Antihepatotoxic activity of monomethyl fumarate isolated from Fumaria indica

Monomethyl fumarate, isolated for the first time from the methanolic extract of the whole plant of Fumaria indica, was characterised and screened for its antihepatotoxic activity in albino rats. The compound showed significant (P<0.01) antihepatotoxic activity against thioacetamide in vitro, and against hepatotoxicities induced by carbon tetrachloride, paracetamol and rifampicin in vivo to an extent almost similar to that of silymarin, a known antihepatotoxic agent.     

M1- and M2-macrophage polarization in rat liver cirrhosis induced by thioacetamide (TAA), focusing on Iba1 and galectin-3

Introduction

Resident and exudate macrophages play an important role in the development of liver cirrhosis. Ionized calcium binding adaptor molecule 1⁺ (Iba1⁺) and galectin-3⁺ (Gal-3⁺) macrophages regulate liver fibrosis probably through pro-inflammatory and pro-fibrotic factors. Macrophages show polarized functions in liver fibrosis; however, M1-/M2-polarization of Iba1⁺ and Gal-3⁺ macrophages remains obscured. This study investigated the M1-/M2-polarized properties of Iba1⁺ and Gal-3⁺ macrophages in chemical-induced liver cirrhosis.

Materials and methods

Cirrhosis was induced in F344 rats by repeated injections of thioacetamide (100 mg/kg BW, twice a week for 25 weeks). Liver samples were collected from post-first-injection (PFI) week 5 to 25. Macrophage immunophenotypes and myofibroblasts in the fibrous bridges (FBs) and pseudolobules (PLs) were analyzed by immunohistochemistry. Expressions of M1- and M2-related factors were analyzed with RT-PCR, separately in FBs and PLs.

Results

Activation of myofibroblasts was most pronounced in livers at week 15. CD68⁺ (M1), CD204⁺ (M2), Iba1⁺ and Gal-3⁺ macrophages in the FBs increased gradually and peaked at week 15, consistent with the upregulation of both M1-(MCP-1, IFN-γ, IL-1β, IL-6, and TNF-α) and M2-(TGF-β1, IL-4, and IL-10) related factors. Iba1⁺ and Gal-3⁺ macrophages showed both M1- and M2-immunophenotypes. CD163⁺ macrophages showed a persistent increase, consistent with TGF-β1 upregulation. MHC class II⁺ macrophages increased in the developing fibrotic lesions, and then reduced in the advanced stage cirrhosis.

Conclusion

Both M1- and M2-macrophage polarizations occur during development of liver cirrhosis. Iba1⁺ and Gal-3⁺ macrophages participate in liver cirrhosis through production of both M1- and M2-related factors.     

Chronic High Doses of Thioacetamide Followed by Vitamin A Modify Dolichol,Dolichol Isoprenoids,and Retinol Content in Rat Liver Cells

Our line of researches follows the hypothesis that dolichol and retinol metabolism might be interrelated and involved in liver fibrosis. To this end, in this study rats were subjected to chronic treatment with thioacetamide (TAA) (300 mg/L liquid diet) for 1 and 2 months and, after liver damage had occurred, supplemented with vitamin A before sacrifice. Dolichol, dolichol isoprene units, and retinol content were determined in isolated parenchymal and sinusoidal liver cells (hepatic stellate cells; Kupffer cells; sinusoidal endothelial cells). Dolichol increased in hepatocytes after TAA treatment, with or without vitamin A. Dolichol decreased in the other cells. Retinol in general decreased. In hepatocytes, retinol decreased only on normal nutrition, while the vitamin A load was taken up normally. The percentages of dolichol isoprene units (Dol-16 to Dol-20, in rats) confirm that Dol-18, which was not modified in percentage by TAA on normal nutrition, did not increase after vitamin A, as it did in control cells (7–12%). The behavior of Dol-18 was similar in all the cells studied. Vitamin A might reveal a latent damage produced by TAA on dolichol homologues. These data support previous hypotheses that the action of TAA depends on the administration modality, the dosage, and the diet, and that Dol-18 might have different functions and compartmentalization in the cells. Furthermore, the resultssupport the hypothesis that dolichol chain length might be interrelated with retinol metabolism, perhaps through their metabolites.     

台盼蓝肝脏原位灌流在研究肝微循环变化中的应用

探讨台盼蓝肝脏原位灌流在研究肝微循环变化中的应用。方法采用台盼蓝肝脏原位灌流的方法观察硫代乙酰胺（ｔｈｉｏａｃｅｔａｍｉｄｅ，ＴＡＡ）所致大鼠肝脏损伤时的微循环状态。结果内毒素血症的严重程度、反映缺血性肝损伤程度的ＬＤＨ活性、以及表示肝微循环灌注状况的台盼蓝在肝内的分布时间与肝的病理形态学改变，在ＴＡＡ损伤作用的不同时间，其变化趋势都是一致的。结论１内毒素血症在肝微循环障碍发生发展中具有重要作用。     

Microarray analysis of thioacetamide-treated type 1 diabetic rats

It is well known that diabetes imparts high sensitivity to numerous hepatotoxicants. Previously, we have shown that a normally non-lethal dose of thioacetamide (TA, 300 mg/kg) causes 90% mortality in type 1 diabetic (DB) rats due to inhibited tissue repair allowing progression of liver injury. On the other hand, DB rats exposed to 30 mg TA/kg exhibit delayed tissue repair and delayed recovery from injury. The objective of this study was to investigate the mechanism of impaired tissue repair and progression of liver injury in TA-treated DB rats by using cDNA microarray. Gene expression pattern was examined at 0, 6, and 12 h after TA challenge, and selected mechanistic leads from microarray experiments were confirmed by real-time RT-PCR and further investigated at protein level over the time course of 0 to 36 h after TA treatment. Diabetic condition itself increased gene expression of proteases and decreased gene expression of protease inhibitors. Administration of 300 mg TA/kg to DB rats further elevated gene expression of proteases and suppressed gene expression of protease inhibitors, explaining progression of liver injury in DB rats after TA treatment. Inhibited expression of genes involved in cell division cycle (cyclin D1, IGFBP-1, ras, E2F) was observed after exposure of DB rats to 300 mg TA/kg, explaining inhibited tissue repair in these rats. On the other hand, DB rats receiving 30 mg TA/kg exhibit delayed expression of genes involved in cell division cycle, explaining delayed tissue repair in these rats. In conclusion, impaired cyclin D1 signaling along with increased proteases and decreased protease inhibitors may explain impaired tissue repair that leads to progression of liver injury initiated by TA in DB rats.     

Protective effect of aqueous extract of Feronia elephantum correa leaves on thioacetamide induced liver necrosis in diabetic rats

Objective

To evalueate hepatoprotective effects Feronia elephantum (F. elephantum) correa against thioacetamide (TA) induced liver necrosis in diabetic rats.

Methods

Male wistar rats were made diabetic with alloxan (160 mg/kg) on day 0 of the study. They were intoxicated with hepatotoxicant (thioacetamide, 300 mg/kg, ip) on day 9 of study to produce liver necrosis. Effects of 7 day daily once administration (day 2 to day 9) of EF (400 and 800 mg/kg, po) were evaluated on necorosis of liver in terms of mortality, liver volume, liver weight, serum aspartate aminotransferase (AST) and serum alanine transaminase (ALT), and histopathology of liver sections (for signs of necorosis and inflammation) on day-9 of the study. Separate groups of rats with treated only with alloxan (DA control), thioacetamide (TA control) and both (TA₊DA control) were maintained.

Results

FE significantly lowered the mortality rate and showed improvement in liver function parameters in TA-induced diabetic rats without change in liver weight, volume and serum glucose levels.

Conclusions

FE showed promising activity against TA-induced liver necorsis in diabetic rats and so might be useful for prevention of liver complications in DM.     

Inhibition of experimentally-induced liver cirrhosis in rats by a nonpeptidic mimetic of the extracellualr matrix-associated Arg-Gly-Asp epitope

Aims/Methods: in the present study, we examined whether a nonpeptidic mimetic of Arg-Gly-Asp (RGD), which specifically inhibits RGD-dependent adhesion of CD4⁺ T lymphocytes or fibroblasts to fibronectin, can prevent thioacetamide-induced liver cirrhosis in rats. The nonpeptidic RGD mimetic, rather than the RGD peptide was utilized, since the peptide is rapidly degraded, and therefore, relatively ineffective for in vivo use.Results: We now report that rats treated with thioacetamide and the RGD analogue SF-6,5 for 12 weeks had lower histopathologic scores than those treated with thioacetamide alone. Further improvement in liver histology was observed after another 8 weeks of treatment with the analogue SF-6,5. Quantitative microscopic analysis by computerized imaging morphometry of liver biopsies from the three groups and controls confirmed the semi-quantitative histopathologic scores (pτ0.001). After 3 months of treatment, the spleen weights in the SF-6,5-treated rats were 30% less than those of rats and that received only thioacetamide, which indicated that the analogue-treated rats were less portal hypertensive.Conclusions: The observed inhibition of the progression of cirrhosis in rats by the nonpeptidic RGD analogue suggests that RGD mimetics may be useful therapeutically in inhibiting pathological processes that involve RGD recognition.     

硫代乙酰胺致小鼠肝性脑病模型的建立及评估

目的：探讨应用硫代乙酰胺（TAA）和C57/BL6小鼠制备肝性脑病模型的方法。方法100只小鼠分成4组,对照组（A组）、实验组B、C、D组,每组25只,分别予生理盐水和TAA 100、200、300 mg/（kg·d）腹腔内注射。注射药物后每24小时通过旷场实验、ROTA ROD实验及神经功能学评分检测神经功能,观察肝组织病理变化。结果旷场试验中,与对照组相比,从造模第1天开始,各实验组小鼠均出现运动总路程和总时间明显下降（P 〈0.05）,而各实验组之间无明显差异。 ROTA ROD实验中,与对照组相比,造模第3天实验D组小鼠运动时间明显下降（P =0.036）;第4天,各实验组小鼠均明显下降（P =0.000）,而实验组内无明显差异。造模第4天,实验C、D组小鼠神经功能学评分平均值分别为6.3和5.1分,与对照组比较,差异有统计学意义（P 〈0.05）。造模第4天实验B、C、D 3组小鼠的累积生存率分别为71.4%、71.4%和42.9%。3组实验组小鼠肝组织病理改变均符合急性肝衰竭改变。结论 C57/BL6小鼠采用TAA可成功制备肝性脑病动物模型,200 mg/（kg＆#183;d）TAA腹腔注射,连续4 d,为推荐方法。     

Ameliorative effect of grape seed proanthocyanidin extract on thioacetamide-induced mouse hepatic fibrosis

The present study was designed to examine the effect of the grape seed proanthocyanidin extract (GSPE) on developing hepatic fibrosis that was induced by thioacetamide (TAA) in mice. Administration of TAA for 9 weeks led to a serious necrosis and apoptosis of the parenchymal cells, which resulted in an accumulation of excessive collagen in the liver and an increase of transformed hepatic stellate cells (HSCs). In addition, the mRNA expression of transforming growth factor β1 (TGF-β1), α-smooth muscle actin (α-SMA), as the marker of the activated HSCs, and α1-(I)-collagen were all up-regulated significantly when compared with the control. However, combined oral administration of GSPE at 100mg/kg suppressed the mRNA expression of TGF-β1 and α-SMA, with decreased collagen accumulation as demonstrated by histomorphological evaluation and quantitative RT-PCR. The mRNA expression of the pro-inflammatory factors, including inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), was remarkably enhanced by TAA treatment. However, their levels displayed a down-regulated trend beyond simultaneous GSPE treatment. Moreover, GSPE administration markedly suppressed lipid peroxidation. In conclusion, as a plant antioxidant, GSPE manifested effective hepatocellular protective action to ameliorate the developing liver fibrosis induced by chronic TAA administration in mice.     

Evaluation of the Developmental Toxicities of Ethanol,Acetaldehyde, and Thioacetamide Using FETAX

Abstract

Potential mechanisms of the developmental toxicities of ethanol, acetaldehyde, and thioacetamide were evaluated using frog embryo teratogenesis assay-Xenopus (FETAX). Early X. laevis embryos were exposed to ethanol and thioacetamide in two separate definitive concentration-response tests with and without differentially induced exogenous metabolic activation systems (MAS) or selectively inhibited MAS. Two concentration-response tests were also performed with ethanol metabolites, acetaldehyde and acetic acid. The MAS was treated with 3,4-amino-1,2,4-triazole to modulate CYP2E1 activity, and heat to inactivate flavin containing monooxygenases (FMO) activity. Results from these studies suggested that thioacetamide may be bioactivated by both CYP2E1 and the FMO systems. Ethanol also appeared to be bioactivated by CYP2E1. Acetaldehyde was markedly more potent as a developmental toxicant than ethanol or acetic acid. Binary joint mixture studies conducted with ethanol and acetaldehyde indicated that the parent compound and metabolite acetaldehyde acted in a response additive manner. These results warrant the continued use of FETAX as a means of evaluating mechanisms of developmental toxicity in vitro.