61±0.44, respectively, in strong and weak osteogenic groups. They were significantly higher compared to the blank group (the mRNA expression of OCN and RUNX2 were 1.00±0.47 and 1.00±0.39, respectively). The expression of OCN and RUNX2 were also higher in strong osteogenic group than that in weak osteogenic group (P<0.05). The mean gray level of the acetylation of H3 in strong osteogenic group (0.76±0.09) and weak osteogenic group (0.54±0.12) were also higher than that in the blank group (0.30±0.04)(P<0.05). qPCR results showed that KAT6A in strong osteogenic group exhibiting higher expression (P<0.05) compared to weak osteogenic group and the blank group, which were corresponding to the changes of acetylation levels. Conclusions: Single-cell clones from healthy periodontium showed heterogeneity in osteogenic differentiation abilities. Single-cell clones with strong osteogenesis abilities had an advantage over others by promoting others' osteogenesis differentiation and this change mediated by cell-to-cell communication might be caused by modulating KAT6A to affect the acetylation level of histone.

PROTEIN EXPRESSION OF METADHERIN AND SENSIBILITY OF BREAST CANCER MDA-MB-231 CELLS TO TREATMENT BEFORE AND AFTER TRANSFECTION.

Breast cancer tends to have an increasing mortality, severely threatening the health of females. The invasion and metastasis of breast cancer are the leading causes of death. It has been reported that breast cancer is caused by the activation of a series of proto-oncogenes and inactivation of anti-oncogenes. In the present study, Real-time PCR and Western blot were used to detect the protein expression level of metadherin before and after transfecting MDA-MB-231 cells to identify the effect, while the sensitivity of MDA-MB-231 cells to 1 mg/L doxorubicin and 8mg/L taxol was measured by methylthiazolyldiphenyl-tetrazolium bromide (MTT). The results demonstrated that mRNA and protein expression level of metadherin both improved after transfection. The inhibition effect of 1 mg/L doxorubicin and 8 mg/L taxol on breast cancer cells decreased after transfection. Detected by flow cytometry, the apoptosis rate of breast cancer cells was 39.68±0.42%, 20.64±0.55%, respectively, under the effect of 1 mg/L doxorubicin; while under the effect of 8 mg/L taxol, the rate was 24.89±0.41% and 13.8±0.63%, respectively. Thus the inhibition effects of 1 mg/L doxorubicin and 8mg/L taxol to breast cancer cells and their effects on apoptosis were different, and the differences were statistically significant (P<0.05). Based on the statistics on the expression level of metadherin after transfecting breast cancer cells MDA-MB-231 and the exploration of the sensitivity of the cells to treatment, the effect of metadherin on breast cancer MDA-MB-232 cells was proved.

Establishment of an acute rejection model by transplanting both renal grafts into two different rats using a modified method of ureterovesical anastomosis.

OBJECTIVE: To establish an acute rejection model after kidney transplantation in the rat using a modified method of ureterovesical anastomosis. METHODS: Thirty-nine Wistar rat donors, were transplanted into 70 male SD rats. Wistar rats (group 1; n = 18) underwent harvest of both kidneys, cold perfusion, and transplantation into 36 SD rats. Wistar rats (group 2; n = 18) underwent left kidney harvest, cold perfusion and transplantation into 18 SD rats. Groups 1 and 2 did not receive immunosuppression after transplantation. Six kidneys were harvested from 3 Wistar rats (group 3), were transplanted into 6 SD rats that were treated with CsA (5 mg/kg per day) postoperatively, and humanely killed at 21 days. There were 10 SD in sham operated rats (group 4). The renal allograft vein was end-to-end anastomosed to the recipient renal vein using an epidural catheter. The renal allograft was anastomosed end-to-side to the recipient abdominal aorta with an abdominal aortic flap. The renal allograft ureterovesical flap was directly inserted into the recipient bladder, and attached by 4-5 interrupted sutures. The recipient's right kidney vessels were ligated at 3 days postoperatively. RESULTS: The success rates were 91.7% and 88.9% in groups 1 and 2, respectively. Except for the time for removal of the renal allografts, the operative durations and warm ischemia times differed insignificantly between both groups (P > .05). Blood creatinine levels increased significantly after kidney transplantation in groups 1 and 2 compared with the sham operated and CsA-treated cohorts (P < .01), but insignificantly between groups 1 and 2 (P > .05). CONCLUSIONS: A dual renal allograft model was established in the rat using a modified ureterovesical anastomosis. The technique can be reproduced reliably, reducing costs and shorten using overall operative duration.

Electrochemical deposition and formation mechanism of single-crystalline cu(2)o octahedra on aluminum.

A simple electrochemical deposition was developed to synthesize the cuprous oxide (Cu(2)O) octahedra on aluminum foils. The average edge length of the octahedra is about 300 nm. The chemical composition of the octahedra was determined using energy dispersive X-ray spectroscopy and electron energy-loss spectroscopy. The microstructure of the octahedra was investigated using transmission electron microscopy. The formation mechanism of the octahedra is proposed.

Combined use of etanercept and MTX restores CD4⁺/CD8⁺ ratio and Tregs in spleen and thymus in collagen-induced arthritis.

OBJECTIVE: To further explore the mechanism of etanercept (ENT, rhTNFR:Fc) and methotrexate (MTX) in the combined treatment of rheumatoid arthritis (RA), we investigated whether thymic and splenic T-cell subsets and their related cytokines imbalance could be restored by ETN/MTX treatment. METHODS: The effect of ETN/MTX on collagen-induced arthritis (CIA) was evaluated by arthritis scores, joint and spleen histopathology, as well as indices of thymus and spleen. T lymphocytes proliferation was determined by [(3)H]-TdR incorporation. Levels of TNF-α, LT-α, IL-1ß, RANKL, IL-10, IL-17, IFN-γ and IL-6 were detected by enzyme linked immunosorbent assay. The subsets of T lymphocytes including CD4(+), CD8(+), CD3(+)CD4(+), CD4(+)CD25(+), CD4(+)CD62L(+) and CD4(+)CD25(+)Foxp3(+) cells were quantified using flow cytometry. RESULTS: Combined administration of ETN/MTX significantly inhibited the proliferation of T lymphocytes, decreased serum IL-6, TNF-α, IL-1ß, RANKL and macrophage supernatant IL-17, LT-α, increased serum IFN-γ and macrophage supernatant IL-10. Moreover, the combined administration could restore CD4(+)/CD8(+) ratio and Treg cells of CIA thymus and spleen. CONCLUSION: Taken together, our findings suggest that ENT/MTX may modify the abnormal T lymphocytes balance from central to peripheral lymphoid organs, which may partially, explained the mechanism of the combined administration.