Month: September 2020

PXR activation was achieved by intraperitoneal injections of pregnenolone 16 a-carbonitrile, a mouse specific PXR activator. In AKR/J mice we demonstrate that PXR activation is capable of regulating lipid metabolism and energy expenditure, and consequently, preventing the development of high-fat diet-induced MK-4827 obesity and insulin resistance. They are also more insulin resistant and therefore, are commonly used as a model for research on diet-induced obesity and obesity-related insulin resistance. As shown in Figure 1A, PCN treatment did not affect the growth rate of mice on regular chow. However, for animals fed with high-fat diet, PCN treatment resulted in a significant decrease in growth rate as compared to those treated with DMSO. Results shown in Figures 4 and 6 demonstrate that PCNmediated PXR activation significantly prevented the lipid storage in adipose tissue and the liver. Except for inhibiting lipogenesis, a more important characteristic of PXR in diet-induced obesity was reduction of lipid uptake in liver and adipose, confirmed by inhibiting up-regulation of Cd36 expression by high-fat diet. Cd36 codes for a scavenger receptor with broad ligand specificity. Activation of Cd36 facilitates free fatty acid uptake from circulation and also contributes to obesity, hepatic steatosis and type-2 diabetes. Previous studies show that Cd36 is a shared transcriptional target of LXR, PXR and PPARc in their regulation of lipid homeostasis. In the PXR-transgenic mice, Cd36 was up-regulated when fed with regular chow, which is different from our result obtained from the high-fat diet-fed mice. In our opinion, as in drug metabolism, PXR may serve as a “sensor” for maintaining energy homeostasis. Unlike the effect of PCN in the liver and WAT, PCN treatment increased Cd36 transcript level by approximately 2-fold compared to the DMSO-treated group in BAT. The main function of BAT is to generate heat for thermogenesis. Bartelt et al. reported that increased BAT activity enhanced triglyceride-rich lipoprotein metabolism in mice. Exposure to low temperatures drastically accelerated plasma clearance of triglycerides as a result of increased uptake into BAT, which was mediated by an increase of transmembrane receptor CD36. Although it remains to be demonstrated for increased energy expenditure, mice with PCN treatment had a much slower decrease in body temperature than the control group, suggesting that the activation of PXR enhanced the thermogenesis activity in BAT. At the same time, data from real-time PCR showed a significant increase in the transcription of genes involved in fatty acid b-oxidization and thermogenesis, indicating increased energy expenditure in BAT, which contributed to cleaning more lipids in BAT. In summary, our study demonstrates that PCN-mediated PXR activation prevented diet-induced obesity in AKR/J mice, decreased lipid accumulation and maintained insulin sensitivity. Additional work is needed to illustrate the precise mechanisms through which PXR modulates energy metabolism and lipid homeostasis.

In conclusion, six serum miRNAs identified in our study can be used to predict sepsis patients’ mortality. Germline mutations in the currently known high risk-breast cancer genes are common in familial breast cancer, but they can explain, at best, 20–25% of the overall excess familial risk.. Still, the large majority of breast cancer cases that arise in families with strong familial aggregation are not explained by mutations in any know breast cancer susceptibility gene, and are designated as BRCAX-type tumors. In the past decade, gene expression profiling by microarray analysis has lead to great advances in classification of human breast tumors, and the identification of five reproducible molecular subtypes of breast cancer, that have distinct biological features, clinical outcomes, and responses to chemotherapy. On the other hand, there have been only a handful of studies focused on familial breast cancer, due to difficulties in collecting the tumor material, demonstrating that BRCA1/2-mutated breast tumors could be distinguished from sporadic ones based on their gene expression signatures. Recently, microRNA expression profiling calls a great attention to define various types of cancers. miRNAs are an abundant class of small,22 nt long single-stranded non-coding RNA molecules acting as negative regulators at post-transcriptional level by binding the 39 untranslated regions of their mRNA-targets. miRNAs are involved in crucial biological processes including development, differentiation, apoptosis and proliferation. Notably, miRNA deregulation has been extensively implicated in cancer pathogenesis in various tumor types. The observed effects of miRNA mis-expression on tumor initiation, maintenance or metastasis can be explained by the mRNA targets and pathways they regulate, which include known tumor suppressors and oncogenes. Specifically, in breast cancer, various studies have identified mis-expressed miRNAs in tumours vs. normal tissue, and shown that changes in their expression seem to define, similarly to what has been found by expression profiling of coding genes, different histological and molecular subtypes described so far. In addition, integration of miRNA and mRNA data of a set of breast cancer samples allowed the association of miRNAs to relevant cellular processes, such as proliferation, cell cycle, immune response or cell adhesion, as well as with molecular characteristics of tumors like TP53 mutations. Still, very little is known about the role of miRNAs in familial breast cancer. The identification of target genes and pathways NVP-BKM120 regulated by miRNAs would be critical to understand their function in tumor development. In this study we sought to establish miRNA expression profiles using microarray technology of familial breast cancer tumors and comparing with normal breast tissues. Interestingly, KRAS has been identified as a target oncogene for down-regulated miRNAs.The identification of miRNAs deregulated in familial breast tumors could provide a better understanding of the biology of familial breast cancer and could indicate novel targets for therapy.

We further found that GEP100 down-regulation with shRNA significantly inhibited the Matrigel invasion ability of PaTu8988 cells, but had no effect on the cell viability, migration and adhesion, indicating that GEP100 is particularly responsible for the invasive ability of cells. In vivo experiment also showed that GEP100 knock-down significantly inhibited the liver metastasis of pancreatic cancer cells in Balb/c nude mice. This result was in line with previous data demonstrating that in breast cancer cells GEP100 was highly expressed in the invasive ones and that GEP100 down-regulation inhibited cell invasion and lung metastasis. These results indicated that GEP100 is not only a CUDC-907 target for breast cancer, it might be a general mechanism used by other cancer types. Several other biological functions for GEP100 have been reported. In HeLa cells, GEP100 regulated cell adhesion through controlling endocytosis and recycling of integrin b. In a liver carcinoma cell line HepG2, GEP100 directly interacted with acatenin and played a role in actin cytoskeleton remodeling. In myoblasts and macrophages, GEP100 was involved in cell-cell fusion. It has also been reported to take part in the regulation of nucleolar architecture and phagocytosis. In this study, we did not observe any significant effect of GEP100 on cell adhesion to collagen matrix or migration activity. Since there is not any report on the role of GEP100 in pancreatic cancer cells, this may be due to the difference in cell type. Unlike the previous report on a close relation between Arf6 expression and breast cancer cell invasive ability, we failed to detect that in the panel of pancreatic cancer cell lines we used. The above mentioned biological functions of GEP100 have been found to be dependent on the activation of Arf6, but it is believed that GEP100 is a multifunctional protein that regulates cellular functions in both an Arf-dependent and -independent manner. For example, GEP100 was involved in apoptotic cell death independent of Arf6 activity. Further studies including determination of the activation status of Arf6 will be necessary to reveal the role of Arf6 in the process of pancreatic cancer cell invasion. We found GEP100 knock-down caused a morphological change of cells from mesenchymal to epithelial phenotype. In pancreatic cancer, shifting of their epithelial features toward a mesenchymal phenotype enhances cell motility and is considered to be a prerequisite for tumor invasion. E-cadherin is the best characterized molecular marker in epithelial cells and localized at the adherens junctions. Loss of E-cadherin expression and/or function is a well-recognized marker of EMT and promotes invasion. Therefore we examined the expression of Ecadherin protein. GEP100 shRNAs treatment increased Ecadherin expression level 2 to 3 -fold. This is consistent with Hiroi’s report showing that in HepG2 cell, down-regulation of GEP100 increased the expression of E-cadherin. In addition, they also demonstrated that GEP100 interacted with a-catenin.

In an animal study, hyper-ammonemia results in increased numbers of swollen astrocytes, increased immuno-reactivity of glutamine synthetase, and some cytoskeletal proteins like the intermediate filament glial fibrillary acidic protein. These increases in swollen astrocytes and GFAP immuno-reactivity can be reduced by GS inhibition. The GFAP is expressed in astrocytes and is involved in cell structure maintenance and functioning of the blood brain barrier. It is also proposed to play a role in astrocyteneuron interactions, which may explain the alteration of astrocyte morphology and function. However, the limited correlation between decreased DMN integrity and ammonia level in this and another small sample-sized studies also imply a more complicated process in HE development or an adaptive change of functional LEE011 network that exists in each individual subject with a diverse clinical profile. Our results reveal significant antero-posterior functional disconnection in the PCC functional maps. Similar results have been suggested in subjects with increasing depth of sleep, wherein the posterior areas strengthen their connectivity, while connections between the frontal and posterior regions are lost. Functional uncoupling between the PCC and ACC and MFC may impair the brain’s ability to integrate information. Results of the present study suggest that integrated DMN activity may reflect ongoing conscious mental activity. In addition, increased MD, especially in the ACC, has good correlation with the disruption of the antero-posterior functional network, which further supports the notion that the PCC/precuneus are the central core within the DMN. Early hepatic encephalopathy is not identified by structural abnormalities. Instead, a comprehensive neurologic examination is usually required. Revealing the subtle functional alteration in “sub-clinical HE” is even more difficult. In an FDG-PET study, mHE is associated with decreased glucose uptake and blood flow in the ACC, medial frontal region, and precuneus. These indicate that brain regions involved in controlling the “attention system” responsible for monitoring is less active in mHE patients than in normal subjects. The NP tests, especially the block design and digit-symbol tests, are attention-demanding. Taken together, these findings support the hypothesis that a part of DMN connectivity fluctuates as it is more closely related to the presence of cognitive processes and can be altered earlier in liver cirrhosis. This study has some limitations. In subject recruitment, significantly ill patients were excluded from MRI study for safety concerns. As such, the preservation of DMN in comatose patients is unknown. Moreover, cirrhotic patients with altered consciousness are difficult to hold motionless during MRI scanning. Since the sample size is small, a relative low threshold for head motion criteria in data analysis has been applied, which might affect the results. Further studies with larger sample populations are warranted to assess the effect of therapy on functional network. Further validation of this diagnostic value on mHE by rs-fMRI is also required. In conclusion, HE patients show connectivity de-coupling between the fronto-posterior areas of DMN, which is associated with the degree of HE and brain oedema. Rs-fMRI can be used to investigate variability among patients with differing symptom profiles, including sub-clinical states.

Vc9Vd2 T-cells are involved in host response to many chronic viral infections, including HCV. As observed in other chronic infection such as HIV, a KRX-0401 decrease of peripheral Vc9Vd2 T cell subset was observed associated to HCV infection. Activated Vc9Vd2 T lymphocytes were found able to inhibit subgenomic HCV replication, and this effect was mediated mainly by IFN-c release. A role of recombinant IFN-c on subgenomic HCV replication was also described. Moreover, several studies showed that the combination of recombinant IFN-c and IFN-a resulted in a strongly enhanced antiviral activity in the HCV replicon model, opening the way to new combined treatment approaches. Thus, IFN-c induced by Vc9Vd2 T-cell stimulation could enhance standard treatment effectiveness. In this work, phenotype and function of Vc9Vd2 T-cells were analyzed during chronic HCV infection, evaluating possible strategies aimed to improve their effector response. This approach was validated in vivo in a non-human primate model. Main aim of our work was to study the effects of chronic HCV infection on Vc9Vd2 T-cell phenotype and function, and on possible strategies aimed to improve their effector activity. Chronic HCV infection induced a slight but significant decrease in the frequency of Vc9Vd2 T-cells. An increased liver tissue compartmentalization of these cells may represent an additional factor. Differentiation and activation profile analysis of Vc9Vd2 T-cells showed an increase in circulating effector and activated cells. These data may be explained in the context of a chronic infection leading to a persistent stimulation of immune cells, driving their activation and differentiation. In our patients, no correlation was found between Vc9Vd2 T-cells dysfunction and any clinical parameter. Vc9Vd2 T-cells play a pivotal role in viral infections, for their ability to mediate broad antiviral and immunomodulating activities. Specifically, antiviral role of activated Vc9Vd2 T-cells, mainly mediated by IFN-c release, has been demonstrated for several viruses such as coronavirus, orthopoxvirus, HIV, and HCV. In our work, a severe functional inability of Vc9Vd2 T-cells to produce IFN-c was shown in HCV patients, independently from viral load and genotype. Such as DC and NK cells, that could be associated to adaptive immune response dysfunction and/or exhaustion. In this context, a complex network of different signals can act to induce immune cell exhaustion, such as chronic inflammation, persistent antigen stimulation, and/or direct viral effects. Chronic inflammation and persistent antigen stimulation, as observed during HIV infection, may result in Vc9Vd2 T-cell exhaustion and anergy through activation-induced cell death, or through a decrease in Vc9Vd2 T-cells response by down-modulating CD3-j chain expression. Finally, although controversial, a possible direct HCV-driven inhibition of NK cell function through HCVE2/CD81 binding has been reported. Interestingly, CD81 expression by cd T-cells was previously reported. A study aimed to define cellular and molecular mechanisms involved in Vc9Vd2 T-cells exhaustion during chronic HCV infection may be useful to evaluate possible strategies to restore their activity. The main result of our work is the demonstration that Vc9Vd2 T-cell function may be improved by IFN-a both in HD and in HCV-infected patients.