ObjectiveThe effects of two microbial fertilizers （Bacillus subtilis fertilizer and Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer） on the growth and development， the accumulation of active ingredients， and the microbial community diversity of rhizosphere soil of Atractylodes chinensis were investigated. MethodsA field experiment was carried out with two-year-old Atractylodes chinensis as the test material. Plant samples were collected during the wilt stage （September 26， 2023） to determine the general agronomic traits of Atractylodes chinensis. High-performance liquid chromatography （HPLC） was utilized to evaluate the effects of microbial fertilizers on the synthesis and accumulation of four active ingredients （atractylodin， atractylon， β-eudesmol， and atractylenolide Ⅰ） in Atractylodes chinensi. PacBio Sequel sequencing technology was used to explore the differences in bacterial community structures and diversity in the rhizosphere soil of Atractylodes chinensis treated with different microbial fertilizers. ResultsThe two microbial fertilizers had significant growth-promoting effects on Atractylodes chinensis. Compared with those of the CK group， the stem diameter， stem and leaf dry and fresh weight， and rhizome dry and fresh weight of Atractylodes chinensis significantly increased by 0.47-1.07 times （P<0.05） after the application of the Bacillus subtilis fertilizer （16 kg/667 m²）， and those significantly increased by 0.62-0.96 times （P<0.05） after the application of the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer （1.5 kg/667 m²）. The effect on plant height was not significant. The application of two microbial fertilizers was beneficial to the accumulation of atractylodin， atractylon， β-eudesmol， and atractylenolide Ⅰ （P<0.01）， and the effect of the Bacillus subtilis fertilizer on the accumulation of active ingredients of Atractylodes chinensis was better than that of the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer. The results of high-throughput sequencing showed that compared with the CK group， the Bacillus subtilis fertilizer （8 kg/667 m²） could significantly increase the diversity of rhizosphere bacterial species by regulating the Simpson index and Shannon index （P<0.05）， and the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer significantly reduced the bacterial diversity （P<0.05）. The relative abundance of dominant bacteria was compared at the phylum and genus levels. The relative abundance of Proteobacteria （45.73%） and Burkholderia_Caballeronia_Paraburkholderia （9.98%） significantly increased after the application of the Bacillus subtilis fertilizer （P<0.01）， and the relative abundance of Acidobacteriota （20.53%） and Sphingomonas （3.63%） increased significantly （P<0.01） after the application of the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer. The relative abundance of beneficial bacteria in the Bacillus subtilis fertilizer was slightly higher than that in the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer. Pearson correlation analysis showed that Burkholderia_Caballeronia_Paraburkholderia and Sphingomonas were positively correlated with the content of atractylodin， atractylon， β-eudesmol， and atractylenolide Ⅰ （P<0.05）. ConclusionThe application of the Bacillus subtilis fertilizer and Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer can increase the yield of medicinal materials and promote the synthesis and accumulation of active ingredients by regulating the rhizosphere microecological diversity of Atractylodes chinensis， and the application effect of the Bacillus subtilis fertilizer is better than that of the Trichoderma harzianum-Purpureocillium lilacinum compound fertilizer.

Objective:Bone morphogenetic protein-4(BMP4)has been proved to be an important regulatory factor for the pathological process of atherosclerosis(AS).However,there are few related clinical studies.This study aims to investigate the levels of plasma BMP4 in patients suffering from the arterial occlusive diseases(ACD)characterized by AS,and further to test the relationship between BMP4 and inflammation and vascular injury. Methods:A total of 38 ACD patients(the ACD group)and 38 healthy people for the physical examination(the control group)were enrolled.The plasma in each subject from both groups was obtained to test the levels of BMP4,tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β),IL-10,and vascular endothelial cadherin(VE-cadherin),and the relationship between BMP4 and the detected indicators above were further analyzed. Results:Compared with the control group,the patients in the ACD group displayed significant elevations in the neutrophil to lymphocyte ratio[NLR,1.63(1.26,1.91)vs 3.43(2.16,6.61)]and platelet to lymphocyte ratio[PLR,6.37(5.26,7.74)vs 15.79(7.97,20.53)],while decrease in the lymphocyte to monocyte ratio[LMR,5.67(4.41,7.14)vs 3.43(2.07,3.74)](all P<0.05).Besides,the ACD patients displayed significant elevations in plasma BMP4[581.26(389.85,735.64)pg/mL vs 653.97(510.95,890.43)pg/mL],TNF-α[254.16(182.96,340.70)pg/mL vs 293.29(238.90,383.44)pg/mL],and VE-cadherin[1.54(1.08,2.13)ng/mL vs 1.85(1.30,2.54)ng/mL],and decrease in IL-10[175.89(118.39,219.25)pg/mL vs 135.92(95.80,178.04)pg/mL](all P<0.05).While the levels of IL-1β remained statistically comparable between the 2 groups(P=0.09).Furthermore,the plasma BMP4 levels were further revealed to be positively correlated with the levels of IL-1β(r=0.35),TNF-α(r=0.31)and VE-cadherin(r=0.47),while they were negatively correlated with the levels of IL-10(r=-0.37;all P<0.01). Conclusion:After ACD occurrence,the patients'plasma concentrations of BMP4 would be upregulated,which may serve as a candidate to indicate the levels of inflammation and vascular injury.

Objective:This study aimed to explore the status of radiological Kashin-Beck disease (KBD) among school-aged children in Chamdo City, Tibet, through a 3-year monitoring survey, providing epidemiological evidence for prevention and control strategies.Methods:The target areas for this study were Luolong, Bianba, and Basu counties in Chamdo City, Tibet Autonomous Region, identified as having the most severe historical cases of KBD. Children aged 7-12 years attending school were enrolled as study subjects. Anteroposterior X-ray films of the right-hand were taken, and radiological diagnoses were made based on the "Diagnosis of Kashin-Beck Disease" criteria (WS/T 207-2010). Two experienced researchers independently reviewed the X-rays, and intra- and inter-group consistency were assessed using weighted Kappa values and percentage agreement. Cross-sectional surveys were conducted in 2017 and 2020 to describe the X-ray detection rates of KBD, and logistic regression analysis was employed to construct a predictive model of risk factors for radiological KBD cases.Results:In 2017, a total of 5,711 children aged 7-12 years in Chamdo City, Tibet, participated in the baseline cross-sectional survey (average age 9.2 years, 48.0% female), with 28 cases of radiological KBD. The age- and gender-standardized prevalence rate was 0.527%. In 2020, 6,771 participants (average age 9.3 years, 49.5% female) underwent a second cross-sectional survey, with 9 cases of radiological KBD and a standardized prevalence rate of 0.134%. Logistic regression analysis indicated that older age [ OR=2.439, 95% CI(1.299, 4.580), P=0.006] and female gender [ OR=8.157, 95% CI(1.016, 65.528), P=0.048] were independent risk factors for radiological KBD cases. Conversely, higher residential altitude, under the premise of Tibet's high altitude, was a protective factor [ OR=0.995, 95% CI(0.990, 0.999), P=0.032). Conclusion:The radiographically positive detection rate of KBD among school-aged children in Chamdo City, Tibet Autonomous Region, is at an extremely low level and showing a declining trend, reaching the historical standard in 2020. Considering the absence of positive signs in affected children, it suggests that local KBD has been effectively eliminated.

Objective:To construct a recombinant bioluminescent bacteriophage (HT7) targeting Escherichia coli, and evaluate its ability to identify Escherichia coli. Methods:Initially, pCRISPR-sg (1-10) and PFN-1000 plasmid strains were constructed by genetic engineering, and the most efficient small guild RNA (sgRNA) were screened by bilayer plate. By the gene editing technique, which comprised homologous recombination and clustered regularly interspaced short palin dromic repeats (CRISPR)-Cas system, the Nanoluc luciferase gene was integrated into the downstream non-coding region of 10A gene of T7 phage, to constructe the bioluminescent phage HT7 successfully. The difference of biological characteristics between HT7 phage and T7 phage was evaluated by plaque assay and liquid amplification assay. In addition, 51 strains of Escherichia coli, 20 strains of Klebsiella pneumoniae, 14 strains of Staphylococcus aureus, 6 strains of Enterococcus faecium, 5 strains of Enterococcus faecalis, 3 strains of Acinetobacter baumannii and 1 strain of Pseudomonas aeruginosa were collected and isolated to evaluate the limit of detection and specificity of HT7 phage. Results:Among the 10 CRISPR-targeted cleavage systems constructed, sgRNA8 exhibited the highest cleavage efficiency, with a cleavage rate of 0.18. After three rounds of recombination screening using the pCas9/pCRISPR/PFN-1000 triple-plasmid system, PCR validation yielded recombinant phage bands at 2 798 bp, indicating the successful construction of the HT7 phage. The recombinant phage showed significant differences in biological characteristics in terms of lysis efficiency ( P<0.001), one-step growth curve ( P=0.001), and infection multiplicity ( P=0.031). Both lysis burst time and log growth node were extended by 10 min, with the optimal infection multiplicity being 0.1. Clinical sample testing identified lysis of 6 strains of Escherichia coli within 4.5 h, while other strains remained unaffected, with detection of pathogenic bacteria below 10 CFU/ml. Conclusions:The developed pCas9/pCRISPR/PFN-1000 triple-plasmid editing system efficiently edits the bacteriophage genome. The constructed HT7 fluorescent bacteriophage enables the detection of Escherichia coli below 10 CFU/ml within 4.5 hours, demonstrating low detection limits and high detection specificity.

Gallbladder carcinoma is a common malignant tumor of the biliary system characterized by poor specificity of early symptoms， a high degree of malignancy， and rapid progression， and it is difficult to make an early diagnosis. Gallstones and gallbladder polyps are considered the most common risk factors for gallbladder carcinoma. Ultrasound is the preferred examination， while CT， MRI， and PET also have their own advantages. There is a lack of radical treatment methods for gallbladder carcinoma， and surgical operation remains the preferred treatment method for gallbladder carcinoma； however， due to the rapid progression of this disease， most patients have lost the opportunity for surgery at the time of diagnosis. A combination of various treatment modalities， such as radiochemotherapy， targeted therapy， and immunotherapy， has improved the prognosis of patients to a certain extent， but with an unsatisfactory long-term therapeutic effect. Therefore， it is of particular importance to give priority to prevention rather than treatment and emphasize early identification and treatment.

Hypertensive disorder of pregnancy (HDP) involves two major public health issues: mother-infant safety and prevention and controlling major chronic disease. HDP poses a serious threat to maternal and neonatal safety, and it is one of the leading causes of maternal and perinatal morbidity and mortality worldwide, as well as an important risk factor for long-term cardiovascular disease (CVD). In order to explore effective strategies to prevent and control the source of CVD and reduce its risk, we have established a cohort of HDPs in Shenzhen for the primordial prevention of CVD. The construction of the HDP cohort has already achieved preliminary progress till now. A total of 2 239 HDP women have been recruited in the HDP cohort. We have established a cohort data management platform and Biobank. The follow-up and assessment of postpartum cardiovascular metabolic risk in this cohort has also been launched. Our efforts will help explore the pathophysiological mechanism of HDP, especially the pathogenesis and precision phenotyping, prediction, and prevention of pre-eclampsia, which, therefore, may reduce the risk of adverse pregnancy outcomes, and provide a bridge to linking HDP and maternal-neonatal cardiovascular, metabolic risk to promote the cardiovascular health of mothers and their infants.

Building a strong public health system has become an urgent task in the new era. Based on more than eight years of systematic research， we believe that five aspects need to be prioritized for a strong system. First， we should change the perspective on public health， using the word “gonggong jiankang” to replace “gonggong weisheng” and the word “gonggong jiankang tixi” to replace “gonggong weisheng tixi”， to lead the public health system development. Second， we should develop a suitable public health system and continuously improve the health capacity for governance. Third， we should make it clear that the goal of building a strong system is not far-fetched， and we need to consolidate the existing institutional advantages of China’s public health system： when encountering major problems， we can maintain a unified goal and mobilize the whole society to cooperate effectively to accomplish the goal. However， we need to make up for shortcomings one by one， especially to solve the key problem of lacking a strong coordination mechanism in daily work. Fourth， we should pursue excellence and consolidate the“suitable” mechanism proven in the process of coping with the COVID-19， so that efficient mechanisms to deal with major issues can be used in routine work， and efforts should be made to consolidate the advantages of prevention and control of infectious diseases and emergency response，so as to achieve the balanced development of regions categories and units.Finally， it is necessary to strengthen the coordination of government and research institutions， in the aspects of technological innovation， talent team building and accurate consulting services， and work together to pursue a suitable and strong system to realize the modernization of the health system and capacity for governance.

Abstract The growth and development of children is related to the future of the country and the nation. In recent years, there have been more and more cohort studies in the field of children s mental health. Bymainly introducing the advantages of cohort studies on children in distress and organizes domestic and foreign cohort studies in the field of mental health of children in distress, this article finds that it is mostly used in depression, anxiety, post traumatic stress disorder, suicidal ideation and attempts, etc, and mainly explores the risk and prevalence of mental health development in children in distress, and identifies long term negative damage. The research aims to promote the long term development and high quality development of such research by analyzing and summarizing the status quo and prospects of cohort research in the field of mental health of children in distress.

The family of voltage-gated potassium Kv2 channels consists of the Kv2.1 and Kv2.2 subtypes. Kv2.1 is constitutively highly phosphorylated in neurons and its function relies on its phosphorylation state. Whether the function of Kv2.2 is also dependent on its phosphorylation state remains unknown. Here, we investigated whether Kv2.2 channels can be phosphorylated by protein kinase C (PKC) and examined the effects of PKC-induced phosphorylation on their activity and function. Activation of PKC inhibited Kv2.2 currents and altered their steady-state activation in HEK293 cells. Point mutations and specific antibodies against phosphorylated S481 or S488 demonstrated the importance of these residues for the PKC-dependent modulation of Kv2.2. In layer II pyramidal neurons in cortical slices, activation of PKC similarly regulated native Kv2.2 channels and simultaneously reduced the frequency of action potentials. In conclusion, this study provides the first evidence to our knowledge that PKC-induced phosphorylation of the Kv2.2 channel controls the excitability of cortical pyramidal neurons.
Action Potentials ; HEK293 Cells ; Humans ; Protein Kinase C/metabolism* ; Pyramidal Cells/enzymology* ; Shab Potassium Channels/genetics*

Objective:To investigate the 2 years’ efficacy of intravesical instillation of domestic BCG versus epirubicin in the prevention of recurrence of intermediate-risk or high-risk non-muscular invasive bladder cancer and predictive factors of BCG instillation.Methods:From July 2015 to June 2020, 18-75 years old patients with moderate to high-risk non muscle invasive bladder cancer (NMIBC) confirmed by pathological examination were involved. The ECOG score was 0-2. Exclusion criteria included ①immune deficiency or impairment (such as AIDS), using immunosuppressive drugs or radiotherapy, suspected allergic to BCG or epirubicin or excipients of the two drugs, fever or acute infectious diseases including active tuberculosis or receiving anti tuberculosis treatment, with severe chronic cardiovascular and cerebrovascular diseases or chronic kidney disease; ②combined with other urogenital system tumors or other organ tumors; ③combined with muscle invasive bladder urothelial carcinoma (≥T 2); ④undergoing chemotherapy, radiotherapy or immunotherapy within 4 weeks (immediate instillation after surgery not included); ⑤ pregnant or lactating women; ⑥ comfirmed or suspected bladder perforation; ⑦gross hematuria; ⑧cystitis with severe bladder irritation that may affect the evaluation; ⑨participat in other clinical trials within 3 months; ⑩alcohol or drug addiction; ?any risk factors that may increasing the risk of patients. Epirubicin 50 mg was irrigated immediately after the operation(TURBT or laser resection). The patients were randomly divided into BCG15 group, BCG19 group and epirubicin group by the ratio of 2∶2∶1, and the patients were maintained intravescical instillation for 1 year. The recurrence and adverse events of the three groups were compared. Univariate and multivariate analysis was performed to predict the risk factors of BCG irrigated therapy failure. Result:By June 15, 2020, the median follow-up duration was 22.1 months(12.1, 32.3), and there was no statistical difference between the groups ( P=0.9024). There were 274 patients enrolled in BCG19 group, 277 patients enrolled in BCG15 group and 130 patients enrolled in the epirubicin group. The drop-off rate was 16.6%(113 cases)and made no difference between groups( P=0.6222). There were no significant difference in age, gender, BMI, or ECOG score( P>0.05). During the follow-up, 116 cases was detected recurrence or progression. The recurrence rate of the three groups was 14.2% and 14.8% in BCG19 group and BCG15 group, and 27.7% in the epirubicin group. There was no difference in recurrence rate between BCG19 and BCG15 group( P=0.9464). The recurrence rate of BCG19 group was lower than that of the epirubicin group ( P=0.0017). The recurrence rate of BCG15 group was lower than that of the epirubicin group ( P=0.0020). There was no difference in the cumulative recurrence free survival rate between BCG19 and BCG15 group (95% CI0.57-1.46, P=0.7173). The cumulative recurrence free survival rate of BCG 19 group was better than that of the epirubicin group( HR=0.439, 95% CI0.26-0.74, P=0.0006), and the cumulative recurrence free survival rate of BCG15 group was better than that of the epirubicin group ( HR=0.448, 95% CI0.29-0.80, P=0.0021). The total incidence of adverse events in 19 BCG19, BCG15 and epirubicin group were 74.5%, 72.6% and 69.8% respectively. There was no difference in the incidence of adverse events between BCG19 and BCG15 group( P=0.6153). The incidence of adverse events in epirubicin group was lower than that of BCG19( P=0.0051) and BCG15( P=0.0167) groups.There was no significant difference in the incidence of serious adverse events (SAE) among the three groups ( P=0.5064). Log rank test univariate analysis and Cox risk regression model multivariate analysis showed that the history of bladder cancer recurrence( HR=6.397, 95% CI1.95-20.94, P=0.0001)was independent risk factor for BCG irrigation failure. Conclusions:The 2 years’ efficacy of intravesical instillation of domestic BCG is better than than of epirubicin with good tolerance and safety. There is no difference between BCG19 and BCG15 group. BCG doesn’t increase SAE compared with epirubicin. Recurrence status was an independent prognostic factor regarding recurrence-free survival.