Biallelic loss-of-function ZFYVE19 mutations are associated with congenital hepatic fibrosis, sclerosing cholangiopathy and high-GGT cholestasis

2020 ◽  
pp. jmedgenet-2019-106706
Author(s):  
Weisha Luan ◽  
Chen-Zhi Hao ◽  
Jia-Qi Li ◽  
Qing Wei ◽  
Jing-Yu Gong ◽  
...  
Keyword(s):  
Hepatic Fibrosis ◽  
Intrahepatic Cholestasis ◽  
Cultured Cells ◽  
High Serum ◽  
Gamma Glutamyl Transferase ◽  
Congenital Hepatic Fibrosis ◽  
Loss Of Function ◽  
Glutamyl Transferase

BackgroundFor many children with intrahepatic cholestasis and high-serum gamma-glutamyl transferase (GGT) activity, a genetic aetiology of hepatobiliary disease remains undefined. We sought to identify novel genes mutated in children with idiopathic high-GGT intrahepatic cholestasis, with clinical, histopathological and functional correlations.MethodsWe assembled a cohort of 25 children with undiagnosed high-GGT cholestasis and without clinical features of biliary-tract infection or radiological features of choledochal malformation, sclerosing cholangitis or cholelithiasis. Mutations were identified through whole-exome sequencing and targeted Sanger sequencing. We reviewed histopathological findings and assessed phenotypical effects of ZFYVE19 deficiency in cultured cells by immunofluorescence microscopy.ResultsNine Han Chinese children harboured biallelic, predictedly complete loss-of-function pathogenic mutations in ZFYVE19 (c.314C>G, p.S105X; c.379C>T, p.Q127X; c.514C>T, p.R172X; c.547C>T, p.R183X; c.226A>G, p.M76V). All had portal hypertension and, at liver biopsy, histopathological features of the ductal plate malformation (DPM)/congenital hepatic fibrosis (CHF). Four children required liver transplantation for recurrent gastrointestinal haemorrhage. DPM/CHF was confirmed at hepatectomy, with sclerosing small-duct cholangitis. Immunostaining for two primary-cilium axonemal proteins found expression that was deficient intraluminally and ectopic within cholangiocyte cytoplasm. ZFYVE19 depletion in cultured cells yielded abnormalities of centriole and axoneme.ConclusionBiallelic ZFYVE19 mutations can lead to high-GGT cholestasis and DPM/CHF in vivo. In vitro, they can lead to centriolar and axonemal abnormalities. These observations indicate that mutation in ZFYVE19 results, through as yet undefined mechanisms, in a ciliopathy.

scholarly journals Use of GP73 in the diagnosis of non-alcoholic steatohepatitis and the staging of hepatic fibrosis

2021 ◽  
Vol 49 (11) ◽  
pp. 030006052110553
Author(s):  
Yadi Li ◽  
Yan Yang ◽  
Yufang Li ◽  
Ping Zhang ◽  
Gaiying Ge ◽  
...  
Keyword(s):  
Hepatic Fibrosis ◽  
High Serum ◽  
Gamma Glutamyl Transferase ◽  
Strong Positive Correlation ◽  
Characteristic Analysis ◽  
Alcoholic Fatty Liver ◽  
Alcoholic Steatohepatitis ◽  
Golgi Protein ◽  
Glutamyl Transferase ◽  
Nafld Activity Score

Objective To evaluate the utility of Golgi protein 73 (GP73) in the diagnosis of non-alcoholic steatohepatitis (NASH) and hepatic fibrosis (HF) staging. Methods Ninety-one patients with non-alcoholic fatty liver disease (NAFLD) were allocated to NAFL (n = 46) and NASH (n = 45) groups according to their NAFLD activity score (NAS), and there were 30 healthy controls. Serum GP73 was measured by ELISA, GP73 protein expression was evaluated using immunohistochemistry, and FibroScan was used to determine liver hardness. Results The serum GP73 concentrations of the NAFL and NASH groups were significantly higher than those of controls. GP73 expression in the liver of the patients gradually progressed from absent or low to moderate or high. Serum GP73 positively correlated with liver expression, and the serum and liver GP73 of the patients positively correlated with FibroScan value and HF stage. There was a strong positive correlation of the combination of alanine aminotransferase, gamma glutamyl transferase and GP73 with NASH. The combination of serum GP73 and FibroScan value was found to predict NASH (NAS > 4) and advanced HF (stage ≥2) in patients with NAFLD using receiver operating characteristic analysis. Conclusion Serum GP73 may be useful in the diagnosis of NASH and the staging of HF.

scholarly journals Flavonoids content of 70% methanolic extract of Erucaria pinnata (Viv.) and its effect as anticancer and antiangiognic: in vitro, in vivo and docking study

2020 ◽  
Vol 11 (1) ◽  
pp. 25-38
Author(s):  
Nadia I. Zakhary ◽  
Emad E.H. El Gemeie ◽  
Adel K. Youssef ◽  
Marwa Abdel-salam Ibrahim Metwaly
Keyword(s):  
Cytotoxic Activity ◽  
Methanolic Extract ◽  
Coastal Region ◽  
Experimental Period ◽  
Docking Study ◽  
Gamma Glutamyl Transferase ◽  
Histopathological Studies ◽  
Glutamyl Transferase

Erucaria pinnata (Viv.) is a wild annual plant growing in North-Western Coastal Region in Egypt. This study reports for the first time the cytotoxic activity of different extracts of Erucaria pinnata plant against HEP-G2 cell line. The 70% methanolic extract (E1) recorded the best potent cytotoxic activity (IC50=13.6 µg/ml), so we analysis the flavonoids constituent of this extract using HPLC, which show that our extract is rich with important flavonoids compounds (rutin, quercetin, leutolin, etc.). We evaluated its antitumor activity against hepatocellular carcinoma (HCC) induced by diethylnitrosamine (DEN) (200mg/Kg. b.wt., i.p, single dose) after two weeks, animals received carbontetrachloride (CCl4) (3ml/Kg. b.wt., SC, once a week for 6 weeks) and the experiment continued for 44 weeks in rats. After the experimental period, the administration of DEN/CCl4 showed significant increase in the levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), total bilirubin, gamma-glutamyl transferase (γGT) and significant decrease in the levels of total proteins and albumin content in the serum with reduction in the liver antioxidants, including superoxide dismutase (SOD) and catalase (CAT). This was accompanied by increases in serum specific tumor markers (AFP). The 70% methanolic plant extract (E1) was orally administrated (400mg/kg/day respectively) for the whole study period, and it showed a significant improvement at the different biological liver functions, remodeled the antioxidant enzymes activity and down-regulated the serum AFP. All these findings were confirmed by histopathological studies of the liver samples obtained from all groups. In addition, we evaluated its antiangeogenic activity by docking study against VEGFR-2 tyrosine kinase after it showed an ability to inhibit the VEGFR-2 expression in vitro and inhibit the concentration of VEGF-A in vivo. The hepatoprotective effect of our extract was attributed to its antioxidant and antiangeogenic activity.

scholarly journals GALNT2 regulates ANGPTL3 cleavage in cells and in vivo of mice

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Xuedan Li ◽  
Yiliang Zhang ◽  
Minzhu Zhang ◽  
Yan Wang
Keyword(s):  
Cultured Cells ◽  
Coiled Coil ◽  
Lipoprotein Metabolism ◽  
Endothelial Lipase ◽  
Loss Of Function ◽  
Proprotein Convertase ◽  
Primary Hepatocytes ◽  
Coiled Coil Domain

Abstract Angiopoietin-like protein 3 (ANGPTL3) is an important inhibitor of lipoprotein lipase and endothelial lipase that plays critical roles in lipoprotein metabolism. It specifically expresses in the liver and undergoes proprotein convertase-mediated cleavage during secretion, which generates an N-terminal coiled-coil domain and C-terminal fibrinogen-like domain that has been considered as the activation step for its function. Previous studies have reported that the polypeptide GalNAc-transferase GALNT2 mediates the O-glycosylation of the ANGPTL3 near the cleavage site, which inhibits the proprotein convertase (PC)-mediated cleavage in vitro and in cultured cells. However, loss-of-function mutation for GALNT2 has no effect on ANGPTL3 cleavage in human. Thus whether GALNT2 regulates the cleavage of ANGPTL3 in vivo is unclear. In present study, we systematically characterized the cleavage of Angptl3 in cultured cells and in vivo of mice. We found that endogenous Angptl3 is cleaved in primary hepatocytes and in vivo of mice, and this cleavage can be blocked by Galnt2 overexpression or PC inhibition. Moreover, suppressing galnt2 expression increases the cleavage of Angptl3 in mice dramatically. Thus, our results support the conclusion that Galnt2 is a key endogenous regulator for Angptl3 cleavage both in vitro and in vivo.

A cullin-RING ubiquitin ligase targets exogenous α-synuclein and inhibits Lewy body–like pathology

2019 ◽  
Vol 11 (495) ◽  
pp. eaau6722 ◽  
Author(s):  
Juan A. Gerez ◽  
Natalia C. Prymaczok ◽  
Edward Rockenstein ◽  
Uli S. Herrmann ◽  
Petra Schwarz ◽  
...  
Keyword(s):  
Lewy Body ◽  
Cultured Cells ◽  
Neuroblastoma Cell ◽  
Lewy Bodies ◽  
Neuroblastoma Cell Line ◽  
Loss Of Function ◽  
Intracerebral Administration ◽  
F Box Protein

Parkinson’s disease (PD) is a neurological disorder characterized by the progressive accumulation of neuronal α-synuclein (αSyn) inclusions called Lewy bodies. It is believed that Lewy bodies spread throughout the nervous system due to the cell-to-cell propagation of αSyn via cycles of secretion and uptake. Here, we investigated the internalization and intracellular accumulation of exogenous αSyn, two key steps of Lewy body pathogenesis, amplification and spreading. We found that stable αSyn fibrils substantially accumulate in different cell lines upon internalization, whereas αSyn monomers, oligomers, and dissociable fibrils do not. Our data indicate that the uptake-mediated accumulation of αSyn in a human-derived neuroblastoma cell line triggered an adaptive response that involved proteins linked to ubiquitin ligases of the S-phase kinase-associated protein 1 (SKP1), cullin-1 (Cul1), and F-box domain–containing protein (SCF) family. We found that SKP1, Cul1, and the F-box/LRR repeat protein 5 (FBXL5) colocalized and physically interacted with internalized αSyn in cultured cells. Moreover, the SCF containing the F-box protein FBXL5 (SCFFBXL5) catalyzed αSyn ubiquitination in reconstitution experiments in vitro using recombinant proteins and in cultured cells. In the human brain, SKP1 and Cul1 were recruited into Lewy bodies from brainstem and neocortex of patients with PD and related neurological disorders. In both transgenic and nontransgenic mice, intracerebral administration of exogenous αSyn fibrils triggered a Lewy body–like pathology, which was amplified by SKP1 or FBXL5 loss of function. Our data thus indicate that SCFFXBL5 regulates αSyn in vivo and that SCF ligases may constitute targets for the treatment of PD and other α-synucleinopathies.

Ultrastructural Correlations between Clonal Human Tumor Colonies and in Vivo Neoplasms

1982 ◽  
Vol 40 ◽  
pp. 210-211
Author(s):  
M.J. Murphy ◽  
R.R. Price ◽  
J.C. Sloman
Keyword(s):  
Cultured Cells ◽  
Human Tumor ◽  
Cell Type ◽  
Tumor Mass ◽  
Initial Cell ◽  
Cloning Assay ◽  
Human Tumor Cloning ◽  
The Relationship

The in vitro human tumor cloning assay originally described by Salmon and Hamburger has been applied recently to the investigation of differential anti-tumor drug sensitivities over a broad range of human neoplasms. A major problem in the acceptance of this technique has been the question of the relationship between the cultured cells and the original patient tumor, i.e., whether the colonies that develop derive from the neoplasm or from some other cell type within the initial cell population. A study of the ultrastructural morphology of the cultured cells vs. patient tumor has therefore been undertaken to resolve this question. Direct correlation was assured by division of a common tumor mass at surgical resection, one biopsy being fixed for TEM studies, the second being rapidly transported to the laboratory for culture.

Preparation of cultured astrocytes for x-ray microanalysis

1989 ◽  
Vol 47 ◽  
pp. 988-989
Author(s):  
N.K.R. Smith ◽  
K.E. Hunter ◽  
P. Mobley ◽  
L.P. Felpel
Keyword(s):  
Cultured Cells ◽  
Elemental Content ◽  
Elemental Distribution ◽  
Sprague Dawley ◽  
X Ray ◽  
Cultured Astrocytes ◽  
Freeze Dried ◽  
Ultimate Objective

Electron probe energy dispersive x-ray microanalysis (XRMA) offers a powerful tool for the determination of intracellular elemental content of biological tissue. However, preparation of the tissue specimen , particularly excitable central nervous system (CNS) tissue , for XRMA is rather difficult, as dissection of a sample from the intact organism frequently results in artefacts in elemental distribution. To circumvent the problems inherent in the in vivo preparation, we turned to an in vitro preparation of astrocytes grown in tissue culture. However, preparations of in vitro samples offer a new and unique set of problems. Generally, cultured cells, growing in monolayer, must be harvested by either mechanical or enzymatic procedures, resulting in variable degrees of damage to the cells and compromised intracel1ular elemental distribution. The ultimate objective is to process and analyze unperturbed cells. With the objective of sparing others from some of the same efforts, we are reporting the considerable difficulties we have encountered in attempting to prepare astrocytes for XRMA.Tissue cultures of astrocytes from newborn C57 mice or Sprague Dawley rats were prepared and cultured by standard techniques, usually in T25 flasks, except as noted differently on Cytodex beads or on gelatin. After different preparative procedures, all samples were frozen on brass pins in liquid propane, stored in liquid nitrogen, cryosectioned (0.1 μm), freeze dried, and microanalyzed as previously reported.

Recovery from LCDD-associated Severe Liver Cholestasis: a Case Report and Literature Review

2016 ◽  
Vol 25 (1) ◽  
pp. 99-103 ◽  
Author(s):  
Benoit Brilland ◽  
Johnny Sayegh ◽  
Anne Croue ◽  
Frank Bridoux ◽  
Jean-François Subra ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Light Chain ◽  
Intrahepatic Cholestasis ◽  
Relevant Literature ◽  
Gamma Glutamyl Transferase ◽  
Light Chain Deposition Disease ◽  
Deposition Disease ◽  
Uncommon Presentation ◽  
Light Chain Deposition ◽  
Glutamyl Transferase

Light chain deposition disease (LCDD) is a rare multisystemic disorder associated with plasma cell proliferation. It mainly affects the kidney, but liver and heart involvement may occur, sometimes mimicking the picture of systemic amyloidosis. Liver disease in LCDD is usually asymptomatic and exceptionally manifests with severe cholestatic hepatitis. We report the case of a 66-year-old female with κ-LCDD and cast nephropathy in the setting of symptomatic multiple myeloma who, after a first cycle of bortezomib-dexamethasone chemotherapy, developed severe and rapidly worsening intrahepatic cholestasis secondary to liver κ-light chain deposition. Intrahepatic cholestasis was attributed to LCDD on the basis of the liver histology and exclusion of possible diagnoses. Chemotherapy was maintained and resulted in progressive resolution of cholestasis. We report here an uncommon presentation of LCDD, with prominent liver involvement that fully recovered with bortezomib-based chemotherapy, and briefly review the relevant literature. Abbreviations: AKI: Acute kidney injury; ALP: alkaline phosphatase; ALT: alanine aminotransferase; AST: aspartate aminotransferase; CMV: Cytomegalovirus; EBV: Epstein–Barr virus; GGT: gamma-glutamyl transferase; HSV: Herpes simplex virus; LC: light chain; LCDD: Light chain deposition disease; MIDD: Monoclonal immunoglobulin deposition disease; MM: Multiple myeloma.

Interactions Between Dental Composite Resins and Saliva A comparative biochemical in vitro study

2019 ◽  
Vol 56 (3) ◽  
pp. 529-533
Author(s):  
Mihaela Pantea ◽  
Diana Andreea Ighigeanu ◽  
Alexandra Totan ◽  
Maria Greabu ◽  
Daniela Miricescu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
In Vitro Study ◽  
Composite Resins ◽  
Vitro Study ◽  
Dental Composite ◽  
Gamma Glutamyl Transferase ◽  
Specific Protocol ◽  
Dental Composite Resins ◽  
Glutamyl Transferase

This in vitro study analyses the biochemical interaction between saliva and three types of dental composite resins (a direct resin, an indirect resin and a dual-cure resin used for cementation of indirect dental restorations). The resin samples were obtained following a specific protocol and in line with the producers� recommendations; the resin samples were incubated with saliva samples collected from 19 healthy volunteers. The obtained results showed that the tested composite resins did not produce significant changes in oxidative stress parameters that were analysed (albumin, uric acid, GGT / gamma glutamyl transferase, OXSR-1 / oxidative stress responsive kinase 1) and do not influence the inflammatory salivary status reflected by the levels of IL-6 - an inflammatory marker.

scholarly journals TRIM27 interacts with Iκbα to promote the growth of human renal cancer cells through regulating the NF-κB pathway

BMC Cancer ◽  
2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Chengwu Xiao ◽  
Wei Zhang ◽  
Meimian Hua ◽  
Huan Chen ◽  
Bin Yang ◽  
...  
Keyword(s):  
Cell Lines ◽  
Prognostic Indicator ◽  
The Cancer Genome Atlas ◽  
Mrna Levels ◽  
Loss Of Function ◽  
Protein Levels ◽  
Kaplan Meier ◽  
Cancer Genome Atlas

Abstract Background The tripartite motif (TRIM) family proteins exhibit oncogenic roles in various cancers. The roles of TRIM27, a member of the TRIM super family, in renal cell carcinoma (RCC) remained unexplored. In the current study, we aimed to investigate the clinical impact and roles of TRIM27 in the development of RCC. Methods The mRNA levels of TRIM27 and Kaplan–Meier survival of RCC were analyzed from The Cancer Genome Atlas database. Real-time PCR and Western blotting were used to measure the mRNA and protein levels of TRIM27 both in vivo and in vitro. siRNA and TRIM27 were exogenously overexpressed in RCC cell lines to manipulate TRIM27 expression. Results We discovered that TRIM27 was elevated in RCC patients, and the expression of TRIM27 was closely correlated with poor prognosis. The loss of function and gain of function results illustrated that TRIM27 promotes cell proliferation and inhibits apoptosis in RCC cell lines. Furthermore, TRIM27 expression was positively associated with NF-κB expression in patients with RCC. Blocking the activity of NF-κB attenuated the TRIM27-mediated enhancement of proliferation and inhibition of apoptosis. TRIM27 directly interacted with Iκbα, an inhibitor of NF-κB, to promote its ubiquitination, and the inhibitory effects of TRIM27 on Iκbα led to NF-κB activation. Conclusions Our results suggest that TRIM27 exhibits an oncogenic role in RCC by regulating NF-κB signaling. TRIM27 serves as a specific prognostic indicator for RCC, and strategies targeting the suppression of TRIM27 function may shed light on future therapeutic approaches.

scholarly journals β-elemene alleviates airway stenosis via the ILK/Akt pathway modulated by MIR143HG sponging miR-1275

2021 ◽  
Vol 26 (1) ◽  
Author(s):  
Guoying Zhang ◽  
Cheng Xue ◽  
Yiming Zeng
Keyword(s):  
Cell Cycle ◽  
Cell Viability ◽  
Cell Model ◽  
Protective Efficacy ◽  
Rabbit Model ◽  
Akt Pathway ◽  
Loss Of Function ◽  
Airway Stenosis

Abstract Background We have previously found that β-elemene could inhibit the viability of airway granulation fibroblasts and prevent airway hyperplastic stenosis. This study aimed to elucidate the underlying mechanism and protective efficacy of β-elemene in vitro and in vivo. Methods Microarray and bioinformatic analysis were used to identify altered pathways related to cell viability in a β-elemene-treated primary cell model and to construct a β-elemene-altered ceRNA network modulating the target pathway. Loss of function and gain of function approaches were performed to examine the role of the ceRNA axis in β-elemene's regulation of the target pathway and cell viability. Additionally, in a β-elemene-treated rabbit model of airway stenosis, endoscopic and histological examinations were used to evaluate its therapeutic efficacy and further verify its mechanism of action. Results The hyperactive ILK/Akt pathway and dysregulated LncRNA-MIR143HG, which acted as a miR-1275 ceRNA to modulate ILK expression, were suppressed in β-elemene-treated airway granulation fibroblasts; β-elemene suppressed the ILK/Akt pathway via the MIR143HG/miR-1275/ILK axis. Additionally, the cell cycle and apoptotic phenotypes of granulation fibroblasts were altered, consistent with ILK/Akt pathway activity. In vivo application of β-elemene attenuated airway granulation hyperplasia and alleviated scar stricture, and histological detections suggested that β-elemene's effects on the MIR143HG/miR-1275/ILK axis and ILK/Akt pathway were in line with in vitro findings. Conclusions MIR143HG and ILK may act as ceRNA to sponge miR-1275. The MIR143HG/miR-1275/ILK axis mediates β-elemene-induced cell cycle arrest and apoptosis of airway granulation fibroblasts by modulating the ILK/Akt pathway, thereby inhibiting airway granulation proliferation and ultimately alleviating airway stenosis.

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