Cytoplasm Of Liver Cells Research Articles

Associated metabolic fatty liver disease and cardiovascular pathology – double causal connection

Introduction. The accumulation of neutral lipids in the cytoplasm of liver cells favors the onset of insulin resistance, the accumulation of lipotoxic metabolites and bile acids in hepatocytes, oxidative stress, the release of cytokines and other inflammatory mediators, as well as the intestinal microbiota, are considered to promote the transition from steatosis to inflammation liver damage and fibrosis through mitochondrial dysfunction, endoplasmic reticulum stress, cell death and immune cell infiltration. These processes are the key factor in explaining the associations between fatty liver disease, metabolic diseases and cardiovascular diseases. Results. The presence of metabolic-associated fatty liver disease (MAFLD) is associated with increased cardiovascular (CV) risk, which is why appropriate management is necessary to improve the outcome of CV diseases and of hepatic ones. According to the recent statements of the American Cardiology Association, MAFLD should be considered as a marker of risk and worsening of cardiovascular pathologies. Conclusions. Currently, there are insufficient data to categorize MAFLD as a CV risk factor because its impact on risk has not been quantified, although MAFLD has been shown to have an additional negative impact on atherosclerotic CV disease risk (ACVDF), in addition to traditional risk factors (such as age, sex, family history of CVD, obesity, DM, hypertension, dyslipidemia, and smoking). Furthermore, studies demonstrating ACVDF reduction as a result of MAFLD treatment are not yet available, but they are essential in integrating an element as a risk factor.

Pathological effects due to metacercariae of Clinostomum piscidium migration in snakeskin gourami (Trichopodus pectoralis) in Thailand.

Clinostomum spp. is a fish-borne pathogen and a digenetic trematode with a global range. Despite its zoonotic relevance, the pathogenic impact of the parasite in Thai aquaculture is currently unclear. The present study deals with the pathogenic changes that fluke causes in their host, Trichopodus pectoralis, and the molecular confirmation of the Clinostomum piscidium by targeting 18 s rDNA and ITS gene. The metacercariae of C. piscidium were discovered in the body cavity of infected fish. The gross pathological examination revealed a few white migratory tracks on the surface of the liver and spleen. The migratory track showed histologically as a primary hemorrhage and necrosis of hepatic cells surrounded by a layer of macrophages and epithelioid cells, inflammatory cells, and eosinophilic granular cells in the cytoplasm of liver cells and close to the epithelial cells of the intestine. Also, the migratory track in the spleen appeared as a marked decrease of Red Blood Cell (RBC) count and changes in the necrotic tissue. Infection with this metacercaria produced hepatic tissue injury, which disrupted hepatic metabolism and decreased body weight in the fish hosts. The findings of the study suggest that the pathological effect of C. piscidium on farm T. pectoralis can cause significant economic loss by stunting fish development and predisposing fish to opportunistic pathogens in the environment. Hence, the treatment and control of C. piscidium infections are crucial for the viability of the aquaculture sector since this parasite has been found to cause pathological damage to the vital organs of fish.

Preparation of a JAZF1 protein polyclonal antibody and its potential role in broiler ascites syndrome

As a novel functional protein, juxtaposed with another zinc finger protein 1 (JAZF1) can regulate the growth and apoptosis through various pathways, and maintain the body's normal physiological metabolism. To explore the important role of JAZF1 in broiler ascites syndrome (BAS), we analysed the expression and distribution of the protein in poultry and mammal tissues based on the prepared polyclonal antibody. In this study, the recombinant plasmid PET32a-JAZF1 was constructed by TA cloning, subcloning and other technical methods, and the fusion protein His-JAZF1 was successfully expressed. After purification, His-JAZF1 was used as the antigen to prepare high-quality chicken-derived antibodies. Subsequently, the results showed that JAZF1 protein in broiler tissues could be specifically recognized by this antibody. Immunofluorescence showed that JAZF1 protein mainly exists in the cytoplasm of pulmonary artery, liver, kidney, heart and lung tissue cells of various animals. The expression of this protein was more obvious in broiler and duck tissues than in mammalian tissues. In addition, western blotting combined with immunofluorescence showed that BAS caused a significant decrease in JAZF1 protein in tissue cells. This effect further indicated that JAZF1 protein was closely related to the occurrence of BAS and provided a new entry point for the functional study of JAZF1 protein.

Acox2 is a regulator of lysine crotonylation that mediates hepatic metabolic homeostasis in mice

Acyl-CoA oxidase 2 (Acox2) is an enzyme involved in peroxisomal bile acid synthesis and branched-chain fatty acid degradation. Acox2 knockout (−/−) mice spontaneously developed liver cancer with marked lymphocytic infiltrate. Tandem-affinity purification coupled with mass spectrometry analysis revealed that Acox2 interacted with methylcrotonoyl-CoA carboxylase followed by co-immunoprecipitation confirmation. Here we reported that non-histone lysine crotonylation (Kcr) levels were downregulated in Acox2−/− mice livers. Interestingly, Kcr signals were concentrated in the nucleus of tumor cells but mostly located in the cytoplasm of adjacent normal liver cells of Acox2−/− mice. Quantitative analysis of the global crotonylome further revealed that 54% (27/50) of downregulated non-histone Kcr sites were located in mitochondrial (11/50) and peroxisomal (17/50) enzymes including Ehhadh, Scp2, Hsd17b4, Crot, Etfa, Cpt1a, Eci1/2, Hadha, Etfdh, and Idh2. Subsequent site-directed mutagenesis and transcriptome analysis revealed that Ehhadh K572cr might have site-specific regulatory roles by downregulating TOP3B expression that lead to increased DNA damage in vitro. Our findings suggested Acox2 is a regulator of Kcr that might play critical role on hepatic metabolic homeostasis.

Cloning and prokaryotic expression of the chicken liver kinase B1 (LKB1) and its localization in liver, heart and hypothalamus

Liver kinase B1 (LKB1) is a member of the serine/threonine kinase family, which plays an indispensable role in the organism of animals. In the current study, the chicken LKB1 protein gene was amplified by PCR based on the primers and cDNA templates. Then, the cloning vector was constructed and the target gene was cloned. After that, the target gene was inserted into the expression vector to construct the recombinant plasmid. The recombinant plasmid was transformed into BL21 (DE3) host cells and the LKB1 recombinant proteins were successfully expressed by using Isopropyl-β-D-thiogalactopyranoside (IPTG). Finally, purified LKB1 proteins were used as antigen and the rabbit-derived antiserums were collected. The antiserum titer determined by ELISA was not less than 1:128000. The results of Western blot suggested that the polyclonal antibody is highly specific to chicken LKB1 protein. Immunofluorescence indicated that the LKB1 protein is mainly expressed in the cytoplasm of liver, heart and hypothalamus cells of chicken. Our study showed that the LKB1 polyclonal antibodies produced by this method are effective and can be used to further study the role of LKB1 in the pathogenesis of chicken disease.

TOXIC EFFECTS OF ULTRA-DISPERSED FORMS OF METALS (Mo AND MoO3) IN THE EXPERIMENT in vivo

TOXIC EFFECTS OF ULTRA-DISPERSED FORMS OF METALS (Mo AND MoO3) IN THE EXPERIMENT in vivo

Molecular Evolution of Apolipoprotein Multigene Family and the Original Functional Properties of Serum Apolipoprotein (LAL2) in Lampetra japonica.

Apolipoprotein (APO) genes represent a large family of genes encoding various binding proteins associated with plasma lipid transport. Due to the long divergence history, it remains to be confirmed whether these genes evolved from a common ancestor through gene duplication and original function, and how this evolution occurred. In this study, based on the phylogenetic tree, sequence alignment, motifs, and evolutionary analysis of gene synteny and collinearity, APOA, APOC, and APOE in higher vertebrates may have a common ancestor, lamprey serum apolipoprotein LAL1 or LAL2, which traces back to 360 million years ago. Moreover, the results of immunofluorescence, immunohistochemistry, and flow cytometry show that LAL2 is primarily distributed in the liver, kidney, and blood leukocytes of lampreys, and specifically localized in the cytoplasm of liver cells and leukocytes, as well as secreted into sera. Surface plasmon resonance technology demonstrates that LAL2 colocalizes to breast adenocarcinoma cells (MCF-7) or chronic myeloid leukemia cells (K562) associated with lamprey immune protein (LIP) and further enhances the killing effect of LIP on tumor cells. In addition, using quantitative real-time PCR (Q-PCR) and western blot methods, we found that the relative mRNA and protein expression of lal2 in lamprey leukocytes and sera increased significantly at different times after stimulating with Staphylococcus aureus, Vibrio anguillarum, and Polyinosinic-polycytidylic acid (Poly I:C). Moreover, LAL2 was found to recognize and bind to gram-positive bacteria (Staphylococcus aureus and Bacillus cereus) and gram-negative bacteria (Escherichia coli) and play an important role in the antibacterial process. All in all, our data reveals a long, complex evolutionary history for apolipoprotein genes under different selection pressures, confirms the immune effect of LAL2 in lamprey sera against pathogens, and lays the foundation for further research regarding biological functions of lamprey immune systems.

Cytochrome P450s regulates aloperine-induced pathological changes in mouse liver and kidney

Aloperine is a major active component in Sophora alopecuroides L that plays diverse pharmacological properties. Recent studies have indicated the potential effect of aloperine against hypertension and cancers. However, possible toxicity of aloperine has not been carefully studied in vivo. The aim of this study was to assess the effect of intraperitoneal aloperine injection on mouse liver and kidney tissues and to investigate the role of CYP450 genes in aloperine-induced toxicity. 72 BALB/c mice were randomly divided into four groups: vehicle control group (normal saline), low-dose group (4 mg/kg), medium-dose group (8 mg/kg), and high-dose group (16 mg/kg). 18 mice in each group were intraperitoneally injected with aloperine daily for 4 weeks, and were then kept for another 1 or 4 weeks without aloperine treatment. Serum was colleted for analysis of serum biochemical indexes including ALT, AST, BUN and CRE. The liver and kidney were collected for analysis of histopathologic changes and CYP450 expression and activity. Vacuolization of cytoplasm in liver cells, swelling in kidney tubular cells, increased levels of ALT, AST, BUN, and CRE, and alteration in the expression and activity of CYP450 were observed in the high-dose group after 4 weeks of treatment. However, all aloperine-induced damages were recovered to a certain degree after maintained without aloperine for 1 week, and fully recovered after maintained without aloperine for 4 weeks. These findings suggested that aloperine regulated the expression of CYP450, which was possibly involved in aloperine-induced reversible toxicity in mouse liver and kidney tissues.

A Novel Gene Therapy Approach for GSD III Using an AAV Vector Encoding a Bacterial Glycogen Debranching Enzyme

Glycogen storage disease type III (GSD III) is an inherited disorder caused by a deficiency of glycogen debranching enzyme (GDE), which results in the accumulation of abnormal glycogen (limit dextrin) in the cytoplasm of liver, heart, and skeletal muscle cells. Currently, there is no curative treatment for this disease. Gene therapy with adeno-associated virus (AAV) provides an optimal treatment approach for monogenic diseases like GSD III. However, the 4.6 kb human GDE cDNA is too large to be packaged into a single AAV vector due to its small carrying capacity. To overcome this limitation, we tested a new gene therapy approach in GSD IIIa mice using an AAV vector ubiquitously expressing a smaller bacterial GDE, Pullulanase, whose cDNA is 2.2 kb. Intravenous injection of the AAV vector (AAV9-CB-Pull) into 2-week-old GSD IIIa mice blocked glycogen accumulation in both cardiac and skeletal muscles, but not in the liver, accompanied by the improvement of muscle functions. Subsequent treatment with a liver-restricted AAV vector (AAV8-LSP-Pull) reduced liver glycogen content by 75% and reversed hepatic fibrosis while maintaining the effect of AAV9-CB-Pull treatment on heart and skeletal muscle. Our results suggest that AAV-mediated gene therapy with Pullulanase is a possible treatment for GSD III.

Changes of lysosomal membrane permeabilization and lipid metabolism in sidt2 deficient mice.

The SID1 transmembrane family member 2 (sidt2) deficient mouse model was used to investigate the function of sidt2 in lysosomal membrane permeabilization and lipid metabolism of liver tissue. The mouse model was established by Cre/LoxP technology. Enzymatic methods were used to analyze the sidt2−/− mouse serum lipids, aspartate transaminase, alanine transaminase and serum bilirubin, compared with sidt2+/+ mice. Defective lipid metabolism and damaged liver functions were observed in the sidt2−/− mice. By using hematoxylin and eosin and Oil Red O staining, changes of morphology were observed in sidt2−/− mice with optical microscopy. Transmission electron microscopy was also used. Hepatic steatosis and partial liver tissue apoptosis were observed. The tissue distribution of sidt2 protein and mRNA was measured in knockout mice. The results indicated that negligible sidt2 mRNA and protein expression were observed in sidt2−/− mice, and that sidt2−/− mice had abnormal liver functions. Transmission electron microscopy revealed membrane lipid droplets in the liver cell cytoplasm, and some apoptotic body formation. These results demonstrated that absence of the lysosomal membrane protein sidt2 led to changes in lysosomal membrane permeabilization and lipid metabolism.

Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent organic pollutant (POP), an unintentional byproduct of various industrial processes, and a human carcinogen. The expression of the cytochrome P450 1A (cyp1a) gene is upregulated in the presence of TCDD through activating the aryl hydrocarbon receptor pathway in a dose-dependent manner. Several essential response elements, including the 8 potential xenobiotic response elements in the cyp1a promoter region, have been identified to be the main functional parts for the response to TCDD. Thus, we aimed to develop a convenient and sensitive biomonitoring tool to examine the level of POPs in the environment and evaluate its potential human health risks by TCDD. Here, we established a transgenic zebrafish model with a red fluorescent reporter gene ( mCherry) using the truncated cyp1a promoter. Under exposure to TCDD, the expression pattern of mCherry in the reporter zebrafish mirrored that of endogenous cyp1a mRNA, and the primary target tissues for TCDD were the brain vessels, liver, gut, cloaca, and skin. Our results indicated that exposure of the embryos to TCDD at concentrations as low as 0.005 nM for 48 h, which did not elicit morphologic abnormalities in the embryos, markedly increased mCherry expression. In addition, the reporter embryos responded to other POPs, and primary liver cell culture of zebrafish revealed that Cyp1a protein was mainly expressed in the cytoplasm of liver cells. Furthermore, our transgenic fish embryos demonstrated that TCDD exposure can regulate the expression levels of several tumor-related factors, including epidermal growth factor, TNF-α, C-myc, proliferating cell nuclear antigen, TGF-β, serine/threonine kinase (Akt), and phosphorylated Akt, suggesting that our transgenic fish can be used as a sensitive model to evaluate the carcinogenicity induced by TCDD exposure.-Luo, J.-J., Su, D.-S., Xie, S.-L., Liu, Y., Liu, P., Yang, X.-J., Pei D.-S. Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

Intracellular localization of pregnane X receptor in HepG2 cells cultured by the hanging drop method

Pregnane X receptor (PXR) is localized in the cytoplasm of liver cells, whereas it is localized in the nucleus of monolayer-cultured HepG2 cells. Since cultured cells are affected by the microenvironment in which they are grown, we studied the effect of three-dimensional (3D) culture on the localization of PXR in HepG2 cells using the hanging drop method. The results showed that PXR was retained in the cytoplasm of HepG2 cells and other human hepatocarcinoma cell lines (FLC5, FLC7 and Huh7) when they were cultured by the hanging drop method. Treatment with rifampicin, a ligand of PXR, translocated PXR from the cytoplasm to nucleus and increased expression levels of CYP3A4 mRNA in HepG2 cells cultured by the hanging drop method. These findings suggest that 3D culture is a key factor determining the intracellular localization of PXR in human hepatocarcinoma cells and that PXR that becomes retained in the cytoplasm of HepG2 cells with 3D culture has functions of nuclear translocation and regulation of target genes in response to human PXR ligands. Three-dimensionally cultured hepatocarcinoma cells would be a useful tool to evaluate induction potency of drug candidates and also to study mechanisms of nuclear translocation of PXR by human PXR ligands.

Abstract LB-139: PRMT6-dependent CRAF/ERK signaling regulates cancer stem cell plasticity in liver cancer

Abstract Hepatocellular carcinoma (HCC), the major type of liver cancer, remains one of the most prevalent and deadliest cancer types in the world. Contemporary challenge in treating HCC has been the common therapy resistance and recurrence after therapy, all of which have been reported to be associated with stem-like behavior of cancer cells. Our group has previously identified a functional liver cancer stem cell (CSC) subset marked by the CD133 cell surface phenotype. Utilizing a PCR array encompassing diverse human chromatin modifiers, protein arginine methyltransferase 6 (PRMT6) was found to be differentially down-regulated in CD133+ liver CSCs of human HCC cells as well as CD133 enriched chemoresistant hepatospheres as compared to their counterparts. Clinically, reduced PRMT6 expression was detected in HCC specimens and correlated with a higher risk of metastasis. PRMT6 negatively regulated diverse in vitro cancer stem cell properties of HCC cells including self-renewal, therapy resistance, metastasis and expression of CSC and pluripotency markers. In addition, PRMT6 also suppressed in vivo tumor initiation and serial transplantation. Surprising, contrary to its usual localization in the nucleus as a chromatin modification enzyme mediating histone H3R2 methylation, we found PRMT6 to be predominantly expressed in the cytoplasm in normal liver and HCC cells. Through tandem affinity purification and subsequent mass spectrometry profiling, we identified CRAF, a serine/threonine-protein kinase, as a novel cytoplasmic protein partner of PRMT6. Binding of PRMT6 to CRAF inhibited its kinase activity through site-specific arginine methylation, resulting in inhibition of ERK-mediated CSC plasticity in HCC, demonstrated through in vivo / in vitro methylation assays, kinase assay and functional rescue experiments with the ERK inhibitor U0126. The link between PRMT6, ERK and cancer stemness was further substantiated in primary human normal liver and HCC organoids with or without PRMT6 modulated. Taken together, we found PRMT6 to be down-regulated in the liver CSC subset and to be functionally involved in regulating liver CSC plasticity via an unprecedented role in the cytoplasm through suppression of CRAF/ERK cascade. Citation Format: Lok Hei Chan, Lei Zhou, Kai Yu Ng, Tin Lok Wong, Stella Chai, Terence K Lee, Xin Yuan Guan, Yick Pang Ching, Chung Mau Lo, Kwan Man, Benedetta Artegiani, Hans Clevers, Helen H Yan, Suet Yi Leung, Stèphane Richard, Michael SY Huen, Stephanie Ma. PRMT6-dependent CRAF/ERK signaling regulates cancer stem cell plasticity in liver cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr LB-139. doi:10.1158/1538-7445.AM2017-LB-139

Drug interaction s in the constituents of street drug mixture“Nyaope”in South Africa: a mini-review

Nyaope is a unique South African street drug mixture thought to contain illicit drugs and other compounds and is usually inhaled after wrapping in the Cannabis leaf. Despite its illegalization in March 2014, abuse of Nyaope is on the increase. While highly addictive, withdrawal symptoms are very severe, unbearable and drive the user to desperately seek for the next fix. Due to the lack of knowledge in its composition and how the constituents interact with each other, treatment for withdrawal symptoms and rehabilitation has been a challenge. A mini-literature review was done to explore how the major constituents of Nyaope relate to each other in their actions and in their path of breaking down (metabolism). The literature suggests that the inside opiate group, in between opiates and benzodiazepines, in between opiates and cannabis group, in between benzodiazepines and phenobarbitals, and also amongst the minor constituents, there are extensively shared the metabolic pathways which lead to longer plasma half-life in each of these drugs and thus synergistic effects. These shared pathways are via the cytochrome P450 family of enzymes in the liver cell cytoplasm and these enzyme actions are to inactivate or detoxify the drugs or convert them to more water soluble compounds in order to excrete them through the kidneys. Not only sharing the metabolic pathways, but also the actions of these drugs at certain receptors in the brain have either opposing or stimulating effects on one another, making the complex nature of their combined actions. Such findings can explain the unique withdrawal symptom complex of Nyaope, which is important for the clinicians and public health workers who are dealing with the users. Understanding the biochemical and metabolic basis of Nyaope drug interactions provides valuable insight towards the development of withdrawal signs and symptoms which may contribute to the targeted treatment program. Keywords: Nyaope, drugs of abuse, withdrawal symptoms, drug synergy, metabolism

Mechanism of hepatotoxicity due to black cohosh (Cimicifuga racemosa): Histological, immunohistochemical and electron microscopy analysis of two liver biopsies with clinical correlation

BackgroundConsumption of herbal supplements in the developed world remains high. Cimicifuga racemosa (C. racemosa) extract, or black cohosh, is widely used as a hormone replacing and an anti-inflammatory agent, and has been shown to cause idiosyncratic hepatitis. The mechanism of acute liver injury in those cases is unclear. To date, hepatotoxic effects of C. racemosa have been studied mostly in vitro and in animal models. Data on human tissue is extremely limited, and mostly confined to histological findings of explanted livers. MethodsWe evaluated clinical data and examined surgical diagnostic liver biopsy specimens obtained from two female patients, who developed acute submassive liver necrosis, following consumption of C. racemosa. Both patients presented with acute elevation of liver enzymes, cholestasis, absence of reactivity to hepatitis A, B and C antibodies, and weak non-specific positivity for autoimmune serological markers. Initial histological interpretation of the biopsies, with focus on hepatic parenchyma and portal tracts, was done by light microscopy, followed by special stain series and immunohistochemical studies, including Cam 5.2, AE1/AE3, reticulin, α-actin, sirius red, and PAS with diastase. Areas of prominent lymphocytic infiltration of the periportal liver plate, observed microscopically, were further evaluated by electron microscopy (EM). 4HNE adduction study, an immunofluorescent assay, was performed to detect products of the oxidative damage and their localization in the liver parenchyma. ResultsOxidative damage was evident by accumulation of 4HNE protein adducts in the cytoplasm of hepatocytes, secondary lysosomes and macrophages. We hypothesize that the adducted proteins, accumulated in the liver parenchyma, serve as autoantigens, which provoke an autoimmune response, and cause migration of lymphocytes to the affected regions. The formation of immunological synapses between hepatocytes and lymphocytes, predominantly T-lymphocytes, is demonstrated by electron microscopy. The autoimmune response induces piecemeal, or troxis necrosis of hepatocytes, a well described biological phenomenon, where lymphocytes gradually remove hepatocytes in a piecemeal fashion, slowly consuming them and leaving fragments of liver cells, or nubbins of anuclear cytoplasm of liver cell, at the interface between lymphocytes and hepatocytes. ConclusionThe pattern of pathological injury of liver cells in both patients, following consumption of black cohosh, is identical to troxis necrosis, seen during autoimmune hepatitis. Recognition of the possibility of the acute hepatic injury by the herbal supplement black cohosh is essential for early accurate diagnosis, and timely patient management.

Differential localization of transferrin receptors and lipocalin-2 in rat hepatic and extra-hepatic organs during acute phase response

The transferrin receptors (TfR1, TfR2) and lipocalin-2 (LCN-2) are mainly involved in iron transport. We previously reported increasing amounts of liver cytoplasmic and nuclear iron content during acute phase response (ARP). Aim of the present study is to investigate intracellular localization of transferrin receptors and lipocalin-2 in liver compared to extra-hepatic organs under physiological and acute phase conditions. Immunohistochemistry was performed on 5 µm thick sections of hepatic and extra-hepatic tissues. Liver, spleen and heart cytoplasmic and nuclear proteins were used for Western blot analysis. By means of immunohistology TfR1 was detected in membranes and cytoplasm of liver and spleen cells while TfR2 showed nuclear expression mostly. In heart TfR1 and TfR2 both showed nuclear expression along with membranous and cytoplasmic localization, colocalization of TfR1 and TfR2 was also observed in spleen and heart. LCN-2 immuno-reactivity was detected in neutrophils and hepatocytes both. However, LCN-2 was detected in macrophages mostly, in spleen. Colocalization of LCN-2 with both transferrin receptors (TfR1, TfR2) was detected in hepatic and extra-hepatic tissues. The data presented demonstrate the differential and colocalization of transferrin receptors and LCN-2 in liver, spleen and heart. Western blot analysis of cytoplasmic and nuclear proteins from hepatic and extra-hepatic organs confirmed the immunihistology findings. Differential localization of iron transport proteins in different organs suggest their diverse role in these organs under physiological and pathological conditions. Colocalization of LCN2 with transferrin receptors clearly demonstrates its role in iron transport during acute phase conditions and presence of TfR1, TfR2 in cell nuclei demonstrate transport of iron to the cell nuclei needed for the integrity of life i.e. DNA repair, DNA replication and RNA metabolism.

The regulation mechanism of protein kinase Cδ on arsenic liver injury caused by coal-burning

To investigate the effects of mRNA transcriptional and protein expressions of protein kinase Cδ (PKCδ) on the development of arsenic liver injury caused by coal-burning. Population study:133 arsenic exposures were selected as arsenic exposure groups including the ward non-patient group (25 cases) , no obvious hepatopathy group (38 cases) , mild (43 cases) and moderate to severe hepatopathy group (27 cases) from the area with endemic arsenism in Guizhou province. Another 34 healthy residents were selected as the control group in non-arsenic pollution village. The urine and peripheral blood were collected from the subjects. The arsenic contents in urine and mRNA expressions of PKCδ in peripheral blood were detected. Animal experiment study:thirty wistar rats were randomly by random number table divided into control group, drinking water arsenic poisoning group and coal-burning arsenic poisoning group (i.e., low, medium and high arsenic contaminated grain group) by random number table method, including 6 rats in each group. The control group was fed normally for 3 months, drinking water arsenic poisoning group and coal-burning arsenic poisoning groups were fed respectively with 10 mg/kg As2O3 solution and different concentrations (25, 50 and 100 mg/kg) of arsenic-containing feed which was persisted 3 months. The arsenic contents in urine, mRNA expression levels of PKCδ in peripheral blood and liver tissue and the protein expression levels of phosphorylated protein kinase Cδ(pPKCδ) in liver tissue were detected. The median(quartile) of arsenic contents in urine were 25.58 (18.62-40.73), 56.66 (38.93-76.77), 64.90 (39.55- 98.37) and 75.47 (41.30-109.70) µg/g Cr respectively for the non-patient group, no obvious hepatopathy group, mild and moderate to severe hepatopathy group. The levels were higher than that in the control group (23.34 (17.84-37.45) µg/g Cr) (P < 0.05), except for the ward non-patient group. The arsenic contents in rat urine were 2223.61 (472.98-3976.73), 701.16 (194.01-1300.27), 1060.94 (246.33-2585.47) and 3101.11 (1919.97-5407.07) µg/g Cr, respectively for the drinking water arsenic poisoning group, the low, medium and high dosage arsenic grain contamination groups, all higher than that in the control group (94.32 (22.65-195.25) µg/g Cr) (P < 0.05) . The protein expressions of pPKCδ in liver tissue were 324.83 ± 25.06, 278.50 ± 30.57, 308.83 ± 34.67 and 326.33 ± 35.09, which were significantly higher than that in the control group (240.17 ± 28.07) (P < 0.05) . The protein expression levels of pPKCδ in liver cell membrane were 0.49 ± 0.06,0.33 ± 0.05,0.37 ± 0.06 and 0.50 ± 0.08, which were significantly higher than that in the control group (0.28 ± 0.04) (P < 0.05) . The protein expression levels of pPKCδ in liver cell cytoplasm were 0.38 ± 0.06,0.31 ± 0.05, 0.35 ± 0.05 and 0.36 ± 0.05, which were significantly higher than that in the control group (0.24 ± 0.05) (P < 0.05). The arsenic may regulate protein expressions of pPKCδ and induce its membrane translocation, and cause the development of arsenic liver injury caused by coal-burning.

Leishmania Dices Away Cholesterol for Survival

Host lipid alterations are centrally involved in Leishmania donovani infection, and infected patients exhibit hypocholesterolemia. In this issue of Cell Host & Microbe, Ghosh etal. (2013) show that the metalloprotease GP63 released by L.donovani in the liver cleaves DICER1, inhibiting miR-122 maturation, which regulates cholesterol metabolism. These events decrease serum cholesterol and promote parasite growth.

Effects of bone morphogenetic protein-7 therapy on E3 ubiquitin ligase expression in mouse liver with experimentally induced fibrosis

This study explored the dynamic expression of the E3 ubiquitin-protein ligase gene, Arkadia, in response to carbon tetrachloride (CCl4)-induced liver fibrosis in a mouse model and investigated the differential expression that occurs following treatment with the anti-fibrotic bone morphogenetic protein-7 (BMP-7). Thirty healthy male imprinting control region (ICR) mice were randomly assigned to three groups: normal (control; n = 6), CCl4-induced model group (model; n = 18), and CCl4-induced model with BMP-7 treatment group (treatment; n = 6). The model group was further divided into three subgroups (n = 6 each) for analysis at 4, 8 and 12 weeks after fibrosis induction. Liver fibrosis was induced by hypodermic injections of 60% CCl4 /peanut oil (5 mL/kg) to the hind legs of mice two-times per week in alternating legs for a period of 12 weeks. At week 9, the treatment group of CCl4-induced mice were given an intraperitoneal injection of BMP-7 (300 pg/g) simultaneously with that day's hypodermic injection of 60% CCl4 /peanut oil, and then every other day for a period of four weeks. The pathological changes in liver tissues were observed after staining with hematoxylin-eosin (HE) and Masson's trichrome. Messenger RNA (mRNA) and protein expression of Arkadia in liver were evaluated using reverse transcription-polymerase chain reaction and immunohistochemistry and Western blotting, respectively. Mouse models of liver fibrosis were successfully established by CCl4 exposure. Arkadia, Smad7 and TGF-beta1 mRNA levels were up-regulated in the model group in a time-dependent manner (vs. control group), and BMP-7 treatment led to significant down-regulation of the CCl4-induced expression of the three genes (vs. control group: F = 812.80, 451.46, and 998.96, respectively; P less than 0.01). At week 12, the mRNA levels of Arkadia, Smad7, and TGF-b1 were significantly lower in the BMP-7 treatment group than in the model group (t = 12.108, 18.737, and 16.364, respectively; P less than 0.01). Arkadia, Smad7, and TGF-b1 protein staining was weak in the portal area of control liver tissue. In contrast, the model group showed significantly stronger staining for all three proteins in the portal area and in the cytoplasm of liver cells. The staining of Arkadia, Smad7, and TGF-b1 proteins was significantly lower in the treatment group (vs. control group: F = 8.399, 609.690, and 900.561, respectively; P < 0.01). At week 12, the protein levels of Arkadia, Smad7, and TGF-b1 were significantly lower in the treatment group than in the model group (t = 23.438, 11.667, and 42.889, respectively; P < 0.01). Arkadia expression gradually increased along with the development of liver fibrosis but was suppressed by treatment with the anti-fibrotic factor, BMP-7.

Activity of nucleoside diphosphate kinase α (NDPK α) capable of binding to outer mitochondrial membrane accounts for less than 10% of total NDPK activity present in cytoplasm of liver cells

During incubation of a constant volume of rat liver cytosol with an increasing quantity of mitochondrial protein in the presence of 3.3 mM MgCl(2), the binding of nucleoside diphosphate kinase (NDPK) from the cytosol to mitochondrial membranes is described by a saturation curve. The highest bound NDPK activity accounts for less than 9% of the added activity. Analysis of the results suggests that only one NDPK isozyme is bound to the membranes. Western blotting showed it to be NDPK α, a homolog of human NDPK-B. Substrates of NDPK, hexokinase, and glycerol kinase, as well as N,N'-dicyclohexylcarbodiimide and palmitate, did not influence the association of NDPK with mitochondrial membranes. We conclude that the sites of NDPK binding to the outer mitochondrial membrane are not identical to those of hexokinase and glycerol kinase.