Characterization of the response of growth and differentiation to lipoproteins and agents affecting cholesterol metabolism in murine neuroblastoma cells

Treatment with mevinolin, a competitive inhibitor of HMGCoAR, the key enzyme of isoprenoid metabolism, causes the arrest of proliferation and the differentiation of a neuroblastoma cell line (N18TG2). Mevalonate and high density lipoproteins partially restore growth.Cholesterol synthesis in the presence of mevinolin remains active, because in these cells the key enzyme HMG-CoA reductase is not completely inhibited by this drug. The fact that cell growth is reduced, while cholesterogenesis remains active, suggests that mevinolin acts by interfering with the synthesis of some unknown compound, other than cholesterol, which is necessary for proliferation.     

Lowering of plasma cholesterol levels in animals by lovastatin and simvastatin

Summary Lovastatin and simvastatin are potent competitive inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase. Key inhibit the synthesis of cholesterol in cultured HepG23 cells, rat hepatocytes and in rats. The primary target organ of cholesterol synthesis inhibition by lovastatin and simvastatin is the liver. Lovastating and simvastatin lower levels of plasma cholesterol in rats, dogs and rabbits by inhibition the endogenous cholesterol synthesis and induction of LDL receptor in the liver.     

Hepatic HMGCoA reductase in human cholelithiasis: effects of chenodeoxycholic and ursodeoxycholic acids*

Abstract. To study further the role of hepatic cholesterol synthesis in patients with gallstones, the activity of the rate-limiting step in cholesterbgenesis, hydroxy-methyl glutaryl co-enzyme A reductase (HMGCoAR), was measured in operative wedge liver biopsies from ten patients with untreated cholesterol gallstones, six with pigment stones and ten controls. Hepatic HMGCoAR was also measured in six patients with cholesterol-rich gallstones treated for 1–24 months with 14-6-18-6 mg chenodeoxycholic acid (CDCA) kg^-1 day^-1, in two patients with radiolucent pigment stones treated with 17-3 and 17-7 mg CDCA kg^-1 day^-1 and in ten other patients with cholesterol-rich stones given 4–5-7-2 mg ursodeoxycholic acid kg^-1 day^-1 for 1–6 months. HMGCoAR activity was also related to the free and esterified cholesterol content of both hepatic homogenates and the microsomal fractions. Compared with the controls (HMGCoAR activity 14-6±1 6 (SEM) pmole mg microsomal protein^-1 min^-1), the mean activity in untreated cholesterol cholelithiasis was 32 2 ±2-0 (P < 0–001), but was near normal in patients with pigment stones (16-2 + 1 5). In cholesterol gallstone patients, chenodeoxycholic acid treatment reduced the mean enzyme activity by 51% compared with the untreated gallstone group (P < 0001) and in smaller doses, ursodeoxycholic acid therapy lowered it by 40% (P < 0001) but in the two patients with pigment stones, CDCA did not seem to affect HMGCoAR activity. Despite this four-fold variation in enzyme activity, there were no significant differences in mean free or esterified cholesterol concentrations in whole liver homogenates or in the microsomal fraction from any of the patient groups. Presented in part at the European Society for Clinical Investigation, Rotterdam, April 1978 (Maton P.N. & Dowling R.H (1978) Eur J Clin Invest 8, 329 (abstract)) and at the V International Bile Acid Meeting, Freiburg, June 1978 (Maton P.N. & Dowling R.H. (1978) in: Biological Effects of Bile Acids (ed. by G. Paumgartner, A. Stiehl and W. Gerok), pp. 91–98. MTP Press, Lancaster).     

The imidazoline-like drug S23515 affects lipid metabolism in hepatocyte by inhibiting the oxidosqualene: lanosterol cyclase activity

Imidazoline-like drugs are centrally-acting antihypertensive agents that inhibit the activity of the sympathetic nervous system by interacting with the alpha2-adrenoreceptor and also with a non-adrenergic imidazoline binding site called the imidazoline 1 receptor. Recently, these molecules were proposed to play an additional role in cardiovascular diseases by acting on glucose and lipid metabolism. We used S23515, a potent imidazoline-like molecule acting selectively on blood pressure through the imidazoline 1 receptor, to decipher the effects of these drugs on lipid metabolism. We found that S23515 inhibited specifically and dose-dependently cholesterol synthesis in cultured rodent and primate hepatocytes. This hypocholesterolemic effect was likely due to the inhibition of the oxido:lanosterol cyclase (OSC), a rate-limiting enzyme in the cholesterol biosynthetic pathway. Partial OSC inhibition induced by S23515 led to the generation of 24(S),25-epoxycholesterol, a potent ligand for the liver X receptor (LXR). Furthermore, S23515 increased in human macrophages the expression of both ABCA1 and G1, the 2 ATP binding cassette transporters, which play a pivotal role in the reverse cholesterol transport. Thus, these results suggest that S23515, and potentially other imidazoline-like drugs, could exert hypolipidemic effects in addition to their hypotensive activities.     

Antihyperlipidemic effect of aqueous extract of Plumbago zeylanica roots in diet-induced hyperlipidemic rat

This study examined the antihyperlipidemic effect of the aqueous extract of Plumbago zeylanica Linn. (Plumbaginaceae) roots in diet-induced hyperlipidemic rats. The oral administration of the aqueous extract at the dose of 20, 40, and 80?mg kg^?1 were found to ameliorate the hyperlipidemic condition as evidenced by a reduction of cholesterol and triglyceride levels. The standards fenofibrate (20?mg kg^?1) and atorvastatin (8?mg kg^?1) were also found to exhibit significant (p?<?0.05) cholesterol and triglyceride lowering effect. Further, the aqueous extract at all doses demonstrated a significant (p?<?0.05) increase in fecal cholesterol excretion indicating a reduction in intestinal cholesterol absorption. Additionally, the activity of lipogenic enzymes like HMGCoA reductase in the liver remained significantly (p?<?0.05) low on treatment of aqueous extract (80?mg kg^?1), thus decreasing the cholesterogenesis. The aqueous extract (20, 40 and 80?mg kg^?1) also significantly (p?<?0.05) reduced the total lipid content in the liver. Moreover, the aqueous extract demonstrated a potential antioxidant capacity in DPPH and TBARS in vitro antioxidant assay. Thus the results suggest a beneficial role of aqueous extract of Plumbago zeylanica roots in ameliorating the hyperlipidemic condition leading to atherosclerosis.     

Influence of rosuvastatin on the NAD(P)H oxidase activity in the retina and electroretinographic response of spontaneously hypertensive rats

Background and purpose:

Retinal complications may be encountered during the development of hypertension as a response to oxidative stress. Statins may reduce the risk of developing hypertension and ocular diseases. We evaluate the effects of rosuvastatin (ROSU) on retinal functionality and oxidative stress levels in normotensive and spontaneously hypertensive rats (SHR).

Experimental approach:

Wistar Kyoto (WKY) and SHR were treated for 3 weeks with rosuvastatin (10 mg kg⁻¹ day⁻¹). Electroretinograms (ERG) were recorded before and after rosuvastatin treatment. Reactive oxygen species (ROS) were determined in the retina with dihydroethidium staining and NAD(P)H oxidase activity was evaluated.

Key results:

Retinal ganglion cell ROS and retinal NAD(P)H oxidase activity were higher in SHR than in WKY rats, respectively (17.1±1.1 vs 10.2±1.2 AU, P<0.01; 38095±8900 vs 14081±5820 RLU mg⁻¹; P<0.05). The ERG b-wave amplitude in SHR was significantly lower than that in WKY rats. Rosuvastatin reduced SBP in SHR but did not change plasma lipid levels. Rosuvastatin treatment in SHR significantly decreased ROS levels (11.2±1.3, P<0.01), NAD(P)H activity in retinal ganglion cells (9889±4290; P<0.05), and increased retinal plasmalogen content in SHR, but did not modify the ERG response.

Conclusions and implications:

Rosuvastatin, beyond lowering cholesterol levels, was able to lower ROS in the retina induced by hypertension, but without improving retinal function in SHR. These findings point to a complex relationship between ROS in the pathogenesis of retinal disease and hypertension.     

Intracellular signal transduction modulating expression of plasminogen activator inhibitor-1 in adipocytes

The concentrations in blood of plasminogen activator inhibitor-1 (PAI-1), an inhibitor of fibrinolysis and proteolysis, are elevated in obese and insulin-resistant subjects, predispose them to the risk of thrombosis, and may accelerate atherogenesis. Adipose tissue is a prominent source. Accordingly, intracellular signaling pathways that may influence PAI-1 expression in adipocytes have been the focus of considerable study. Rho, a small GTP binding and GTPase protein, when activated in turn activates its target, Rho-associated coiled-coil forming protein, to yield an active kinase, Rho-kinase, an effector in the Rho pathway. Rho-kinase exerts calcium-sensitizing effects in vascular smooth muscle cells and inhibitory effects on transforming growth factor-beta (TGF-beta) expression in chicken embryonic heart cells. Because TGF-beta is a powerful agonist of PAI-1 expression, we characterized the effects of inhibition of Rho-kinase in 3T3-L1 adipocytes. PAI-1 mRNA was determined by Northern blotting, and PAI-1 protein was determined by Western blotting. The Rho-kinase inhibitor, Y-27632 [(R)-(+)-trans-N-(4-pyridyl)-4-(1-aminoethyl)-cyclohexanecarboxamide], increased PAI-1 expression markedly. Although genistein, a flavonoid tyrosine kinase, attenuated the increase of PAI-1 induced by Y-27632, other non-flavonoid tyrosine kinase inhibitors did not. However, another flavonoid, daidzein, which lacks tyrosine kinase activity, decreased basal PAI-1 expression and attenuated the induction of PAI-1 expression by Y-27632. Thus, the Rho/Rho-kinase system inhibits PAI-1 expression by a flavonoid-sensitive mechanism in adipocytes. Therefore, flavonoids may be useful in decreasing elevated PAI-1 expression in adipose tissue and its consequent pathophysiologic sequelae.     

The inhibition of hepatic S-3-hydroxy-3-methylglutaryl-CoA reductase by 3,3,5-trimethylcyclohexanol and its mandelic acid ester,cyclandelate

Rat hepatic HMGCoA reductase was found to be at least 50% inhibited 17 hr after administration of a single oral dose of 3,3,5-trimethylcyclohexanyl mandelate (cyclandelate), a vasoactive substance. This inhibition was also found in rats given the 3,3,5-trimethylcyclohexanol component but only slight inhibition was seen after an equimolar dose of mandelate. The inhibition of HMGCoA was observed both around the high point and near the low point of the diurnal activity cycle. The effect did not persist to 41 hr after treatment. There was no direct inhibition of HMGCoA reductase by trimethylcyclohexanol when added to the assay system in vitro. The in vivo effect of these inhibitors was specific for HMGCoA reductase. There was no change, neither elevation nor depression, of the amount of microsomal membrane components cytochromes b₅ and P-450, not was the activity of another microsomal enzyme, arylesterase, affected by dosing with cyclandelate or trimethylcyclohexanol.