[Explorations about the correlation between biological changes of meninges in periodontitis mice and cognitive impairment via single-cell RNA sequencing].

Objective: To clarify the potential correlation between biological changes of meninges in periodontitis mice and cognitive impairment by analyzing the biological changes of meninges in periodontitis mice using single-cell RNA sequencing. Methods: Thirty C57BL/6 mice were divided into two groups by using random number table method (15 mice in each group). Mice in the control group were locally administered 2% carboxyl methyl cellulose (CMC) without Porphyromonas gingivalis (Pg) on both buccal sides. A mixture of Pg W83 and 2% CMC was applied on both buccal sides in the experimental group mice three times a week, lasting for 16 weeks in total. The absorption of alveolar bone, locomotor activity and cognitive function, the activation of microglia and astrocytes in the cortex were observed and assessed. The mRNA expression levels of Occludin in meninges and brain were detected in two groups. Single-cell RNA sequencing data of meninges were processed by uniform manifold approximation and projection (UMAP). Differential genes expressions of endothelial cells were processed by gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. In addition, real-time fluorescence quantitative PCR (RT-qPCR) was used to verify the expressions of transcription activating factor 3 (Atf3) and apolpoprotein L domain-containing 1 (Apold 1). Results: Methylene blue staining found the distances of buccal and palatal cement-enamel junction-alveolar bone crest in experimental mice [(185.60±17.60), (206.90±13.37) µm] increased significantly compared with the control group [(135.33±9.57), (163.05±14.98) µm] (t=5.02, P=0.002; t=4.37, P=0.005). Open field experiment showed the total distance and average speed of mice in the experimental group [(971.88±164.57) cm, (3.25±0.55) cm/s] were not statistically significant compared with the control group [(914.24±278.81) cm, (3.05±0.93) cm/s] (t=0.65, P=0.525; t=0.65, P=0.520). The recognition index of the experimental group [(48.02±16.92) %] was lower than the control group [(66.27±17.90) %] (t=2.40, P=0.027) by novel object recognition tests. Compared with the control group [(63.56±11.88) %], the alternation of experimental group [(50.99±14.17) %] was significantly decreased in Y maze tests (t=2.33, P=0.030). Immunohistochemistry results showed microglia and astrocytes were activated in the cortex of experimental mice. Compared with the control group (1.02±0.25, 1.04±0.31), the relative mRNA expressions of Occludin decreased significantly in the meninges and brain of periodontitis mice, respectively (0.61±0.10, 0.64±0.20) (t=3.47, P=0.010; t=2.66, P=0.024). By single-cell RNA sequencing, meninges cells were divided into 11 types, such as endothelial cells, fibroblasts, immune cells and so on. Endothelial cells were the main cell types in meninges [the control group: 26.47% (1 589/6 004), the experimental group: 26.26% (807/3 073)]. Compared with the control group [5.56% (334/6 004)], the percentage of granulocytes increased in the periodontitis mice [11.65% (358/3 073)]. Using clustering analysis to further focus on endothelial cells, GO enrichment analysis revealed differential genes were mainly related to angiogenesis, cell adhesion, apoptosis and so on. KEGG enrichment analysis revealed that differential genes were related to signaling pathways of interleukin-17, relaxin and so on. The relative mRNA expressions of Atf3 and Apold1 in meninges of periodontitis mice (0.42±0.24, 0.54±0.27) were significantly lower than the control group (1.03±0.26, 1.02±0.23) (t=3.88, P=0.005; t=3.02, P=0.017). Conclusions: The mice chronically infected with Pg W83 occurred memory impairment, neuroinflammation and changes of barrier function. In the meninges of periodontitis mice, there were infiltration of immune cells and down-regulation expressions of Atf3 and Apold1 by single-cell RNA sequencing. Meningeal immunity and changes of barrier function may play an important role in the cognitive impairment caused by periodontitis.

[Standard analysis and implementation suggestions of "General Principles for Diagnosis of Occupational Radiation Diseases"].

The national standard "General Principles for the Diagnosis of Occupational Radiation Diseases" (GBZ112-2017) was issued and implemented to replace "General Principles for the Diagnosis of Occupational Radiation Diseases" (GBZ112-2002). In this paper, the significance of the new standard, the background of revision, the revision basis of important indicators and the matters needing attention in the application of the standard were interpretated in detail. Especially for the revision basis of important indicators, such as the determination of the basic principles of diagnosis, the diagnostic basis, the principle of dose evaluation and the principle of treatment are explained in detail, so as to avoid the possible misunderstandings and misgiving in the use of the standard.

[Effect of hormones on keratoconus and its mechanism].

Keratoconus is an ectatic corneal disorder and characterized by central and/or inferior temporal corneal thinning in a cone shape. Studies have shown that its pathogenesis is related to changes in tissue proteins, enzymes, immune functions and specific genes, but the exact pathogenesis has not been clearly clarified yet. Hormones such as sex hormones have been found to be associated with keratoconus. This review aims to summarize and analyze the research progress of the effect of hormones on keratoconus and its mechanism, so as to explore the role of hormones in the early diagnosis of keratoconus and find new targets in the treatment of keratoconus, providing reference for clinical work.

[Treatment and prognostic analysis of isolated chest wall recurrence of breast cancer after mastectomy].

Objective: To analyze the prognostic factors of breast cancer patients with isolated chest wall recurrence (ICWR) after mastectomy, and investigate the optimal treatment. Methods: A total of 201 breast cancer patients with ICWR after mastectomy who were treated in Cancer Hospital, Chinese Academy of Medical Sciences and the Fifth Medical Center Chinese PLA General Hospital from October 1998 to April 2018 were retrospectively analyzed. The median follow-up was 92.8 months and survival data were obtained. Results: Among 201 patients with ICWR, 103 patients developed subsequent locoregional recurrence (sLRR) and 5-year cumulative sLRR rate was 49.1%; 134 patients developed distant metastasis (DM) and 5-year DM rate was 64.4%; 103 patients died, the median progression-free survival (PFS) was 17.4 months and the 5-year PFS rate was 23.2%; the median overall survival (OS) was 62.5 months and the 5-year OS rate was 52.1%. Multivariate analysis showed that the recurrence interval (HR=2.17, 95% CI: 1.26-3.73) and the locoregional treatment (HR=1.59, 95% CI: 1.05-2.40) were the independent prognostic factors for sLRR. The initial HER2 status (HR=1.60, 95% CI: 1.03-2.48) was the independent prognostic factor for DM. The recurrence interval (HR=1.99, 95% CI: 1.30-3.04), the locoregional treatment (HR=1.99, 95% CI: 1.43-2.76) and the treatment modalities after recurrence (HR=1.70, 95% CI: 1.18-2.46) were the independent prognostic factors for PFS. The initial HER2 status (HR=1.69, 95% CI: 1.02-2.81), the recurrence interval (HR=1.85, 95% CI: 1.15-2.98) and the treatment modalities after recurrence (HR=2.48, 95% CI: 1.56-3.96) were the independent prognostic factors for OS. Conclusions: Breast cancer patients after ICWR have an optimistic OS until now, but the risk of sLRR and DM is high. Comprehensive treatment modalities including surgery, radiotherapy and systemic therapy improve the outcome of breast cancer patients with ICWR after mastectomy.

MicroRNA-27a-3p aggravates renal ischemia/reperfusion injury by promoting oxidative stress via targeting growth factor receptor-bound protein 2.

Renal ischemia-reperfusion (RI/R) injury with high morbidity and mortality is one common clinical disease. Development of drug targets to treat the disorder is critical important. MiR-27a-3p plays important roles in regulating oxidative stress. However, its effects on RI/R injury have not been reported. In this paper, hypoxia/reoxygenation (H/R) models on NRK-52E and HK-2 cells, and RI/R model in C57BL/6 mice were established. The results showed that H/R in vitro decreased cell viability and increased ROS levels in cells, and RI/R caused renal injury and oxidative damage in mice. The expression levels of miR-27a-3p were up-regulated based on real-time PCR and FISH assays in model groups compared with control groups, which directly targeted Grb2 based on dual luciferase reporter assay and co-transfaction test. In addition, miR-27a- 3p markedly reduced Grb2 expression to down-regulate the expression levels of p-PI3K, p-AKT, Nrf2, HO-1, and up-regulate Keap1 expression in model groups. MiR-27a-3p mimics in vitro enhanced H/R-caused oxidative stress via increasing ROS levels and decreasing Grb2 expression to down-regulate PI3K-AKT signal. In contrary, miR-27a-3p inhibitor in vitro significantly reduced H/R-caused oxidative damage via decreasing ROS levels and increasing Grb2 expression to up-regulate PI3K-AKT signal. In vivo, miR-27a- 3p agomir exacerbated RI/R-caused renal damage by decreasing SOD level and increasing Cr, BUN, MDA levels via suppressing Grb2 expression to down-regulate PI3K- AKT signal. However, miR-27a -3p antagomir alleviated RI/R-caused oxidative damage via increasing Grb2 expression to up-regulate PI3k-AKT signal. Grb2siRNA in mice further enhanced RI/R-caused renal injury by increasing Cr, BUN, MDA levels and decreasing SOD level via inhibiting the expression levels of Grb2, Nrf2, HO-1, and increasing Keap1 expression. Our data showed that miR-27a-3p aggravated RI/R injury by promoting oxidative stress via targeting Grb2, which should be considered as one new drug target to treat RI/R injury.

[Detection and Analysis of the peripheral lymphocytes micronucleus rate of radiation workers in a city].

Objective: To perform lymphocyte micronucleus analysis on radiation workers with long-term exposure to low doses ionizing radiation, Evaluate the health condition of radiation workers, and provide the evidence for strengthening surveillance of radiation workers. Methods: From January 1, 2013 to December 21, 2016, a statistical analysis and evaluation was conducted of the peripheral lymphocytes micronucleus rate in 5 901 radiation workers who had undergone medical examinations of employees at Chinese Academy of Medical Sciences Institute of Radiation Medicine. Results: The micronucleus rates in radiation workers of the on-job group were higher than the pre-job group (P<0.01) . Significant difference was found among the different sex (t=5.97) , different types (χ(2)=378.69) , different levels of work units (χ(2)=115.48) . Significant difference was found among the micronucleus rates of 672 radiation workers of the on-job group from 2013 to 2016 (χ(2)=92.57, P<0.01) . Conclusion: The peripheral lymphocytes micronucleus rate of radiation workers were significantly higher than non-contact workers. Significant increasing trend of micronucleus rates was noted among the radiation worker with increasing exposure time. The peripheral lymphocytes micronucleus rates of interventional therapy workers were highest. The peripheral lymphocytes micronucleus rates of Private hospitals workers were highest. This phenomenon deserves attention. Protection needs to be strengthened to ensure the health of radiation workers.

Anticandidal activity and biocompatibility of a rechargeable antifungal denture material.

OBJECTIVES: Candida-associated denture stomatitis is a recurrent and debilitating oral mucosal disease. Development of anticandidal denture materials represents a promising strategy to manage this condition. We have previously shown that miconazole incorporated in methacrylic acid (MAA) copolymerized diurethane dimethacrylate (UDMA) denture materials has long-term anticandidal activity. In this study, we examined the ability of culture medium conditioned with drug-free- or miconazole-MAA-UDMA discs to prevent Candida infection in an in vitro oral epithelial cell/Candida albicans coculture system. MATERIALS AND METHODS: Candida albicans (C. albicans)-induced OKF6/TERT-2 cell damage was quantified by the release of lactate dehydrogenase from epithelial cells, cytokine production was quantified using protein cytokine arrays, and the expression of C. albicans genes was measured by RT-qPCR. RESULTS: Candida albicans had limited growth with altered expression levels of secreted aspartyl proteinase-2 and -5 in culture medium conditioned by miconazole-MAA-UDMA discs. Significantly, the ability of C. albicans to induce oral epithelial cell damage and trigger epithelial proinflammatory cytokine production was also inhibited by miconazole disc conditioned media. CONCLUSION: Miconazole released from MAA-UDMA denture materials effectively prevents the development of candidal infection in an in vitro oral epithelial system. Further characterization of this drug-rechargeable denture material is warranted.

Induction of apoptosis in oral epithelial cells by Candida albicans.

During infection, interactions between Candida albicans and oral epithelial cells result in oral epithelial cell death. This is clinically manifested by the development of oral mucosal ulcerations generally associated with discomfort. In vitro studies have shown that C. albicans induces early apoptotic alterations in oral epithelial cells; however, these studies have also shown that treatment of infected cells with caspase inhibitors does not prevent their death. The reasons for these contradictory results are unknown and it is still not clear if C. albicans stimulates oral epithelial signaling pathways that promote apoptotic cell death. Activation of specific death pathways in response to microbial organisms plays an essential role in modulating the pathogenesis of a variety of infectious diseases. The aim of this study was to (i) characterize C. albicans-induced apoptotic morphological alterations in oral epithelial cells, and (ii) investigate the activation of apoptotic signaling pathways and expression of apoptotic genes during infection. Candida albicans induced early apoptotic changes in over 50% of oral epithelial cells. However, only 15% of those showed mid-late apoptotic alterations. At the molecular level, C. albicans caused a loss of the mitochondrial transmembrane potential and translocation of mitochondrial cytochrome c. Caspase-3/9 activities increased only during the first hours of infection. Moreover, poly[ADP ribose] polymerase 1 was cleaved into apoptotic and necrotic-like fragments. Finally, five anti-apoptotic genes were significantly upregulated and two pro-apoptotic genes were downregulated during infection. Altogether, these findings indicate that epithelial apoptotic pathways are activated in response to C. albicans, but fail to progress and promote apoptotic cell death.

Candida albicans induces early apoptosis followed by secondary necrosis in oral epithelial cells.

The capacity of Candida albicans to invade and damage oral epithelial cells is critical for its ability to establish and maintain symptomatic oropharyngeal infection. Although oral epithelial cells are reported dead after 18 h of candidal infection, activation of specific epithelial cell-death pathways in response to C. albicans infection has not yet been demonstrated. Considering the key role of oral epithelial cell damage in the pathogenesis of oropharyngeal candidiasis, the aim of this study was to characterize this event during infection. Using an oral epithelial-C. albicans co-culture system, we examined the ability of C. albicans to induce classic necrotic, pyroptotic and apoptotic cellular alterations in oral epithelial cells such as osmotic lysis, exposure of phosphatidylserine on the epithelial cell plasma membrane and internucleosomal DNA fragmentation. It was found that the ability of C. albicans to kill oral epithelial cells depends on its capacity to physically interact with and invade these cells. Caspase-dependent apoptotic pathways were activated early during C. albicans infection and contributed to C. albicans-induced oral epithelial cell death. Earlier apoptotic events were followed by necrotic death of infected oral epithelial cells. Hence, C. albicans stimulates oral epithelial signaling pathways that promote early apoptotic cell death through the activation of cellular caspases, followed by late necrosis.

Polybrominated diphenyl ethers in mudsnails (Cipangopaludina cahayensis) and sediments from an electronic waste recycling region in South China.

Mudsnails and sediments from an electronic waste recycling region in South China were chosen to study the polybrominated diphenyl ethers (PBDEs) bioavailability of mudsnail in different ambient levels. Significant (p < 0.05) correlations of biota-sediment accumulation factor (BSAF) versus the reciprocal of PBDE concentration in sediment (1/Sed) occurred in all quantitative PBDE congeners except BDE-209, showed that the BSAFs of PBDEs in mudsnails were increased with reciprocal increasing ambient levels. The BDE-183 correlation of mudsnail versus sediment (r = 0.580) was much lower than the correlation of BDE-209 versus BDE-183 in mudsnails (r = 0.812), indicated the main source of BDE-183 in mudsnails was from the debromination of higher brominated PBDEs.

Declining polychlorinated dibenzo-p-dioxins and dibenzofurans levels in the sediments from Dongting Lake in China.

Dongting Lake is the second largest freshwater lake in China. Technical sodium pentachlorophenate (Na-PCP) had been sprayed since 1960s to control the spread of snailborne schistosomiasis up to the middle of 1990s. As one of the by-products of Na-PCP, polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) may enter the environment. It has been reported that the concentrations of PCDD/Fs in sediments in Dongting Lake were 130-891 pg I-TEQ g(-1) in 1995. High toxicity of sediment to exposed microorganisms and fish may be risk on environment. In order to determine the present levels of the contamination, eight sediment samples from Dongting Lake were analyzed for PCDD/Fs using HRGC-HRMS. Total I-TEQ values for these samples were at a ranged of 0.7-11 pg g(-1), with a mean value of 4.5 pg g(-1). The results show that PCDD/Fs concentrations of contemporary sediment have declined since 1995. Certain PCDD/F congeners as well as OCDD, typical for Na-PCP, predominated in lake sediment indicating that the using of PCP-Na is the main source of PCDD/Fs in the lake. These variations in levels can reflect changes in PCDD/F sources to the environment over time.

Polybrominated diphenyl ethers in leaves and soil from typical electronic waste polluted area in South China.

Polybrominated diphenyl ethers (PBDEs) in leaves and soil from typical e-waste polluted area in South China were investigated. The concentrations (ng/g dry weight) of PBDE congeners and summation operatorPBDE of five leaf samples were much lower than those in soil sample. The general patterns of summation di-BDEs to summation hepta-BDEs percentage distribution in leaf samples were similar to those of the soil sample, except the percentage of BDE209 which were lower than in soil. The percentages of summation di-BDEs to summation hepta-BDEs in soil were in the range of those in leaf samples. The results showed that the contamination of PBDEs in the leaf samples had good correlation with the soil around them.

Glucose regulated proteins 78 protects insulinoma cells (NIT-1) from death induced by streptozotocin, cytokines or cytotoxic T lymphocytes.

Endoplasmic reticulum stress-mediated apoptosis plays an important role in the destruction of pancreatic beta-cell, and contributes to the development of type 1 diabetes. The chaperone molecule, glucose regulated proteins 78 (GRP78), is required to maintain ER function during toxic insults. In this study, we investigated the effect of GRP78 on the beta-cell apoptosis. We first measured GRP78 protein expression in different phase of streptozotocin-affected beta-cell by immunoblotting analysis. An insulinoma cell line, NIT-1, transfected with GRP78 was established, named NIT-GRP78, and used to study apoptosis, which was induced by streptozotocin or inflammatory cytokines. Apoptosis of NIT-1 or NIT-GRP78 cells was detected by flow cytometry, the transcription of C/EBP homologous protein (CHOP) was monitored by real-time PCR, the concentration of nitric oxide and the activity of superoxide dismutase were measured by colorimetric method. We found that, in comparison to NIT-1 cells, NIT-GRP78 cells responded to the streptozotocin or cytokines treatments with decreased concentration of nitric oxide, but increased activity of superoxide dismutase. In addition, the level of CHOP was also decreased in the NIT-GRP78 cells, which may mediate the resistance of the GRP78 overexpressed NIT-1 cells from apoptosis. Finally, we found that NIT-GRP78 cells were also more resistant than NIT-1 cells to cytotoxic T lymphocyte (CTL) specific killing detected by flow cytometry through target cells expressing green fluorescent protein cultured with effector cells and finally stained with propidium iodide. The data suggest that modulating GRP78 expression could be useful in preventing pancreatic beta-cell from the immunological destruction in type 1 diabetes individuals.

[The effects of exogenous p16 expression on CDK4, Cyclin D1 and pRb in nasopharyngeal carcinoma cell lines].

OBJECTIVE: To investigate the effect of exogenous p16 on Cyclin D1, CDK4 and pRb, and to explore the mechanism of the growth suppression of p16 in nasopharyngeal carcinoma cell lines. METHODS: The curve of cell growth rate in three kinds of HNE1, # 3-2 and # 4-2 cell lines was analyzed and their double time was compared. Then the distribution of the cell cycle was detected by flow cytometry. The expression of p16 and the effect of exogenous p16 expression on CDK4, Cyclin D1 and pRb are studied by means of Western Blot. RESULTS: As compared with HNE1, # 3-2 and # 4-2 showed a longer double time(23.4 h vs 28.8 h, 31.2 h). # 4-2 showed a significant accumulation of cells in G0/G1 phase(P < 0.01) and decreasing in S phase(P < 0.05) while HNE1 and # 3-2 had no obvious difference(P > 0.05). Cyclin D1 expression was upregulated in # 4-2 but downregulated in # 3-2 by exogenous expressed p16. No obvious difference on CDK4 expression was found. Hypophosphorylated pRb was detected in three cell lines. The expression was stronger in # 4-2, and # 3-2 than that in HNE1 and Hela. Hyperphosphorylated pRb was also detected in HNE1. CONCLUSION: Exogenous p16 expression may arrest cell cycle in G0/G1 phase and suppress cell growth. The major mechanism is not to regulate the level of the expression of CDK4. There might be a threshold in p16 regulating Cyclin D1 expression. However, the final result contributes to the inhibition of pRb phosphorylation.