Assessment on pollution of Ochratoxin A in grain in China and its apoptosis effect on vitro-cultured human tubular kidney cells

Ochratoxin A (OTA) is nephrotoxic, immunosuppressive, and teratogenic in many species and is a possible human carcinogen. In this study, we investigated the OTA pollution situations of grains in northern China and the signaling pathway that mediated OTA-induced apoptosis in human tubular kidney cells (HKCs). Samples of grains collected from three representative areas were determined by using high-performance liquid chromatography fluorescence method. The effects of OTA on cell apoptosis, caspase-3, Bax, and Bcl-2 expression, and phosphorylation of c-Jun NH(2) terminal kinase (JNK) were detected in cultured HKCs via flow cytometry (FCM), Hoechst 33258 staining, and Western blot. It showed that OTA pollution of edible grains was very common in north China. OTA could affect caspase-3, Bax, and Bcl-2 expression and increased cell apoptosis in cultured HKCs. The JNK signalling pathway might play an important role during these cellular events.     

α‐ketoglutarate delays age‐related fertility decline in mammals

The fecundity reduction with aging is referred as the reproductive aging which comes earlier than that of chronological aging. Since humans have postponed their childbearing age, to prolong the reproductive age becomes urgent agenda for reproductive biologists. In the current study, we examined the potential associations of α‐ketoglutarate (α‐KG) and reproductive aging in mammals including mice, swine, and humans. There is a clear tendency of reduced α‐KG level with aging in the follicle fluids of human. To explore the mechanisms, mice were selected as the convenient animal model. It is observed that a long term of α‐KG administration preserves the ovarian function, the quality and quantity of oocytes as well as the telomere maintaining system in mice. α‐KG suppresses ATP synthase and alterations of the energy metabolism trigger the nutritional sensors to down‐regulate mTOR pathway. These events not only benefit the general aging process but also maintain ovarian function and delay the reproductive decline. Considering the safety of the α‐KG as a naturally occurring molecule in energy metabolism, its utility in reproduction of large mammals including humans deserves further investigation.     

Protective Effect of Phosphatidylcholine on Restoration of Ethanol-Injured Hepatocytes Related with Caveolin-1

The absorption of phospholipid may improve the fluidity of membrane and enzyme activities. Phospholipids also play a role in promoting Caveolae formation and membrane synthesis. Caveolin-1 has a significant effect on signaling pathways involved in regulating cell proliferation and stress responsiveness. Thus, we can speculate that Caveolin-1 could affect the sense of environmental stress. We use Chang liver cell line to investigate the ability of Caveolin-1 to modulate the cellular response to ethanol injury. Caveolin-1 downregulate cells (Cav-1^?/?) were established by stable transfecting with psiRNA-CAV1 plasmids, which were more sensitive to toxic effects of ethanol than the untransfected parental cells (WT). Releasing of ALT and electric conductivity were changed significantly in Cav-1^?/? cells compared with WT. Caveolin-1 gene silencing could obviously down-regulate the activities of protein kinase C-α (PKC-α) and phospho-p42/44 MAP kinase, indicating cell proliferation and self-repairing abilities were inhibited. However, the levels of Caveolin-1 and PKC-α were increased by phosphatidylcholine administration. The results indicated that the inhibition of lipid peroxidation by phosphatidylcholine could lead to the prevention of membrane disruption, which closely correlated with the level of Caveolin-1. Since the protective effects of phosphatidylcholine against ethanol-induced lipid peroxidation might be regulated by phospholipid-PKC-α signaling pathway, related with Caveolin-1, the potential effects of phosphatidylcholine on membranes need to be verified.     

The Antiretroviral Lectin Cyanovirin-N Targets Well-Known and Novel Targets on the Surface of Entamoeba histolytica Trophozoites

Entamoeba histolytica, the protist that causes amebic dysentery and liver abscess, has a truncated Asn-linked glycan (N-glycan) precursor composed of seven sugars (Man₅GlcNAc₂). Here, we show that glycoproteins with unmodified N-glycans are aggregated and capped on the surface of E. histolytica trophozoites by the antiretroviral lectin cyanovirin-N and then replenished from large intracellular pools. Cyanovirin-N cocaps the Gal/GalNAc adherence lectin, as well as glycoproteins containing O-phosphodiester-linked glycans recognized by an anti-proteophosphoglycan monoclonal antibody. Cyanovirin-N inhibits phagocytosis by E. histolytica trophozoites of mucin-coated beads, a surrogate assay for amebic virulence. For technical reasons, we used the plant lectin concanavalin A rather than cyanovirin-N to enrich secreted and membrane proteins for mass spectrometric identification. E. histolytica glycoproteins with occupied N-glycan sites include Gal/GalNAc lectins, proteases, and 17 previously hypothetical proteins. The latter glycoproteins, as well as 50 previously hypothetical proteins enriched by concanavalin A, may be vaccine targets as they are abundant and unique. In summary, the antiretroviral lectin cyanovirin-N binds to well-known and novel targets on the surface of E. histolytica that are rapidly replenished from large intracellular pools.Entamoeba histolytica causes amebic dysentery and liver abscess in the developing world (10, 20, 29). We are interested in E. histolytica glycoproteins containing Asn-linked glycans (N-glycans) for numerous reasons. E. histolytica makes an N-glycan precursor that contains 7 sugars (Man₅GlcNAc₂-PP-dolichol) rather than 14 sugars (Glc₃Man₉GlcNAc₂-PP-dolichol) made by most animals, plants, and fungi (21, 31, 44). E. histolytica N-glycans are used for quality control of glycoprotein folding in the endoplasmic reticulum (ER) lumen, and there is positive selection for sites of N-linked glycosylation in secreted and membrane proteins of E. histolytica (5, 11, 53).Unprocessed Man₅GlcNAc₂, by far the most abundant E. histolytica N-glycan, is present on the plasma membrane and vesicular membranes (31). The antiretroviral lectin cyanovirin-N, which is specific for α-1,2-linked mannose present on unprocessed N-glycans, binds E. histolytica N-glycans and forms aggregates or caps on the surface of E. histolytica trophozoites (1, 25, 31, 44, 45). E. histolytica glycoproteins are also capped by the plant lectin concanavalin A (ConA), which has a broader carbohydrate specificity (mannose and glucose) than cyanovirin-N (3, 16, 18, 19). Heavy subunits of the Gal/GalNAc lectin, the most important E. histolytica vaccine candidate, have 7 to 10 potential sites for N-linked glycosylation (32, 39, 43). Inhibition of N-glycan synthesis results in Gal/GalNAc lectins that are unable to bind to sugars on host epithelial cells.Carbohydrates appear to be an important target on the surface of E. histolytica as anti-proteophosphoglycan (PPG) monoclonal antibodies bind to O-phosphodiester-linked glycans and protect animal models from amebic infection (6, 33, 35, 40, 48). Lectin affinity columns are a powerful method for enriching unique parasite glycoproteins that may be identified by mass spectrometry (MS) of tryptic fragments (17, 55). For example, we recently used the plant lectin wheat germ agglutinin to dramatically enrich glycoproteins with short N-glycans of Giardia (42).The goal of the present studies was to explore further the interaction of the antiretroviral lectin cyanovirin-N with E. histolytica trophozoites in vitro. Questions asked included the following: Are E. histolytica glycoproteins with N-glycans replenished on the plasma membrane after capping with cyanovirin-N? What is the effect of cyanovirin-N capping on other amebic virulence factors and/or vaccine candidates (e.g., the Gal/GalNAc lectin and PPG)? Is capping by cyanovirin-N mediated by actin, as described for capping by the Gal/GalNAc lectin and ConA? What is the effect of the cyanovirin-N on amebic phagocytosis of mucin-coated beads, a surrogate assay for virulence? Which trophozoite glycoproteins are potential targets of cyanovirin-N (identified by mass spectrometry of lectin-enriched E. histolytica proteins)? Are any of them potential vaccine candidates?     

piR-823 inhibits cell apoptosis via modulating mitophagy by binding to PINK1 in colorectal cancer

Mitophagy plays a vital role in the maintenance of mitochondrial homeostasis and tumorigenesis. Noncoding RNA piR-823 contributes to colorectal tumorigenesis. In this study, we aim to evaluate piR-823-mediated mitophagy and its mechanistic association with colorectal cancer (CRC). Digital gene expression analysis was performed to explore the potential functions of piR-823. A piR-823 antagomir (Ant-823) was used to inhibit piR-823 expression, and piR-823 mimics (mimics-823) were used to increase piR-823 expression. Mitophagy was measured in vivo and in vitro by immunofluorescence and western blot analysis. JC-1 staining, ATP production, real-time PCR, and western blot analysis were used to measure changes in mitochondrial quality and number. siRNA transfection was used to inhibit mitophagy, and CCCP was used to induce mitophagy. RNA pull-down assays and RNA-binding protein immunoprecipitation assays were conducted to investigate the molecular mechanisms. Here, we found that CRC cells transfected with Ant-823 presented an altered expression of autophagic and mitophagy genes by Digital gene expression analysis. Ant-823 could promote Parkin activation and mitophagy in vitro and in vivo, followed by mitochondrial loss and dysfunction of some mitochondria, whereas mimics-823 exerted the opposite effects in CRC cells. The inhibition of mitophagy by siParkin alleviated Ant-823-induced mitochondrial loss and dysfunction, as well as apoptosis to a certain extent. Furthermore, piR-823 was found to interact with PINK1 and promote its ubiquitination and proteasome-dependent degradation, thus alleviating mitophagy. Finally, these findings were verifed in samples obtained by patients affected by colorectal cancer. In conclusion, we identify a novel mechanism by which piR-823 regulates mitophagy during CRC tumorigenesis by increasing PINK1 degradation. Subject terms: Colorectal cancer, Gastrointestinal cancer     

甘油二酯激酶α在结直肠癌中的表达及其与PKC、TNFα的相关性

目的探讨甘油二酯激酶α(DGKα)在结直肠癌中的表达及其与蛋白激酶C(PKC)、肿瘤坏死因子α(TNFα)表达的相关性。方法应用免疫组化方法检测DGKα、PKC和TNFα在48例结直肠癌、癌旁正常组织和9例腺瘤性息肉中的表达。结果 DGKα在结直肠癌组织和癌旁正常组织有表达,结直肠癌组织中DGKα阳性表达率(79.2%)显著高于腺瘤性息肉(阴性)和癌旁正常组织(33.3%,P0.05);PKC主要分布在结直肠癌和癌旁正常组织中,结直肠癌组织中PKC阳性表达率(35.4%)显著高于腺瘤性息肉(阴性),与癌旁正常组织(20.8%)相比无显著性差异(P0.05);TNFα在三种组织中均表达阳性,结直肠癌组织中TNFα阳性表达率(95.8%)显著高于腺瘤性息肉(55.6%,P0.05),与癌旁正常组织(87.5%)相比无显著性差异(P0.05);结直肠癌组织中,DGKα与PKC的表达呈负相关(r=-0.437,P0.05),与TNFα的表达没有相关性(r=0.185,P0.05)。结论DGKα在结直肠癌组织中的表达高于腺瘤性息肉,DGKα可能抑制了PKC的活性,但对TNFα没有明显的抑制作用,在临床病理鉴别诊断中有辅助价值。     

I型马立克氏病毒38kD磷蛋白与Ⅱ和Ⅲ型病毒间的交叉免疫反应

由致病性Ⅰ型马立克氏病病毒(MDV)特异性的单克隆抗体H_(19)制备亲和层析柱,从感染重组杆状病毒BP38Ⅱ的昆虫细胞系Sf9细胞中提纯MDV的pp38基因表达产物,以此免疫小鼠制备抗血清,测定其与不同型MDV毒株感染的鸡胚成纤维细胞免疫反应性,同时也用不同型MDV毒株(HVT,Z4)制备的抗血清和自然感染发病鸡血清分别与感染重组病毒的Sf9细胞进行免疫交叉反应.试验结果表明,完整的Ⅰ型MDV来源的pp38基因产物与所试Ⅱ型Z4株和Ⅲ型HVT Fc126毒株在抗原性上有显著交叉反应.这一结果表明,在Ⅱ型和Ⅲ型MDV毒株中可能存在着pp38的类似物.     

<Superscript>1</Superscript>H NMR metabolic profiling of gastric cancer patients with lymph node metastasis

Introduction

Gastric cancer (GC) is a malignant tumor worldwide. As primary pathway for metastasis, the lymphatic system is an important prognostic factor for GC patients. Although the metabolic changes of gastric cancer have been investigated in extensive studies, little effort focused on the metabolic profiling of lymph node metastasis (LNM)-positive or negative GC patients.

Objectives

We performed ¹H NMR spectrum of GC tissue samples with and without LNM to identify novel potential metabolic biomarkers in the process of LNM of GC.

Methods

¹H NMR-based untargeted metabolomics approach combined with multivariate statistical analyses were used to study the metabolic profiling of tissue samples from LNM-positive GC patients (n?=?40), LNM-negative GC patients (n?=?40) and normal controls (n?=?40).

Results

There was a clear separation between GC patients and normal controls, and 33 differential metabolites were identified in the study. Moreover, GC patients were also well-classified according to LNM-positive or negative. Totally eight distinguishing metabolites were selected in the metabolic profiling of GC patients with LNM-positive or negative, suggesting the metabolic dysfunction in the process of LNM. According to further validation and analysis, especially BCAAs metabolism (leucine, isoleucine, valine), GSH and betaine may be as potential factors of diagnose and prognosis of GC patients with or without LNM.

Conclusion

To our knowledge, this is the first metabolomics study focusing on LNM of GC. The identified distinguishing metabolites showed a promising application on clinical diagnose and therapy prediction, and understanding the mechanism underlying the carcinogenesis, invasion and metastasis of GC.