Reverse effect of curcumin on CDDP-induced drug-resistance via Keap1/p62-Nrf2 signaling in A549/CDDP cell

Objective: To assess the effect of curcumin on CDDP-induced drug resistance and explore the underlying molecular mechanism through Nrf2 system and autophagy pathway.Methods: A drug-resistant cell model was established by exposing A549/CDDP cell to2 μg/mL CDDP. A549/CDDP cell was treated with 20 μg/mL CDDP and 10 μM curcumin. The cell viability and apoptosis level, the signals of Keap1/P62-Nrf2 and autophagy pathway were analyzed.Results: CDDP induction promoted drug-resistant phenotype in A549/CDDP cell and activated autophagy as well as Nrf2 signals in A549/CDDP cell. Meanwhile, curcumin combination attenuated autophagy and Nrf2 activation induced by CDDP, and reversed the drug-resistant phenotype. Notably, curcumin combination augmented Keap1 transcription. Furthermore, Keap1 ablation with short hairpin RNAs hampered the efficacy of curcumin, suggesting Keap1 played a crucial role on reversal effect of curcumin.Conclusions: The present findings demonstrate that CDDP promotes abnormal activation of Nrf2 pathway and autophagy, leading to drug resistance of A549/CDDP cell.Curcumin attenuates this process and combat drug-resistance through its potent activation on Keap1 transcription, which is essential for interplay between oxidative stress induced Nrf2 activation and autophagy/apoptosis switch.

Comparison of extended spectrum β-lactamasesproducing Escherichia coli with non-ESBLsproducing E.coli:drug-resistance and virulence

The virulent factors of Escherichia coil （E.cofi） play an important role in the process of pathopoiesis. The study aimed to compare drug-resistant genes and virulence genes between extended spectrum β-1actamases （ESBLs）-producing E.coli and non-ESBLs-producing E.cofi to provide a reference for physicians in management of hospital infection. From October 2010 to August 2011,96 drug-resistant strains of E. coli isolated were collected from the specimens in Qingdao Municipal Hospital, Qingdao, China. These bacteria strains were divided into a ESBLs-producing group and a non-ESBLs-producing group. Drug sensitivity tests were performed using the Kirby-Bauer （K-B） method. Disinfectant gene, qacEAl-sull and 8 virulence genes （CNF2, hlyA, eaeA, VT1, est, bfpA, elt, and CNF1） were tested by polymerase chain reaction （PCR）. Among the 96 E.coli isolates, the ESBLs-producing E.coli comprised 46 （47.9%） strains and the non-ESBLs-producing E.cofi consisted of 50 （52.1%） strains. The detection rates of multiple drug-resistant strain, qacEAl-sull, CNF2, hlyA, eaeA,VT1, est, bfpA, elt, and CNF1 in 46 ESBLs-producing E.coli isolates were 89.1%, 76.1%, 6.5%, 69.6%, 69.6%, 89.1%, 10.9%, 26.1%, 8.7%, and 19.6%, respectively. In the non-ESBLs-producing E.cofi strains, the positive rates of multiple drug-resistant strain, qacEAl-sull, CNF2, hlyA, eaeA, VT1, est, bfpA, elt, and CNF1 were 62.0%, 80.0%, 16.0%, 28.0%, 64.0%, 38.0%, 6.0%, 34.0%, 10.0%, and 24.0%, respectively. The difference in the detection rates of multiple drug-resistant strain, hlyA and VT1 between the ESBLs-producing E.cofi strains and the non-ESBLs-producing E.cofi strains was statistically significant （P〈0.05）. The positive rate of multiple drug-resistant strains is higher in the ESBLs-producing strains than in the non-ESBLs-producing strains. The expression of some virulence genes hlyA and VT1 varies between the ESBLs-producing strains and the non-ESBLs-producing strains. Increased awareness of clinicians and enhanced testing by laboratories are required to reduce treatment failures and prevent the spread of multiple drug-resistant strains.

Three-dimensional collagen-based scaffold model to study the microenvironment and drug-resistance mechanisms of oropharyngeal squamous cell carcinomas

Objective:Squamous cell carcinoma(SCC)represents the most common histotype of all head and neck malignancies and includes oropharyngeal squamous cell carcinoma(OSCC),a tumor associated with different clinical outcomes and linked to human papilloma virus(HPV)status.Translational research has few available in vitro models with which to study the different pathophysiological behavior of OSCCs.The present study proposes a 3-dimensional(3 D)biomimetic collagen-based scaffold to mimic the tumor microenvironment and the crosstalk between the extracellular matrix(ECM)and cancer cells.Methods:We compared the phenotypic and genetic features of HPV-positive and HPV-negative OSCC cell lines cultured on common monolayer supports and on scaffolds.We also explored cancer cell adaptation to the 3 D microenvironment and its impact on the efficacy of drugs tested on cell lines and primary cultures.Results:HPV-positive and HPV-negative cell lines were successfully grown in the 3 D model and displayed different collagen fiber organization.The 3 D cultures induced an increased expression of markers related to epithelial–mesenchymal transition(EMT)and to matrix interactions and showed different migration behavior,as confirmed by zebrafish embryo xenografts.The expression of hypoxia-inducible factor 1α(1α)and glycolysis markers were indicative of the development of a hypoxic microenvironment inside the scaffold area.Furthermore,the 3 D cultures activated drug-resistance signaling pathways in both cell lines and primary cultures.Conclusions:Our results suggest that collagen-based scaffolds could be a suitable model for the reproduction of the pathophysiological features of OSCCs.Moreover,3 D architecture appears capable of inducing drug-resistance processes that can be studied to better our understanding of the different clinical outcomes of HPV-positive and HPV-negative patients with OSCCs.

Molecular Characterization and Drug-resistance of Mycobacterium tuberculosis Strains in Xuzhou, China

To understand the genetic diversity and drug resistance status of Mycobocterium tuberculosis （M. tuberculosis） circulating in Xuzhou of China, the spacer-oligonucleotide typing （Spoligotyping） and multi-loci VNTRs （variable number tandem repeats） analysis （MLVA） were utilized for the genotyping of the isolates. Drug susceptibility test （DST） was performed by the proportion method on the Lowenstein-Jensen （L-J） medium using isoniazid, rifampicin, ethambutol, and streptomycin. By Spoligotyping, 287 M. tuberculosis isolates were differentiated into 14 clusters. Then with 15-1oci MLVA, these strains could be divided into 32 clusters, 228 genotypes. Of 15 VNTRs, 6 loci had the highly discriminatory powers, 6 loci presented moderate discrimination and 3 loci demonstrated less polymorphism. The DST results showed that 46 strains were resistant to at least one first-line anti-tuberculosis agent. There was a difference in the isoniazid resistance between Beijing and non-Beijing genotype strains. We concluded that the combination of Spoligotyping and 15 VNTR loci as the genotyping in our study was applicable for this region, the drug resistant isolates were identified, and the Beijing family was the most prevalent genotype in the rural counties of Xuzhou.

Investigation of immune escape-associated mutations of hepatitis B virus in patients harboring hepatitis B virus drug-resistance mutations

It is unclear whether immune escape-associated mutations in the major hydrophilic region of hepatitis B virus surface antigen(HBsAg)are associated with nucleoside/nucleotide analog resistance.AIM To evaluate the association between immune escape-associated mutations and nucleoside/nucleotide analog resistance mutations.METHODS In total,19440 patients with chronic hepatitis B virus infection,who underwent resistance testing at the Fifth Medical Center of Chinese PLA General Hospital between July 2007 and December 2017,were enrolled.As determined by sequence analysis,6982 patients harbored a virus with resistance mutations and 12458 harbored a virus lacking resistance mutations.Phenotypic analyses were performed to evaluate HBsAg production,replication capacity,and drug-induced viral inhibition of patient-derived drug-resistant mutants with or without the coexistence of sA159V.RESULTS The rate of immune escape-associated mutation was significantly higher in 9 of the 39 analyzed mutation sites in patients with resistance mutations than in patients without resistance mutations.In particular,these mutations were sQ101H/K/R,sS114A/L/T,sT118A/K/M/R/S/V,sP120A/L/Q/S/T,sT/I126A/N/P/S,sM133I/L/T,sC137W/Y,sG145A/R,and sA159G/V.Among these,sA159V was detected in 1.95%(136/6982)of patients with resistance mutations and 1.08%(134/12,458)of patients lacking resistance mutations(P<0.05).The coexistence of sA159V with lamivudine(LAM)and entecavir(ETV)-resistance mutations in the same viral genome was identified during follow-up in some patients with drug resistance.HBsAg production was significantly lower and the replication capacity was significantly higher,without a significant difference in LAM/ETV susceptibility,in sA159V-containing LAM/ETV-resistant mutants than in their sA159V-lacking counterparts.CONCLUSION In summary,we observed a close link between the increase in certain immune escape-associated mutations and the development of resistance mutations.sA159V might increase the fitness of LAM/ETV-resistant mutants under environmental pressure in some cases.

Dihydroartemisinin inhibits plasmid transfer in drug-resistant Escherichia coli via limiting energy supply

Conjugative transfer of antibiotic resistance genes(ARGs)by plasmids is an important route for ARG dissemination.An increasing number of antibiotic and nonantibiotic compounds have been reported to aid the spread of ARGs,highlighting potential challenges for controlling this type of horizontal transfer.Development of conjugation inhibitors that block or delay the transfer of ARG-bearing plasmids is a promising strategy to control the propagation of antibiotic resistance.Although such inhibitors are rare,they typically exhibit relatively high toxicity and low efficacy in vivo and their mechanisms of action are inadequately understood.Here,we studied the effects of dihydroartemisinin(DHA),an artemisinin derivative used to treat malaria,on conjugation.DHA inhibited the conjugation of the IncI2 and IncX4 plasmids carrying the mobile colistin resistance gene(mcr-1)by more than 160-fold in vitro in Escherichia coli,and more than two-fold(IncI2 plasmid)in vivo in a mouse model.It also suppressed the transfer of the IncX3 plasmid carrying the carbapenem resistance gene bla_(NDM-5)by more than twofold in vitro.Detection of intracellular adenosine triphosphate(ATP)and proton motive force(PMF),in combination with transcriptomic and metabolomic analyses,revealed that DHA impaired the function of the electron transport chain(ETC)by inhibiting the tricarboxylic acid(TCA)cycle pathway,thereby disrupting PMF and limiting the availability of intracellular ATP for plasmid conjugative transfer.Furthermore,expression levels of genes related to conjugation and pilus generation were significantly down-regulated during DHA exposure,indicating that the transfer apparatus for conjugation may be inhibited.Our findings provide new insights into the control of antibiotic resistance and the potential use of DHA.

A STUDY OF DECREASING DRUG-RESISTANCE OF HUMAN OVARIAN CANCER CELLS IN VITRO

A chemosensitivity test for ovarian cancer using tritiated thymidine incorporation assay was carried out. A dose-response relationship for cisplatin and potentiation of verapamil in increasing vincristine inhibition to ovarian cancer were investigated. A 5- fold increase of cisplatin density converted the tumors which were initially resistance to standard-dose cisplatin Into drug-sensitive ones. Vera-pamil was found to be able to overcome vincristine-resistance of some tumors in vitro. These results suggest that using high dose cisplatin therapy or increasing local drug concentration by using other administration way, we could expect some ovarian cancers that had failed to standard dose cisplatin therapy to be effective. Combination of vincristine with verapamll may be helpful in treating some vincristineresistant cases.

Global burden of HIV-negative multidrug-and extensively drug-resistant tuberculosis based on Global Burden of Disease Study 2021

Background:Tuberculosis(TB),caused by Mycobacterium tuberculosis,remains the second leading cause of death from a single infectious disease globally and poses a significant economic and clinical burden in the world in 2022.Of particular concern is the emergence of drug-resistant TB,accounting for 15%-20%of TB deaths.It is imperative to delve into the global trends of incidence and death rate for multidrug-resistant tuberculosis(MDRTB)and extensively drug-resistant tuberculosis(XDR-TB),drawing upon the comprehensive Global Burden of Disease(GBD)2021 drug-resistant tuberculosis dataset.Methods:From the GBD 2021,data on incidence,prevalence,disability-adjusted life years(DALYs),and death of MDR-TB and XDR-TB from 1990 to 2021 were collected.We calculated the estimated annual percentage changes in age standardized incidence rate(ASIR)and age-standardized death rate(ASDR),segmented by age,sex,and socio-demographic index(SDI).The impacts of various risk factors on MDR-TB and XDR-TB were also analyzed.Results:In 2021,there were an estimated 443,680(95%uncertainty interval[UI]:259,196-766,545)incident cases of MDR-TB,and an estimated 106,818(95%UI:41,612-211,854)death cases of MDR-TB,while there were an estimated 24,036(95%UI:17,144-34,587)incident cases of XDR-TB and 7,946(95%UI:3,326-14,859)death cases of XDR-TB.The incidence and death cases of MDR-TB were lowest in high SDI regions,whereas the incidence rates of XDR-TB in high-middle SDI regions were higher than those in middle SDI and high SDI regions.Conclusion:This study reported the disease burden of drug-resistant TB from 1990 to 2021.Until 2021,drugresistant TB is still a serious problem in low SDI countries,especially for high-risk age populations with highrisk factors.Controlling drug-resistant TB requires effective control strategies and healthcare systems.

Mobile genetic elements facilitate the transmission of antibiotic resistance genes in multidrug-resistant Enterobacteriaceae from duck farms

Multidrug-resistant(MDR)Enterobacteriaceae critically threaten duck farming and public health.The phenotypes,genotypes,and associated mobile genetic elements(MGEs)of MDR Enterobacteriaceae isolated from 6 duck farms in Zhejiang Province,China,were investigated.A total of 215 isolates were identified as Escherichia coli(64.65%),Klebsiella pneumoniae(12.09%),Proteus mirabilis(10.23%),Salmonella(8.84%),and Enterobacter cloacae(4.19%).Meanwhile,all isolates were resistant to at least two antibiotics.Most isolates carried tet(A)(85.12%),blaTEM(78.60%)and sul1(67.44%)resistance genes.Gene co-occurrence analysis showed that the resistance genes were associated with IS26 and integrons.A conjugative IncFII plasmid pSDM004 containing all the above MGEs was detected in Proteus mirabilis isolate SDM004.This isolate was resistant to 18 antibiotics and carried the blaNDM-5 gene.MGEs,especially plasmids,are the primary antibiotic resistance gene transmission route in duck farms.These findings provide a theoretical basis for the rational use of antibiotics in farms which are substantial for evaluating public health and food safety.

blaNDM-1 Carried by a Transferable Plasmid in a Salmonella Strain Isolated from Healthy Individuals

Objective Our study aimed to conduct genomic characterization of Salmonella strains carrying the blaNDM-1 gene in the intestinal tract of healthy individuals.The objectives were to underscore the importance of genomic surveillance for drug resistance in both commensal and pathogenic bacteria among healthy populations,and to establish protocols for regulating drug resistance plasmids based on the completion of a comprehensive map of drug resistance plasmid genomes.Methods We performed antimicrobial susceptibility testing and employed second-and third-generation sequencing techniques to analyze Salmonella strains harboring the blaNDM-1 gene,to surveil drug-resistant bacteria in the intestines of healthy subjects.Sequence comparison was conducted using both core-and pan-genome approaches.Concurrently,conjugation experiments were carried out to assess the efficiency of plasmid transfer.Results We isolated a carbapenem-resistant Salmonella enterica serovar Typhimurium strain from a healthy food worker in China.This strain harbored an IncHI2/IncHI2A plasmid carrying blaNDM-1 along with multiple antibiotic resistance genes(ARGs).Our findings highlight the potential for asymptomatic carriers to facilitate the transmission of ARGs.Pan-genomic analysis revealed that blaNDM-1-positive plasmids could traverse bacterial species barriers,facilitating cross-host transmission.Conclusion This study marks the first detection of blaNDM-1 in Salmonella strains isolated from healthy individuals.We underscore the risk associated with the transmission of conjugative hybrid plasmids carrying blaNDM-1,which have the potential to be harbored and transmitted among healthy individuals.Enhanced surveillance of drug-resistant pathogens and plasmids in the intestinal microbiota of healthy individuals could provide insights into the risk of ARG transmission and pathways for population-wide dissemination via ARG transfer factors.

Observation of Two Novel mcr-10.1–Carrying Plasmids in Clinical Escherichia coli Strains

Polymyxins are a class of last-resort antibiotics used to treat infections caused by multidrug-resistant bacteria. Mobile resistance determinants for polymyxins, including mcr-1 to mcr-10 genes, have previously been reported. Among them, mcr-10 has commonly beenobserved in Enterobacter and Klebsiella species. However, the presence of mcr-10 in Escherichia coli, an important opportunistic pathogen, has rarely been reported. This work describes the observation of mcr-10.1 in two clinical E. coli strains, with mcr-10.1 hosted ontwo novel plasmids, one of which carries transconjugation genes. These strains were isolated from anal fistula–suffering patients, suggesting their close relation to human bacterial infection. The genetic context of mcr-10.1 was also found to differ from those previouslyidentified in E. coli strains. This work is the first observation of mcr-10.1–carrying E. coli in clinical settings, expanding our knowledge ofthis important antibiotic resistance gene.

Plasmid DNA Analysis of Pathogenic Escherichia coli in Musk Deer

[Objective] The pathogenic Escherichia coli in musk deer was classified at molecular level to provide basic materials for molecular epidemiology of pathogenic Escherichia coli in musk deer. [Method] Plasmids from 24 pathogenic Escherichia coli in musk deer were extracted by the Lysis Triton method, and then identified by single enzyme digestion with three endonucleases of Hind Ⅲ, EcoR Ⅰ and BamH Ⅰ. [Result] The yield rate of plasmids was 91.6%, and 24 pathogenic Escherichia coli in musk deer had the identical or similar plasmid profiles. [Conclusion] Plasmid DNA analysis offers scientific basis for molecular epidemiology of pathogenic Escherichia coli in musk deer in Sichuan Institute of Musk Deer Breeding.

Researches of Agrobacterium rhizogenes Ri Plasmid rol Genes

Expression of rol genes from Ri plasmid of Agrobacterium rhizogenes not only leads to the excessive formation of adventitious roots, but also exhibits various genetically modified characteristics that have broad prospects for the application of plant genetic improvement. Since the 1980s of the last century, much progress has been made in the studies of A. rhizogenes, in particular the agropine type Ri plasmid rol genes and their applications for plant genetic improvement, which involves the structure and function of Ri plasmid, the characters of rol genes, the influence of rol genes expression on plants growth and development, and the applications of rol genes for genetic improvement of forest tree. In this paper, the advances in this field are reviewed and the existing problems about the application of rol genes for genetic improvement of forest tree are also discussed.

Curing of the Bacillus subtilis Plasmid Using Sodium Dodecyl Sulfate

Curing of Bacillus subtilis plasmid using sodium dodecyl sulfate (SDS)was studied in order to obtain a host strain. An overnight culture of Bacillus subtilis 24/pMX45 was used to inoculate fresh LB containing SDS (0-0.008%). No growth of 24/pMX45 was observed when LB contained an SDS concentration of 0.006% or greater, and the sublethal concentration (w/v) of SDS was 0.005% with a killing rate of 99%. Samples were diluted and plated on LB agar, individual colonies were randomly picked to a selective agar medium by tooth to screen for loss of plasmid-encoded erythomycin resistance. CsCl-EtBr gradient centrifugation and plasmid DNA profile demonstrated that plasmid-cured derivative A7 has completely lost its plasmid. A7 had a shorter lag, and its cell concentration was consistently higher than that of the 24/pMX45. Elimination of the plasmid was first observed after 24/pMX45 had been treated with SDS for 8 h. The percent elimination then continued to increase until about 22 h, after which the fraction of cured cell in the population remained constant. Plasmid cured cell numbers were measured in a separate control culture of 24/pMX45 untreated by SDS. No spontaneous loss of pMX45 was observed after 24/pMX45 were incubated for 24 h and 48 h with shaking at 37 ℃.These results suggested that SDS can be used as curing agent to eliminate the plasmid of Bacillus subtilis.

Construction of the Plasmid Reference Molecule for Detection of Transgenic Soybean MON89788

[Objective] The aim was to construct a plasmid reference molecule (PRM) for detection of transgenic soybean MON89788. [Method] the lectin gene sequence,3'-junction and 5'-junction sequence between host plant DNA integrated DNA of MON89788 soybean were amplified independently,and the three fragments were cloned into the cloning vector pMD18-T in order through molecular manipulation method to construct pMD-LM3M5,the applicability of the constructed novel PRM was tested. [Result] Sequencing confirmation result showed that the PRM was 3 700 bp in length,containing 1 029 bp of recombined DNA fragment. The limits of qualitative detection of the PRM were 10 copies. [Conclusion] The PRM constructed in this study was suitable for the identification of MON89788 event.

质粒介导的利奈唑胺耐药基因optrA在粪肠球菌的水平转移机制

目的研究临床上耐利奈唑胺粪肠球菌(LREf)携带的optrA基因通过质粒介导在粪肠球菌间的水平转移机制。方法收集昆明市某三级甲等医院2022年8月—2023年8月留取的非重复LREf,使用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF MS)仪进行鉴定,VITEK 2 Compact、纸片扩散法、微量肉汤稀释法进行药敏试验,聚合酶链式反应(PCR)检测optrA基因,全基因组测序(WGS)分析LREf的分子生物学特征,以LREf为供体菌、临床来源粪肠球菌为受体菌进行接合实验。结果共收集17株LREf,12株LREf的质粒检测出optrA基因,检测出多种耐药基因及毒力基因。12株LREf菌株以ST16(50.0%)为主要分型。接合试验的24株接合子中有8株接合成功,接合率为33.3%,进一步分析发现,菌株接合前后位于质粒上的optrA基因上下游均存在IS1216E插入序列,且形成IS1216E-fexA-optrA-erm(A)-IS1216E样的转座单元。结论optrA基因可通过同时携带erm(A)、fexA基因的质粒介导在临床粪肠球菌间进行水平转移,而插入序列IS1216E在其水平转移过程中发挥着重要作用。

环境中抗生素抗性基因的健康风险分级

细菌耐药性的快速传播和蔓延是全球公共健康领域的重大挑战。除了医疗和动物养殖业外,抗生素抗性基因可在多种环境介质中传播扩散进而转移到人类病原菌中,影响人类健康。甄别环境中高风险抗性基因是实现抗性基因有效环境监测和风险评估的基础。对环境中抗性基因的研究现状进行了总结,聚焦质粒携带的抗性基因,提出了环境中抗性基因风险分级框架,进而对制药厂和养殖厂场地环境样品进行了质粒组分析,形成了风险分级清单。结果表明,按抗性基因的移动性(与可移动遗传元件共存)、质粒类型、在人类病原菌中的流行度、致病潜力(与毒力因子共存)等4个标准,从制药厂和养殖厂场地环境样品中共检测到425个质粒携带的抗性基因,其中最高风险Ⅰ级抗性基因0种,Ⅱ级2种,Ⅲ级33种,Ⅳ级89种,最低风险Ⅴ级301种,形成了制药厂和养殖厂场地环境抗性基因风险分级清单。所检测到的高风险抗性基因,为环境抗性基因污染监测及人群暴露健康风险评估提供了理论和数据支撑。

钙/钙调蛋白依赖性蛋白激酶Ⅱ质粒的构建和表达及其与Ca_(V)1.2通道结合的鉴定

目的构建钙/钙调蛋白依赖性蛋白激酶Ⅱ(CaMKⅡ)长片段的融合蛋白质粒,表达、提取、纯化,并鉴定其与Ca_(V)1.2通道蛋白的结合。方法将提取的pGEX-6p-1/CaMKⅡ长片段质粒转化到原核大肠杆菌BL21感受态细胞,摇床培养箱中培养12 h,添加异丙基-β-D-硫代半乳糖苷,促进GST融合蛋白的表达。采用DTT联合超声破碎法,谷胱甘肽-琼脂糖凝胶4B(GS-4B)分离、纯化GST-CaMKⅡ长片段后,经过PreScission蛋白酶处理,切除GST标签,得到CaMKⅡ长片段蛋白。采用15%SDS-PAGE鉴定CaMKⅡ长片段蛋白的分子量和相对纯度;采用Bradford方法测定纯化后蛋白的浓度;采用pull-down结合Western blotting鉴定其与Ca_(V)1.2通道蛋白的结合能力。结果测序结果表明,CaMKⅡ长片段构建成功。通过DTT联合超声破碎法,得到了较高纯度和浓度的长片段CaMKⅡ蛋白,并能够与心肌Ca_(V)1.2钙通道末端CT1蛋白结合。结论本研究成功构建了CaMKⅡ长片段钙调蛋白激酶融合蛋白质粒,提取、纯化了能够与Ca_(V)1.2通道蛋白结合并具有生物活性的CaMKⅡ长片段蛋白,为深入研究CaMKⅡ蛋白的功能奠定了基础。

The role of axon guidance molecules in the pathogenesis of epilepsy

Current treatments for epilepsy can only manage the symptoms of the condition but cannot alter the initial onset or halt the progression of the disease. Consequently, it is crucial to identify drugs that can target novel cellular and molecular mechanisms and mechanisms of action. Increasing evidence suggests that axon guidance molecules play a role in the structural and functional modifications of neural networks and that the dysregulation of these molecules is associated with epilepsy susceptibility. In this review, we discuss the essential role of axon guidance molecules in neuronal activity in patients with epilepsy as well as the impact of these molecules on synaptic plasticity and brain tissue remodeling. Furthermore, we examine the relationship between axon guidance molecules and neuroinflammation, as well as the structural changes in specific brain regions that contribute to the development of epilepsy. Ample evidence indicates that axon guidance molecules, including semaphorins and ephrins, play a fundamental role in guiding axon growth and the establishment of synaptic connections. Deviations in their expression or function can disrupt neuronal connections, ultimately leading to epileptic seizures. The remodeling of neural networks is a significant characteristic of epilepsy, with axon guidance molecules playing a role in the dynamic reorganization of neural circuits. This, in turn, affects synapse formation and elimination. Dysregulation of these molecules can upset the delicate balance between excitation and inhibition within a neural network, thereby increasing the risk of overexcitation and the development of epilepsy. Inflammatory signals can regulate the expression and function of axon guidance molecules, thus influencing axonal growth, axon orientation, and synaptic plasticity. The dysregulation of neuroinflammation can intensify neuronal dysfunction and contribute to the occurrence of epilepsy. This review delves into the mechanisms associated with the pathogenicity of axon guidance molecules in epilepsy, offering a valuable reference for the exploration of therapeutic targets and presenting a fresh perspective on treatment strategies for this condition.

Construction and Application of Plasmid pUC19-CM-D

[Objective] The aims were to construct a new suicide plasmid of Lactobacillus and gene deletion engineering bacteria of Lactobacillus with pUC19 vector. [Methods] pUC19-CM was constructed by inserting a chloramphenicol resistant gene into the multi-cloning site of pUC19,and then two homologous fragments were cloned into each side of the pUC19-CM to construct suicide plasmid pUC19-CM-D. [Results] A replacement mutant strain,whose target gene was replaced by resistant gene,could be obtained by transforming the suicide plasmid pUC19-CM-D into Lactobacillus for resistance screening. [Conclusion] The construction and application of pUC19-CM-D provided a fast and efficient means of construction of gene deletion engineering bacteria of Lactobacillus,and laid a foundation for study of gene function of Lactobacillus.