Metabolites are important determinants of disease progression. In this study, we aimed to screen caries-related metabolites in saliva samples and explore their effect on Streptococcus mutans (S. mutans). In total, 957 individuals were enrolled, including 637 individuals with dental caries. Metabolomic and microbiome-focused analyses were then performed on a subset of the study population (100 caries-free individuals and 100 individuals with caries). We then explored the impact of the metabolites on the pathogenicity of the important cariogenic bacterium S. mutans. Our results showed a significant difference in metabolic and microbiome profiles between the two groups. The saliva metabolite 4-acetamidobutanoic acid was enriched in healthy individuals and exhibited the greatest area under the curve value > 0.9. Further studies showed that 4-acetamidobutanoic acid inhibited growth, early adhesion, and biofilm formation of S. mutans and caused negligible cytotoxicity. In addition, 4-acetamidobutanoic acid destroyed the internal structure of S. mutans by inhibiting the transcription and translation of the SloABC operon, which is important in metal ion transport in S. mutans. In conclusion, we suggest that 4-acetamidobutanoic acid is a candidate target for new antibacterial agent development, and the correlation between dental caries and foods rich in 4-acetamidobutanoic acid merits further study.

The Hedgehog (Hh) signaling pathway is involved in T cell differentiation and development and plays a major regulatory part in different stages of T cell development. A previous study by us suggested that prenatal exposure to staphylococcal enterotoxin B (SEB) changed the percentages of T cell subpopulation in the offspring thymus. However, it is unclear whether prenatal SEB exposure impacts the Hh signaling pathway in thymic T cells. In the present study, pregnant rats at gestational day 16 were intravenously injected once with 15μg SEB, and the thymi of both neonatal and adult offspring rats were aseptically acquired to scrutinize the effects of SEB on the Hh signaling pathway. It firstly found that prenatal SEB exposure clearly caused the increased expression of Shh and Dhh ligands of the Hh signaling pathway in thymus tissue of both neonatal and adult offspring rats, but significantly decreased the expression levels of membrane receptors of Ptch1 and Smo, transcription factor Gli1, as well as target genes of CyclinD1, C-myc, and N-myc in Hh signaling pathway of thymic T cells. These data suggest that prenatal SEB exposure inhibits the Hh signaling pathway in thymic T lymphocytes of the neonatal offspring, and this effect can be maintained in adult offspring via the imprinting effect.

BACKGROUND The methylation of IFN-γ and IL-4 genes is regarded as an epigenetic regulation that maintains the Th1 or Th2 phenotype. OBJECTIVE To explore the influence of prenatal administration of the staphylococcal enterotoxin B (SEB) in pregnant rats, on the IFN-γ or IL-4 expression in the offspring spleen. METHODS The SEB or PBS was administered intravenously to pregnant rats on the embryo-day 16. After normal delivery, the spleens from the fifth-day neonates and adult offspring were isolated under anesthesia. Quantitative PCR, western blot, ELISA and MeDIP-qPCR were applied to determine the levels of the splenic IFN-γ or IL-4 mRNAs, their protein levels, and methylation status, respectively. RESULTS Prenatal administration of the SEB in pregnant rats decreased the levels of the splenic IFN-γ and IL-4 proteins in neonates, but increased their mRNA levels. However, prenatal administration of the SEB significantly augmented both mRNA and protein levels of the IFN-γ and IL-4 in the adult spleen. In addition, the prenatal SEB administration decreased the methylation of the splenic IFN-γ and IL-4 in adult but not neonatal offspring. CONCLUSION The prenatal administration of SEB in pregnant rats can cause a mixed Th1 and Th2 cytokines response in the offspring spleen, and alter the cytokine expression of the Th1 and Th2 via decreasing the methylation in adult but, not neonatal offspring spleen.

目的:探讨"立德树人"视域下思政元素融入医学超声影像学"混合式教学模式"的应用效果.方法:选取蚌埠医学院2017级184名医学影像学专业学生作为研究对象.采用"互联网+课程思政"、案例教学等教学方法,将思政元素与超声医学专业知识有机结合.应用医学超声影像学课程思政教学调查问卷、关怀能力评价量表及加利福尼亚批判性思维倾向量表,测评教学效果.结果:教学实施后,90％以上的学生认为有效提高了爱国主义意识、提升了人生观与价值观,增强了专业认同感、治病救人的责任担当、职业道德与职业素养、人文关怀理念及医患沟通技巧等;学生的关怀能力评价量表和批判性思维倾向量表各维度得分及总分均高于教学前(P<0.05～P<0.01).结论:"立德树人"视域下将思政元素润物无声地融入医学超声影像学"混合式教学模式",可在培养卓越的社会主义医学影像人才过程中发挥重要作用.

We have developed an easy and fast method to semiquantify low levels of mRNA from small amounts of brain tissues based on nonradioactive reverse transcription-polymerase chain reaction (RT-PCR). The regulation of mRNA for the growth associated protein GAP-43/B-50 and the homeodomain protein islet-1 was examined in the facial nucleus of the rat after a unilateral transection of the nerve. In both cases a similar sensitivity for radioactive and nonradioactive RT-PCR methods was found. The expression of the housekeeping gene, cyclophilin A was used to normalize total mRNA amounts and PCR conditions. After amplification the PCR products were separated electrophoretically on polyacrylamide gels. For nonradioactive semiquantification gels were stained with ethidium bromide and recorded using a CCD camera and transillumination. The recordings were evaluated with specialized software. Using nonradioactive RT-PCR, the increase in GAP-43/B-50 mRNA in response to axotomy was easily detectable in the small volume of tissue obtained from the facial nucleus. In contrast, the low expression of islet-1 mRNA made it necessary to develop a two-step amplification procedure in order to provide a reliable semiquantitative analysis. The procedure included preamplification of the cDNA and subsequent purification of the cDNA. Using this method, the down-regulation of islet-1 could be demonstrated with a similar sensitivity to that previously shown with radioactive RT-PCR.

Objective To study the effect of outer membrane vesicles (OMVs) derived from Salmonella typhimurium (ST) on the ultrastructural features and immune function of dendritic cells (DC). Methods Mice bone marrow cells were collected aseptically, and myeloid DC were generated by the combined induction and amplification with recombinant mouse granulocyte-macrophage colony-stimulating factor (GM-CSF) and recombinant mouse interleukin-4 (rm IL-4). Cell morphology was observed under inverted phase contrast microscope and the phenotype was identified with flow cytometry. ST-OMVs were isolated through ultracentrifugation. The survival rate of DC was assessed with CCK-8 assay, and the stimulus concentration of OMVs was henceforth determined. The ultrastructural characteristics of DC loaded with OMVs were observed with transmission electron microscopy. The cytokine secretion, surface molecule expression and phagocytic capacity of DC were examined with flow cytometry. Results The DC induced and amplified in vitro displayed typical DC phenotype in morphological analysis and the purity of DC exceeded 85%. Transmission electron microscopy showed that there were large numbers of protrusions on the cell surface. After stimulation with ST-OMVs, it was observed that the dendritic structures on the surface of DC were reduced and a large number of phagolysosomes were found in the cytoplasm. In addition, increased numbers of mitochondria, swelling and typical apoptosis were observed. After treatment with ST-OMVs at 5 μg/mL and 10 μg/mL, the secretion of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) of DC increased significantly ( P<0.05). Furthermore, the immature DC could differentiate into mature DCs after stimulation with ST-OMVs, which were characterized by a decrease in phagocytic capacity ( P<0.05) and an upregulation of phenotypic markers ( P<0.05). Conclusion ST-OMVs can stimulate DC to produce TNF-α and IL-1β and promote DC maturation and antigen presentation.

Introduction. Streptococcus mutans is an important cariogenic microbe. Hypothesis/Gap Statement. The potential characteristics of S. mutans isolates from site-specific dental plaque are still not clear. Aim. This study aimed to investigate the phenotypic and genetic characteristics of S. mutans isolates from site-specific dental plaque in China. Methodology. We used S. mutans isolated from children with early-childhood caries (ECC) and caries-free children to compare the phenotypic and genetic characteristics of S. mutans from site-specific dental plaque samples. The ECC subjects presented two sites: a cavitated lesion and a sound surface. The caries-free subjects presented one sound surface. Growth pattern, biofilm, decrease in pH, extracellular polysaccharide, expression levels of virulence-related genes, multilocus sequence typing (MLST) and phylogenetic trees were evaluated among these three sites. Results. The phenotypes detected between the cavitated and sound surfaces of ECC children were similar. However, the capacity for biofilm formation, pH drop and expression levels of genes ( gtfB and spaP ) of S. mutans in the caries-free group were lower compared with those of the ECC group. We identified 44 new alleles and 77 new sequence types. More than 90 % of the children with ECC shared an identical sequence type. The distribution of sequence types among different subjects showed diversity, and child-to-child transmission was detected. Conclusions. This is the first report of MLST on site-specific dental plaques in a single subject, and indicates that S. mutans isolated from site-specific dental plaque of a single subject showed similar phenotypes as a result of the isolates were closely related.

Staphylococcus aureus is an infectious pathogen that is relatively common, but that can cause severe disease in pregnant women and their fetus. We previously demonstrated that exposing pregnant rats to staphylococcal enterotoxin B (SEB) altered splenic CD4/CD8 T cell frequencies in their offspring. Whether prenatal SEB exposure impacts Tregs in these offspring, however, remains to be determined. As such, in this study, we intravenously injected pregnant rats with 15 μg of SEB on gestational day 16. Splenic tissue was then harvested from 1-, 3-, and 5-day-old neonatal rats and analyzed via flow cytometry to assess Treg numbers. In addition, FoxP3 expression levels were assessed via qPCR and western blotting, while FoxP3 methylation status was evaluated via methyl-DNA immunoprecipitation qPCR. Immunosuppression assays were additionally used to gauge Treg suppressive functionality. We found that exposing pregnant rats to SEB resulted in a significant increase in Treg numbers, FoxP3 expression, and Treg suppressive capacity in the spleens of both neonatal and adult offspring. In addition, total T cell, CD4+T cell, and non-Treg CD4+ T cell numbers were elevated in the spleens of offspring following prenatal SEB exposure. We additionally determined that SEB exposure resulted in a significant reduction in FoxP3 DNA methylation. Together, our results indicate that prenatal SEB exposure can markedly enhance offspring splenic Treg numbers and functionality at least in part by decreasing FoxP3 methylation.

Introduction. Staphylococcal enterotoxin B (SEB) is an extensively studied super-antigen. A previous study by us suggested that SEB exposure during pregnancy could alter the percentage of CD4+ and CD8+ T cells in the peripheral blood of neonatal offspring rats.Aim. It is unknown whether SEB exposure during pregnancy can influence the development of regulatory T cells (Tregs) in the peripheral blood of neonatal offspring rats.Methodology. Pregnant rats at gestational day 16 were intravenously injected with 15 µg SEB. Peripheral blood was acquired from neonatal offspring rats on days 1, 3 and 5 after delivery and from adult offspring rats for determination of Treg number by cytometry, cytokines by ELISA, and FoxP3 expression by real-time PCR and western blot.Results. SEB given to pregnant rats significantly increased the absolute number of Tregs and the expression levels of FoxP3, IL-10 and TGF-β (P<0.05, P<0.01) in the peripheral blood of not only neonatal but also adult offspring rats. Furthermore, repeated SEB exposure in adult offspring rats significantly decreased the absolute number of Tregs (P<0.01), and the expression levels of FoxP3, IL-10 and TGF-β (P<0.05, P<0.01) in their peripheral blood.Conclusion. Prenatal SEB exposure attenuates the development and function of Tregs to repeated SEB exposure in the peripheral blood of adult offspring rats.

Regulatory T cells (Tregs) play an essential role during homeostasis and tolerance of the immune system. Based on our previous study that exposure of pregnant rats to staphylococcal enterotoxin B (SEB) can alter the percentage of CD4/CD8 subsets in the thymus of the offspring, in this study, we focus on the influence of exposure of pregnant rats to SEB on number, function and response of Tregs in the thymus of the offspring. Pregnant rats at gestational day of 16 were intravenously injected with 15 μg SEB and the thymuses of the neonatal and adult offspring were harvested for this study. We found that exposure of pregnant rats to SEB could significantly increase the absolute number of Tregs and the FoxP3 expression level in the thymus of not only neonatal but also adult offspring. Re-exposure of adult offspring to SEB remarkably reduced the suppressive capacity of Tregs to CD4+ T cells and the expression levels of TGF-β and IL-10 in the thymus, but had no effect on production of IL-4 and IFN-γ. Furthermore, it also notedly decreased the absolute number of Tregs and the FoxP3 expression level. These data suggest that prenatal exposure of pregnant rats to SEB attenuates the response of increased Tregs to re-exposure to SEB in the thymus of adult offspring.

Streptococcus mutans (S. mutans) is a serious microbe causing dental caries. Mutacin IV is an effective bacteriocin produced by S. mutans to antagonize numerous non-mutans streptococcal species. However, the posttranscriptional regulation of mutacin IV remains unclear. This study aimed to analyze the effect of small RNA srn225147 on mutacin IV. The functional prediction suggested that srn225147 is involved in the production of mutacin IV, an important secondary metabolite. According to RNAhybrid and RNAPredator prediction, the mutacin IV formation-associated gene comD is a target of srn225147. We further analyzed the roles of srn225147 and comD in 20 S. mutans clinical strains with high production of mutacin IV (High-IV group) and lacking mutacin IV (None-IV group). Levels of comD expression were significantly higher in the High-IV group, whereas the Non-IV group showed relatively higher expression of srn225147, with a negative correlation observed between srn225147 and comD. Moreover, compared to the mimic negative control (NC) group, comD expression was decreased at 400-fold srn225147 overexpression but increased at approximately 1400-fold overexpression. Although the production of mutacin IV in the 1400-fold change srn225147 mimic group was larger than that in the 400-fold change mimic group, there was no significant difference in the production of mutacin IV between the srn225147 mimic group and mimic NC group. These results indicate that srn225147 has a two-way regulation effect on the expression of comD but that its regulation in the production of mutacin IV is weak.

The authors have withdrawn this preprint from Research Square.

结合国内外医学教育标准和我校的办学特色,我校打破传统医学教育模式,开设了"卓越人才培养实验班",突出"以学生为中心"的教学理念,基础医学院探索适合我校临床医学本科专业人才培养的医学基础课程教学模式.本文介绍了我校基础医学院在"卓越人才培养实验班"基础医学课程教学和考核的具体实施.

OBJECTIVE:To study the influences of ureaplasma urealitycum (UU) infection on testicular tissue structure and secretion function in rats.METHODS:Forty clean grade male SD rats were randomly divided into the experiment group A (at 7 d after surgery), experiment group B (at 14 d after surgery), control group C (at 7 d after surgery) and control group D (at 14 d after surgery). There were 10 rats in each group. The experimental groups were injected with 0.6 mL UU4 through bladder. In the same way, the control groups were injected with the same volume of UU liquid medium. At day 7 and 14 after injection, the structures of testis of all rats were observed by light microscopy and spermatogenic cells by transmission electron microscopy. The content of testosterone in plasma and testicular fluid were detected by chemiluminescence method.RESULTS:The changes of inflammatory pathology (including the layer and amount of spermatogenic cell decreasing, inflammatory cell infiltrating and mature sperms decreasing) in the testis of group A and group B were found by light microscopy, and the inflammatory changes in group B were lighter than those in group A. The structures of testicular tissue in group C and group D were normal. The apoptosis performances of germ cell (including the cell membrane corrugated, nuclear chromatin concentration and nuclear rupture) in the testis of group A and group B were found by transmission electron microscopy, and the changes in group B were lighter than those in group A. The structures of germ cell in group C and group D were normal. The levels of plasma testosterone in group A and group B were significantly lower than that in group C and group D (P<0.01), the difference between group A and group B was not statistically significant. The testosterone level in testis interstitial fluid in group A was significantly lower than that in other groups (P<0.01), the differences between other groups were not statistical significant.CONCLUSIONS:The testicular tissue of UU infected rats can have various pathological damage and functional changes, further confirming that UU infection can cause male infertility. The pathological damage and functional changes of the testicular tissue of rats after UU infection can be gradually restored with the extension of the duration of the disease.

当前,病原体感染在世界各国仍是严重问题,尤其是在医学不发达的发展中国家.病原体种类繁多,其中寄生虫、细菌、真菌、病毒与人类的健康和安全息息相关.而随着生物、医学等学科的不断发展,人们对病原体感染引起的免疫防御反应及免疫机制有了更深入的认识.T淋巴细胞具有介导细胞免疫和调节免疫应答的作用,主要可分为CD4+、CD8+T细胞两大类.CD4+T细胞又可根据功能不同分成不同亚群,每个亚群具有不同的分化和调节机制,产生不同的细胞因子,发挥不同的功能.研究[1]证明,Th17细胞和Treg细胞在寄生虫、细菌、真菌、病毒感染的免疫应答中至关重要,尤其是Th17/Treg的平衡紊乱与病原体的感染有着密不可分的联系,且在不同环境下可以互相转化.本文就Th17细胞和Treg细胞在病原体感染与免疫中的作用作一综述.

Dickkopf-1 (DKK-1) plays an important role in tumorigenesis. It has been previously demonstrated that DKK-1 is overexpressed in human pancreatic cancer tissues. This present study aimed to investigate the roles of DKK-1 in pancreatic carcinogenesis. Knockdown of DKK-1 by siRNA inhibited proliferation, migration, and apoptosis in PANC-1 human pancreatic cancer cells. Additionally, knockdown resulted in a decrease in mRNA and protein expression levels of DKK-1, c-Myc, and cyclin D1. Positive correlation between DKK-1, c-Myc and cyclin D1 expression was observed. Present data suggests that DKK-1, c-Myc and cyclin D1 promote pancreatic carcinogenesis.

The aim was to study hyphae growth characteristics and polysaccharide yield of Cordyceps militaris strains (150707, 150715, 150725, 150808, 52244, 51762) from different genetic resources. Hyphae growth rate and conidiospores yield of different Cordyceps militaris strains were measured. Crude polysaccharide from different strains was extracted by water extraction and alcohol deposition method. Hyphae growth rate, conidiospores yield and polysaccharide yield rate of these six strains from different genetic resources were different. The hyphae growth rate was faster on enrichment PDA medium, while the conidiospores yield and polysaccharide yield rate of strains was better on blood plate medium. It can be analysed that the strains 150707 and 52244 were dominant strains because their yield of conidiospores and polysaccharide was superior. Blood plate medium was superior medium for the yield of conidiospores and polysaccharide of Cordyceps militaris strains.

OBJECTIVE To study the effect of cordycepin on cell cycle, apoptosis and autophagy of human tongue cancer TCA-8113 cells and explore the mechanism of cordycepin for inhibiting the occurrence of tongue cancer. METHODS CCK-8 method was used to assess the inhibitory effect of cordycepin on TCA-8113 cell proliferation in vitro. The cell cycle and cell apoptosis of TCA-8113 cells treated with different concentrations of cordycepin were analyzed using flow cytometry. The expressions of apoptosis-related genes caspase-3, caspase-9, Bcl-2, and Bax were examined using quantitative real-time PCR and Western blotting, and immunohistochemistry was used to detect the expressions of autophagy-related proteins LC-3β, P62, p-mTOR, and AMPK. RESULTS CCK-8 assay showed that cordycepin significantly inhibited the proliferation of TCA-8113 cells in a concentration-dependent manner with an IC50 of 3.548 mg/mL at 24 h and an IC50 of 1.185 mg/mL at 48 h. Flow cytometric analysis showed that cordycepin caused cell cycle arrest at S phase and dose-dependently increased the apoptotic rate of TCA-8113 cells. Treatment of the cells with cordycepin enhanced the expressions of Bax, caspase-3 and caspase-9 at both the mRNA and protein levels and inhibited the expression of the antiapoptotic gene Bcl-2. Immunohistochemistry demonstrated that cordycepin promoted the expression of LC-3β and AMPK and inhibited the expression of P62 and p-mTOR. CONCLUSION Cordycepin inhibits the proliferation and induces apoptosis of HCT-116 cells through the mitochondrial pathway and induces autophagy via the AMPK/mTOR pathway.

医学检验技术是临床医学与现代实验室科学技术的结合,是一门发展十分迅速、多技术和多学科交叉的临床实践性很强的应用学科.现状下医学检验专业教学中遇到一些瓶颈亟待突破,因此寻找恰当的教学方法、探索专业建设发展方向已是当务之急.虽然我国医学专业认证刚刚起步,但专业认证已成为高等医学教育改革与发展的必然趋势.为探索这一背景下医学院校检验医学教学的客观规律,我们以医学检验专业为试点,进行了《临床微生物学检验》课程体系改革的探索和实践,从教学内容、教学手段、实践教学体系及考核方案等方面进行了系列改革与尝试.