Background & Aims Hepatocyte cellular dysfunction and death induced by lipids and macrophage-associated inflammation are characteristics of nonalcoholic steatohepatitis (NASH). The fatty acid ...palmitate can activate death receptor 5 (DR5) on hepatocytes, leading to their death, but little is known about how this process contributes to macrophage-associated inflammation. We investigated whether lipid-induced DR5 signaling results in the release of extracellular vesicles (EVs) from hepatocytes, and whether these can induce an inflammatory macrophage phenotype. Methods Primary mouse and human hepatocytes and Huh7 cells were incubated with palmitate, its metabolite lysophosphatidylcholine, or diluent (control). The released EV were isolated, characterized, quantified, and applied to macrophages. C57BL/6 mice were placed on chow or a diet high in fat, fructose, and cholesterol to induce NASH. Some mice also were given the ROCK1 inhibitor fasudil; 2 weeks later, serum EVs were isolated and characterized by immunoblot and nanoparticle-tracking analyses. Livers were collected and analyzed by histology, immunohistochemistry, and quantitative polymerase chain reaction. Results Incubation of primary hepatocytes and Huh7 cells with palmitate or lysophosphatidylcholine increased their release of EVs, compared with control cells. This release was reduced by inactivating mediators of the DR5 signaling pathway or rho-associated, coiled-coil-containing protein kinase 1 (ROCK1) inhibition. Hepatocyte-derived EVs contained tumor necrosis factor-related apoptosis-inducing ligand and induced expression of interleukin 1β and interleukin 6 messenger RNAs in mouse bone marrow–derived macrophages. Activation of macrophages required DR5 and receptor-interacting protein kinase 1. Administration of the ROCK1 inhibitor fasudil to mice with NASH reduced serum levels of EVs; this reduction was associated with decreased liver injury, inflammation, and fibrosis. Conclusions Lipids, which stimulate DR5, induce release of hepatocyte EVs, which activate an inflammatory phenotype in macrophages. Strategies to inhibit ROCK1-dependent release of EVs by hepatocytes might be developed for the treatment of patients with NASH.
COVID 19 is a novel pandemic affecting globally. Although no reliable data suggests that patients of well controlled Type 1 Diabetes Mellitus (T1DM) being at increased risk of becoming severely ill ...with SARS-CoV2, but lockdown may impact patients with T1DM requiring regular medications and follow up. Hence this study was planned to see the impact of lockdown on glycemic control in patients with T1DM.
A cross sectional study was done in T1DM patients in whom a structured questionnaire was administered on follow up within 15 days after lockdown. Data regarding hypoglycemic and hyperglycemic episodes, Diabetic ketoacidosis (DKA), insulin dose missed, regular glucose monitoring, dietary compliance, physical activity, hospitalization during the phase of lockdown was taken. Average blood glucose and HbA1C of lockdown phase was compared with the readings of prelockdown phase.
Out of 52 patients, 36.5% had hyperglycemic and 15.3% had hypoglycemic episodes. Insulin dose was missed in 26.9%, glucose monitoring not done routinely in 36.5% and 17.4% were not diet compliant during lockdown. Average blood glucose during lockdown phase was 276.9 ± 64.7 mg/dl as compared to 212.3 ± 57.9 mg/dl during prelockdown phase. Mean HbA1c value of lockdown (10 ± 1.5%) which was much higher that of pre lockdown (8.8 ± 1.3%) and the difference was statistically significant (p < 0.05).
Glycemic control of T1DM patients has worsened mainly due to non availability of insulin/glucostrips during lockdown period. There is a need for preparedness in future so that complications can be minimised.
•Lockdown in COVID 19 may impact patients with Type 1 Diabetes Mellitus (T1DM) requiring regular medications and follow up.•Glycemic control of T1DM patients has worsened during lockdown period.•Main reasons were non availability of insulin / glucostrips, poor dietary compliance and less physical activity in lockdown.
Exosomes are cell-derived extracellular vesicles thought to promote intercellular communication by delivering specific content to target cells. The aim of this study was to determine whether ...endothelial cell (EC)-derived exosomes could regulate the phenotype of hepatic stellate cells (HSCs). Initial microarray studies showed that fibroblast growth factor 2 induced a 2.4-fold increase in mRNA levels of sphingosine kinase 1 (SK1). Exosomes derived from an SK1-overexpressing EC line increased HSC migration 3.2-fold. Migration was not conferred by the dominant negative SK1 exosome. Incubation of HSCs with exosomes was also associated with an 8.3-fold increase in phosphorylation of AKT and 2.5-fold increase in migration. Exosomes were found to express the matrix protein and integrin ligand fibronectin (FN) by Western blot analysis and transmission electron microscopy. Blockade of the FN-integrin interaction with a CD29 neutralizing antibody or the RGD peptide attenuated exosome-induced HSC AKT phosphorylation and migration. Inhibition of endocytosis with transfection of dynamin siRNA, the dominant negative dynamin GTPase construct Dyn2K44A, or the pharmacological inhibitor Dynasore significantly attenuated exosome-induced AKT phosphorylation. SK1 levels were increased in serum exosomes derived from mice with experimental liver fibrosis, and SK1 mRNA levels were up-regulated 2.5-fold in human liver cirrhosis patient samples. Finally, S1PR2 inhibition protected mice from CCl4-induced liver fibrosis. Therefore, EC-derived SK1-containing exosomes regulate HSC signaling and migration through FN-integrin-dependent exosome adherence and dynamin-dependent exosome internalization. These findings advance our understanding of EC/HSC cross-talk and identify exosomes as a potential target to attenuate pathobiology signals.
Liver fibrosis is characterized by the activation and migration of hepatic stellate cells (HSCs), followed by matrix deposition. Recently, several studies have shown the importance of extracellular ...vesicles (EVs) derived from liver cells, such as hepatocytes and endothelial cells, in liver pathobiology. While most of the studies describe how liver cells modulate HSC behavior, an important gap exists in the understanding of HSC‐derived signals and more specifically HSC‐derived EVs in liver fibrosis. Here, we investigated the molecules released through HSC‐derived EVs, the mechanism of their release, and the role of these EVs in fibrosis. Mass spectrometric analysis showed that platelet‐derived growth factor (PDGF) receptor‐alpha (PDGFRα) was enriched in EVs derived from PDGF‐BB‐treated HSCs. Moreover, patients with liver fibrosis had increased PDGFRα levels in serum EVs compared to healthy individuals. Mechanistically, in vitro tyrosine720‐to‐phenylalanine mutation on the PDGFRα sequence abolished enrichment of PDGFRα in EVs and redirected the receptor toward degradation. Congruently, the inhibition of Src homology 2 domain tyrosine phosphatase 2, the regulatory binding partner of phosphorylated tyrosine720, also inhibited PDGFRα enrichment in EVs. EVs derived from PDGFRα‐overexpressing cells promoted in vitro HSC migration and in vivo liver fibrosis. Finally, administration of Src homology 2 domain tyrosine phosphatase 2inhibitor, SHP099, to carbon tetrachloride–administered mice inhibited PDGFRα enrichment in serum EVs and reduced liver fibrosis. Conclusion: PDGFRα is enriched in EVs derived from PDGF‐BB‐treated HSCs in an Src homology 2 domain tyrosine phosphatase 2–dependent manner and these PDGFRα‐enriched EVs participate in development of liver fibrosis. (Hepatology 2018;68:333‐348).
Extracellular vesicles (EVs) are nanometer‐sized, membrane‐bound vesicles released by cells into the extracellular milieu. EVs are now recognized to play a critical role in cell‐to‐cell ...communication. EVs contain important cargo in the form of proteins, lipids, and nucleic acids and serve as vectors for delivering this cargo from donor to acceptor or target cell. EVs are released under both physiologic and pathologic conditions, including liver diseases, and exert a wide range of effects on target cells. This review provides an overview on EV biogenesis, secretion, cargo, and target cell interactions in the context of select liver diseases. Specifically, the diverse roles of EVs in nonalcoholic steatohepatitis, alcoholic liver disease, viral hepatitis, cholangiopathies, and hepatobiliary malignancies are emphasized. Liver diseases often result in an increased release of EVs and/or in different cargo sorting into these EVs. Either of these alterations can drive disease pathogenesis. Given this fact, EVs represent a potential target for therapeutic intervention in liver disorders. Because altered EV composition may reflect the underlying disease condition, circulating EVs can be exploited for diagnostic and prognostic purposes as a liquid biopsy. Furthermore, ex vivo modified or synthesized EVs can be engineered as therapeutic nano‐shuttles. Finally, we highlight areas that merit further investigation relevant to understanding how EVs regulate liver disease pathogenesis. (Hepatology 2016;64:2219‐2233).
The efficacy and durability studies of the dimensionally stable anode (DSA) have been carried out for the treatment of human urine metabolite, creatinine using electro-oxidation process. The ...influence of various input parameters such as current density, treatment time, pH and concentration of electrolyte on responses like % degradation and energy consumption were explained using Box-Behnken design. 85.41% pollutant degradation along with complete mineralization was achieved at optimum conditions like pH-(2.4), current density-(10.29mAcm−2), NaCl concentration-(0.75gL−1) and treatment time-(85min). Interestingly, a treatment time of was significantly lower than the reported values with similar kind of advanced technologies. Further reduction in treatment time i.e. 60min was achieved by incorporating synergistic effect through Photoelectrocatalysis for the same %degradation. The electrodes used were durable enough even after forty-five cycles as confirmed through SEM/EDS and Raman Spectra. Further, mineralization studies were carried out by measuring the reduction in COD (81.25%) and TOC (84.37%) along with generation of various ions such as nitrite, nitrate and ammonium ion under the optimized conditions. Based upon the detected intermediates analyzed through FT-IR and LC-MS, a degradation mechanism for creatinine has been proposed.
•Electro-oxidation process was used for the treatment of the creatinine.•Pt-Ru-Ir-Ti quaternary DSA anode was used for the present study.•Response surface methodology was used for the optimization of process parameters.•Durability of electrodes was confirmed through SEM/EDS and Raman Spectra.•Degradation pathway has been proposed through LC-MS studies.
Background and Aims
Interleukin‐22 has beneficial effects on inflammation and impaired hepatic regeneration that characterize alcohol‐associated hepatitis (AH). F‐652 is a recombinant fusion protein ...of human interleukin‐22 and immunoglobulin G2 fragment crystallizable. This study aims to assess the safety and efficacy signals of F‐652 in patients with moderate and severe AH.
Approach and Results
A phase‐2 dose‐escalating study was carried out. F‐652 (10 μg/kg, 30 μg/kg, or 45 μg/kg) administered on days 1 and 7 was tested in 3 patients each with moderate (Model for End‐Stage Liver Disease MELD scores: 11‐20) and severe AH (MELD scores: 21‐28). Safety was defined by absence of serious adverse events and efficacy was assessed by Lille score, changes in MELD score, and serum bilirubin and aminotransferases at days 28 and 42. Three independent propensity‐matched comparator patient cohorts were used. Plasma extracellular vesicles and multiplex serum cytokines were measured to assess inflammation and hepatic regeneration. Eighteen patients (9 moderate and 9 severe AH) were enrolled, 66% were male, and the mean age was 48 years. The half‐life of F‐652 following the first dose was 61‐85 hours. There were no serious adverse events leading to discontinuation. The MELD score and serum aminotransferases decreased significantly at days 28 and 42 from baseline (P < 0.05). Day‐7 Lille score was 0.45 or less in 83% patients as compared with 6%, 12%, and 56% among the comparator cohorts. Extracellular vesicle counts decreased significantly at day 28 (P < 0.013). Cytokine inflammatory markers were down‐regulated, and regeneration markers were up‐regulated at days 28 and 42.
Conclusions
F‐652 is safe in doses up to 45 μg/kg and associated with a high rate of improvement as determined by Lille and MELD scores, reductions in markers of inflammation and increases in markers of hepatic regeneration. This study supports the need for randomized placebo‐controlled trials to test the efficacy of F‐652 in AH.
The efficacy of electro-oxidation has been checked for the deterioration of synthetic urine (SU) using mixed metal oxide anode along with the potential of harnessing the commercially useful byproduct ...i.e. molecular hydrogen gas. The results from batch have been used to execute the scale-up studies for the continuous electro-oxidation treatment of SU in a photovoltaic driven reactor. The effect of different operational variables like pH, time, current density and N/Cl ratio on process efficiency was evaluated in terms of %COD removal and specific energy consumption using response surface methodology. The results showed that 87.25% removal in COD and 85.88% in TOC were achieved in 8.8 h. The complete deactivation of E. coli spiked synthetic urine wastewater was achieved in 45 min only. The main strength lies in the demonstration of the significant reduction in treatment time to 6 h by incorporating dual effect i.e. Photo-electrocatalysis. The anode used was proven to be stable and effective even after 100 recycles (207.5 h). The intermediates formed during the treatment process were analyzed through LC-MS. The techno-economic analysis for the proposed technology under optimized conditions was calculated to be 0.85 $/kg of COD removed.
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•Quaternary MMO was used for the electrochemical oxidation/disinfection of synthetic urine.•BBD under response surface methodology was used for the parametric optimization.•Stability of anodes after 100 recycles was confirmed through XRD and SEM/EDS.•Photo-electrocatalysis was performed for assessing the synergistic effect.•The potential harnessing of molecular hydrogen generation in a photovoltaic driven electrolytic reactor.
Liver microenvironment is a critical determinant for development and progression of liver metastasis. Under transforming growth factor beta (TGF‐β) stimulation, hepatic stellate cells (HSCs), which ...are liver‐specific pericytes, transdifferentiate into tumor‐associated myofibroblasts that promote tumor implantation (TI) and growth in the liver. However, the regulation of this HSC activation process remains poorly understood. In this study, we tested whether vasodilator‐stimulated phosphoprotein (VASP) of HSCs regulated the TGF‐β‐mediated HSC activation process and tumor growth. In both an experimental liver metastasis mouse model and cancer patients, colorectal cancer cells reaching liver sinusoids induced up‐regulation of VASP and alpha‐smooth muscle actin (α‐SMA) in adjacent HSCs. VASP knockdown in HSCs inhibited TGF‐β‐mediated myofibroblastic activation of HSCs, TI, and growth in mice. Mechanistically, VASP formed protein complexes with TGF‐β receptor II (TβRII) and Rab11, a Ras‐like small GTPase and key regulator of recycling endosomes. VASP knockdown impaired Rab11 activity and Rab11‐dependent targeting of TβRII to the plasma membrane, thereby desensitizing HSCs to TGF‐β1 stimulation. Conclusions: Our study demonstrates a requirement of VASP for TGF‐β‐mediated HSC activation in the tumor microenvironment by regulating Rab11‐dependent recycling of TβRII to the plasma membrane. VASP and its effector, Rab11, in the tumor microenvironment thus present therapeutic targets for reducing TI and metastatic growth in the liver. (Hepatology 2015;61:361–374)
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