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关键词： colorectal cancer   lipid nanoparticle   mRNA   neoantigen   personalized cancer vaccine   post-surgery recurrence  

摘要： Personalized cancer vaccines (PCVs) represent a promising frontier in cancer immunotherapy;however, challenges in neoantigen prediction and treatment optimization persist. This study aims to introduce an innovative mRNA-based PCV platform that addresses these limitations. Co-administration of our major histocompatibility complex (MHC)-I and MHC-II-restricted neoantigens increases antigen-specific T cell responses and exhibits strong anti-cancer efficacy, significantly inducing antigen-specific CD8+ T cell-immune responses. The mRNA-based vaccine targeting the novel antigens demonstrates anti-tumor efficacy in a murine model of colorectal cancer and reduces post-surgery recurrence in mRNA-vaccinated mice. Notably, early-stage vaccination induces a striking anti-cancer effect, underscoring the critical role of MHC-II neoantigens alongside MHC-I antigen prediction in shaping effective anti-tumor immunity, in the activation of antigen-specific CD8+ T cells. In addition, the combination of immune checkpoint inhibitors and the vaccine synergistically inhibits tumor growth by inducing robust T cell responses and promoting favorable alterations in the tumor microenvironment. Taken together, the results provide a strong rationale for the clinical investigation of rapid-turnaround co-administration of MHC-I/II-restricted neoantigens-based mRNA vaccines in colorectal cancer, as supported by the anti-tumor efficacy of early-stage application and combination immunotherapy approaches.

关键词： 慢病毒   痛风性关节炎   NLRP3炎症小体   Caspase-1   IL-1   IL-18  

摘要： 目的 利用shRNA 慢病毒干扰载体调控痛风性关节炎(Gouty arthritis， GA)滑膜细胞蛋白表达抑制Caspase-1、IL-1、IL-18的表达。构建pNLRP3shRNA慢病毒干扰载体，观察其在阻断NLRP3基因表达调控痛风性关节炎(GA)发展机制的影响。方法 以大鼠NLRP3基因mRNA序列为靶标设计4条短发夹RNA干扰序列，并设计 1 条阴性对照序列。完成化学合成后重组构建，获得重组慢病毒载体质粒。经测序无误后，分别将质粒转染到CHO（中国仓鼠卵母细胞）中，采用Western blot（WB）技术分别检测NLRP3基因转录产物及翻译产物的表达。筛选最优干扰慢病毒载体，再次感染滑膜细胞。设置Con组：正常大鼠滑膜细胞；MSU组：通过注入单钠尿酸盐晶体（MSU）溶液制成GA大鼠滑膜细胞；shNLRP3组：pNALRP3shRNA 转染的GA大鼠滑膜细胞 ，感染 72 h 后检测各组中 Caspase-1、IL-1、IL-18 蛋白表达水平的变化。结果 与Con组相比，MSU组细胞中 IL-1β、 IL-18含量、Caspase-1蛋白表达、 NLRP3蛋白表达明显升高；与MSU组相比，shNLRP3组细胞内 IL-1β、 IL-18 含量、Caspase-1蛋白表达、 NLRP3蛋白表达下降，差异具有统计学意义 (P＜0.05)。结论 pNLRP3shRNA慢病毒干扰载体能阻断NLRP3基因表达，抑制NLRP3炎症小体的激活，减少IL-1、IL-18等炎症因子的释放，减轻GA的炎症反应。

关键词： Bioinformatics   Endothelial cells   Inflammation   Molecular pathology   Pulmonology   Vascular biology  

摘要： The pathobiology of pulmonary hypertension (PH) is complex and multiple cell types contribute to disease pathogenesis. We sought to characterize the molecular crosstalk between endothelial and mesenchymal cells that promote PH in the tumor necrosis factor alpha-transgenic (TNF-Tg) model of PH. Pulmonary endothelial and mesenchymal cells were isolated from WT and TNF-Tg mice and underwent single-cell RNA sequencing. Data were analyzed using clustering, differential gene expression and pathway analysis, ligand-receptor interaction, transcription factor binding, and RNA velocity assessments. Significantly altered ligand-receptor interactions were confirmed with immunofluorescent staining. TNF-Tg mice had increases in smooth muscle cells and Col14+ fibroblasts, and reductions in general capillary (gCAP) endothelial cells, Col13+ fibroblasts, pericytes, and myofibroblasts. Pathway analysis demonstrated NF-kappa B-, JAK/STAT-, and interferon-mediated inflammation, endothelial apoptosis, loss of vasodilatory pathways, increased TGF-beta signaling, and smooth muscle cell proliferation. Ligand-receptor analysis demonstrated a loss of BMPR2 signaling in TNF-Tg lungs and establishment of a maladaptive BMP signaling cascade, which functional studies revealed stemmed from endothelial NF-kappa B activation and subsequent endothelial SMAD2/3 signaling. This system highlights a complex set of changes in cellular composition, cell communication, and cell fate driven by TNF signaling that lead to aberrant BMP signaling that is critical for development of PH.

关键词： inborn errors of immunity   IEI   IL-2Rβ   IL-2   IL-15   STAT5 signaling   CD8 T cells   regulatory T cells   Tregs   pro-inflammatory cytokines  

摘要： Here, we describe the use of a homologous knockin mouse model to further decipher the mechanism(s) of a novel human homozygous IL2RB hypomorphic mutation. Our model recapitulates the human immune dysregulation phenotype, showing decreased mutant interleukin-2RD (IL-2RD) cell-surface expression, impaired IL-2/15-dependent STAT5 signaling, elevated serum IL-2/15 levels, expanded effector memory CD8+ T cells, and severely reduced regulatory T cells (Tregs). Using mixed bone marrow chimeras (BMCs) and wild-type (WT) Treg transfers, we distinguish receptor-intrinsic from receptor-extrinsic immunopathogenesis. Both approaches suppress abnormal serum cytokine levels and autoimmunity without affecting endogenous mutant Tregs. Mutant animals receiving WT Tregs neonatally exhibit almost complete restoration of conventional T cell distribution, IL-2RD receptor surface expression, and STAT5 signal transduction, while BMC animals exhibit only partial restoration. Our findings demonstrate that CD8+ T cells and Tregs have distinct IL-2/15 ligand/receptor ratios and signaling thresholds required for proper development/function, revealing mechanistic insights applicable to immunotherapy for autoimmunity.

关键词： 产气荚膜梭菌   α毒素   铁蛋白   纳米颗粒   免疫原性  

摘要： 本研究旨在制备产气荚膜梭菌α毒素-铁蛋白纳米颗粒抗原并评价其对小鼠的免疫原性。将带有D56G、H68G点突变和Spytag序列的α毒素全长基因和带有SpyCatcher序列的铁蛋白（Ferritin）基因片段分别克隆至原核表达载体pET 28a，后进行表达并对表达产物进行鉴定和纯化。对纯化后的αm2ST进行细胞毒性、卵磷脂酶活性以及溶血活性分析。将纯化的αm2ST和FeSC在体外对接，随后，将α毒素-铁蛋白(αm2-Fe)对接产物和αm2ST分别与佐剂混合后免疫小鼠。免疫后每周采集小鼠血清，检测IgG、IgG亚型抗体水平和中和效价，并在二免后28 d取脾细胞分析T细胞亚群和IFN-γ分泌水平。结果显示，在28 ℃条件下，αm2ST和FeSC均以可溶性表达为主。αm2ST未被检测出细胞毒性、卵磷脂酶活性和溶血活性，这说明αm2ST已无毒性。将αm2ST和FeSC体外对接后，经SDS-PAGE、TEM和DLS分析，证实αm2-Fe形成直径32.7～50.7 nm的纳米颗粒，主峰为37.8 nm。小鼠免疫后，αm2-Fe组能产生较高水平的IgG2a抗体（P<0.001）；小鼠二免后21 d，αm2-Fe组血清中和效价为64倍，显著高于αm2ST组的32倍（P<0.05）；二免后28 d，αm2-Fe组脾细胞刺激后上清中的IFN-γ也高于αm2ST组（P<0.05），流式细胞术分析显示αm2-Fe组产生了较高比例的效应性CD8+T细胞（P<0.001）。小鼠试验证实αm2-Fe纳米颗粒抗原能有效刺激小鼠产生体液免疫和细胞免疫。

关键词： porcine epidemic diarrhea virus   PEDV   coronavirus   live attenuated vaccine   recombination resistance   interferon antagonist   IFN   transcriptional regulatory sequence   TRS  

摘要： Porcine epidemic diarrhea virus (PEDV) is a deadly coronavirus (CoV) for neonatal piglets, and no effective vaccine is available. Live attenuated vaccines (LAVs) show promise, but risks of recombination and reversion to virulence hinder their application. For LAV development, we previously engineered a recombination-resistant PEDV mutant RMT by rewiring the transcriptional regulatory sequence (TRS)-core sequences (TRS-CSs). Because TRS is critical for CoV replication, the incompatibility between wild-type TRS-CS and the rewired TRS-CS should disrupt the structural and accessory protein-encoding mRNA transcription from a recombinant PEDV genome, preventing the production of progeny infectious viruses. However, the RMT contained a 189-nt insertion upstream of the E gene TRS-CS and a missing guanine in the N gene TRS-CS. Here, we generated RMTv1 via correcting these mutations and removing the EGFP gene in RMT. Using the RMTv1 as a backbone, we generated a series of PEDV mutants carrying one or two attenuating mutations of non-structural protein 1 (nsp1), nsp15, and nsp16. Their replication efficiency, sensitivity to interferons (IFNs), induction of IFNs, and genetic stability were tested in Vero and/or LLC-PK1 cells. We selected RMTv1 and RMTv1-nsp1 + nsp15 for the pathogenesis studies in neonatal gnotobiotic pigs and tested the immunogenicity of RMTv1-nsp1 + nsp15. The RMTv1-nsp1 + nsp15 was further attenuated, causing no pig mortality, compared with RMTv1. All the pigs infected with RMTv1-nsp1 + nsp15 were protected from severe diarrhea and death post-challenge with virulent PEDV at 21 days post-inoculation, whereas 50% of mock-challenged piglets died. These findings establish RMTv1-nsp1 + nsp15 as a promising PEDV LAV candidate and can be further *** continues to cause devastating economic losses in the global swine industry. Exposing pregnant sows to feedback materials from infected pigs is still one of the main methods used to control PEDV out

关键词： α-synucleinopathy   blood-brain barrier   BBB   brain endothelial cells   TNF-α signaling pathway   vasculopathy   axonal degeneration  

摘要： Dysfunction of the blood-brain barrier (BBB) is recognized as a key factor in the progression of neurodegenerative diseases (NDs), but the detailed mechanisms behind its pathogenesis and impact on neurodegeneration remain elusive. This study aimed to reveal the pathological effects of alpha-Synuclein (alpha-Syn), an aggregation protein in synucleinopathy, on BBB integrity and function and identify therapeutic targets for alpha-Syn-related vasculopathy. Using a brain endothelial cell model, we investigated the pathological effect of preformed fibril alpha-Syn (PFF) on BBB integrity, employing generative adversarial network (GAN) deep learning to analyze pathological changes. We found that PFF activates immune responses, increasing endothelial monolayer permeability via the TNF-alpha-NF-kappa B pathway. Further in vivo studies with PFF induced alpha-synucleinopathy, and a transgenic animal model (G2-3) revealed that alpha-Syn aggregation disrupts the BBB, leading to axonal degeneration that was mitigated by treatment with a non-BBB-penetrating TNF-alpha inhibitor, etanercept. These findings suggest that targeting brain endothelial TNF-alpha signaling could be a potential therapeutic approach for synucleinopathy-related NDs.

关键词： 麻黄   荨麻疹   超敏反应   临床应用   机制探讨  

摘要： 荨麻疹作为临床常见的局限性水肿性皮肤疾病，其发病与超敏、炎症反应等生理病理过程密切相关。麻黄作为中医治疗荨麻疹的常用药，具有疏风透疹、宣肺利水的功效，现代研究进一步揭示其多成分、多靶点的药效特征。本文系统综述了麻黄治疗荨麻疹的古今应用脉络，整合其生物碱类、黄酮类及挥发油等活性物质的相关研究进展，重点探讨其在治疗荨麻疹方面抗过敏、免疫抑制、抗炎等核心作用机制。通过整合药理机制与临床证据，进行系统地梳理与综述，旨在为麻黄在不同复方中针对荨麻疹的药效及其作用机制以及基于有效性合理评价麻黄资源提供理论基础，以促进麻黄治疗荨麻疹的现代化研究进程。