Objective To develop and evaluate a nucleic acid amplification test for spectinomycin-resistant Neisseria gonorrhoeae(N.gonorrhoeae).Methods N.gonorrhoeae-specific primers NG1/NG2 and primers specific to the N.gonorrhoeae rpsE gene mutation(80_82 delTTA)were designed.Genomic nucleic acids of spectinomycin-sensitive and resistant N.gonorrhoeae,Escherichia coli,Pseudomonas aeruginosa,and Salmonella typhi were used as templates to be amplified by PCR and quantitative real-time PCR(qPCR).The sensitivity and specificity of the method were evaluated accordingly.Results The NG1/NG2 primers could effectively amplify specific fragments of N.gonorrhoeae,yielding negative results for the nucleic acid amplification test of the other types of bacteria tested.E64/E175R and E-87/E95R could effectively differentiate the wild type and mutant(80_82 delTTA)rpsE genes.In PCR reactions,the minimum limits of NG1/NG2,E64/E175R,and E87/E95R for the target genes were 414.8 copies,414.8 copies,and 4.1 copies/μL,respectively,while those for qPCR reactions were 41.5,41.5,and 4.1×10-2 copies/μL,respectively.Conclusion A nucleic acid amplification test for spectinomycin-resistant N.gonorrhoeae with high specificity and sensitivity was successfully established in this study,which is expected to provide support for the rapid diagnosis of N.gonorrhoeae infection and treatment decision-making in clinical settings.

Objective To investigate the role of a novel rpsE gene mutation in mediating high-level spectinomycin resistance in Neisseria gonorrhoeae and to evaluate its effect on the biological fitness of the bacteria.Methods Spectinomycin-containing medium was used to screen for Neisseria gonorrhoeae strains with spontaneous mutations that conferred spectinomycin resistance.Minimum inhibitory concentrations(MIC)were determined,and the rpsE gene was sequenced.Changes in the growth rates of spectinomycin-resistant strain were assessed using the drop plate method and growth curves.Additionally,in vitro competition experiments were conducted with spectinomycin at different concentrations to assess changes in the biological fitness of the spectinomycin-resistant strain.Results A Neisseria gonorrhoeae strain with high-level spectinomycin resistance mediated by a novel rpsE gene mutation(88_90delGTT)was successfully identified and designated NG-SPTR.Compare with the wild-type strain,the NG-SPTR exhibited reduced growth rate(optical density[OD]comparison,P<0.05).In addition,in vitro competition experiments showed a competitive index(CI)<1 in gonococcal base liquid(GCBL)without or with low-concentration spectinomycin(≤16 μg/mL).In the GCBL with 32 μg/mL spectinomycin,the CI value gradually increased from<1 before 18 h to>1 after 18 h.The mutant strain showed CI>1 in GCBL with spectinomycin concentrations≥64 μg/mL.Conclusion The rpsE gene mutation(88_90delGTT)mediates high-level spectinomycin resistance in Neisseria gonorrhoeae,and imposes a fitness cost on the bacteria.The biological fitness of the mutant strain is influenced by the concentration of spectinomycin.

Objective:To evaluate the item properties and applicability of the eHealth Literacy Scale in Web 3.0 context (eHLS-Web 3.0) among diabetic patients by applying classical test theory (CTT) and item response theory (IRT) to provide a new effective tool for assessing the level of eHealth literacy among diabetic patients.Methods:A cross-sectional study design was adopted and 230 diabetic patients from the inpatient endocrinology department of the Affiliated Yantai Yuhuangding Hospital of Qingdao University were selected by convenience sampling from November 2022 to May 2023 and were surveyed by using the self-general information questionnaire and eHLS-Web 3.0. The correlation coefficient method, Cronbach α coefficient, Guttman split-half reliability coefficient, and structural validity in CCT were used, and discriminant validity, convergent validity, and Samejima Graded Response model in IRT were used to evaluate the scale items. Results:The CTT results showed that Cronbach α coefficient for the eHLS-Web 3.0 was 0.973, the Guttman split-half reliability coefficient was 0.894, and the correlation coefficient between the items and the total score of the scale ranged from 0.708 to 0.845 (all P<0.01). Exploratory factor analysis extracted three common factors with a cumulative variance contribution rate of 76.75%, and the confirmatory factor analysis model had a good fit. The reliability of each dimension combination was >0.60, and the average variance extracted was >0.50. The square root of the average variance extracted from each dimension (0.817, 0.875, 0.845) was greater than the correlation coefficient between this dimension and other dimensions (0.742, 0.763, 0.715). The IRT results showed that the discrimination parameter of all the items ranged from 2.174 to 4.054, the difficulty coefficients increased with the increase of the level (- 2.471 - 2.187), and the average information content of all the items ranged from 1.217 to 2.609. Conclusions:The eHLS-Web 3.0 has good characteristics among diabetic patients as tested by CTT and IRT, and can be used as an effective tool to assess the eHealth literacy level of diabetic patients in China.

Background and purpose:Colorectal cancer(CRC)is one of the major malignant tumors threatening human health worldwide,with long-term high incidence and mortality rate.Potassium channel modulatory factor 1(KCMF1)is a member of the E3 ubiquitin ligase family.It binds to target proteins through the RING domain and participates in the regulation of a variety of biological processes in vivo.However,the function of KCMF1 in CRC remains unclear.This study aimed to investigate the expression level of E3 ubiquitin ligase KCMF1 in colorectal tumor,and to explore the effects of KCMF1 on the proliferation of CRC cells and its underlying molecular mechanism.Methods:The The Cancer Genome Atlas(TCGA)and Genotype-Tissue Expression(GTEx)databases were used to analyze the expression level of KCMF1 in CRC tissues and adjacent tissues and the association between the KCMF1 expression and the prognosis of CRC patients.Furthermore,immunohistochemical staining was performed to detect the protein level of KCMF1 in 90 paired human CRC tissues and adjacent non-tumor tissues.Lentiviral shRNA delivery system was employed to specifically target the KCMF1 gene(shKCMF1)in HCT116 and HCT15 CRC cell lines.The effects of KCMF1 knockdown on cell proliferation,apoptosis and cell cycle distribution were assessed by methyl thiazoyl terazolium(MTT)assay,colony formation assay,Western blot and flow cytometry.Changes in the transcriptional profile in HCT116 cells upon KCMF1 knockdown were identified by RNA sequencing(RNA-Seq),and the affected signaling pathways were evaluated by bioinformatics analysis.Real-time fluorescence quantitative polymerase chain reaction(RTFQ-PCR),Western blot,luciferase reporter assay and cell immunofluorescence assay were utilized to validate the alteration of the affected signaling pathway.Results:The TCGA and GTEx databases and IHC results showed that the mRNA and protein expression levels of KCMF1 in CRC tissues were significantly upregulated compared with adjacent tissues(P＜0.01).KCMF1 expression level was negatively correlated with the survival time of patients with CRC(P＜0.01),and was positively associated with CRC clinical stage(P＜0.05).Compared with control cells,KCMF1 knockdown significantly inhibited the proliferation of HCT116 and HCT15 cells(P＜0.001),induced cell apoptosis(P＜0.001),and led to cell cycle arrest in G1 phase(P＜0.01).RNA-Seq analysis showed that KCMF1 was involved in the regulation of several signaling pathways,including nuclear factor-κB(NF-κB)signaling pathway.KCMF1 knockdown reduced the transcription levels of the target genes of NF-κB signaling pathway,including BCL-XL,XIAP and CIAP(P＜0.05),and suppressed the expression of phosphorylated p65 and nuclear translocation of p65(P＜0.01).Meanwhile,the activity of NF-κB reporter was reduced in tumor cells upon KCMF1 knockdown(P＜0.01).Conclusion:The expression of KCMF1 is significantly upregulated in human CRC tissues and positively associated with advanced clinical stage and poor prognosis.KCMF1 may promote the proliferation of CRC cells by activating the NF-κB signaling pathway.KCMF1 may be a potential new therapeutic target for CRC.

Objective To establish and evaluate a microbial sensitivity test method for Neisseria gonorrhoeae based on resazurin coloration.Methods Based on the broth microdilution method,resazurin was added as a live bacteria indicator.WHO G,a WHO gonococcal reference strain,was used to optimize the incubation time for resazurin-stained bacteria and the color change was visually observed to obtain the results.Agar dilution method(the gold standard)and resazurin-based microdilution assay were used to determine the minimum inhibitory concentration(MIC)of azithromycin,ceftriaxone,and spectinomycin for 3 reference strains and 32 isolates of Neisseria gonorrhoeae.The results were analyzed based on essential agreement(EA),which reflected the consistency of the MIC values,category agreement(CA),which reflected the consistency in the determination of drug resistance,intermediary,and sensitivity,very major error(VME),which reflected false sensitivity,and major error(ME),which reflected pseudo drug resistance,to evaluate the accuracy of resazurin-based microdilution assay as a microbial sensitivity test of of Neisseria gonorrhoeae.CA and EA rates≥90%and VME and ME rates≤3%were found to be the acceptable performance rates.Results The results obtained 6 hours after resazurin was added were consistent with those of the agar dilution method and the resazurin-based microdilution assay was established accordingly based on this parameter.The EA of resazurin-based microdilution assay for measuring the MIC results of azithromycin,ceftriaxone,and spectinomycin was 97.1%,91.5%,and 94.3%,respectively,and the CA was 88.6%,94.3%,and 94.3%,respectively.The VME was 0%for all three antibiotics,while the ME was 11.4%,5.7%,and 5.7%,respectively.Conclusion The resazurin-based microdilution assay established in this study showed good agreement with agar dilution method for measuring the MIC of antibiotics against Neisseria gonorrhoeae.Moreover,the sensitivity results of this method were highly reliable and could be easily obtained through naked eye observation.Nonetheless,the results of drug resistance should be treated with caution and the optimization of parameters should be continued.

Objective:To study the risk factors of adverse pregnancy outcomes for the diagnosis and treatment of pregnancy after cesarean section complicated with placenta previa.Methods:A national multicenter retrospective study was conducted to select a total of 747 pregnant women with the third trimester singleton pregnancy after cesarean section complicated with placenta previa from 12 tertiary hospitals in January 1st to December 31st, 2018. The risk factors of severe adverse outcomes [hysterectomy, intraoperative blood loss ≥1 000 ml, intraoperative diagnosis of placenta accreta spectrum disorders (PAS)] in pregnant women with second pregnancy complicated with placenta previa after cesarean section were investigated by logistic regression analysis. The roles of prenatal ultrasonography and magnetic resonance imaging (MRI) in the prediction of PAS and severe adverse outcomes were observed. According to whether vascular intervention was performed (uterine artery embolization or abdominal aortic balloon occlusion), the pregnant women were divided into the blocked group and the unblocked group, and the maternal and infant perinatal outcomes between the two groups were compared.Results:(1) General information: the hysterectomy rate of 747 pregnant women with second pregnancy complicated with placenta previa after cesarean section was 10.4% (78/747), the intraoperative blood loss ≥1 000 ml in 55.8% (417/747), and PAS was confirmed in 47.5% (355/747). The incidence of uterine rupture was 0.8% (6/747). (2) Analysis of risk factors for severe adverse outcomes: based on binary unconditioned logistic regression univariate and multivariate analysis, the risk factors for hysterectomy were the mode of vascular embolization and intraoperative blood loss. The probability of hysterectomy with uterine artery embolization was 5.319 times higher than that with abdominal aortic balloon occlusion (95% CI: 1.346-21.018). The risk factors of intraoperative blood loss ≥1 000 ml were the number of cesarean section delivery, ultrasonography indicated PAS and suspected PAS, intraoperative PAS and complete placenta previa. The risk factors for intraoperative PAS were uterine scar thickness, ultrasonography indicated PAS and suspected PAS, MRI indicated PAS and suspected PAS, and complete placenta previa. (3) The roles of ultrasonography and MRI in predicting PAS: the sensitivity and specificity of ultrasonography in predicting PAS were 47.5% and 88.4%; the kappa value was 0.279 ( P<0.001), with fair agreement. The sensitivity and specificity of MRI to predict PAS were 79.2% and 97.8%, respectively. The kappa value was 0.702 ( P<0.001), indicating a good agreement. The intraoperative blood loss and hysterectomy rate of pregnant women with PAS indicated by ultrasonography and MRI were significantly higher than those with PAS only by ultrasonography or MRI. (4) Influence of vascular occlusion on pregnancy outcome: there were no significant differences in intraoperative blood loss and incidence of intraoperative bleeding ≥1 000 ml between the blocked group and the unblocked group (all P>0.05). There was no significant difference in intraoperative blood loss between the pregnant women with abdominal aortic balloon occlusion, uterine artery embolization and those without occlusion ( P=0.409). The hysterectomy rate of pregnant women with uterine artery embolization was significantly higher than those with abdominal aortic balloon occlusion [39.3% (22/56) vs 10.0% (5/50), P=0.001]. Conclusions:In the third trimester of pregnancy with placenta previa after cesarean section, MRI examination has better consistency in predicting PAS than ultrasonography examination. Ultrasonography examination combined with MRI examination could effectively predict the hysterectomy rate and intraoperative blood loss. Vascular occlusion could not reduce the amount of intraoperative blood loss. The hysterectomy rate of pregnant women with uterine artery embolization is higher than those with abdominal aortic balloon occlusion.

Sulforaphane (SFN), a natural anti-tumor compound from cruciferous vegetables, has been reported to induce protective autophagy to cancer cells, which might impair the anti-tumor efficiency of SFN. However, the accurate function and mechanism of SFN inducing autophagy in cancers are still obscure, especially in esophageal squamous cell carcinoma (ESCC), one of malignancies with high incidence in North China. Here, we mainly explored the potential function of autophagy upon SFN treatment in ESCC and molecular mechanism. We demonstrated that SFN could inhibit cell proliferation and induce apoptosis by activating caspase pathway. Moreover, we found activation of NRF2 pathway by SFN was responsible for the induction of autophagy and also a disadvantage element to the anti-tumor effects of SFN on ESCC, indicating that SFN might induce protective autophagy in ESCC. We, therefore, investigated effects of autophagy inhibition on sensitivity of ESCC cells to SFN and found that chloroquine (CQ) could neutralize the activation of SFN on NRF2 and enhance the activation of SFN on caspase pathway, thus improved the anti-tumor efficiency of SFN on ESCC

Objective:To study the risk factors of adverse pregnancy outcomes for induced abortion of cesarean scar pregnancy in midtrimester.Methods:A national multicenter retrospective study was conducted. A total of 154 singletons pregnant women with cesarean scar pregnancy during the second trimester induced abortion by various reasons in 12 tertiary A hospitals were selected, their pregnant outcomes were observed and the risk factors of serious adverse outcomes were analyzed with univariate and multivariate logstic regression; the role of ultrasound and MRI in predicting placenta accreta and severe adverse outcomes was evaluated, the effectiveness of uterine artery embolization (UAE) in preventing hemorrhage in pregnant women with and without placenta accreta was compared.Results:Among 154 subjects, the rate of placenta accreta was 42.2% (65/154), the rate of postpartum hemorrhage≥1 000 ml was 39.0% (60/154), the rate of hysterectomy was 14.9% (23/154), the rate of uterine rupture was 0.6% (1/154). The risk factor of postpartum hemorrhage≥1 000 ml and hysterectomy was placenta accreta ( P<0.01). For each increase in the number of parity, the risk of placenta accreta increased 2.385 times (95% CI: 1.046-5.439; P=0.039); and the risk of placenta accreta decreased with increasing ultrasound measurement of scar myometrium thickness ( OR=0.033, 95% CI: 0.001-0.762; P=0.033). The amount of postpartum hemorrhage and hysterectomy rate in the group with placenta accreta diagnosed by ultrasound combined with MRI were not significantly different from those in the group with placenta accreta diagnosed by ultrasound only or MRI only (all P>0.05). For pregnant women with placenta accreta, there were no significant difference in the amount of bleeding and hysterectomy rate between the UAE group [median: 1 300 ml; 34% (16/47)] and the non-embolization group (all P>0.05); in pregnant women without placenta accreta, the amount of bleeding in the UAE group was lower than that in the non-embolization group (median: 100 vs 600 ml; P<0.01), but there was no significant difference in hysterectomy rate [2% (1/56) vs 9% (3/33); P>0.05]. Conclusions:(1) Placenta accreta is the only risk factor of postpartum hemorrhage≥1 000 ml with hysterectomy for induced abortion of cesarean scar pregnancy in midtrimester; multi-parity and ultrasound measurement of scar myometrium thickness are risk factors for placenta accreta. (2) The technique of using ultrasound and MRI in predicting placenta accreta of cesarean scar pregnancy needs to be improved. (3) It is necessary to discuss of UAE in preventing postpartum hemorrhage for induced abortion of cesarean scar pregnancy in midtrimester.

Objective:To explore the effect of intervention plan based on a shared decision-making model on the diabetes knowledge, self-efficacy, self-management behavior, and blood glucose level of patients with diabetic ketosis.Methods:From November 2019 to March 2020, a total of 212 patients with diabetic ketosis who were admitted to the Endocrinology Department of a ClassⅢGrade A hospital in Yantai were recruited by convenience sampling. Among them, 104 patients admitted from July to October 2019 were involved into the control group, and 108 patients for the experimental group. The control group was given regular health education, and the experimental group was given the education based on the shared decision-making model. The changes of diabetes knowledge, diabetes management self-efficacy, self-management behavior, fasting blood glucose (FBG) , 2-hour postprandial blood glucose (2hPG) , glycosylated hemoglobin value (HbA1c) in two groups were compared before intervention, 3 months and 6 months after intervention.Results:3 months and 6 months after the intervention, the scores of diabetes knowledge with 85 (89.3, 70.0) and 88 (71.3, 92) , diabetes self-efficacy with 77.5 (70.0, 88.0) and 79.2 (63.8, 92.4) , self-management behavior with [64.5 (55.8, 74.8) ] and [62.5 (53.9, 74.2) ]in the experimental group were significantly higher than those of the control group ( P<0.05) . The level of FBG, 2hPG, and HbA1c of the experimental group were lower than those of the control group, and the differences were statistically significant ( P<0.05) . There was no significant difference in the recurrent cases of ketosis between the two groups within 6 months ( P>0.05) . Conclusions:The self-management intervention based on the shared decision-making model as the starting point may improve the level of diabetes knowledge, diabetes management self-efficacy and self-management behavior and stabilize the level of blood glucose of patients with diabetic acidosis.

Dysregulation of mTORC1/mTORC2 pathway is observed in many cancers and mTORC1 inhibitors have been used clinically in many tumor types; however, the mechanism of mTORC2 in tumorigenesis is still obscure. Here, we mainly explored the potential role of mTORC2 in esophageal squamous cell carcinoma (ESCC) and its effects on the sensitivity of cells to mTOR inhibitors. We demonstrated that RICTOR, the key factor of mTORC2, and p-AKT (Ser473) were excessively activated in ESCC and their overexpression is related to lymph node metastasis and the tumor-node-metastasis (TNM) phase of ESCC patients. Furthermore, we found that mTORC1/ mTORC2 inhibitor PP242 exhibited more efficacious anti-proliferative effect on ESCC cells than mTORC1 inhibitor RAD001 due to RAD001-triggered feedback activation of AKT signal. Another, we demonstrated that down-regulating expression of RICTOR in ECa109 and EC9706 cells inhibited proliferation and migration as well as induced cell cycle arrest and apoptosis. Noteworthy, knocking-down stably RICTOR significantly suppresses RAD001-induced feedback activation of AKT/PRAS40 signaling, and enhances inhibition efficacy of PP242 on the phosphorylation of AKT and PRAS40, thus potentiates the antitumor effect of RAD001 and PP242 both and . Our findings highlight that selective targeting mTORC2 could be a promising therapeutic strategy for future treatment of ESCC.