Bile reflux and bile acids in the progression of gastric intestinal metaplasia

Gastric intestinal metaplasia (GIM) is a precancerous lesion of gastric cancer (GC) and is considered an irreversible point of progression for GC. Helicobacter pylori infection can cause GIM, but its eradication still does not reverse the process. Bile reflux is also a pathogenic factor in GIM and can continuously irritate the gastric mucosa, and bile acids in refluxed fluid have been widely reported to be associated with GIM. This paper reviews in detail the relationship between bile reflux and GIM and the mechanisms by which bile acids induce GIM.     

Nur77通过NF-κB/IL-6信号途径促进胃癌细胞的侵袭与迁移

目的探讨孤儿核因子受体（Nur77）在胃癌组织中表达及其与患者预后状况之间联系，研究Nur77在胃癌细胞侵袭与迁移进程中的作用机制。方法利用Oncomine数据库在线分析Nur77在胃癌及胃黏膜组织中mRNA表达情况；基于Human ProteinAtlas网站数据对比胃癌及正常胃组织中Nur77蛋白表达分布特征；GEPIA2在线分析工具评估Nur77与患者总生存期之间联系；蛋白免疫印迹法(Western blot)比较正常胃黏膜上皮细胞GES-1与胃癌细胞AGS、MKN-45中Nur77蛋白表达差异；分别通过siRNA靶向干扰以及质粒转染上调Nur77表达和靶向下调IL-6验证Nur-77与IL-6之间表达调控关系；运用划痕实验检测干扰Nur77表达前后胃癌细胞的迁移能力改变；将胃癌细胞分空载体组和过表达Nur77组以及空载体转染后干扰IL-6处理组，过表达Nur77结合干扰IL-6处理组（质粒转染24 h后siRNA继续处理24 h）经Transwell小室实验检测Nur-77及IL-6在胃癌细胞迁移和侵袭过程中作用；利用Western blot检测Nur77表达对NF-κB/IL-6信号通路活化的影响。结果Oncomine数据库在线分析显示，与正常胃黏膜组织相比胃癌组织中Nur77的mRNA表达水平显著升高（P < 0.05）；免疫组化结果显示，胃癌组织中Nur77多表达于胃癌细胞细胞核内；Nur77的高表达与胃癌患者不良预后相关（P < 0.05）；胃癌细胞中高表达的Nur77参与IL-6的表达调控；划痕实验和Transwell小室结果显示，Nur77可能通过IL-6参与胃癌细胞的迁移和侵袭进程（P < 0.05）；Western blot结果显示，Nur77通过调控p-p65、p65、p-Stat3、Stat3的表达参与NF-κB/IL-6信号通路活化。结论胃癌中高表达Nur77与患者不良预后密切相关；Nur77可能通过调控NF-κB/IL-6信号途径活化参与胃癌细胞侵袭与迁移进程。     

miR-16-5p通过靶向YWHAQ调控乳腺癌耐药细胞的凋亡与迁移能力

目的探究miR-16-5p对乳腺癌紫杉醇耐药细胞生物学活性的影响及分子机制。方法以人乳腺癌亲本细胞（SKBR-3）与乳腺癌紫杉醇耐药细胞（SKBR-3/PR）为研究对象。实验设空白对照组、阴性对照组、miR-16-5p类似物组（miR-16-5p mimics）、miR-16-5p抑制剂组（miR-16-5p inhibitor），YWHAQ干扰组（si-YWHAQ），以及联合组（miR-16-5p mimics+si-YWHAQ）。利用qRT-PCR检测乳腺癌组织与癌旁组织、SKBR-3与SKBR-3/PR间miR-16-5p的表达水平。使用生信数据对miR-16-5p的靶基因做预测，双荧光素酶实验验证靶向结合。免疫印迹检测细胞YWHAQ、Bcl-2和Bax表达。CCK-8和Transwell检测细胞增殖迁移能力。流式细胞术检测耐药细胞周期凋亡改变。结果qRT-PCR显示乳腺癌组织中miR-16-5p水平低于癌旁组织（-1.19± 1.90 vs 1.59±1.76，P < 0.01）。生物信息预测YWHAQ是miR-16-5p的靶基因，荧光素酶实验证实miR-16-5p能够靶向调节YWHAQ（P < 0.01）。相对于SKBR-3细胞，SKBR-3/PR细胞中miR-16-5p表达水平降低（P < 0.01），YWHAQ表达水平增高（P < 0.05）。Western blot结果显示miR-16-5p类似物能够抑制YWHAQ表达，miR-16-5p抑制剂能够促进YWHAQ表达（P < 0.01）。相对于对照组，miR-16-5p类似物能够将耐药细胞阻滞在G0/G1期（[55.61±1.99）% vs（43.06±1.53）%，P < 0.01]，抑制细胞增殖和迁移能力（P < 0.01），促进细胞凋亡（[10.37±0.23）% vs（3.81±0.88）%，P < 0.01]，YWHAQ干扰组细胞迁移能力下降，凋亡率升高，miR-16-5p类似物组与YWHAQ干扰组细胞的Bax蛋白表达增加，YWHAQ与Bcl-2蛋白水平降低。相对于miR-16-5p类似物组，联合组细胞迁移能力被抑制（75.75±29.85 vs 181.11±11.71，P < 0.01），凋亡率显著升高（[24.20±2.43）% vs（14.10±4.47）%，P < 0.01]。结论miR-16-5p能够通过靶向调节YWHAQ调控Bcl-2/Bax的表达，从而改变乳腺癌紫杉醇耐药细胞的生物学活性。     

DNAM-1通过IL-2/STAT-5通路调节Ⅰ型调节性T细胞的增殖和功能

目的探讨DNAM-1对Ⅰ型调节性T细胞（Tr1细胞）活化、增殖和功能的影响及相关分子机制。方法利用anti-CD3/CD28激活小鼠T细胞，采用流式细胞术分别检测静息和激活状态下CD4⁺ T细胞和Tr1细胞DNAM-1分子表达变化；分离DNAM-1基因敲除小鼠（KO小鼠）脾脏初始CD4⁺ T细胞并体外诱导Tr1细胞，流式细胞术检测CD25和CD69活化分子表达水平，CFSE标记后检测增殖能力，IL-2刺激前后检测KO小鼠Tr1细胞分泌IL-10和转录激活蛋白（p-STAT5）水平变化。结果流式细胞术结果显示：与静息状态下相比较，激活状态的CD4⁺ T细胞和Tr1细胞表达DNAM-1分子均增高（P＜0.05）；敲除DNAM-1不影响小鼠脾脏Tr1细胞的数量和比例，但KO小鼠Tr1细胞表达细胞激活分子CD25和CD69均降低，差异有统计学意义（P＜0.05）；与WT小鼠比较，KO小鼠诱导型Tr1细胞体外增殖能力降低（P＜0.05）；与WT组Tr1细胞比较，KO小鼠Tr1细胞分泌抑制性细胞因子IL-10水平降低（P＜0.05），给予IL-2刺激后仍无法逆转，表达Il-10 mRNA和Gzmb mRNA水平降低（P＜0.05）；给予不同剂量IL-2刺激Tr1细胞后，KO小鼠Tr1细胞表达p-STAT5水平相比较WT组均降低（P＜0.05）。结论DNAM-1参与Tr1细胞的活化和增殖，并通过IL-2/STAT5信号通路影响Tr1细胞抑制功能。     

非小细胞肺癌细胞外泌体源性FZD10促进体外血管生成

目的探讨非小细胞肺癌（NSCLC）细胞外泌体源性FZD10在血管生成中的作用及其机制。方法采用超速离心法分离外泌体，并利用Western blot和RT-qPCR技术分析NSCLC细胞（95D和H1299）、正常人支气管上皮细胞（BEAS-2B）及其外泌体中FZD10的表达；通过转染FZD10-siRNA敲低FZD10表达，用FZD10未敲低和敲低的NSCLC细胞外泌体分别处理HUVEC细胞，利用体外血管生成实验观察其成管能力，采用ELISA和RT-qPCR技术分析血管生成相关因子VEGFA和Ang-1的表达；进一步利用Western blot分析外泌体源性FZD10对信号通路PI3K、Erk1/2和YAP/TAZ激活的影响。结果同BEAS-2B细胞及其外泌体相比较，FZD10在95D和H1299细胞及其外泌体中高表达（P < 0.01）；95D和H1299细胞来源的外泌体可促进HUVEC的微管形成及VEGFA、Ang-1的蛋白分泌、mRNA的表达（P < 0.01），但在95D和H1299细胞敲低FZD10后这些效果受到抑制。FZD10的敲低可抑制PI3K、Erk1/2信号通路的激活，但对YAP/TAZ信号通路的影响不显著。结论NSCLC细胞外泌体源性FZD10可促进体外血管生成，其机制可能与PI3K、Erk1/2信号通路的激活有关。     

Epigallocatechin gallate ameliorates tetrahydrochloride-induced liver toxicity in rats via inhibition of TGFβ / p-ERK/p-Smad1/2 signaling,antioxidant, anti-inflammatory activity

Chronic liver disease is a worldwide health problem. Carbon tetra hydrochloride is an environmental toxin which is regarded as highly toxic and a potential human carcinogen. It can cause liver damage through the generation of metabolites and production of free radicals. Green tea contains catechins such as Epigallocatechin gallate which has been found to reduce the inflammation, oxidative stress, and fibrosis in experimental animal models. Hence, it represents a good source to prevent or ameliorate several chronic diseases. Silymarin is extracted from milk thistle seeds and has been found to be an effective agent to reduce the oxidative stress and free radical production and thereby exert protective effects in chronic liver conditions. The present study was planned to keep in view the above-mentioned facts. We included thirty rats in our study and divided them into five groups, each having six rats and the study continued for 8 weeks. Group I received normal saline; Group 2 received i.p. CCl4 injections; Group 3 received CCl4 i.p. injection and Epigallocatechin gallate (EGCG) oral gavage, Group 4 received CCl4 i.p. injection and silymarin by oral gavage; and Group 5 received CCl4 i.p. injection and combined EGCG + silymarin by oral gavage. The study found that in group 2, CCl4 induced significant elevation of ALT and MDA and reduced GSH thereby signifying increased oxidative stress. CCl4 also significantly increased inflammatory (TNFα, NFκB, IL1β, and TGFβ) as well as fibrotic markers (p-ERK and p-Smad1/2 protein expression). EGCG and silymarin significantly reversed the previously mentioned parameters either alone or in combination; however, the effect was more pronounced in case of EGCG. We conclude that EGCG and silymarin possess liver protective effects through their antioxidant, anti-inflammatory, and antifibrotic action.     

AZD9291通过抑制PI3K-AKT-mTOR通路抑制鼻咽癌细胞的增殖和迁移

目的探讨AZD9291对鼻咽癌细胞增殖和迁移的影响。方法体外培养鼻咽癌HNE1和CNE2Z细胞，在HNE1细胞加入浓度分别为0、0.5、1、2、4、8 μmol/L的AZD9291，CNE2Z细胞加入分别为0、1、2、4、8、16 μmol/L的AZD9291。采用CCK8法检测细胞存活率；集落克隆实验检测AZD9291对细胞的增殖抑制作用；细胞划痕实验和Transwell实验检测细胞修复和迁移能力；Western blot法检测EGFR相关信号通路蛋白及迁移相关蛋白的表达。结果CCK8和集落克隆实验结果显示AZD9291可显著抑制HNE1和CNE2Z细胞增殖（P < 0.01）；细胞划痕实验和Transwell实验结果显示AZD9291抑制HNE1和CNE2Z细胞迁移能力（P < 0.01）；Western blot结果显示，随着浓度增加，AZD9291可通过调控EGFR下游PI3K-AKT-mTOR信号通路磷酸化蛋白的下调（P < 0.01），抑制HNE1和CNE2Z细胞迁移（P < 0.01）。结论AZD9291可通过抑制EGFR/PI3K/AKT/mTOR信号通路，抑制鼻咽癌HNE1和CNE2Z细胞的增殖并降低其修复和迁移能力，为后续AZD9291尝试用于鼻咽癌的治疗提供依据。     

Resveratrol Protects against Zearalenone-Induced Mitochondrial Defects during Porcine Oocyte Maturation via PINK1/Parkin-Mediated Mitophagy

Mitochondria hold redox homeostasis and energy metabolism as a crucial factor during oocyte maturation, while the exposure of estrogenic mycotoxin zearalenone causes developmental incapacity in porcine oocyte. This study aimed to reveal a potential resistance of phytoalexin resveratrol against zearalenone during porcine oocyte maturation and whether its mechanism was related with PTEN-induced kinase 1 (PINK1)/Parkin-mediated mitophagy. Porcine oocytes were exposed to 20 μM zearalenone with or without 2 μM resveratrol during in vitro maturation. As for the results, zearalenone impaired ultrastructure of mitochondria, causing mitochondrial depolarization, oxidative stress, apoptosis and embryonic developmental incapacity, in which mitophagy was induced in response to mitochondrial dysfunction. Phytoalexin resveratrol enhanced mitophagy through PINK1/Parkin in zearalenone-exposed oocytes, manifesting as enhanced mitophagy flux, upregulated PINK1, Parkin, microtubule-associated protein light-chain 3 beta-II (LC3B-II) and downregulated substrates mitofusin 2 (MFN2), voltage-dependent anion channels 1 (VDAC1) and p62 expressions. Resveratrol redressed zearalenone-induced mitochondrial depolarization, oxidative stress and apoptosis, and accelerated mitochondrial DNA copy during maturation, which improved embryonic development. This study offered an antitoxin solution during porcine oocyte maturation and revealed the involvement of PINK1/Parkin-mediated mitophagy, in which resveratrol mitigated zearalenone-induced embryonic developmental incapacity.     

An Eight-Year Survey on Aflatoxin B1 Indicates High Feed Safety in Animal Feed and Forages in Northern Italy

Aflatoxins (AFs) remain the main concern for the agricultural and dairy industries due to their effects on the performances and quality of livestock production. Aflatoxins are always unavoidable and should be monitored. The objective of this paper is to bring to light a significant volume of data on AF contamination in several animal feed ingredients in Northern Italy. The Regional Breeders Association of Lombardy has been conducting a survey program to monitor mycotoxin contamination in animal feeds, and in this paper, we present data relating to AFB1 contamination. In most cases (95%), the concentrations were low enough to ensure compliance with the European Union’s (EU’s) maximum admitted levels for animal feed ingredients. However, the data show a high variability in AF contamination between different matrices and, within the same matrix, a high variability year over year. High levels of AFs were detected in maize and cotton, especially in the central part of the second decade of this century, i.e., 2015–2018, which has shown a higher risk of AF contamination in feed materials in Northern Italy. Variability due to climate change and the international commodity market affect future prospects to predict the presence of AFs. Supplier monitoring and control and reduced buying of contaminated raw materials, as well as performing analyses of each batch, help reduce AF spread.     

Small junction,big problems: Neuromuscular junction pathology in mouse models of amyotrophic lateral sclerosis (ALS)

Amyotrophic lateral sclerosis (ALS) is a motor neuron disease with an extremely heterogeneous clinical and genetic phenotype. In our efforts to find therapies for ALS, the scientific community has developed a plethora of mouse models, each with their own benefits and drawbacks. The peripheral nervous system, specifically the neuromuscular junction (NMJ), is known to be affected in ALS patients and shows marked dysfunction across mouse models. Evidence of pathology at the NMJ includes denervated NMJs, changes in endplate size and loss of terminal Schwann cells. This review compares the temporal disease progression with severity of disease at the NMJ in mouse models with the most commonly mutated genes in ALS patients (SOD1, C9ORF72, TARDBP and FUS). Despite variability, early NMJ dysfunction seems to be a common factor in models with SOD1, TARDBP and FUS mutations, while C9ORF72 models do not appear to follow the same pattern of pathology. Further work into determining the timing of NMJ pathology, particularly in newer ALS mouse models, will confirm its pivotal role in ALS pathogenesis and therefore highlight the NMJ as a potential therapeutic target.