[This corrects the article DOI: 10.3892/ol.2019.10255.].

Ulcerative colitis (UC), a chronic inflammatory bowel disease, significantly increases the risk of colon adenocarcinoma (COAD). Disulfidptosis, a novel form of programmed cell death, has been implicated in various diseases, including UC. This study investigates the expression of disulfidptosis-related genes, particularly CD2AP and MYH10, in UC and COAD. Through analysis of public datasets, we found MYH10 significantly upregulated and CD2AP downregulated in UC compared to healthy controls, with consistent patterns in COAD. Immune infiltration analysis revealed correlations between these genes and specific immune cell types, suggesting their roles in immune modulation. Molecular docking showed strong binding affinities of UC drugs such as budesonide and sulfasalazine with CD2AP and MYH10. Connectivity Map analysis identified additional drug candidates, including simvastatin and mephenytoin, which may be repurposed for UC and COAD therapy. These findings suggest disulfidptosis-related genes as potential biomarkers and therapeutic targets, linking chronic inflammation to cancer progression.

SARS-CoV-2 Omicron sublineages escape most preclinical/clinical neutralizing antibodies in development, suggesting that previously employed antibody screening strategies are not well suited to counteract the rapid mutation of SARS-CoV-2. Therefore, there is an urgent need to screen better broad-spectrum neutralizing antibody. In this study, a comprehensive approach to design broad-spectrum inhibitors against both SARS-CoV-1 and SARS-CoV-2 by leveraging the structural diversity of nanobodies is proposed. This includes the de novo design of a fully human nanobody library and the camel immunization-based nanobody library, both targeting conserved epitopes, as well as the development of multivalent nanobodies that bind nonoverlapping epitopes. The results show that trivale B11-E8-F3, three nanobodies joined tandemly in trivalent form, have the broadest spectrum and efficient neutralization activity, which spans from SARS-CoV-1 to SARS-CoV-2 variants. It is also demonstrated that B11-E8-F3 has a very prominent preventive and some therapeutic effect in animal models of three authentic viruses. Therefore, B11-E8-F3 has an outstanding advantage in preventing SARS-CoV-1/SARS-CoV-2 infections, especially in immunocompromised populations or elderly people with high-risk comorbidities.

PDF file - 429K, Fig. S1: GMCSFR expression in primary and transformed CEC. Fig S2: Receptors for IL-1, IL-6 and VEGF in CT26 cells. Fig. S3: GMCSF elicits IL-1β and IL-6. Fig. S4: Blocking GMCSF or VEGF decreases the number of proliferative epithelial cells. Fig. S5: Blocking GMCSF or VEGF regulates cell cycle gene expression in CEC. Fig. S6: GMCSF elicits VEGF expression in HCT116 cells. Fig. S7: TLR4 expression in primary and transformed CEC.

Abstract Programmed cell death–ligand 1 (PD-L1)/PD-1 axis is crucial for maintenance of immune homeostasis and its impairment partially accounts for the pathogenesis of inflammatory diseases. Hence, augmenting PD-L1/PD-1 signals represents a novel strategy to prevent destructive inflammation and induce immune tolerance. Recently, we developed a new cargo by conjugating the ectodomain of PD-L1 with pHLIP, a low pH-responding and membrane-inserting peptide, and demonstrated its potent immune-suppressive activity under acidic conditions in vitro. Herein, we further showed that PD-L1-pHLIP well responded to relatively high acidic buffer, while it could not inhibit T cell expansion in weakly acidic solutions. Furthermore, in a mouse model of acute intestinal inflammation, PD-L1-pHLIP treatment prolonged survival time and attenuated colitis in mice subjected to 6% dextran sulfate sodium (DSS) instead of 3% DSS. The different efficacy was due to the distinct acidity in the lesions, which facilitated PD-L1-pHLIP accumulation in the niche of 6% DSS-triggered severe inflammation. Mechanistic investigations revealed that PD-L1-pHLIP inhibited the release of proinflammatory cytokines in infiltrating macrophages and other immune cells in a PD-1-dependent and/or –independent fashion. Taken together, this study highlights PD-L1-pHLIP as a novel therapeutic avenue for inflammatory diseases.

目前呼气检测在临床中得到越来越广泛的应用,质谱技术作为呼吸气体检测的一项主流技术,随着直接质谱技术的发展,呼气检测质谱仪逐步走向商业化.文章阐述了呼吸标志物与人体疾病和代谢异常的关系、直接质谱技术、直接质谱技术在呼吸分析中的研究进展、质谱呼吸检测临床应用需要解决的问题.质谱呼气检测的未来应该着重于解决以下问题:建立更多新的生物标志物系统;建立疾病和呼出的呼吸生物标志物浓度之间的标准相关性,建立标准的呼吸收集和存储程序;区分呼出气体中的外源性和内源性气体;开发用于选择性检测呼吸气体的新型特异性纳米材料;开发操作简单、可重复、可靠、实时、质量轻、价格低廉的手持设备;进行大量适当的临床试验,包括尽可能多的受试者和实验验证等.目前商用的呼气检测质谱仪依然是气相色谱质谱(GC-MS)居多.随着各方面的研究进展,在疾病的筛查上各方面会逐步形成共识,低成本、操作便捷的非侵入式呼气检测质谱仪正处于产业爆发的前夕;呼气质谱技术发展应用过程中前景与挑战并存.

PDF file - 171K, Primer list.

Background: Mucosal-associated invariant T (MAIT) cells are nonconventional T cells with protection from infection. Sepsis is caused by severe life-threatening infection with organ dysfunction; however, the relationships between host immune responses and disease severity are not fully understood. Here, we explore the phenotype, function and clinical implications of MAIT cells in sepsis immunopathogenesis. Methods: Blood samples were collected from 35 sepsis patients and 46 healthy donors. The phenotypic and functional properties of MAIT cells were analyzed by flow cytometry and single-cell RNA sequencing (scRNA-seq). The mechanisms of MAIT cell differentiation were assessed using cell-culture experiments. The levels of lactate and IL-18 in plasma were examined by colorimetric and ELISA. Findings: Our data showed that circulating MAIT cells in sepsis patients, preferentially express IL-17A and RORγt (MAIT17). The abundance of MAIT17 cells were correlated with the severity of sepsis. Analysis of scRNA-seq further revealed that MAIT17 cells in sepsis patients exhibited enhanced level of lactate dehydrogenase A (LDHA) expression with a hyper inflammatory phenotype. Mechanistic studies demonstrated that RORγt expression of MAIT17 cells required PI3K-AKT-LDHA signaling, and was associated with plasma lactate levels in patients. Moreover, the elevated levels of lactate and IL-18 in plasma may promote the differentiation of MAIT17 cells in sepsis. Interpretation: Our study reveals that circulating MAIT17 cells play a significant role in the severity of sepsis, targeting MAIT17 cells could be a potential therapeutic strategy for the treatment of sepsis.Funding: This work was supported by the National Natural Science Foundation of China (Grant no. 82200668, 82372778, 82370605). Anhui Province Key Research and Development Plan (Grant no. 2022e07020039), Natural Science Foundation of the Anhui Education Department (Grant no. KJ2021A0281).Declaration of Interest: The authors declare no competing interest.Ethical Approval: This study was approved by the Ethics Committee of Clinical Research of the First Affiliated157 Hospital of Anhui Medical University (2022027). All participants gave informed consent.

AIM:To investigate the impact of deficiency of LIG4 gene on site-specific integration in CHO cells.RESULTS:CHO cells are considered the most valuable mammalian cells in the manufacture of biological medicines, and genetic engineering of CHO cells can improve product yield and stability. The traditional method of inserting foreign genes by random integration (RI) requires multiple rounds of screening and selection, which may lead to location effects and gene silencing, making it difficult to obtain stable, high-yielding cell lines. Although site-specific integration (SSI) techniques may overcome the challenges with RI, its feasibility is limited by the very low efficiency of the technique. Recently, SSI efficiency has been enhanced in other mammalian cell types by inhibiting DNA ligase IV (Lig4) activity, which is indispensable in DNA double-strand break repair by NHEJ. However, this approach has not been evaluated in CHO cells. In this study, the LIG4 gene was knocked out of CHO cells using CRISPR/Cas9-mediated genome editing. Efficiency of gene targeting in LIG4-/--CHO cell lines was estimated by a green fluorescence protein promoterless reporter system. Notably, the RI efficiency, most likely mediated by NHEJ in CHO, was inhibited by LIG4 knockout, whereas SSI efficiency strongly increased 9.2-fold under the precise control of the promoter in the ROSA26 site in LIG4-/--CHO cells. Moreover, deletion of LIG4 had no obvious side effects on CHO cell proliferation.CONCLUSIONS:Deficiency of LIG4 represents a feasible strategy to improve SSI efficiency and suggests it can be applied to develop and engineer CHO cell lines in the future.

ABSTRACT CRISPR/Cas9 mediated precise gene editing requires homology-directed repair (HDR), which occurs less frequently than non-homologous end-joining (NHEJ) including the canonical NHEJ and alternative NHEJ (Alt-EJ) in mammalian cells, especially in CHO cells that inherent resist HDR. To solve the above hurdle, here we for the first time show that knockout the DNA polymerase θ (POLθ), which is essential for Alt-EJ, significantly increases the knock-in efficiency by nearly forty-fold in CHO cells via eGFP reporter system and does not affect the normal growth and proliferation of cells. Meanwhile, even when transfecting simple circular, without negative element homologous template DNA donor and CRISPR/Cas9 plasmid to two different genomic sites, the knock-in rate of 4kb donor integration can still reach a mean of over 80% (29/36) and 2.7% (1/36) of the selected cell colonies in POLQ -/- CHO cells, however, no positive knock-in cell colonies was obtained in wild-type CHO cells which respectively selected 62 cell colonies and 36 cell colonies. Furthermore, we show that POLQ promotes random integration in CHO cells. Finally, RNA-sequence analysis reveals not significant altered DNA repair, metabolism, apoptosis, and cell cycle in POLQ -/- cells. These findings open a new target gene POLQ to overcome bottlenecks of the precision genome editing.

目的 探讨全切片图像分割在肿瘤生物学效应空间分布分析中的准确性和可行性.方法 采用Lab色彩空间K均值聚类分别获取苏木精-伊红(HE)、TdT介导的dUTP缺口末端标记(TUNEL)和血小板内皮细胞黏附因子(PECAM-1/CD31)染色图像分割的颜色阈值关系式和亮度阈值,与基于颜色反卷积等分割方法比较,验证其分割性能[性能参数包括归一化互信息(NMI)、Kappa系数、平均交互比(mIoU)、平均精准率(mPr)、平均召回率(mRe)和平均准确度(mA)].以光动力治疗小鼠乳腺癌为例,对来自科学数据银行(ScienceDB)的数据集,运用核密度估计热力图和分布密度的空间分析方法分别可视化和量化肿瘤生物学效应,并对坏死、凋亡和血管密度与光通量进行相关性分析.结果 图像分割速率约为54.5 s/G;以基于颜色反卷积和Otsu的分割结果为金标准,HE、TUNEL和CD31染色图像分割的NMI为0.45～ 0.58,Kappa系数为0.60～0.80,mIoU为0.70～0.83,mPr为0.73～0.91,mRe为0.81～0.97,mA为0.94～0.96.光通量与坏死密度线性相关,其Pearson相关系数为0.88;进行线性回归分析,可得到关系式为Y=0.1014X+ 22.4700(其中:Y为坏死密度;X为光通量).结论 基于Lab色彩空间K均值聚类的方法实现了图像的精准分割,结合空间分析和统计学方法实现了肿瘤生物标记物的空间数据分析及可视化,在肿瘤图谱生物效应分析中具有巨大的潜力.

Gastric cancer is one of the significant lethal cancers in China. Most patients are diagnosed at an advanced stage, and if gastric cancer can be detected at an early stage through large-scale screening, patient survival can be considerably improved. In China, there are two obstacles toward the large-scale screening of early gastric cancer. One is that endoscopy is overly invasive, resulting in low patient acceptance, and the other is that the number of endoscopists is too small compared with China's large population. A capsule endoscopic robot can alleviate the first obstacle, and the second obstacle is expected to be solved using artificial intelligence. We transferred the state-of-the-art Big Transfer (BiT) to a small dataset of early gastric cancer endoscopic images and built an early gastric cancer classification model based on white-light endoscopic images. We customized the BiT hyperparameter rules in transfer learning based on local situations. The batch size was selected according to the GPU memory limit, and based on the batch size, the linear scale rules were used to adjust the optimizer's initial learning rate dynamically. The total number of training images for the small dataset was set at 256000, on which other hyperparameters of the transfer learning were set. This study experimented with multiple models having the same structure of ResNet-v2 but different depths and widths. The best model has a depth of 101 and a width three times the original one. It has an accuracy of 97.14%, an F1 score of 94.77%, a sensitivity of 90.67%, and a specificity of 99.73% on the test set. Furthermore, the results show that the effect of batch size on the model training is statistically insignificant. This paper transferred a large model to a small dataset of endoscopic images with the BiT customization. This will promote the use of large-scale models in the field of endoscopic image analysis, which can help realize a large-scale screening of early gastric cancer.

光动力疗法是目前临床上出现的一种新型治疗肿瘤的方法,利用光敏剂吸收可见-近红外光并在组织氧的参与下发生光化学反应,产生活性氧物质进而诱导肿瘤细胞凋亡或坏死.光动力疗法对肿瘤侵袭性低,具有较高的选择性和良好的患者依从性,因此被广泛用于各种消化道恶性肿瘤的姑息性治疗和挽救性治疗.本文对近年来国内外应用光动力疗法治疗消化道肿瘤的文献进行综述,对所报道的疾病类型、治疗病例数、光敏剂和光源、疗效和安全性等信息进行分类整理,并对所面临的挑战和近期的研究进展进行了汇总,以期全面和客观地了解光动力疗法在治疗消化道肿瘤中的应用和研究进展,探讨目前的新应用及存在的问题和可能的发展方向.

[目的]对排除自身影响因素条件下(饮食、药物和吸烟)肺癌患者呼吸标志物进行了研究,探讨呼出气中特征性挥发性化合物对肺癌无创诊断的价值.[方法]本实验通过气相色谱-质谱联用仪对20例初诊初治肺癌患者和11例健康志愿者空腹状态时呼出气体中挥发性化合物进行定性定量分析.[结果]通过曼-惠特尼U检验对肺癌患者以及健康对照组的成分均值比较,筛选出二硫化碳(对照组24.21±19.94ppb,肺癌组1.28±3.95ppb)、正己烷(对照组16.75±16.93ppb,肺癌组0.46±1.18ppb)和乙苯(对照组6.30±2.27ppb,肺癌组1.46±2.12ppb).3种挥发性化合物浓度差异具有统计意义(P值均＜0.05).二硫化碳、正己烷和乙苯三种物质联合指标的受试者工作特征曲线曲线下的面积为0.932,敏感度为95％,特异性为90.9％,P＜0.01.[结论]本研究初步探讨了在排除自身影响因素条件下(饮食、药物和吸烟)的肺癌呼吸特征标志物,为肺癌呼吸标志物研究的进一步发展提供了参考依据.

Diffuse optical tomography (DOT) is a noninvasive biomedical imaging method to reconstruct optical property distribution. Since the underdetermined characteristic of reconstruction process, a priori information such as the structure provided by multimodal images are beneficial for imaging quality. We introduce a deep convolutional neural network-based method to rapidly calculate the heterogenous region by the diffusive intensity distribution measured by the same device used for DOT imaging. The process is based on a convolutional forward model which can accurately calculate the diffusive light intensity distribution with known structure and corresponding optical properties. The heterogeneous region imaging network is the inverse of the forward model and trained with Monte Carlo simulation results. The trained inverse network achieves the imaging sensitivity and specificity of 0.91 and 0.89 for validation data-set and the reconstruction speed is under 0.1s peer image.

Alzheimer's disease (AD) is a dementia-type neurodegenerative disease with an increasing incidence in elderly population and a poor prognosis. Therefore, the early diagnosis technology of AD urgently needs to be improved. In this paper, the laboratory diagnostic technologies of Alzheimer's disease were reviewed in the field of neuropsychological assessment, neuroimaging technology, and biomarker detection, including the simple intelligence state scale, the Montreal cognitive assessment scale, the memory and executive function screening scale, structural MRI, and functional MRI, positron emission computed tomography, MRI-based artificial intelligence analysis, and β amyloid (Aβ), homocysteine, S100B protein, Aβ 42, tau protein, urine AD-related neurofilament protein (AD7c-NTP) and Aβ plaques in the retinas. The limitations of these technologies were analyzed, and the development trends of the technologies were summarized. In order to improve the efficiency of AD screening, it is necessary to build an early diagnosis system for AD, in which multimodal diagnosis technology should be used to distinguish different types of AD.

This study was designed to investigate the effect of 650-nm low-level laser irradiation (LLLI) as an adjunctive treatment of experimental periodontitis. To investigate possible LLLI-mediated anti-inflammatory effects, we utilized an experimental periodontitis (EP) rat model and analyzed c-Jun, c-Fos, ICAM-1, and CCL2 gene expressions on PB leukocytes and in the gingival tissue. Total RNA was isolated from the gingivae and peripheral blood (PB) leukocytes of normal, EP, scaling, and root planing (SRP)-treated EP and LLLI + SRP-treated EP rats, and gene expressions were analyzed by real-time PCR. The productions of c-Jun, c-Fos, ICAM-1, and CCL2 in gingivae were analyzed immunohistochemically. Tartrate-resistant acid phosphatase (TRAP) staining was used to determine osteoclast activity in alveolar bone. The c-Jun and ICAM-1 messenger RNA (mRNA) levels were significantly decreased in the EP rat gingival tissue treated by SRP + LLLI than by SRP, the c-Jun, ICAM-1, and c-Fos mRNA levels on PB leukocytes reduced after LLLI treatment but did not show any significant differences in both groups. There was no significant difference in CCL2 mRNA levels on PB leukocytes and in gingivae between the SRP + LLLI and the SRP groups. The c-Fos mRNA levels in gingivae did not show significant difference in both groups. Immunohistochemistry showed that the CCL2, ICAM-1, c-Jun, and c-Fos productions were significantly reduced in rats of the SRP + LLLI group compared with the only SRP group. LLLI significantly decreased the number of osteoclasts as demonstrated by TRAP staining. The 650-nm LLLI might be a useful treatment modality for periodontitis.

An interesting puzzle is the fact that an infection of a tumor necrosis factor α (TNF)-deficient host with pathogens such as bacteria or parasites that reside intracellularly inevitably ends fatally. Is this due to one specific role of TNF in the immune defense or are different functions responsible for this outcome? In this review we provide an update of the functions of TNF in the defense against the intracellular pathogens Listeria monocytogenes, Mycobacterium tuberculosis, and Leishmania major. Furthermore, we discuss the role of TNF in the generation of proinflammatory macrophages in mouse models of infection and summarize briefly the potential consequences of anti-TNF treatment for infectious diseases.