Primary open-angle glaucoma (POAG) is the leading cause of irreversible blindness worldwide, primarily due to the degeneration of retinal ganglion cells (RGCs). In this study, we reported vav guanine nucleotide exchange factor 2 (VAV2) as a POAG-associated gene. Through whole exome sequencing (WES) of 398 Han Chinese POAG patients and 2,010 controls, we discovered nine rare VAV2 variants linked to POAG (P_burden=1.40×l0−6). Functional analyses revealed that these variants disrupted normal VAV2 protein function, leading to compromised cytoskeletal organization in human trabecular meshwork cells and impaired axonal growth in the 661W cell line. In vivo, Vav2 knockout mice exhibited key POAG features, including increased intraocular pressure (IOP), abnormal trabecular meshwork structure, reduced visual sensitivity, and RGC loss. This study also implicated VAV2 in the modulation of the Rho signaling pathway, which is essential for maintaining trabecular meshwork integrity and neuronal function. Taken together, this research identified VAV2 as a candidate gene for POAG and suggests VAV2 as a potential target for genetic screening of POAG diagnostics.

Age-related macular degeneration (AMD), particularly its atrophic (dry) form, is a leading cause of irreversible blindness in the elderly. Limited treatment efficacy stems from its complex pathogenesis, highlighting an urgent need for novel therapeutic targets. This study investigates the contribution of the choroidal immune microenvironment, focusing on intercellular communication involving resident fibroblasts-a cell type whose role in AMD remains poorly defined. By analyzing single-cell RNA sequencing data from human choroid, we interrogated crosstalk between fibroblasts, macrophages, and NK/T cells, identifying interferon-gamma (IFNγ) and tumor necrosis factor-alpha (TNFα) signaling pathways as central mediators. We demonstrate that activated choroidal fibroblasts release key inflammatory mediators, including IL6, CCL2, CSF1, CXCL9, and CXCL10, which functionally recruit macrophages and CD8+ T cells, thereby shaping the local immune landscape. Critically, targeting these pathways in vivo using TAPI-1 (inhibiting TNFα processing) and Tofacitinib (inhibiting IFNγ signaling) significantly ameliorated retinal, RPE, and choroidal pathology in a NaIO3-induced murine model of dry AMD. Our findings underscore the pathogenic role of fibroblast-mediated choroidal inflammation driven by TNFα and IFNγ signaling in dry AMD, presenting these pathways as promising therapeutic targets.

The Frizzleds (FZDs) receptors on the cell surface belong to the class F of G protein-coupled receptors (GPCRs) which are the major receptors of WNT protein that mediates the classical WNT signaling pathway and other non-classical pathways. Besides, the FZDs also play a core role in tissue regeneration and tumor occurrence. With the structure and mechanism of FZDs activation becoming clearer, a series of FZDs modulators (inhibitors and agonists) have been developed, with the hope of bringing benefits to the treatment of cancer and degenerative diseases. Most of the FZDs inhibitors (small molecules, antibodies or designed protein inhibitors) block WNT signaling through binding to the cysteine-rich domain (CRD) of FZDs. Several small molecules impede FZDs activation by targeting to the third intracellular domain or the transmembrane domain of FZDs. However, three small molecules (FZM1.8, SAG1.3 and purmorphamine) activate the FZDs through direct interaction with the transmembrane domain. Another type of FZDs agonists are bivalent or tetravalent antibodies which activate the WNT signaling via inducing FZD-LRP5/6 heterodimerization. In this article, we reviewed the FZDs modulators reported in recent years, summarized the critical molecules’ discovery processes and the elucidated relevant structural and pharmacological mechanisms. We believe the summaried molecular mechanisms of the relevant modulators could provide important guidance and reference for the future development of FZD modulators.

Immunosenescence increases the risk and severity of diseases in elderly individuals and leads to impaired vaccine-induced immunity. With aging of the global population and the emerging risk of epidemics, developing adjuvants and vaccines for elderly individuals to improve their immune protection is pivotal for healthy aging worldwide. Deepening our understanding of the role of immunosenescence in vaccine efficacy could accelerate research focused on optimizing vaccine delivery for elderly individuals. In this review, we analyzed the characteristics of immunosenescence at the cellular and molecular levels. Strategies to improve vaccination potency in elderly individuals are summarized, including increasing the antigen dose, preparing multivalent antigen vaccines, adding appropriate adjuvants, inhibiting chronic inflammation, and inhibiting immunosenescence. We hope that this review can provide a review of new findings with regards to the impacts of immunosenescence on vaccine-mediated protection and inspire the development of individualized vaccines for elderly individuals.

6-mercaptopurine (6 -MP) serves as the backbone in the maintenance regimens of acute lymphoblastic leukemia (ALL). We aimed to evaluate the influence of NUDT15 gene polymorphism on the risk of myelosupression, hepatotoxicity and interruption of 6 -MP, as well as treatment efficacy and dose of 6 -MP in ALL patients. A total of 24 studies with 3,374 patients were included in this meta -analysis. We found 9 -fold higher risk of 6 -MP induced leukopenia (odds ratio [OR] =9.00, 95% confidence interval [CI]: 3.73-21.74) and 2.5 -fold higher risk of 6 -MP -induced neutropenia (OR=2.52, 95% CI: 1.72-3.69) for NUDT15 c.415C>T variant carriers in the dominant model. Moreover, we found that the dose intensity of 6 -MP in ALL patients with one NUDT15 c.415C>T variant alleles (CT) was 19% less than that in wild -type patients (CC) (mean differences: 19.43%, 95% CI: -25.36 to -13.51). The tolerable dose intensity of 6 -MP in NUDT15 c.415C>T homozygote variant (TT) and heterozygote variant (CT) carriers was 49% and 15% less than that in wild -type patients, respectively. The NUDT15 c.415C>T variant group (CT+TT) had seven times (OR=6.98, 95% CI: 2.83-17.22) higher risk of developing 6 -MP intolerance than the CC group. However, NUDT15 c.415C>T polymorphism did not appear significantly associated with hepatotoxicity, treatment interruption or relapse incidence. We concluded that NUDT15 c.415C>T was a good predictor for 6 -MP -induced myelosuppression in ALL patients. The dose intensity of 6 -MP in ALL patients with NUDT15 c.415C>T variants was significantly lower than that in wild -type patients. This research provided a basis for further investigation into relations between NUDT15 gene and adverse reaction, treatment efficacy and dose intensity of 6 -MP.

Inflammation and elevated expression of high temperature requirement A serine peptidase 1 (HTRA1) are known high risk factors for age-related macular degeneration (AMD). However, the specific mechanism that HTRA1 causes AMD and the relationship between HTRA1 and inflammation remains unclear. We found that lipopolysaccharide (LPS) induced inflammation enhanced the expression of HTRA1, NF-κB, and p-p65 in ARPE-19 cells. Overexpression of HTRA1 up-regulated NF-κB expression, and on the other hand knockdown of HTRA1 down-regulated the expression of NF-κB. Moreover, NF-κB siRNA has no significant effect on the expression of HTRA1, suggesting HTRA1 works upstream of NF-κB. These results demonstrated that HTRA1 plays a pivotal role in inflammation, explaining possible mechanism of overexpressed HTRA1-induced AMD. Celastrol, a very common anti-inflammatory and antioxidant drug, was found to suppress inflammation by inhibiting phosphorylation of p65 protein efficaciously in RPE cells, which may be applied to the therapy of age-related macular degeneration.

Introduction: As demonstrated in pivotal clinical trials, brolucizumab can be used to treat neovascular age-related macular degeneration (nAMD) because it antagonizes vascular endothelial growth factor (VEGF) in the vitreous. However, brolucizumab may cause retinal vasculitis obliterans in the presence of inflammation in the eyes. In the present study, a meta-analysis of randomized controlled trials (RCTs) was conducted to evaluate the efficacy and safety of brolucizumab. Methods: ClinicTrail.gov., Embase, Cochrane Library, and PubMed were retrieved from inception until 31 December 2021 for RCTs assessing the efficacy and safety of brolucizumab. Changes in best corrected visual acuity (BCVA) and central sub-field thickness (CSFT) and incidence of adverse events, serious adverse events, and serious ocular adverse events were extracted from eligible RCTs. A meta-analysis was performed using RevMan 5.4.1. Results: A total of six RCTs with 3,574 participants were finally involved in this meta-analysis. The changes of best corrected visual acuity (BCVA) showed no statistically significant difference between the brolucizumab-treated group and aflibercept-treated group. Brolucizumab induced higher central sub-field thickness (CSFT) reduction than the control agent (aflibercept). The incidence of adverse events was similar between the brolucizumab group and control group (OR 0.63, 95% CI 0.37 to 1.08, p = 0.09), and brolucizumab caused fewer serious adverse events (OR 0.78, 95% CI 0.63 to 0.95, p = 0.01). However, brolucizumab could lead to more serious ocular adverse events than Lucentis and aflibercept (OR 2.15, 95% CI 1.11 to 4.16, p = 0.02). Conclusion: Brolucizumab was non-inferior to other anti-VEGF agents in improving BCVA and decreasing CSFT. But it caused more serious ocular adverse events which is worthy of special attention by ophthalmologists.

目的 观察小儿盐酸普鲁卡因口服溶液对消化系统疾病等所致呕吐患儿的疗效,为该药的后续研究提供参考.方法 收集2018年3月—2019年3月四川省人民医院儿科使用小儿盐酸普鲁卡因口服溶液患儿的临床资料,对其使用年龄、适应证、用法用量、不良反应、治疗效果等进行统计.结果 共收集使用小儿盐酸普鲁卡因口服溶液患儿550例,年龄25 d～13岁;适应证以胃肠炎引起的呕吐为主,也适用于胃炎、扁桃体炎、肠系膜淋巴结炎、上呼吸道感染、胃肠功能紊乱等引起的呕吐;用法用量为新生儿及1岁以下儿童口服用量<1 mL,1岁儿童用量平均为1 mL,每增加1岁用量增加1 mL,临床最大使用剂量为10 mL;不良反应为变态反应、食欲下降;治疗效果为治愈467例,好转78例,无效5例.结论 小儿盐酸普鲁卡因口服溶液对儿童消化系统疾病等引起的呕吐疗效好,不良反应少.

Background: Roxadustat is a new oral drug for anemia in chronic kidney disease (CKD). This study aimed to synthesize the evidence from randomized controlled trial (RCT)-based studies that estimated the efficacy and safety of roxadustat in anemia patients with non-dialysis-dependent (NDD) and dialysis-dependent (DD) CKD. Methods: We searched the PubMed, Web of Science, and Cochrane Central Register of Controlled Trials (CENTRAL) databases for related published studies. Moreover, we manually searched relevant pharmaceutical company websites and two international clinical trial registers to search for published and unpublished RCTs comparing roxadustat with erythropoietin-stimulating agents (ESAs) or placebo. Results: Fifteen RCTs (seven for DD-CKD patients, eight for NDD-CKD patients) were included in the meta-analysis, with 10,189 patients, 4,810 DD-CKD patients, and 5,379 NDD-CKD patients enrolled. Compared with ESAs (epoetin alfa or darbepoetin alfa) and placebo, roxadustat raised the hemoglobin level [weighted mean difference (WMD): 0.82 g/dL; 95% confidence interval (CI): 0.43–1.21], transferrin level (WMD: 0.5 g/L; 95% CI: 0.34–0.65), and TIBC level (WMD: 41.79 μg/dL; 95% CI: 38.67–44.92) and lowered the hepcidin level (WMD: −37.38 ng/ml; 95% CI: −46.63– −28.12) in both the DD-CKD and NDD-CKD patients with renal anemia. Roxadustat improved hemoglobin response and lowered the ferritin and TAST levels in the NDD-CKD patients but not in the DD-CKD patients. Furthermore, there was no difference between the treatment-emergent adverse events (TEAEs) of roxadustat and that of ESAs or placebo. But the incidence of serious TEAEs in the roxadustat group was significantly higher with NDD-CKD patients (OR: 1.15; 95% CI: 1.02–1.29). Conclusion: This study confirmed that roxadustat therapy could alleviate the anemia of DD-CKD and NDD-CKD patients by raising the hemoglobin level and regulating iron metabolism, but increased serious incidences of treatment-emergent adverse events (TEAEs) in NDD-CKD patients.

目的:利用等温滴定量热技术研究VAV2蛋白的DH结构域(VAV2-DH)与CDC42的相互作用.方法:首先分别将VAV2-DH和CDC42蛋白的编码基因克隆至pGEX-6p-1和pET28a载体上,构建VAV2-DH和CDC42蛋白的重组表达载体.再将重组表达载体转入大肠杆菌中诱导表达蛋白,并亲和层析、凝胶过滤层析纯化得到VAV2-DH和CDC42蛋白,最后利用鸟嘌呤核苷酸交换试验来验证VAV2-DH的鸟苷酸交换活性,并采用等温滴定量热技术检测这2种蛋白质相互作用的亲和力和热力学参数.结果:VAV2-DH和CDC42蛋白的平衡解离常数KD=(11.44±2.12) μmol/L,其中摩尔结合焓DH=(-224.33±95.48) kJ/mol、-TDS=(195.97±95.95) kJ/mol、吉布斯自由能DG=(-28.30±0.49)kJ/mol.结论:VAV2-DH和CDC42蛋白属于中等强度的相互作用,是一个焓驱动的结合过程.

Small GTPase cycled between the GDP-bound inactive state and GTP-bound active state, catalyzed by guanine nucleotide exchange factors (GEFs). Guanine nucleotide exchange assay was a direct way to investigate the specificity, activity, and kinetics of GEFs. The N-methylanthraniloyl derivative of GDP (mantGDP), which was bound to small GTPase, served as a substitution for labeled small GTPase involved in bioluminescent, colorimetric, or radioactive methods due to its safety and sensitivity. In this study, we present an economical and efficient approach to prepare qualified mantGDP-bound CDC42, a member of the Rho GTPase family. In our protocol, with a Kd value of 0.048 μM, alkaline phosphatase hydrolysis of CDC42 increased mantGDP binding affinity to CDC42, allowing mant-nucleotide associating onto CDC42 more easily. Only 1.5-fold molar excess of mantGDP was required to prepare mantGDP-bound CDC42 without nonhydrolyzable GTP analog and high performance liquid chromatography. The mantGDP-bound CDC42 was verified to be efficient for measuring the guanine nucleotide exchange activity of VAV2.

Objective . To evaluate the diagnostic value of apolipoprotein E ( APOE ) gene in Alzheimer’s disease (AD). Methods . Databases including PubMed, EMBASE, Google Scholar, Wanfang Med online, China National Knowledge Infrastructure (CNKI), and China Biomedical Literature Database (CBM) were searched for literatures in English or Chinese. No limitations on the date. The sensitivity, specificity, likelihood ratio, and diagnostic odds ratio were pooled for meta-analysis. The symmetric receiver operator characteristic curve (SROC) and Fagan’s Nomogram were drawn, and metaregression and subgroup analysis were used to explore the source of heterogeneity. Results . A total of 13 studies, including 2662 cases and 8843 controls, were analyzed. The combined sensitivity (SEN) was 0.62 (95% CI (0.58-0.66)), specificity (SPE) was 0.84 (95% CI (0.81-0.86)), the positive likelihood ratio was 3.8 (95% CI (3.3-4.3)), and the negative likelihood ratio was 0.45 (95% CI (0.41-0.49)). The area under the ROC curve was 0.80, and the diagnostic ratio (DOR) was 8. Neither publication bias was detected in Deeks’ funnel plot, nor threshold effect was shown in the SROC. Metaregression analysis showed that the diagnostic methods, experimental design, and sample size contributed to the heterogeneity in SEN, while the diagnostic methods, experimental design, blind evaluation on test results, and sample size contributed to the heterogeneity in SPE. When the pretest probability was set as 50%, the posterior probability in Fagan’s Nomogram was 79%, the positive likelihood ratio (LRP) was 5, and the negative likelihood ratio (LRN) was 0.42. Conclusions . AD could neither be confirmed nor excluded by the APOE genotype test. The sensitivity and specificity of the APOE gene test were relatively low in the diagnosis of AD. The diagnostic value of APOE ε 4 gene in AD was moderate; it might play an important role in the prevention of AD.

>Dear Editor,Infection with the novel coronavirus (SARS-CoV-2, which is the virus responsible for the coronavirus disease 2019(COVID-19)) was first reported in Wuhan, China on December 31, 2019. The outbreak of COVID-19 remains ongoing and was linked to more than 80,000 infected patients and more than 3,000 deaths in China as of March 7, 2020

FGD2, a member of FGD family, contains a Dbl homology domain (DH) and two pleckstrin homology domains segregated by a FYVE domain. The DH domain has been deduced to be responsible for guanine nucleotide exchange of CDC42 to activate downstream factors. Our aim was to build a prokaryotic expression system for the DH domain and to examine its guanine nucleotide exchange activity toward CDC42 in vitro. A recombinant vector, which was successfully constructed based on pGEX-6P-1, was employed to express the DH domain of human FGD2 (FGD2-DH) in E. coli BL21 (DE3). Purified FGD2-DH behaved as a homogeneous monomer with an estimated molecular weight that corresponded to the theoretical molecular weight and was predicted to be an α-helix protein by circular dichroism spectroscopy. FGD2-DH displayed weak guanine nucleotide exchange activity in vitro and very weak interactions with CDC42 following glutaraldehyde cross-linking.

Published studies indicate the MTHFR C677T and A1298C polymorphisms are associated with abnormal homocysteine levels, which may cause various pregnancy complications and birth defects. However, the results obtained from different studies have been inconsistent. Therefore, this meta-analysis explores the association between MTHFR polymorphisms and birth defects and adverse pregnancy outcomes. The PubMed, ScienceDirect, Embase, and China Biology Medicine literature databases and ClinicalTrials were searched. Analyses of public bias, meta-regression, subgroups, and sensitivity were used to ensure the robustness of our results. MTHFR C677T was significantly associated with recurrent pregnancy loss in developing countries (odds ratio [OR], 1.34; 95% confidence interval [CI], 1.20-1.50) but not in developed countries (OR, 0.87; 95% CI, 0.68-1.11). No significant relationship was found between MTHFR A1298C and recurrent pregnancy loss (OR, 1.04; 95% CI, 0.93-1.18). MTHFR C677T and A1298C were not associated with preeclampsia (OR, 1.06; 95% CI, 0.97-1.16 and OR, 1.16; 95% CI, 0.97-1.39, respectively), and C677T was not associated with placental abruption (OR, 1.03; 95% CI, 0.87-1.21), intrauterine growth retardation (OR, 1.02; 95% CI, 0.90-1.15), or congenital heart disease (OR, 1.05; 95% CI, 0.89-1.25). MTHFR C677T, but not A1298C, was associated with neural tube defects (OR, 1.24; 95% CI, 1.08-1.42) and Down syndrome (OR, 1.65; 95% CI, 1.39-1.95). Conclusion Although MTHFR C677T and A1298C are significantly associated with some types of congenital defects and adverse pregnancy outcomes, the impact of these polymorphisms is moderate.

Objective:To construct a prokaryotic expression plasmid for the large extracellular loop (LEL) of human TSPAN12 protein and obtain highly purified soluble recombinant proteins.Methods:In this experimental study,the coding sequence of Tspan12 LEL was first cloned into pMa1-c2x to link with the coding sequence of the maltose binding protein (MBP).Then the DNA of MBP-tagged Tspan12 LEL (MBP-TSPAN12 LEL) was cloned to multiple cloning site 1 of vector pETDuet-1 after PCR amplification,restriction of enzyme digestion and T4 ligase reaction.DNA of DsbC was cloned into multiple cloning site 2 of vector pETDuet-1 and co-expressed with MBP-TSPAN12 LEL in order to facilitate disulfide bond formation.After transforming the recombinant plasmid into OrigamiB (DE3),MBP-TSPAN12 LEL was expressed by isopropyl-β-d-thiogalactoside induction and purified by amylose resin affinity chromatography and anion exchange chromatography.Results:Sequencing results suggested that recombinant plasmid was successfully constructed.SDS-PAGE showed that the molecular weight of the soluble MBP-TSPAN12 LEL was about 60 kD.Abundant,soluble and highly purified fusion protein was acquired after affinity and anion exchange chromatography.Conclusions:The experimental results prove that co-expression DsbC with MBP-TSPAN12 LEL is a practicable way to produce soluble large extracellular loop of human Tspan12 protein in Escherichia coli.

目的 探讨亚甲基四氢叶酸还原酶(MTHFR)C677 T、A1298C基因多态性与急性淋巴细胞白血病(ALL)患儿使用大剂量甲氨蝶呤(HD-MTX)后不良反应的相关性.方法 纳入70例经HD-MTX治疗的ALL患儿,检测患儿MTHFR C677 T、A1298C基因多态性和MTX血药浓度,收集各项生理生化指标和HD-MTX化疗后不良反应,分析基因多态性与血药浓度及不良反应发生的相关性.结果 C677 T各基因型间胃肠道反应的发生呈显著性差异(P=0.003),C677 T野生型(CC)vs突变型(CT+TT)胃肠道反应呈显著性差异(P=0.001),Logistic回归分析显示患儿72小时血药浓度水平、C677 T位点多态性,体质指数(BMI),免疫分型及体重可能是导致患儿不良反应(ADR)发生的影响因素(P<0.05).结论 检测MTHFRC677 T基因型可以有效指导临床HD-MTX个体化给药,为减少不良反应的发生提供参考指标.

Objective: To investigate the role of CYP3A4*18B (rs2242480) and CYP3A5*3 (rs776746) genetic polymorphism in predicting the efficacy of tacrolimus (FK506) and the recovery of hepatorenal function after liver transplantation. Methods: Thirty eight patients who received liver transplantation and treated with FK506 were enrolled. The genetic polymorphism of CYP3A4/5 was determined by DNA microarray and the blood trough concentration of FK506 was measured by enzyme multiplied immunoassay technique (EMIT). The adjusted concentration/dosage (C/D) ratio of FK506 and indicators of hepatorenal function were measured one, two and four weeks after transplantation. Results: Among 38 patients, there were 14 patients carrying CYP3A4 C/C (37.8%), 19 carrying CYP3A4 C/T (51.4%), 4 carrying CYP3A4 T/T (10.8%), and 1 undetectable genotype. The corrected C/D ratio was highest for CYP3A4 C/C group one week after operation (P<0.05). The recovery of ALT, Cr and INR of the CYP3A4 C/C group was slower than that of the CYP3A4 C/T group and CYP3A4 T/T group. Among 38 patients, we found 5 carriers of CYP3A5 A/A (13.1%), 15 carriers of CYP3A5 A/G (39.5%) and 18 carriers of CYP3A5 G/G (47.4%). CYP3A5 G/G group had higher corrected C/D ratio and slower recovery of Cr, ALT and INR than CYP3A5 A/G group and CYP3A5 A/A group. Conclusion: The gene polymorphism of CYP3A4/5 was an important factor for the significant individual pharmacokinetics differences of FK506. Slow metabolism genotypes (CYP3A4 C/C and CYP3A5 G/G) required a lower dose of FK506 to reach the proper blood concentration than fast metabolizers (CYP3A4 C/T, T/T and CYP3A5 A/A, A/G).

Increased aggregation of -amyloid (A) peptides induces oxidative stress, which is considered a major contributor in the development of Alzheimer's disease (AD). Prevention of A-induced neurotoxicity is proposed as a possible modality for treatment of AD. The present study aimed to elucidate possible effects of ethyl vanillin (EVA), an analog of vanillin isolated from vanilla beans, on the A(1-42)-induced oxidative injury in PC12 cells. EVA restrained the decrease in PC12 cell viability and apoptosis induction caused by treatment with A(1-42). In addition, EVA markedly alleviated intracellular lipid peroxidation as demonstrated by malondialdehyde levels and reactive oxygen species production in A(1-42)-treated PC12 cells. In addition, the reduction in the activity levels of the antioxidative enzymes superoxide dismutase, catalase and glutathione peroxidase was detected in A(1-42)-treated PC12 cells. This effect was partially reversed by treatment with EVA. Furthermore, the results indicated that EVA attenuated A(1-42)-induced caspase-3 activation and the increase noted in the apoptosis regulator Bcl-2/apoptosis regulator Bax ratio of PC12 cells. These results indicated that EVA could be used as an efficient and novel agent for the prevention of neurodegenerative diseases via inhibition of oxidative stress and cell apoptosis.