关键词： Cell death  

摘要： 5-Fluorouracil (5-FU) is a commonly used chemotherapeutic agent for breast cancer. Its efficacy relies on the function of p53, and mutations in p53 contribute to the development of resistance during 5-FU chemotherapy. Here, we report that microcurrent stimulation (MCS) of a p53-mutant breast cancer cell line induces p53-mediated cell death. Although MDA-MB-231 and MDA-MB-468 cells, both human breast cancer cell lines, are less sensitive to 5-FU due to p53 mutations, MCS (300 μA for 30 min) induced apoptosis in these cells and improved the antitumor effect of 5-FU in tumor-bearing mice. MCS-induced apoptosis was mediated by an increase in intracellular Cu2+ ions and reactive oxygen species, along with the concurrent transcriptional enhancement of pro-apoptotic genes by p53. Furthermore, MCS induced apoptosis in MDA-MB-231 cells that had developed resistance to 5-FU and inhibited tumor growth in tumor-bearing mice with reduced 5-FU sensitivity. These findings suggest that an approach involving MCS could serve as a foundation for developing breast cancer treatment strategies to overcome p53 mutations. © 2025 The Authors

关键词： Hill equation   p53 target protein expression   protein accumulation   stability of gene expression   steady state   transcription factor pulsing  

摘要： The central dogma indicates the basic direction of gene expression pathways. For activated gene expression, the quantitative relationship between various links from the binding of transcription factors (TFs) to DNA to protein synthesis remains unclear and debated. There is consensus that at a steady state, protein levels are largely determined by the mRNA level. How can we find this steady state? Taking p53 as an example, based on the previously discovered Hill-type equation that characterizes mRNA expression under p53 pulsing, I proved that the same equation can be used to describe the average steady state of target protein expression. Therefore, at steady state, the average fold changes in mRNA and protein expression under TFs pulsing were the same. This consensus has been successfully demonstrated. For the p53 target gene BAX, the observed fold changes in mRNA and protein expression were 1.40 and 1.28, respectively;the fold changes in mRNA and protein expression calculated using the Hill-type equation were both 1.35. Therefore, using this equation, we can not only fine-tune gene expression, but also predict the proteome from the transcriptome. Furthermore, by introducing two quantitative indicators, we can determine the degree of accumulation and stability of protein expression.

关键词： Calcium/calmodulin-dependent protein kinase II   kilohertz frequency electrical stimulation   N-methyl-D-aspartate receptor 2B   neuropathic pain  

摘要： Objectives: Kilohertz high-frequency electrical stimulation (KHES), an avant-garde neuromodulation strategy, is progressively emerging in the field of neuropathic pain management, showing its unique therapeutic potential. This study delves into the mechanisms by which KHES exerts therapeutic effects on neuropathic pain induced by chronic constriction injury (CCI) in mice through modulation of the calcium/calmodulin-dependent protein kinase II (CaMKII)/N-methyl-D-aspartate receptor 2B (NMDAR2B) signaling pathway. Materials and Methods: In this study, mice were randomly assigned to groups and received intrathecal injections of CaMKII activator BayK8644, CaMKII inhibitor KN93, and N-methyl-D-aspartate (NMDA). Subsequently, mice underwent a week-long KHES treatment, with each session lasting 30 minutes. The impact of KHES on mechanical allodynia and thermal hyperalgesia in mice was assessed through paw withdrawal threshold and thermal withdrawal latency measurements, respectively. In addition, anxiety and depressive-like behaviors in mice were evaluated using pole climbing, open field, and forced swim tests. Quantitative reverse transcription polymerase chain reaction, Western blot, and immunofluorescence techniques were used to detect the expression levels of CaMKII, phosphorylated CaMKII (p-CaMKII), and NMDAR2B in the spinal cord. Results: Results indicated that KHES not only significantly reduced mechanical allodynia in CCI mice, with a sustained analgesic effect lasting up to six hours, but also somewhat alleviated anxiety and depressive-like symptoms. KHES inhibited the expression of p-CaMKII and NMDAR2B in the spinal cord. This inhibitory effect was reversed in the presence of BayK8644 and NMDA, suggesting that activation of CaMKII and NMDAR2B may contribute to the maintenance of neuropathic pain. Conversely, KN93 enhanced the analgesic effect of KHES by reducing mechanical allodynia and downregulating p-CaMKII and NMDAR2B expression, further confirming the signif

关键词： gain-of-function   mutant p53   p53 expression   p53 immunohistochemistry   TP53 mutation   triple-negative breast cancer  

摘要： AimsImmunohistochemical (IHC) staining of p53 is a potential marker for TP53 mutations in various cancers. However, criteria for predicting TP53 mutations in triple-negative breast cancer (TNBC) using p53 IHC staining are not yet established. We aim to correlate p53 IHC expression patterns with TP53 mutation status in *** and ResultsA total of 113 TNBC cases were analysed for p53 IHC staining pattern and somatic TP53 mutation using whole-exome sequencing. Functional properties of TP53 mutations were determined using the National Cancer Institute (NCI) TP53 database. P53 IHC patterns were categorized as nuclear overexpression (n = 58), null pattern (n = 40), wildtype (n = 15), cytoplasmic (n = 5), and subclonal (n = 5). The cutoff for predictive p53 nuclear overexpression was determined to be 80%, which strongly correlated with TP53 mutations. Notably, p53 overexpression had a positive predictive value (PPV) of 83% for missense or in-frame mutations, while the null pattern showed a PPV of 85% for detecting nonsense, frameshift, or splicing mutations. P53 overexpression was significantly linked to missense mutations within the DNA-binding domain (DBD) exhibiting gain-of-function (GOF) or dominant-negative effect (DNE). Cases exhibiting cytoplasmic expression correlated with nonsense or frameshift mutations in the DBD, nuclear localization signal (NLS), or splice sites. Cases with subclonal p53 staining patterns were associated with TP53 *** study proposes newly defined criteria for interpreting p53 immunostaining patterns in TNBC, potentially allowing for the prediction of TP53 mutation types and their functional implications.

关键词： Hexavalent chromium   HIF-1 alpha   MCT1   Lactate utilization  

摘要： Hexavalent chromium [Cr (VI)] is a known environmental pollutant, which promotes tumorigenesis. Hypoxiainducible factor-1 alpha (HIF-1 alpha) is crucial for cancer development. Here, we found that Cr (VI) treatment promoted lactate utilization by increasing monocarboxylate transporter 1 (MCT1) and monocarboxylate transporter 4 (MCT4) expression, while increasing the expression of HIF-1 alpha in A549 cells but reducing HIF-1 alpha and MCT1 in HELF cells. CoCl2, an HIF-1 alpha inducer, increased MCT1, while the HIF-1 alpha inhibitor YC-1 and MCT1 inhibitor AZD3965 suppressed Cr (VI)-induced lactate utilization and cell growth. Chromatin immunoprecipitation (ChIP) assay revealed HIF-1 alpha bound to the MCT1 promoter to enhance its transcription. Using Reactivating p53 and Inducing Tumor Apoptosis (RITA), which can increase the protein level of p53, we discovered that the low level of p53 protein in A549 cells determined the effect of Cr (VI)-induced HIF-1 alpha. These findings highlighted the role of p53 protein level in the effects of Cr (VI) on HIF-1 alpha/MCT1 to induce lactate utilization and cell growth. Targeting the p53/HIF-1 alpha/MCT1 pathway could inhibit Cr (VI)-mediated tumorigenesis.

关键词： alternative splicing   RNA editing   ion channel   calcium   calcium channel   calmodulin   neurophysiology   neurobiology  

摘要： Alternative splicing and adenosine-to-inosine (A-to-I) RNA editing are post-transcriptional mechanisms that promote proteomic diversity in the brain. The impact of RNA editing on brain physiology is underscored by its association with the etiology and pathogenesis of various neurological and neurodegenerative diseases. Alternative splicing of exon 43 within the CaV1.3 C-terminus produces long (CaV1.3L) and short (CaV1.343S) variants exhibiting distinct gating and pharmacological properties. While the physiological role of CaV1.3 A-to-I RNA editing in brain function is still unfolding, recent findings indicate that unedited CaV1.3 channels display larger calcium (Ca2+) influx and enhanced neuronal excitability, suggesting that A-to-I RNA editing may act as a negative regulator of neuronal Ca2+ signaling. However, it remains unclear which specific splice variants of CaV1.3 undergo RNA editing. We discovered an unexpected link between alternative splicing and RNA editing, where generation of the CaV1.343S variant results in a threefold increase in RNA editing at the IQ site. The CaV1.343S variant undergoes A-to-I RNA editing at two sites within its calmodulin-binding IQ domain, giving rise to three different edited variants. Functional analysis revealed that A-to-I RNA editing markedly decreases current density and induces a depolarizing shift in the current window, aligning the edited short variant's properties with those of the long variant. Our findings suggest that A-to-I RNA editing serves as a physiological mechanism regulating the "short" gating properties of the CaV1.343S variant and position A-to-I RNA editing of CaV1.343S variant as a neuroprotective physiological mechanism that prevents Ca2+ overload, especially in neurons susceptible to Ca2+ toxicity.

关键词： 1型鸭圆环病毒(DuCV)   Cap基因   原核表达   VLPs   多克隆抗体  

摘要： 利用原核表达系统重组表达鸭圆环病毒（DuCV）基因1型Cap蛋白病毒样颗粒（VLPs），纯化后免疫试验兔，检测制备的DuCV-1 Cap VLPs的免疫原性。以DuCV-1型Cap基因为研究对象，应用在线网站对其进行生物信息学分析。对Cap基因序列进行***适应性密码子优化，通过PCR技术扩增不含核定位信号（NLS）的Cap基因，构建重组质粒pET28a-ΔNLS-Cap。IPTG诱导Cap重组蛋白表达，经SDS-PAGE分析及Western blot鉴定，并采用镍柱亲和层析进行纯化。将纯化后的重组蛋白与弗氏佐剂等比例乳化后免疫试验兔，制备多克隆抗体。采用间接ELISA测定抗体效价，采用Western blot检测多克隆抗体特异性。本实验成功构建了重组质粒PET-28a-ΔNLS-Cap，主要以包涵体形式表达，大小约为41.7 kDa。透射电镜观察发现，纯化后的重组蛋白可形成直径约15 nm的病毒样颗粒（VLPs）；间接ELISA法检测重组蛋白兔源多克隆抗体的效价达到1∶160 000以上。Western-blot结果表明，兔抗Cap抗体可以与重组蛋白发生特异性反应。免疫荧光结果显示，兔抗Cap多克隆抗体可以特异性识别由Sf9昆虫细胞表达系统重组表达的DuCV-1Cap蛋白。本试验成功制备了具有免疫原性的DuCV-1Cap VLPs及其兔源多克隆抗体，为进一步研究DuCV亚单位疫苗和相关诊断检测方法奠定了基础。

关键词： 糖尿病   黄斑水肿   血管内皮生长因子类   严重程度  

摘要： 目的 探讨血清血管内皮生长因子（VEGF）水平与糖尿病黄斑囊样水肿（CME）严重程度的相关性。方法前瞻性选取2023年6月—2024年6月唐山市眼科医院玻璃体视网膜外科收治的糖尿病CME患者（病例组）及糖尿病无眼部并发症患者（对照组）各57例（57眼），均接受系统的眼科专科检查及血清VEGF检测。采用多因素Logistic回归明确糖尿病CME发病的危险因素，并绘制受试者工作特征（ROC）曲线分析VEGF对糖尿病CME的诊断价值，采用Spearman相关性分析VEGF水平与糖尿病CME患者病情程度相关指标[矫正视力、囊样水肿个数、黄斑中心凹无血管区（FAZ）非圆度指数]的关系。结果 与对照组相比，病例组糖尿病病程较长，合并高血压占比较高，糖化血红蛋白（HbA1c）、血肌酐、血清VEGF水平较高，估算的肾小球滤过率（eGFR）较低（P<0.05）。多因素Logistic回归结果显示，糖尿病病程长，HbA1c、VEGF水平升高是糖尿病CME发病的危险因素，eGFR水平升高是保护因素（P<0.05）。ROC曲线分析显示，VEGF诊断糖尿病CME的曲线下面积（AUC）为0.758(95%CI:0.669～0.834)。糖尿病病程、HbA1c、e GFR、VEGF联合应用诊断糖尿病CME的AUC为0.916(95%CI:0.877～0.941)，较VEGF单独应用时有明显提升（P<0.05）。VEGF水平与糖尿病CME患者矫正视力[0.34(0.24,0.44)]呈负相关（rs=-0.771），与黄斑囊样水肿检出[3(2,5)个]和FAZ非圆度指数[1.16(1.08,1.20)]呈正相关（rs分别为0.700和0.788，均P<0.05）。结论 血清VEGF水平与CME病情严重程度密切相关，可作为诊断CME发病的可靠参考指标。

关键词： 西尼罗病毒   传播   候鸟   Meta分析  

摘要： 西尼罗病毒（WNV）作为一种蚊媒人畜共患病原体，其传播动力学与鸟类群落结构密切相关。候鸟被认为是病毒大范围扩散的关键媒介，而城市生态系统中的本地蚊种（如埃及伊蚊）和具有病毒宿主能力的鸟类则会加剧局部疫情爆发的风险。本研究综合了生态流行病学数据，以量化鸟类群落中WNV传播的时空驱动因素。本研究采用随机效应模型对全球48项研究（样本量n=52575，感染事件3213例）进行Meta分析，以估算合并患病率（95%置信区间）。通过亚组分析评估区域（Q=3.99，p=0.26）与方法学（Q=2.24，p=0.32）异质性，并利用线性回归分析发病率时间趋势（β=-0.01，SE=0.01，p=0.30）。结果表明，鸟类种群WNV合并效应量为7.28%（95% CI：5.82%–9.06%），检测方法之间无显著差异。区域分析显示，不同地理区域的患病率估计值具有差异性，欧洲的患病率为6.0% (95% CI：5.0%-9.0%)，美洲为7.0% (95% CI：4.0%-10.0%)，非洲为10.0% (95% CI：5.0%-21.0%)，亚洲为11.0% (95% CI：6.0%-19.0%)。时间分析显示，2000至2023年间发病率无显著时间趋势（t=-1.03）。本研究从数据上明确了候鸟携带西尼罗病毒的情况，亚洲地区的候鸟患病率居于高位，带来地区流行风险。除此之外，候鸟携带西尼罗病毒没有显著的时间变化趋势。