Tooth pulpitis is a prevalent oral disorder. Understanding the underlying mechanisms of pulpitis and developing effective treatment strategies hold great significance. Ferroptosis has recently emerged as a new form of cell death, but the role of ferroptosis in pulpitis remains largely unknown. In our study, single-cell RNA sequencing (scRNA-seq) was used to identify cellular heterogeneity between 3 pulpitis tissue and 3 healthy pulp tissue, and explored ferroptosis occurrence in pulpitis tissue and inflamed dental pulp cells (DPCs). In scRNA-seq, 40 231 cells (Pulpitis: 17 814; Healthy pulp: 22 417) were captured, and visualized into 12 distinct cell clusters. Differentially expressed ferroptosis-related genes (DE-FRGs) were almost presented in each cluster in pulpitis vs healthy pulp. ROS and Fe²⁺ levels significantly rose, and immunohistochemistry showed low expression of GPX4 and high expression of PTGS2 in pulpitis. In LPS-stimulated DPCs, thymosin α1 increased the expression of GPX4 and FTL, and decreased expression of TNF-α, IL-1β, IL-6, and Fe²⁺ levels. In rat pulpitis models, both prothymosin α (PTMA, precursor of thymosin α1) gelatin sponge placed at the hole of pulp (LPS-P(gs)) and PTMA injection in pulp (LPS-P(i)) significantly reduced infiltration of inflammatory cells and expression of PTGS2, and increased the expression of GPX4. In RNA sequencing, the expression of DE-FRGs were reversed when thymosin α1 were added in LPS-stimulated DPCs. Collectively, single-cell atlas reveals cellular heterogeneity between pulpitis and healthy pulp, and ferroptosis occurrence in pulpitis. Thymosin α1 may reduce ferroptosis in DPCs to alleviate pulpitis and thus potentially has the ability to treat pulpitis.
Ferroptosis/drug effects* ; Dental Pulp/drug effects* ; Animals ; Pulpitis/pathology* ; Rats ; Thymalfasin/pharmacology* ; Humans ; Male ; Thymosin/pharmacology* ; Disease Models, Animal ; Rats, Sprague-Dawley

OBJECTIVE: Hepatocellular carcinoma (HCC) is a leading cause of mortality worldwide. Huachansu (Cinobufacini) is active extract isolated from the dry skin of Bufo Bufo gargarizans. It has now been widely used in clinical treatment of cancer, this study is to clarify the material basis of down-regulation of thymidylate synthase (TYMS) induced by Huachansu. METHODS: Our study utilized UPLC-MS/MS to identify major bioactive components from Huachansu. Cell Counting Kit 8 (CCK-8) assay and clone formation assay were used to examine the cell viability of tumor cells. TYMS and γ-H2AX level were detected by using quantitative real-time RT-PCR and/or western blotting. Small interfering RNA (siRNA) transfection was used to explore whether inhibition of TYMS could enhance the suppressive effect of Huachansu on cell growth of HCC cells. RESULTS: In our study, firstly, we identify 21 major bioactive components from Huachansu. CCK-8 assay results showed that Huachansu and its bioactive bufadienolides (Bufalin, Bufotalin, Cinobufotalin, Desacetylcinobufagin, Arenobufagin, Telocinobufagin, and Resibufogenin) significantly inhibited the proliferation of HepG2 and SK-HEP-1 cells in a dose- and time-dependent manner. Further molecular mechanistic investigation demonstrates that Huachansu significantly suppresses thymidylate synthase (TYMS), the enzyme which provides the sole de novo source of thymidylate for DNA synthesis. The inhibition of TYMS could lead to cell-cycle block and DNA damage of HCC cells. Furthermore, we identified that Huachansu markedly increased γ-H2AX expression, which indicated the presence of DNA damage. Moreover, we confirmed that transfection of cells with small interfering RNA specific to TYMS could increase the suppressive effects of Huachansu on the HCC cells proliferation. Quantitative RT-PCR analysis showed that Huachansu treatment had no effect on the transcription level of TYMS. Furthermore, proteasomal inhibitor MG132 could block TYMS inhibition induced by Huachansu, and concomitant administration of protein synthesis inhibitor cycloheximide (CHX) with Huachansu could further suppress the protein level of TYMS, indicating that Huachansu promotes proteasome-dependent degradation of TYMS in liver cancer cells. More importantly, the bioactive bufadienolides of Huachansu such as Bufalin, Bufotalin, Cinobufotalin, Desacetylcinobufagin, Arenobufagin, Telocinobufagin, and Resibufogenin could also significantly restrain the protein level of TYMS, revealing the material basis of inhibition of TYMS exposed to Huachansu. 5-Fluorouracil (5-FU) is a TYMS inhibitor, we also evaluate the effects of the combined treatment of Huachansu with 5-FU, the results show that interactions between Huachansu and 5-FU are synergistic or antagonistic. Thus, in clinical, attention should be paid to the dosage of Huachansu in combination with 5-FU. CONCLUSION Huachansu inhibits the growth and induces DNA damage of human HCC cells through proteasome-dependent degradation of TYMS, bioactive bufadienolides are the material basis of down-regulation of TYMS induced by Huachansu.

Background and purpose:It is still a great challenge to clarify the signal molecules that mediate the communication between cancer-associated fibroblasts(CAFs)and tumor cells.These signal molecules are very important for cancer metastasis.The purpose of this study was to explore the communication mechanism of pleckstrin-2/miR-196a signal axis mediated by lung cancer cells in tumor microenvironment.Methods:Human lung adenocarcinoma cell line H1299 and human embryonic lung cell MRC-5 were selected as the research objects.H1299 cells were transfected with lentivirus(PLEK2)expressing PLEK2 and Vector control,and exosomes(Vector_exo,PLEK2_exo)were isolated after 24 h of transfection.MRC-5 cells were transfected with miR-196a mimetic or inhibitor.The expressions of PLEK2 and epithelial-mesenchymal transition(EMT)-related proteins were analyzed by Western blot.The expression of miR-196a was analyzed by polymerase chain reaction(PCR),and the metastasis and invasion ability of cells were determined by transwell assay.Six female BALB/c-nu mice were randomly divided into Vector group and PLEK2 group,with 3 mice in each group.Mice in each group were injected with H1299 cells transfected with Vector or PLEK2 through the tail vein.After 4 weeks,lung tissue was taken out for H-E staining and immunohistochemical staining to analyze the expression of α-smooth muscle actin(α-SMA).All animal experiments were approved by the ethics committee of First Hospital of Changsha City(Changsha Hospital,Xiangya School of Medicine,Central South University)(ethics number:EI-2021-103).Results:Compared with the Vector group,the number of pulmonary metastatic nodules and the expression of α-SMA in metastatic cancer in PLEK2 group increased significantly(P＜0.001).Compared with Vector group,the expression level of miR-196a in H1229 cells in PLEK2 group increased significantly(P＜0.05),and the expression level of miR-196a was significantly higher in PLEK2_exo than in Vector_exo(P＜0.05).Compared with Vector_exo group,the expression levels of miR-196a,α-SMA and fibroblast activation protein(FAP)in MRC-5 cells in PLEK2_exo group increased significantly(P＜0.05).Compared with the negative control(NC),the expression levels of α-SMA and FAP in MRC-5 cells transfected with miR-196a increased significantly(P＜0.05).On the contrary,by transfection with miR-196a inhibitors(si-miR-196a#1 and si-miR-196a#),the expression levels of α-SMA and FAP were significantly inhibited(P＜0.05).Compared with NC-CM group,the number of metastatic cells,invasive cells and the expression of vimentin in miR-196a-CM group increased significantly(P＜0.001),and the expression of E-cadherin decreased significantly(P＜0.001).In addition,compared with Vector_exo-CM group,PLEK2_exo-CM group had significant increase in number of metastatic and invasive cells and the expression of vimentin(P＜0.01),and significant decrease in the expression of E-cadherin(P＜0.001).Conclusion:Upregulation of PLEK2 can enhance the level of exosomes miR-196a derived from lung cancer cells,thereby promoting the activation of CAFs.The activated CAFs can further enhance the invasive ability of lung cancer cells.

Objective: To investigate the epidemiological characteristics of foodborne disease outbreaks in Shaoxing City, Zhejiang Province, from 2012 to 2022, so as to provide the evidence for improving the foodborne disease control strategy. Methods: Foodborne disease outbreaks in Shaoxing City from 2012 to 2022 were collected from National Foodborne Disease Outbreak Monitoring System in China, including populations, places of outbreak, pathogenic factors and suspected foods. The temporal distribution, regional distribution, distribution of outbreak places and pathogenic factors of foodborne disease outbreaks were descriptively analyzed. Results: A total of 89 foodborne disease outbreaks were reported in Shaoxing City from 2012 to 2022, covering totally 699 patients, with an average annual attack rate of 6.35%. The outbreak peaked during the period between June and October (73 outbreaks, 82.02%), and family was the predominant place of outbreak (41 outbreaks, 46.07%). There were 83 outbreaks with known pathogenic factors, including 51 outbreaks caused by microbial factors, with Vibrio parahaemolyticus, Salmonella and norovirus as predominant pathogens, and 29 outbreaks caused by fungi and their toxins, which were all poisonous mushrooms poisoning, resulting in 2 deaths. In addition, there were 3 outbreaks caused by chemical factors. Conclusions The outbreak of foodborne diseases predominantly occurred in summer and autumn in Shaoxing City from 2012 to 2022. Family was the predominant place of outbreak, and toxic mushroom poisoning was the most lethal pathogenic factor.

Objective:To explore the relationship between low serum albumin levels and its duration on first episode of peritonitis in peritoneal dialysis (PD) patients.Methods:PD patients who were regularly followed up in the Pearl River Delta region from September 1, 2000 to July 6, 2021 in Shunde Hospital of Southern Medical University, Nanfang Hospital of Southern Medical University, and Foshan First People′s Hospital were retrospectively selected. The patients were divided into low serum albumin group (LSA group, mean albumin<35 g/L), moderate serum albumin group (MSA group, 35 g/L≤mean albumin<40 g/L) and high serum albumin group (HSA group, mean albumin≥40 g/L) according to the mean albumin of the patients, and the differences among the three groups were compared. The Kaplan-Meier survival analysis method was used to compare the risk of peritonitis events in different mean albumin groups and different durations of hypoalbuminemia. The multivariate Cox regression model was used to analyze the relationship between serum albumin levels and duration of hypoalbuminemia and new-onset peritonitis.Results:A total of 1 853 PD patients were included in this study, aged (49.72±15.34) years, and 1 036(55.9%) males. There were 551 patients (29.7%) in the LSA group, 920 patients (49.7%) in the MSA group, and 382 patients (20.6%) in the HSA group. The median follow-up was 37 (15, 66) months and there were 508 patients (27.4%) with new-onset peritonitis during the follow-up. Compared with the LSA group, the incidence of new peritonitis in the MSA group and HSA group was lower ( χ2=14.053, P<0.001; χ2=21.857, P<0.001), but there was no significant difference in the incidence of new peritonitis between the HSA group and MSA group. The Kaplan-Meier survival analysis showed that the cumulative incidence of peritonitis in the LSA group was significantly higher than that in the MSA group and HSA group (Log-rank χ2=22.128, P<0.001). Compared with PD patients with normal serum albumin, the patients with longer duration of hypoalbuminemia tended to have a higher incidence of new peritonitis. Multivariate Cox regression analysis showed that the mean albumin<35 g/L (LSA group/MSA group, HR=1.495, 95% CI 1.198-1.866, P<0.001; LSA group/HSA group, HR=1.459, 95% CI 1.104-1.928, P=0.008) was an independent risk factor of new-onset peritonitis in PD patients and the prolongation of duration of hypoalbuminemia had a significantly higher risk of new-onset peritonitis ( HR=1.013, 95% CI 1.003-1.024, P=0.014). Conclusion:The mean albumin<35 g/L and prolong duration of hypoalbuminemia are independent risk factors of PD-related peritonitis in PD patients.

Human infections with novel avian influenza A(H7N9)virus was an emerging infectious disease discovered in March, 2013. As of June30, 2016, 770 cases of H7N9 were reported in worldwide including 315 deaths with 40.9% of high case fatality rate. Yangtze River Delta and Pearl River Delta were the high-prevalence area. Formerly, the cases of H7N9 were concentrated on the municipalities. However, most of the case-patients were from smaller cities or rural areas nowadays. The H7N9 human infections mainly occurred in winter and spring every waves as similar as seasonal and H5N1 human infections. Middle aged and old (the median age was 61 years) male patients were occupied the large proportion among the cases of H7N9. In addition, the phenomenon of the limited and unsustained human-to-human transmission were existed. At present, the 4 major epidemic waves had happened and human infections with novel avian influenza A (H7N9) virus could be outbreak regularly in China. In this paper, the pathogenic characteristics and disease distribution of H7N9 influenza A viruses were elaborated, with both transmission factors and control measures, which were helpful to provide the scientific evidence for prevention and control in H7N9avian influenza epidemic.

Objective To explore the association between clinical pathological characteristics and the expressions of epidermal growth factor receptor (EGFR) and protein kinase B (AKT) in gastric carcinomas.Methods The expressions of EGFR and AKT were measured with immunohistochemical method in the cancer tissues and cancer-adjacent normal tissues from 153 cases of patients with gastric cancer.The association between clinical pathological characteristics and their expressions were analyzed.Results The expressions of AKT and EGFR in gastric cancer tissues had no relationship with gender,age,pathological type,and the degree of differentiation (P ＞ 0.05).A positive correlation was existed between the EGFR and TNM stages (x2 =5.43,P ＜0.05).The AKT was positively related to the size,T stage,and TNM stage of the tumor,respectively (x2 =4.73,4.95,5.32,P ＜0.05 orP ＜0.01).The levels of AKT (x2=4.83,4.75,P ＜0.05) and EGFR(x2 =4.67,4.58,P ＜0.05) in the gastric cancer tissues with lymph node and/or distant metastasis were significantly higher than the gastric cancer tissues without metastasis,respectively.Conclusions The over-expressions of AKT and EGFR would benefit the diagnosis and stages of a gastric cancer and the determination of its metastasis.

Objective:To improve the sensitivity and the linear range of electrochemical immunosensor to detect Schistosoma japonicum (S.japonicum) antibody.Methods:Carbon inks and silver/silver chloride inks were printed on a polyethylene terephthalate (PET) board to make a two-electrode test strip,where carbon was the working electrode and S.japonicum soluble egg antigen (SEA) was fixed at one end of working electrode by different methods; silver/silver chloride electrode was used as control.We tested the valency of the antibody by cyclic voltammetry (CV) and differential pulse voltammetry (DPV) in an electrochemistry workstation,and conducted comparison with the results of ELISA.Two new immunosensing electrodes have been developed,based on glutaraldehyde cross-linked (GA) or chitosan-glutaraldehyde cross-linked (Chit-GA) transducer fixing S.japonicum antigen.We tested the titer of the antibody by means of CV and DPV.Results:Our experimental S.japonicum antigen (50 μg/L) is the optimal test concentration for the GA sensor,and 10 μg/L for Chit-GA sensors.The immune reaction time of both electrodes is all essentially complete in 1 minute.The linear range for S.japonicura antibody in human positive serum sample detection by the glutaraldehyde cross-linked immunosensor is 1∶1000 to 1∶400,and by the chitosan-glutaraldehyde cross-linked immunosensor is 1∶1000 to 1∶500.As the concentration of dilution ratio of S.japonicum antibody in human positive serum sample increased,the test value of DPV increased proportionally.Conclusion:GA sensor and Chit-GA cross-linked S.japonicum sensors have high sensitivity and broad linear range response,and both exhibited a good linear relationship between the DPV signal and the test antibody titer.

OBJECTIVE: To investigate the relationship between the expression of leptin, p-mTOR protein and the pathogenesis, development and clinicopathological features in colon carcinoma. METHODS: The expression of leptin and p-mTOR protein was evaluated by immunohistochemical methods in 40 normal colon mucosas, 40 colon adenomatous polyps and 108 cases of colon carcinomas. The relationship between the staining pattern and clinicopathogical features was examined. RESULTS: The positive rates of detection of leptin in normal colon mucosa, adenomatous polyps and colon carcinomas were 10% (4/40), 27.5% (11/40), and 71.3% (77/108), respectively; with significant differences among the three groups (P<0.05). The positive rates of p-mTOR protein in the normal colon mucosa, the adenomatous polyps, and the colon carcinomas were 2.5% (1/40), 20% (8/40), and 61.1% (66/108), respectively; with significant differences among the three groups (P<0.05). The expression of leptin and p-mTOR proteins were related to invasive depth, TNM stages, lymph node metastasis, distant metastasis and tumor differentiation (P<0.05), but not to age, sex, or site (P>0.05). In colon carcinoma tissues, leptin expression was positively correlated with p-mTOR expression (P<0.01). CONCLUSION Leptin and p-mTOR proteins may play important roles in the occurrence and development of colon carcinoma. The detection of leptin and p-mTOR may be helpful for evaluation of the prognosis of the patient with colon carcinoma.
Adenocarcinoma ; metabolism ; pathology ; Adenomatous Polyps ; metabolism ; pathology ; Aged ; Colon ; metabolism ; Colonic Neoplasms ; metabolism ; pathology ; Female ; Humans ; Intestinal Mucosa ; metabolism ; Leptin ; metabolism ; Male ; Middle Aged ; Neoplasm Metastasis ; Phosphorylation ; Prognosis ; TOR Serine-Threonine Kinases ; metabolism

Objective To prepare the infected Oncomelania hupensis by artificial method for the research on the activity, vaccine, and genetic variation of Schistosoma Japonicum (S. Japonicum).Methods The mature eggs of S. Japonicum were collected by Nylon silk method and the miracidia were incubated under appropriate conditions. Negative snails were infected with miracidia in different proportion by means of individual or collective infection to seek the best method and proportion of infection between miracidia and snails. Infected snails were divided into 12 groups in total. Ⅰ-Ⅵ groups were for individual infection and Ⅶ-Ⅻ groups were for collective infection. There were 200 snails in each group. The infection ratios between snails and miracidia in Group Ⅰ-Ⅵ or screened, numbered, and reared singly. The amount of cercariae was calculated once every 10 days until the infected snails died. Then cercariae shedding quantity, infection quantity, and mortality of infected snails in every group were compared to find the best infection method and the best infection proportion between miracidia and snails. The cercariae were collected from the first generation of infected snails and were used to infect experimental animals. The mature eggs of S. Japonicum were saved from the infected experimental animals and incubated to get miracidia. The snails were artificially infected by miracidium to get the second generation of infected snails. The developmental rates of adult worms, the egg density in fecal and liver were compared between artificially and naturally infected snails. Results In individual infection GroupⅠ-Ⅵ,the average infection value of snails were 0±0,22.7±4.2,31.7±4.5,53.0±5.3,39.3±5.9,32.7±4.7,the average fatality of snails were 21.7±3.1,25.0±3.6,31.3±4.9,44.7±6.5,78.3±9.5,89.7±13.6, and the average value of cercariae shedding from infected snails were 0.0±0.0,308.0±96.6,428.1±146.2,527.0±171.1,571.4±148.9,602.9±356.3, respectively. In collective infection Group Ⅶ-Ⅻ,the average infection value of snails were 0±0,12.3±2.5,18.7±4.7,28.3±4.2,33.3±4.7,29.3±5.5,and the average fatality of snails were 22.7±3.8,23.7±4.5,28.3±5.5,47.0±9.5,75.7±8.5,86.3±12.2, and the average value of cercariae shedding from infected snails were 0±0,244.5±57.3,292.3±74.8,347.1±100.8,477.2±142.1,447.3±161.4, respectively. The second generation of artificially infected snails was obtained successfully. The average infection rate and fatality rate for the second generation of artificially infected snails were 24.65% and 24.50%, both of which were not obviously different from that of the first generation of artificially infected snails (P>0.05). In the animal experiment, the worm growth rate for the naturally infected snails, the first or second generation of artificially infected snails were 68.50%,73.50% or 71.00%. There was no obvious difference among them (P>0.05). The fecal (or liver) eggs per gram for the naturally infected snails, the first or the second generation of artificially infected snails were 1 503±269,1 683±233, or 1 541±117 (or 6 641±1 819,6 272±1 419, or 7 263±1 643). There was no significant difference among the 3 groups (P>0.05). Conclusion Infected snails can be obtained through the artificial method by using S. Japonicum miracidia to infect snails. Individual infection has the advantage over collective infection. The optimal proportion of infection between first and the second generation of artificially infected snails in the average of cercariae shedding, infection, and fatality average of snails. There was no significant difference between artificially and naturally infected snails in the developmental rate of adult worms, fecal and liver eggs per gram.