Disruption of intestinal epithelial permeability in the Co-culture system of Caco-2/HT29-MTX cells exposed individually or simultaneously to acrylamide and ochratoxin A.

Mycotoxins and thermal processing hazards are common contaminants in various foods and cause severe problems in terms of food safety and health. Combined use of acrylamide (AA) and ochratoxin A (OTA) would result in more significant intestinal toxicity than either toxin alone, but the underlying mechanisms behind this poor outcome remain unclear. Herein, we established the co-culture system of Caco-2/HT29-MTX cells for simulating a real intestinal environment that is more sensitive to AA and OTA, and showed that the combination of AA and OTA could up-regulate permeability of the intestine via increasing LY permeabilization, and decreasing TEER, then induce oxidative stress imbalance (GSH, SOD, MDA, and ROS) and inflammatory system disorder (TNF-α, IL-1ß, IL-10, and IL-6), thereby leading a rapid decline in cell viability. Western blot, PAS- and AB-staining revealed that AA and OTA showed a synergistic effect on the intestine mainly through the disruption of tight junctions (TJs) and a mucus layer. Furthermore, based on correlation analysis, oxidative stress was more relevant to the mucus layer and TJs. Therefore, our findings provide a better evaluation model and a potential mechanism for further determining or preventing the combined toxicity caused by AA and OTA.

PAIP1 regulates expression of immune and inflammatory response associated genes at transcript level in liver cancer cell.

Poly(A) binding protein interacting protein 1 (PAIP1) is a translation regulator and also regulate the decay of mRNA. PAIP1 has also been reported to be a marker of increased invasive potential of liver cancer. However, the roles and underlying molecular mechanism of PAIP1 in liver cancer is still unclear. Here, cell viability and the gene expression profile of liver cancer line HepG2 transfected with PAIP1 siRNA was compared with cells transfected with non-targeting control siRNA. The results showed that PAIP1 knockdown inhibited cell viability, and extensively affects expression of 893 genes at transcriptional level in HepG2 cells. Gene function analysis showed that a large number of PAIP1 up-regulated genes were enriched in term of DNA-dependent transcription and the down-regulated genes were enriched in some pathways including immune response and inflammatory response. qPCR confirmed that PAIP1 knockdown positively regulated the expression of selected immune and inflammatory factor genes in HepG2 cells. Expression analysis of TCGA revealed that PAIP1 had positive correlations with two immune associated genes IL1R2 and PTAFR in liver tumor tissue. Taken together, our results demonstrated that PAIP1 was not only a translation regulator, but also a transcription regulator in liver cancer. Moreover, PAIP1 could function as a regulatory factor of immune and inflammatory genes in liver cancer. Thus, our study provides important cues for further study on the regulatory mechanism of PAIP1 in liver cancer.

Hepatitis C virus core protein induces dysfunction of liver sinusoidal endothelial cell by down-regulation of silent information regulator 1.

Hepatic fibrosis is a frequent feature of chronic hepatitis C virus (HCV) infection. Some evidence has suggested the potential role of silent information regulator 1 (SIRT1) in organ fibrosis. The aim of this study was to investigate the effect of HCV core protein on expression of SIRT1 of liver sinusoidal endothelial cell (LSEC) and function of LSEC. LSECs were co-cultured with HepG2 cells or HepG2 cells expressing HCV core protein and LSECs cultured alone were used as controls. After co-culture, the activity and expression levels of mRNA and protein of SIRT1 in LSEC were detected by a SIRT1 fluorometric assay kit, real time-PCR (RT-PCR), Western blot, respectively. The levels of adiponectin receptor 2 (AdipoR2), endothelial nitric oxide synthase (eNOS) and vascular endothelial growth factor (VEGF) were measured by Western blot. Cluster of differentiation 31 (CD31), CD14, and von Willebrand factor (vWf) of LSECs was performed by flow cytometry. The level of reactive oxygen species (ROS) was assayed. Malondialdehyde (MDA), superoxide dismutase (SOD), adiponectin, nitric oxide (NO), and endothelin-1 (ET-1) levels in the co-culture supernatant were measured. The co-culture supernatant was then used to cultivate LX-2 cells. The levels of α-smooth muscle actin (ASMA) and transforming growth factor-ß1 (TGF-ß1) protein in LX-2 cells were measured by Western blot. Compared with LSEC co-cultured with HepG2 cells group, in LSEC co-cultured with HepG2-core cells group, the activity and expression level of mRNA and protein of SIRT1 reduced; the level of adiponectin reduced and the expression level of AdipoR2 protein decreased; ROS levels increased; the expression level of eNOS, VEGF protein decreased; and the expression level of CD14 decreased; the expression level of vWf and CD31 increased; NO and SOD levels decreased; whereas ET-1 and MDA levels increased; the levels of ASMA and TGF-ß1 protein in LX-2 cells increased. SIRT1 activator improved the above-mentioned changes. HCV core protein may down-regulate the activity and the expression of SIRT1 of LSEC, then decreasing synthesis of adiponectin and the expression of AdipoR2, thus inducing contraction of LSEC and hepatic sinusoidal capillarization and increasing oxidative stress, ultimately cause hepatic stellate cell (HSC) activation. Treatment with SIRT1 activator restored the function of LSEC and inhibited the activation of HSC.