scholarly journals Mir-382 Promotes Differentiation of Rat Liver Progenitor Cell WB-F344 by Targeting Ezh2

2018 ◽  
Vol 48 (6) ◽  
pp. 2389-2398
Author(s):  
Yongxia Zheng ◽  
Jiansheng Zhou ◽  
Xuebo Li ◽  
Guangtao Xu ◽  
Mingliang Jin ◽  
...  
Keyword(s):  
Rat Liver ◽  
Progenitor Cell ◽  
Cell Model ◽  
Periodic Acid ◽  
Luciferase Reporter ◽  
Hepatic Differentiation ◽  
Periodic Acid Schiff ◽  
Functional Studies ◽  
Apo E ◽  
Liver Progenitor Cell

Background/Aims: Liver progenitor cells (LPCs) were considered as a promising hepatocyte source of cell therapy for liver disease due to their self-renewal and differentiation capacities, while little is known about the mechanism of LPC differentiate into hepatocytes. This study aims to explore the effect of miR-382, a member of Dlk1-Dio3 microRNA cluster, during hepatic differentiation from LPCs. Methods: In this study, we used rat liver progenitor cell WB-F344 as LPC cell model and HGF as inducer to simulate the process of LPCs hepatic differentiation, then microRNAs were quantified by qPCR. Next, WB-F344 cell was transfected with miR-382 mimics, then hepatocyte cell trait was characterized by multiple experiments, including that periodic acid schiff staining and cellular uptake and excretion of indocyanine green to evaluate the hepatocellular function, qPCR and Western Blotting analysis to detect the hepatocyte-specific markers (ALB, Ttr, Apo E and AFP) and transmission electron microscopy to observe the hepatocellular morphology. Moreover, Luciferase reporter assay was used to determine whether Ezh2 is the direct target of miR-382. Results: We found that miR-382 increased gradually and was inversely correlated with the potential target, Ezh2, during WB-F344 hepatic differentiation. In addition, functional studies indicated that miR-382 increased the level of hepatocyte-specific genes. Conclusions: This study demonstrates that miR-382 may be a novel regulator of LPCs differentiation by targeting Ezh2.

scholarly journals Spatial orientation of glycoproteins in membranes of rat liver rough microsomes. II. Transmembrane disposition and characterization of glycoproteins.

1978 ◽  
Vol 78 (3) ◽  
pp. 894-909 ◽  
Author(s):  
E Rodriguez Boulan ◽  
D D Sabatini ◽  
B N Pereyra ◽  
G Kreibich
Keyword(s):  
Rat Liver ◽  
Concanavalin A ◽  
Gel Electrophoresis ◽  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Microsomal Protein ◽  
Outer Face ◽  
Membrane Glycoproteins ◽  
Periodic Acid Schiff ◽  
Low Concentrations

Rat liver microsomal glycoproteins were purified by affinity chromatography on concanavalin A Sepharose columns from membrane and content fractions, separated from rough microsomes (RM) treated with low concentrations of deoxycholate (DOC). All periodic acid-Schiff (PAS)-positive glycoproteins of RM showed affinity for concanavalin A Sepharose; even after sodium dodecyl sulfate (SDS) acrylamide gel electrophoresis, most of the microsomal glycoproteins bound [125I]concanavalin A added to the gels, as detected by autoradiography. Two distinct sets of glycoproteins are present in the membrane and content fractions derived from RM. SDS acrylamide gel electrophoresis showed that RM membranes contain 15--20 glycoproteins (15--22% of the total microsomal protein) which range in apparent mol wt from 23,000 to 240,000 daltons. A smaller set of glycoproteins (five to seven polypeptides), with apparent mol wt between 60,000 and 200,000 daltons, was present in the microsomal content fraction. The disposition of the membrane glycoproteins with respect to the membrane plane was determined by selective iodination with the lactoperoxidase (LPO) technique. Intact RM were labeled on their outer face with 131I and, after opening of the vesicles with 0.05% DOC, in both faces with 125I. An analysis of iodination ratios for individual proteins separated electrophoretically showed that in most membrane glycoproteins, tyrosine residues are predominantly exposed on the luminal face of the vesicles, which is the same face on which the carbohydrate moieties are exposed. Several membrane glycoproteins are also exposed on the cytoplasmic surface and therefore have a transmembrane disposition. In this study, ribophorins I and II, two integral membrane proteins (mol wt 65,000 and 63,000) characteristic of RM, were found to be transmembrane glycoproteins. It is suggested that the transmembrane disposition of the ribophorins may be related to their possible role in ribosome binding and in the vectorial transfer of nascent polypeptides into the microsomal lumen.

scholarly journals FACTORS AFFECTING ENZYMATIC REMOVAL OF GLYCOGEN FROM TISSUE SECTIONS AND FROM ENTIRE RAT LIVER BLOCKS

1965 ◽  
Vol 13 (6) ◽  
pp. 488-497 ◽  
Author(s):  
RONALD D. PAEGLE
Keyword(s):  
Enzymatic Hydrolysis ◽  
Rat Liver ◽  
Sodium Phosphate ◽  
Periodic Acid ◽  
Distilled Water ◽  
Periodic Acid Schiff ◽  
Fixed Tissue ◽  
Factors Affecting ◽  
Tissue Sections ◽  
Different Types

The rates of enzymatic glycogen removal from two different types of rat liver preparations were determined histochemically, employing the periodic acid-Schiff method to visualize glycogen. The first preparation consisted of elongated liver blocks (approximately 2 x 2 x 10 mm) which were fixed, then exposed to the alpha amylases before dehydration and embedding. The second type of preparation consisted of 5-8 µ thick, routinely prepared sections. The glycogen was removed considerably slower from the formaldehyde fixed tissue blocks than from the sections. The rate of glycogen hydrolysis was enchanced in the liver blocks by brief periods of autolysis, by removal of lipids and by washing before exposure to the enzyme. Hydrolysis by amylase was slower when 0.18 M phosphate was used with the formaldehyde during fixation than when veronal acetate or 0.018 M phosphate buffers were used at the same pH 7.2-7.4. Many peripherally located cells retained glycogen even after incubation of the entire blocks for 24 hours in alpha amylase when the buffer during fixation was 0.18 M sodium Phosphate. The contaminating ribonuclease present in malt diastase preparations penetrated readily into the cells of the elongated liver blocks but the protease affected only the periphery during the 24 hour incubation. The differences in resistance to enzymatic hydrolysis were not confined to the tissue blocks. The type of buffer and fixative used for tissue fixation determined the subsequent susceptibility of glycogen to hydrolysis in sections also. The list is arranged in order of progressively decreasing resistance to enzymatic hydrolysis: 3.8% formaldehyde or 3% glutaraldehyde with 0.18 M phosphate, 3.8% formaldehyde with 0.018 M phosphate or 0.18 M veronal acetate, 3% glutaraldehyde with distilled water, 3.8% formaldehyde with 0.129 M veronal acetate or distilled water, and 70-95% ethanol. In addition, glycogen was removed slower from the peripheral zones of the sections than from the central portions.

scholarly journals Isolation and characterization of multiple forms of rat liver UDP-glucuronate glucuronosyltransferase

1986 ◽  
Vol 233 (3) ◽  
pp. 827-837 ◽  
Author(s):  
J Roy Chowdhury ◽  
N Roy Chowdhury ◽  
C N Falany ◽  
T R Tephly ◽  
I M Arias
Keyword(s):  
Rat Liver ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Wistar Rat ◽  
Transferase Activity ◽  
Periodic Acid Schiff ◽  
Isolation And Characterization ◽  
Udp Glucuronosyltransferase ◽  
Fraction I

UDP-glucuronosyltransferase (EC 2.4.1.17) activity was solubilized from male Wistar rat liver microsomal fraction in Emulgen 911, and six fractions with the transferase activity were separated by chromatofocusing on PBE 94 (pH 9.4 to 6.0). Fraction I was further separated into Isoforms Ia, Ib and Ic by affinity chromatography on UDP-hexanolamine-Sepharose 4B. UDP-glucuronosyltransferase in Fraction III was further purified by rechromatofocusing (pH 8.7 to 7.5). UDP-glucuronosyltransferases in Fractions IV and V were purified by UDP-hexanolamine-Sepharose chromatography. The transferase isoforms in Fractions II, III, IV and V were finally purified by h.p.l.c. on a TSK G 3000 SW column. Purified UDP-glucuronosyltransferase Isoforms Ia (Mr 51,000), Ib (Mr 52,000), Ic (Mr 56,000), II (Mr 52,000), IV (Mr 53,000) and V (Mr 53,000) revealed single Coomassie Blue-stained bands on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Isoform III enzyme showed two bands of Mr 52,000 and 53,000. Comparison of the amino acid compositions by the method of Cornish-Bowden [(1980) Anal. Biochem. 105, 233-238] suggested that all UDP-glucuronosyltransferase isoforms are structurally related. Reverse-phase h.p.l.c. of tryptic peptides of individual isoforms revealed distinct ‘maps’, indicating differences in primary protein structure. The two bands of Isoform III revealed distinct electrophoretic peptide maps after limited enzymic proteolysis. After reconstitution with phosphatidylcholine liposomes, the purified isoforms exhibited distinct but overlapping substrate specificities. Isoform V was specific for bilirubin glucuronidation, which was not inhibited by other aglycone substrates. Each isoform, except Ia, was identified as a glycoprotein by periodic acid/Schiff staining.

The pleiotropic actions of rosuvastatin confer renal benefits in the diabetic Apo-E knockout mouse

2010 ◽  
Vol 299 (3) ◽  
pp. F528-F535 ◽  
Author(s):  
Sara Giunti ◽  
Anna C. Calkin ◽  
Josephine M. Forbes ◽  
Terri J. Allen ◽  
Merlin C. Thomas ◽  
...  
Keyword(s):  
Creatinine Clearance ◽  
Combination Treatment ◽  
Plasma Lipids ◽  
Periodic Acid ◽  
Combined Treatment ◽  
Lipid Lowering ◽  
Advanced Glycation ◽  
Periodic Acid Schiff ◽  
Nadph Oxidase 4 ◽  
Apo E

Diabetic nephropathy is a leading cause of end-stage renal disease. Statins may exert renoprotective effects independently of lipid-lowering properties. We investigated the pleiotropic effects of rosuvastatin on renal structure and function in streptozotocin diabetic apolipoprotein-E knockout (Apo-E−/−) mice, a model of progressive nephropathy in which dyslipidemia is resistant to statin treatment. These effects were compared with those observed with conventional renin-angiotensin system blockade (candesartan) or combined treatment. Nondiabetic and diabetic Apo-E−/− mice were randomized to no treatment or treatment with candesartan (2.5 mg/kg), rosuvastatin (5 mg/kg), or their combination per gavage for 20 wk. Urine and blood samples were collected for assessment of albuminuria, creatinine clearance, plasma lipids, glucose, and glycated hemoglobin. Renal sclerosis was analyzed on paraffin-embedded kidney sections stained with periodic acid-Schiff. Renal expression of collagen IV, fibronectin and advanced glycation end products (AGEs), receptor for advanced glycation and products (RAGE), NADPH oxidase 4 (NOX4), and nitrotyrosine was assessed by real-time PCR and/or immunohistochemistry. Diabetes-induced albuminuria was not affected by rosuvastatin and combination treatment but was prevented by candesartan. Diabetes resulted in increased creatinine clearance, which was not modified by the treatments. Rosuvastatin and/or candesartan prevented diabetes-associated renal extracellular matrix accumulation. Rosuvastatin reduced accumulation of AGEs and expression of RAGE, NOX4, and nitrotyrosine. In conclusion, in the diabetic Apo-E−/− mouse, rosuvastatin confers renal benefits that are independent of lipid lowering and equivalent or greater to those observed with candesartan. The combination treatment is not superior to monotherapies.

scholarly journals Mechanism of lncRNA-ANRIL/miR-181b in autophagy of cardiomyocytes in mice with uremia by targeting ATG5

PLoS ONE ◽  
2021 ◽  
Vol 16 (9) ◽  
pp. e0256734
Author(s):  
Ying Xu ◽  
Jing Chen ◽  
Minmin Wang ◽  
Rizhen Yu ◽  
Wenly Zou ◽  
...  
Keyword(s):  
Serum Creatinine ◽  
Periodic Acid ◽  
Renal Tissue ◽  
Luciferase Reporter ◽  
Luciferase Reporter Gene ◽  
Periodic Acid Schiff ◽  
Urea Nitrogen ◽  
Tissue Sections ◽  
Related Proteins ◽  
Hematoxylin Eosin

Objectives This study is to investigate whether the cardiac microvascular endothelial cells (CMECs) can regulate the autophagy of cardiomyocytes (CMs) by secreting lncRNA-ANRIL/miR-181b exosomes, thus participating in the occurrence of uremic cardiovascular disease (CVD). Methods A 5/6 nephrectomy uremia model was established, with the mice injected with ANRIL-shRNA lentivirus vector, miR-181b agomir, and related control reagents, containing the serum creatinine and urea nitrogen measured. The renal tissue sections of mice were stained with Periodic Acid-Schiff (PAS), TUNEL, and Hematoxylin-Eosin (HE) performed on myocardial tissue sections of mice. ANRIL-shRNA, miR-181b mimics, and related control reagents were transfected into CMECs, in which the exosomes were extracted and co-cultured with CMs. The expressions of ANRIL, miR-181b and ATG5 were detected by qRT-PCR, and the expressions of autophagy related proteins by Western blot, as well as the binding of ANRIL and miR-181b by the double luciferase reporter gene experiment. Results ANRIL down-regulation or miR-181b up-regulation can increase the weight of mice with uremia, as well as the expressions of p62 and miR-181b, and reduce the content of serum creatinine and urea nitrogen, the damage of kidney and myocardial tissues, the number of apoptotic cells in myocardial tissues, as well as the expressions of ANRIL, ATG5, Beclin1, and LC3. CMs can absorb the exosomes of CMECs. Compared with IS+ CMEC-Exo group, the expressions of ANRIL and ATG5 in CMs of IS+ CMEC-Exo + sh lncRNA ANRIL and IS+CMEC-Exo+miR-181b mimics groups was down-regulated, as well as the expressions of ATG5, Beclin1, and LC3, while miR-181b expression was up-regulated as well as P62 expression. Conclusions CMECs can regulate autophagy of CMs by releasing exosomes containing ANRIL and miR-181b.

Cytochemical localization of glycogen phosphorylase activity in rat liver

1993 ◽  
Vol 51 ◽  
pp. 298-299
Author(s):  
John E. Michaels ◽  
Robert R. Cardell
Keyword(s):  
Rat Liver ◽  
Glycogen Phosphorylase ◽  
Periodic Acid ◽  
Glycogen Synthesis ◽  
Liver Glycogen ◽  
Periodic Acid Schiff ◽  
Cytochemical Localization ◽  
Key Enzyme ◽  
Staining Methods ◽  
Glycogen Phosphorylase Activity

Glycogen phosphorylase (GP) is a key enzyme in liver glycogen breakdown. GP activity is altered with its state of phosphorylation. In the current study, the intralobular distribution of GP activity was observed histochemically in frozen sections of rat liver during fasting and after stimulation of glycogen synthesis.Normal and adrenalectomized (ADX) rats were fasted overnight to reduce liver glycogen to minimal levels. Fasted ADX rats received 2 mg dexamethasone (DEX) 0-8 h prior to sacrifice to stimulate glycogen synthesis. Liver was removed, rapidly frozen in isopentane cooled in liquid nitrogen, then cryostat sectioned. In order to determine sites of GP activity, sections were incubated in medium that contained glucose 1-phosphate as substrate. Under the incubation conditions used, GP synthesized glycogen as the reaction product. Glycogen was identified by two staining methods: 1) iodine staining has been shown to be rather specific for newly synthesized glycogen produced during the histochemical procedure (Figs.1-6), whereas 2) periodic acid-Schiff (PAS) stained both native and nascent glycogen.

scholarly journals Novel rat liver progenitor cell surface markers

2006 ◽  
Vol 20 (4) ◽  
Author(s):  
Mladen I Yovchev ◽  
Petar N Grozdanov ◽  
Mariana D Dabeva
Keyword(s):  
Cell Surface ◽  
Rat Liver ◽  
Progenitor Cell ◽  
Surface Markers ◽  
Cell Surface Markers ◽  
Liver Progenitor Cell

scholarly journals Tissue fixation with diimidoesters as an alternative to aldehydes. II. Cytochemical and biochemical studies of rat liver fixed with dimethylsuberimidate.

1976 ◽  
Vol 24 (9) ◽  
pp. 1000-1011 ◽  
Author(s):  
A R Hand ◽  
J R Hassell
Keyword(s):  
Rat Liver ◽  
Biochemical Analysis ◽  
Periodic Acid ◽  
Residual Activity ◽  
Paraffin Sections ◽  
Electron Microscopic ◽  
Periodic Acid Schiff ◽  
Biochemical Studies ◽  
Electron Microscopic Cytochemistry ◽  
Background Reaction

Rat liver fixed with dimethylsuberimidate (DMS) was studied to investigate the use of diimidoesters as dixatives for light and electron microscopic cytochemistry. Paraffin sections of liver fixed with DMS at pH 9.5 were weakly stained with the ninhydrin-Schiff procedure, indicating extensive reaction of NH3+ groups with the fixative. Nuclei were strongly strained by the Feulgen procedure, with no background [corrected] reaction. In contrast, glutaraldehyde fixation resulted in a significant background reaction in the cytoplasm and nuclei in controls for the Schiff-based stains. DMS-fixed liver stained intensely for glycogen with the Periodic acid-Schiff procedure, and biochemical analysis of glycogen retention and extractability indicated that DMS retained considerably more glycogen in sections than glutaraldehyde. DMS-fixed liver incubated for thiamine pyrophosphatase activity revealed reaction product in ER cisternae, Goli saccules and bile canaliculi. Peroxisomes were strongly reactive for catalase activity after incubation in diaminobenzidine medium, and reaction product of glucose-6-phosphatase activity was considerably greater following DMS fixation than after glutaraldehyde. Biochemical studies revealed up to twice as musch residual activity of glucose-6-phosphatase after DMS fixation. These results suggest that DMS may be useful as a primary fixative for certain cytochemical procedures.

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