Hepatocyte Peroxisome Research Articles

Abstract 14039: Therapeutic Potential of Beta Hydroxybutyrate in Nonalcoholic Fatty Liver Disease (NAFLD)-Induced Cardiovascular Disease

Background: Cardiovascular disease (CVD) is the leading cause of mortality in non-alcoholic fatty liver disease (NAFLD). Beta hydroxybutyrate (BHOB), a liver metabolite, is the major ketone that serves as an alternative fuel source in the body. Our previous studies show that hepatocyte peroxisome proliferator activator receptor-α knockout (Ppara HEPKO ) mice, model of NAFLD-induced CVD, exhibit reduced circulating levels of BHOB. Hypothesis: We hypothesized that restoring plasma BHOB levels will improve cardiovascular function in Ppara HEPKO mice. Aims: To determine the therapeutic role of BHOB in NAFLD-induced CVD. Methods: 30 weeks old male Ppara HEPKO mice were given 1,3 butanediol (20% in drinking water) (Ppara HEPKO + BHOB) or vehicle (Ppara HEPKO ) (n=6) for 6 weeks. Plasma BHOB was measured at baseline and after treatment. Cardiac structure and function were measured by high resolution ultrasound echocardiography (VEVO 3100), using the B-mode, M-mode, pulse wave and tissue doppler at baseline and after treatment. Mean arterial blood pressure was measured by radiotelemetry. Liver fat was determined by EchoMRI, Oil Red O staining and hepatic triglyceride levels. Results: After 6 weeks of BHOB treatment, Ppara HEPKO exhibit increased plasma BHOB compared to baseline (0.5 ± 0.01 vs. 0.2 ± 0.02mmol/L), attenuated arterial blood pressure compared to control (109 ± 3 vs. 121 ± 4mmHg), improved systolic function: cardiac output (13.8 ± 0.8 vs. 11.1 ± 0.7ml/min), stroke volume (31.1 ± 2.1 vs. 23.4 ± 1.3ml/min), improved diastolic function: isovolumic relaxation time (18.7 ± 0.8 vs. 20.6 ± 0.9m/sec), E/e’ (-29.6 ± 0.9 vs. -32.4 ± 1.4), and attenuated vascular stiffness. Interestingly, BHOB did not alleviate hepatic fat compared to control as demonstrated by EchoMRI (0.8 ± 0.3 vs. 0.7 ± 0.3), hepatic triglyceride (1.4 ± 0.3 vs. 1.3 ± 0.2mM) and Oil Red O staining. Conclusion: Our findings indicate that BHOB treatment improves arterial blood pressure, systolic, diastolic, and vascular function in, Ppara HEPKO knockout mice. However, BHOB did not alleviate hepatic steatosis suggesting that BHOB improves cardiovascular function in Ppara HEPKO mice independent of changes in hepatic fat content.

Maternal protein deficiency impairs peroxisome biogenesis and leads to oxidative stress and ferroptosis in liver of fetal growth restriction offspring

Maternal protein malnutrition leads to liver dysfunction and increases susceptibility to nonalcoholic fatty liver disease in adult fetal growth restriction (FGR) offspring, yet the underlying mechanism remains unknown. Peroxisomes play vital roles in fatty acid β-oxidation (FAO) and detoxification of reactive oxygen species (ROS). Using a well-defined rat model, the peroxins (PEXs), fatty acid metabolic enzymes, and oxidase stress regulators were investigated in the liver of FGR offspring. The results revealed that PEX3, 11b, 14, and 19 were obviously reduced in the fetal liver and lasted to adulthood, suggesting a decrease in the biogenesis and division of peroxisomes. FA metabolism enzymes and ferroptosis regulators were deregulated. To further investigate this association, small interfering RNA was employed to achieve knockdown (KD) of PEX14 in BRL cells (a rat hepatocyte line). PEX14 KD led to dysregulation of PEXs and long-chain FAs accumulation. PEX14 deficiency caused ROS accumulation and lipid peroxidation, finally induced regulated cell death (including apoptosis, autophagy, and ferroptosis). Double knock down (DKD) of PEX14 and fatty acyl-CoA reductase 1 (FAR1) revealed that PEX14 KD-induced ferroptosis was related with enhanced FAR1 level. DKD of PEX14 and Atg5 further confirmed that PEX14 KD-induced cell death was partly autophagy-dependent. Overall, these data demonstrate a vital role for PEX14 in maintaining peroxisome function and liver physiology, and suggest that hepatocyte peroxisome defects partly explain liver dysplasia and lipid metabolism disorders in fetal original liver disease.

Hepatocyte peroxisome proliferator-activated receptor α regulates bile acid synthesis and transport

Peroxisome proliferator-activated receptor alpha (PPARα) controls lipid homeostasis through regulation of lipid transport and catabolism. PPARα activators are clinically used for hyperlipidemia treatment. The role of PPARα in bile acid (BA) homeostasis is beginning to emerge. Herein, Ppara-null and hepatocyte-specific Ppara-null (Ppara∆Hep) as well as the respective wild-type mice were treated with the potent PPARα agonist Wy-14,643 (Wy) and global metabolomics performed to clarify the role of hepatocyte PPARα in the regulation of BA homeostasis. Levels of all serum BAs were markedly elevated in Wy-treated wild-type mice but not in Ppara-null and Ppara∆Hep mice. Gene expression analysis showed that PPARα activation (1) down-regulated the expression of sodium-taurocholate acid transporting polypeptide and organic ion transporting polypeptide 1 and 4, responsible for the uptake of BAs into the liver; (2) decreased the expression of bile salt export pump transporting BA from hepatocytes into the bile canaliculus; (3) upregulated the expression of multidrug resistance-associated protein 3 and 4 transporting BA from hepatocytes into the portal vein. Moreover, there was a notable increase in the compositions of serum, hepatic and biliary cholic acid and taurocholic acid following Wy treatment, which correlated with the upregulated expression of the Cyp8b1 gene encoding sterol 12α-hydroxylase. The effects of Wy were identical between the Ppara∆Hep and Ppara-null mice. Hepatocyte PPARα controlled BA synthesis and transport not only via direct transcriptional regulation but also via crosstalk with hepatic farnesoid X receptor signaling. These findings underscore a key role for hepatocyte PPARα in the control of BA homeostasis.

PPARs and the Cardiovascular System

Peroxisome proliferator-activated receptors (PPARs) belong to the nuclear hormone-receptor superfamily. Originally cloned in 1990, PPARs were found to be mediators of pharmacologic agents that induce hepatocyte peroxisome proliferation. PPARs also are expressed in cells of the cardiovascular system. PPAR gamma appears to be highly expressed during atherosclerotic lesion formation, suggesting that increased PPAR gamma expression may be a vascular compensatory response. Also, ligand-activated PPAR gamma decreases the inflammatory response in cardiovascular cells, particularly in endothelial cells. PPAR alpha, similar to PPAR gamma, also has pleiotropic effects in the cardiovascular system, including antiinflammatory and antiatherosclerotic properties. PPAR alpha activation inhibits vascular smooth muscle proinflammatory responses, attenuating the development of atherosclerosis. However, PPAR delta overexpression may lead to elevated macrophage inflammation and atherosclerosis. Conversely, PPAR delta ligands are shown to attenuate the pathogenesis of atherosclerosis by improving endothelial cell proliferation and survival while decreasing endothelial cell inflammation and vascular smooth muscle cell proliferation. Furthermore, the administration of PPAR ligands in the form of TZDs and fibrates has been disappointing in terms of markedly reducing cardiovascular events in the clinical setting. Therefore, a better understanding of PPAR-dependent and -independent signaling will provide the foundation for future research on the role of PPARs in human cardiovascular biology.

Peroxisome Proliferator-Activated Receptors: Mediators of Phthalate Ester-Induced Effects in the Male Reproductive Tract?

Many phthalate ester plasticizers are classified as peroxisome proliferators (PP), a large group of industrial and pharmaceutical chemicals. Like PP, exposure to some phthalates increases hepatocyte peroxisome and cellular proliferation, as well as the incidence of hepatocellular adenomas in mice and rats. Most effects of PP are mediated by three nuclear receptors called peroxisome proliferator-activated receptors (PPARalpha,beta,gamma). An obligate role for PPARalpha in PP-induced events leading to liver cancer is well-established. Exposure of rats in utero or in the neonate to a subset of phthalate esters causes profound, sometimes irreversible malformations in the male reproductive tract. We review here the data that supports or discounts roles for PPARs in phthalate-induced testis toxicity including (1) toxic effects of phthalates on the male reproductive tract, (2) expression of PPARs in the testis, (3) activation of PPARs by phthalates, (4) role of PPARalpha in testis toxicity, (5) gene targets of phthalates involved in steroid biosynthesis and catabolism, and (6) interactions between PPARs and other nuclear receptors that play roles in testis development and homeostasis. Critical research needs are identified that will help determine the significance of PPARs in phthalate-induced effects in the rat male reproductive tract and the relevance of toxicity to humans.

Peroxisome Proliferator-activated Receptor α-mediated Pathways Are Altered in Hepatocyte-specific Retinoid X Receptor α-deficient Mice

Retinoid x receptor alpha (RXRalpha) serves as an active partner of peroxisome proliferator-activated receptor (PPARalpha). In order to dissect the functional role of RXRalpha and PPARalpha in PPARalpha-mediated pathways, the hepatocyte RXRalpha-deficient mice have been challenged with physiological and pharmacological stresses, fasting and Wy14,643, respectively. The data demonstrate that RXRalpha and PPARalpha deficiency are different in several aspects. At the basal untreated level, RXRalpha deficiency resulted in marked induction of apolipoprotein A-I and C-III (apoA-I and apoC-III) mRNA levels and serum cholesterol and triglyceride levels, which was not found in PPARalpha-null mice. Fasting-induced PPARalpha activation was drastically prevented in the absence of hepatocyte RXRalpha. Wy14,643-mediated pleiotropic effects were also altered due to the absence of hepatocyte RXRalpha. Hepatocyte RXRalpha deficiency did not change the basal acyl-CoA oxidase, medium chain acyl-CoA dehydrogenase, and malic enzyme mRNA levels. However, the inducibility of those genes by Wy14,643 was markedly reduced in the mutant mouse livers. In contrast, the basal cytochrome P450 4A1, liver fatty acid-binding protein, and apoA-I and apoC-III mRNA levels were significantly altered in the mutant mouse livers, but the regulatory effect of Wy14,643 on expression of those genes remained the same. Wy14,643-induced hepatomegaly was partially inhibited in hepatocyte RXRalpha-deficient mice. Wy14,643-induced hepatocyte peroxisome proliferation was preserved in the absence of hepatocyte RXRalpha. These data suggested that in comparison to PPARalpha, hepatocyte RXRalpha has its unique role in lipid homeostasis and that the effect of RXRalpha, -beta, and -gamma is redundant in certain aspects.

Are hepatocyte polarity and peroxisome expression related?

Are hepatocyte polarity and peroxisome expression related?

Comparison of the Effects of Various Peroxisome Proliferators on Peroxisomal Enzyme Activities, DNA Synthesis, and Apoptosis in Rat and Human Hepatocyte Cultures

Peroxisome proliferators (PPs) are a class of rodent nongenotoxic hepatocarcinogens that cause hepatocyte peroxisome proliferation, increased DNA synthesis, and decreased spontaneous apoptosis. We examined the effects of various PPs such as the hypolipidemic agents clofibric acid (CLO), bezafibrate (BEZA), ciprofibrate (CIPRO), and nafenopin (NAFE) and the plasticizer di-(2-ethylhexyl)phthalate (DEHP) on the various parameters in vitro in rat and human hepatocyte cultures. In rat hepatocyte cultures, after 72 h of treatment with the various PPs at 100–500 μM, a compound-dependent increase in acyl CoA oxidase (ACO) and carnitine acetyl transferase (CAT) activities, markers of peroxisome proliferation, was observed with the following potencies: CIPRO = NAFE > BEZA > CLO > DEHP. A minor (120–150%), but significant, no concentration-dependent increase in DNA synthesis and a marked, no compound-dependent and, with the exception of NAFE, no concentration-dependent 60–80% decrease in spontaneous apoptosis was observed with all tested compounds (50–250 μM) after 48 h of treatment. Inhibition of spontaneous apoptosis in PP-treated versus control rat hepatocyte cultures was also observed morphologically. Furthermore, PPs inhibited transforming growth factor β (TGFβ)-induced apoptosis but not tumor necrosis factor α (TNFα)/α Amanitine (αAma)-induced apoptosis in rat hepatocyte cultures. In human hepatocyte cultures, the various PPs at 50–500 μM did not affect peroxisomal enzyme activities, DNA synthesis, or spontaneous and induced (TGFβ or TNFα/αAma) apoptosis. The compound-dependent peroxisome proliferation but no compound-dependent disruption of the mitogenic/apoptotic balance elicited by PPs in primary rat hepatocyte cultures supports the hypothesis that oxidative stress is directly linked to the hepatocarcinogenic potential of a given PP in rodents and that disruption of the mitogenic/apoptotic balance contributes to the development of PP-induced hepatocarcinogenesis. In addition, the absence of effects of all PPs on both peroxisome proliferation-associated parameters and mitogenic/apoptotic balance supports the hypothesis that human liver cells are refractory to PP-induced hepatocarcinogenesis.

Transcriptional modulation of hepatocyte nuclear factor 4a (HNF-4a) and/or peroxisome proliferator-activated receptor a (PPARa)-controlled genes by low molecular weight compounds

Transcriptional modulation of hepatocyte nuclear factor 4a (HNF-4a) and/or peroxisome proliferator-activated receptor a (PPARa)-controlled genes by low molecular weight compounds

Tumour necrosis factor alpha (TNF alpha) suppresses apoptosis and induces DNA synthesis in rodent hepatocytes: a mediator of the hepatocarcinogenicity of peroxisome proliferators?

Peroxisome proliferators (PPs) are a class of non-genotoxic rodent hepatocarcinogens that cause increased hepatocyte DNA synthesis, peroxisome proliferation and liver enlargement. We have demonstrated previously that PPs suppress both spontaneous rat hepatocyte apoptosis and that induced by the physiological negative regulator of liver growth, transforming growth factor beta (TGF beta1). Evidence suggests that the suppression of apoptosis by PPs is mediated via activation of the peroxisome proliferator activated receptor-alpha (PPAR alpha), a member of the nuclear hormone receptor superfamily. Here, we investigate the effects of tumour necrosis factor alpha (TNF alpha) on cultured rat or mouse hepatocytes to determine whether TNF alpha influences hepatocyte growth in a manner analogous to that seen with PPs. Rat recombinant TNF alpha was found to stimulate DNA synthesis and suppress apoptosis in isolated rat hepatocyte monolayers (P < or = 0.01). These effects were seen in the range of 500-5000 U/ml with a maximum effect at 5000 U/ml. Similarly, mouse recombinant TNF alpha was able to stimulate DNA synthesis in mouse hepatocyte monolayers (P < or = 0.01) with a maximal effect at 1000 U/ml. Suppression of mouse hepatocyte apoptosis by TNF alpha was not detected, possibly because of the low levels of apoptosis under control conditions. However, when the levels of mouse hepatocyte apoptosis were augmented using TGF beta1, TNF alpha caused a significant suppression (P < or = 0.01). The neutralization of TNF alpha using anti-TNF alpha antibodies abrogated significantly (P < or = 0.01) the suppression of apoptosis by the PP, nafenopin. These data that suggest TNF alpha may mediate, at least in part, the growth perturbation, liver enlargement and hepatocarcinogenesis seen in response to the PP class of non-genotoxic hepatocarcinogens.

Hepatocellular proliferation in ibuprofen-treated mice.

Female B6C3F1 mice were treated with ibuprofen for 2 wk or 90 days to monitor effects on hepatocellular proliferation during acute and subchronic exposure. Proliferation was assessed by bromodeoxyuridine labeling. Mice treated with 100, 200, or 400 mg/kg ibuprofen for 2 wk had significantly increased liver weights. A dose-related increase in the number of labeled hepatocytes per 1,000 hepatocytes suggested that the weight increases were in part a result of hepatocellular proliferation. Hepatocellular hypertrophy also contributed to the increased liver size as indicated by decreases in the number of hepatocytes per high power field. Ultrastructural evaluation indicated that hepatocyte peroxisome size increased significantly in treated mice. Mice treated with 100 or 200 mg/kg ibuprofen for 90 days and given bromodeoxyuridine during the last 2 wk of ibuprofen exposure had statistically significant increases in relative liver weights. However, the number of labeled hepatocytes per 1,000 hepatocytes was not increased, and there was no evidence of hepatocellular hypertrophy. Mice given 200 mg/kg ibuprofen for 90 days had significantly decreased serum triglycerides. These findings indicate that ibuprofen treatment of mice results in hepatomegaly characterized by hepatocellular hypertrophy and hyperplasia. Peroxisomal changes may be contributory to the pathogenesis of this lesion.