The effects of streptozotocin diabetes on hepatic triglyceride lipase activity in the rat

The function of the hepatic triglyceride lipase (H-TGL) is not yet clear. The purpose of the present study was to investigate the possible hormonal regulation of H-TGL. Postheparin plasma was obtained 3 min after the intravenous injection of 50 U/250 g body weight of heparin into male Wistar rats. The lipase activities were measured using substrate containing [¹⁴C] triolein emulsified with gum arabic and were expressed in μmoles of free fatty acid released/ml/hour (mean ± SD). H-TGL was the lipase activity remaining after inhibition of lipoprotein lipase (LPL) by 1.0 M NaCl. Diabetic rats were prepared by intravenous injection of streptozotocin (STZ), 65 mg/kg body weight. The contributions of H-TGL and LPL to the total plasma triacylglycerol hydrolase (TGH) activity depend on the amount of heparin injected and the time of blood withdrawal after heparin injection. H-TGL was maximally released at higher heparin (50 U/250 g body weight) concentrations, compared to LPL which was maximally released at lower heparin (5 U/250 g body weight) concentrations. H-TGL was significantly higher at 3 min after the injection of 50 U of heparin/250 g body weight than at 20 min. Twenty-four-hour fasting produced a significant fall in H-TGL compared to H-TGL in fed rats. Total TGH was significantly lower in diabetic rats 3 days after STZ injection. In diabetic rats 3, 5, and 7 days after STZ injection, H-TGL were significantly lower than those in control rats. H-TGL and H-TGL/total TGH were 9.49 ± 0.99 and 0.551 ± 0.071, respectively, in rats 3 days after STZ injection, compared to H-TGL (13.46 ± 0.69) and H-TGL/total TGH (0.739 ± 0.052) in control nondiabetic rats. When diabetic rats were treated with insulin, total TGH (14.37 ± 3.01) and H-TGL (6.77 ± 4.12) rose to 25.16 ± 1.02 (total TGH) and 16.46 ± 1.13 (H-TGL), that were comparable to activities in control nondiabetic rats. Separation of H-TGL and LPL was performed using heparin-Sepharose affinity chromatography of postheparin plasma. The enzyme activity of peak I from STZ rats, which is eluted by 0.72 M NaCl-Veronal buffer, pH 7.4 and corresponds to H-TGL, was approximately half the activity from control rats. TGH released by heparin from isolated rat liver parenchymal cells was investigated. The enzyme activities released from isolated liver parenchymal cells prepared from STZ rats was approximately half that from control rats. The role of insulin in the regulation of LPL has been well documented. Our findings suggest that H-TGL also is under hormonal regulation by insulin in rats.     

Testosterone substitution increases the activity of lipoprotein lipase and hepatic lipase in hypogonadal males

We studied the effects of testosterone substitution on serum concentrations of lipids, lipoproteins, apoproteins and on the activity of hepatic lipase (HL) and lipoprotein lipase (LPL) in postheparin plasma and on the activity of LPL in adipose tissue (AT-LPL) in 13 male hypopituitary patients. The activities of LPL and HL in postheparin plasma were markedly increased by 1 week after a testosterone enanthate injection (P less than 0.001). The HL activity remained elevated (P less than 0.05) after 1 month's treatment, but the LPL activity declined to presubstitution levels. The prolonged substitution decreased serum apoproteins A-I and A-II (P less than 0.05). The changes of apo A-I and A-II correlated inversely with those of the free testosterone index (FTI) (r = -0.74, r = -0.67, P less than 0.05). Serum HDL-cholesterol level decreased slightly by 1 week and it correlated inversely with the increase in testosterone and the FTI (r = -0.67, r = -0.85, P less than 0.05). The results suggest that testosterone increases the activity of both lipolytic enzymes in postheparin plasma. The effect on HL appears to be more persistent than that on LPL. The data support a role for androgens in the regulation of serum lipoprotein and HDL-cholesterol levels.     

The effect of thyroid dysfunction and fasting on placenta inner ring deiodinase activity in the rat

The placenta contains iodothyronine 5-deiodinase activity (P5-Dase) that probably acts on iodothyronines in the fetal circulation to convert T4 to rT3 and T3 to 3,3'-T2. Since thyroid status and fasting have profound effects on iodothyronine deiodinases in other tissues, the present studies were performed to determine if these perturbations affected P5-Dase. Control and treated rats were mated and killed near term on the 20th day of gestation. P5-Dase was determined in placenta homogenates enriched with dithiothreitol by measuring the conversion of T4 to rT3. In four of five studies, P5-Dase was similar in dams that underwent thyroidectomy (Tx) on day 7 of gestation and sham Tx dams. P5-Dase was not altered in dams that were treated with methimazole (MMI) to induce maternal and fetal hypothyroidism. Treatment of dams with supraphysiological doses of T4, beginning on the seventh day of gestation, did not significantly affect P5-Dase. In three of four studies, P5-Dase was similar in fed dams to values in dams fasted for the last 5 days of pregnancy. Placenta iodothyronine 5'-deiodinase activity (P5'-Dase) was also measured in some studies. P5'-Dase was not decreased in Tx rats and was modestly decreased in MMI-treated rats. However, the effect of MMI was not reversed by the administration of supraphysiological doses of T4, Tx, MMI treatment, and fasting all decreased hepatic T4 5'-deiodinase activity in pregnant rats. These results strongly suggest that thyroid status and fasting do not alter P5-Dase activity.     

The effect of Synacthen administration on lipoprotein lipase activity in the epididymal fat pad of the rat

Conflicting data have been reported on the influence of (excess) glucocorticoids on lipoprotein lipase (LPL) activity in adipose tissue. To solve this problem hypercorticism was induced in rats by treatment for varying periods with Synacthen, a synthetic corticotrophin-1-24 preparation, and LPL was measured in the epididymal fat pads using different methods. In extracts of defatted tissue preparations from overnight fasted rats treated for 3 days with Synacthen we observed an increase in LPL activity (acetone-ether powder LPL) to values similar to those found in normally fed controls. In contrast, the heparin-elutable part of LPL activity in the tissue was not influenced by the Synacthen treatment. This activity remained significantly lower in overnight fasted animals, Synacthen treated or not, than in normally fed rats. Adrenalectomy lowered the acetone-ether powder LPL activity of the epididymal adipose tissue in fasted as well as in fed rats. In fasted rats it prevented the stimulation of the LPL activity by Synacthen.     

A hepatic lipase gene promoter polymorphism attenuates the increase in hepatic lipase activity with increasing intra-abdominal fat in women

High hepatic lipase (HL) activity is associated with an atherogenic lipoprotein profile of small, dense LDL particles and lower HDL(2)-C. Intra-abdominal fat (IAF) is positively associated with HL activity. A hepatic lipase gene (LIPC) promoter variant (G-->A(-250)) is associated with lower HL activity, higher HDL(2)-C, and less dense LDL particles. To determine whether the LIPC promoter polymorphism acts independently of IAF to regulate HL, 57 healthy, premenopausal women were studied. The LIPC promoter A allele was associated with significantly lower HL activity (GA/AA=104+/-34 versus GG=145+/-57 nmoles x mL(-1) x min(-1), P=0.009). IAF was positively correlated with HL activity (r=0.431, P<0.001). Multivariate analysis revealed a strong relationship between both the LIPC promoter genotype (P=0. 001) and IAF (P<0.001) with HL activity. The relationship between IAF and HL activity for carriers and noncarriers of the A allele was curvilinear with the carriers having a lower apparent maximum level of plasma HL activity compared with noncarriers (138 versus 218 nmoles x mL(-1) x min(-1), P<0.001). In addition, the LIPC A allele was associated with a significantly higher HDL(2)-C (GA/AA=16+/-7 versus GG=11+/-5 mg/dL, P=0.003). We conclude that the LIPC promoter A allele attenuates the increase in HL activity due to IAF in premenopausal women.     

Triglyceride lipase activity in the diabetic rat heart

In diabetes, glucose oxidation is strongly suppressed and the energy demand of the isolated perfused beating heart is met by the oxidation of fatty acids delivered by lipolysis of stored triglycerides. Regulation of lipolysis and triglyceride lipase activity involved are, however, poorly understood. Lipolysis was therefore studied in isolated, perfused hearts from control and streptozotocin-diabetic rats (glycerol release) and triglyceride lipase determined in heart homogenates using [¹⁴C]triolein as substrate. (a) Triglyceride lipase in hearts from control and diabetic rats showed two optima of activity (pH 4.5 and 7.4). Only at neutral pH, glycerol formation and release of fatty acids from triolein agreed well. At acidic pH, fatty acids were released from triolein, but no glycerol formation could be observed. Together with differences in the intracellular distribution of triglyceride lipase, these data might indicate two different triglyceride lipases involved in myocardial lipolysis. (b) In hearts of acutely and chronically diabetic rats triglyceride lipases were reduced, at least partly restored by insulin therapy. (c) Rate of lipolysis in isolated, perfused hearts and triglyceride lipases were, however, strongly correlated with the amount of available triglycerides. (d) Epinephrine had a stimulating, insulin an inhibiting effect on lipolysis in isolated perfused hearts of control and diabetic rats. Whereas the stimulating effect of epinephrine on lipolysis could be inhibited by propranolol, neither propranolol, nicotinic acid, carbocromen nor reserpin could reduce the rate of lipolysis increased in diabetic hearts. (e) There was, however, no direct evidence for a hormonal regulation of triglyceride lipases by cAMP, epinephrine or glucagon in the absence or presence of proteinkinase. (f) Chloroquine did not inhibit lipolysis in isolated, perfused hearts of diabetic rats. Thus, the increased lipolysis in diabetic hearts might not be mediated by alterations in the hormonal regulation of triglyceride lipases, but might reflect more presumably changes in the metabolic regulation (increased substrate availability, oxidation capacity for fatty acids, carnitin). Not the lysosomal (acidic), but the neutral triglyceride lipase is involved in myocardial lipolysis of diabetic rats.     

Lipoprotein lipase and hepatic lipase activity after heparin administration in abetalipoproteinemia and hypobetalipoproteinemia

The purpose of this study was to examine whether an absence of triglyceride-rich lipoproteins (chylomicrons and very-low-density lipoproteins) in plasma is associated with any changes in the enzyme activity of lipoprotein lipase or hepatic lipase after heparin administration. To study this, the activities of hepatic lipase and lipoprotein lipase were determined in control subjects, in two patients with heterozygous hypobetalipoproteinemia, and in three patients with phenotypic abetalipoproteinemia after administration of heparin. Both enzymes showed normal activity in the patients with hypobetalipoproteinemia, but showed consistently reduced activity in the patients with abetalipoproteinemia. Hepatic lipase activity in plasma samples from these three patients obtained 15 minutes after intravenous injection of heparin was 55%, 87%, and 46% of that of the controls, whereas corresponding values in plasma samples obtained 30 minutes after heparin were 47%, 70%, and 57%, respectively. Lipoprotein lipase activity in the three patients with abetalipoproteinemia was 46%, 29%, and 34% of that of the controls in the samples obtained 15 minutes after heparin injection, whereas the values obtained after 30 minutes were 53%, 64%, and 47% of that of the controls. We conclude that an inherent absence of triglyceride-rich lipoproteins, as occurs in abetalipoproteinemia, is associated with reduced enzyme activity of both hepatic lipase and lipoprotein lipase in plasma after heparin administration.     

Comitogenic effect of catecholamines on rat cardiac fibroblasts in culture

OBJECTIVE: We studied the ability of norepinephrine and of other catecholamines to affect the proliferation of cardiac fibroblasts isolated from adult rat hearts. Furthermore, we investigated the possible adrenergic receptor involved in this process. METHODS: Norepinephrine (NE), phenylephrine (PE), isoproterenol (ISO), forskolin (FO), epidermal growth factor (EGF), platelet-derived growth factor AA (PDGF-AA) and specific inhibitors of the alpha(1)-, alpha(2)-, beta(1)- and beta(2)-adrenoceptors and of the protein kinase A (PKA) were applied to cardiac fibroblasts in culture. Cell number was measured by use of a Coulter Counter. Activation of the cAMP response element binding protein (CREB) was measured by Western blotting and subsequent use of a phospho-specific antibody. Activation of the p42- and the p44-mitogen activated protein kinase (p42/p44(MAPK)) was assessed by detection of phosphorylation shifts and by incorporation of 32P-labelled phosphate into myelin basic protein. RESULTS: Fibroblasts isolated from hearts of adult rats were grown in 10% serum-containing media which induced an increase in cell number by 94%. After 48 h, treatment with 10 microM NE caused an even greater increase in cell number by 222%, i.e. another 128% (comitogenic effect). In contrast, NE alone had no effect on the growth of serum-deprived cells. EGF and PDGF-AA did not replace serum as the basic mitogen. After addition of NE to proliferating cells under serum conditions, there was a rapid, time-dependent significant activation of the p42/p44(MAPK) and of CREB for up to 60 and 120 min, respectively. In both cases, the maximum of activation was reached after 5 min. Application of FO (0.1-20 microM) caused a strong activation of CREB, while no increase in the phosphorylation of the p42/p44(MAPK) was detected. Treatment with 20 microM FO led to an identical increase in cell number as application of NE. Specific blockade of PKA with RpcAMPS prevented the activation of CREB and also the comitogenic effect of FO as well as of NE. The alpha- and beta-adrenergic receptor blocker carvedilol (10 microM) normalized all NE-induced effects. Prazosin and yohimbine, inhibitors of alpha(1)- and alpha(2)-adrenoceptor activation, respectively, did not influence the NE-evoked increase in cell number. In contrast, the non-selective beta-adrenoceptor blocker propranolol (1 microM) completely suppressed the comitogenic effect of NE. A similar effect was obtained with the specific beta(2)-adrenoceptor blocker ICI 118,551 (5 microM), while the beta(1)-adrenoceptor blocker metoprolol did not influence the increase in cell number. CONCLUSIONS: NE elicits a comitogenic effect on cultured rat cardiac fibroblasts which is prevented by beta(2)-adrenergic blockade. The activation of CREB contributes to the increase in proliferation. The p42/p44(MAPK) which was also found to be activated by NE might as well be involved in the regulation of the comitogenic effect.     

Effect of lipoprotein lipase and hepatic triglyceride lipase activity on the distribution of apolipoprotein E among the plasma lipoproteins

The independent roles of human lipoprotein lipase (LPL) and hepatic triglyceride lipase (HTGL) in determining the distribution of apolipoprotein E (apo E) among the plasma lipoproteins has been studied in vitro. In one series of three studies, postheparin plasma (10%) was incubated for 2 h with autologous plasma and the changes in the lipoprotein association of apo E after lipase exposure were determined after lipoprotein fractionation on 4% agarose columns. Specificity for LPL or HTGL was achieved by inhibition with goat anti-human HTGL or with 1 M NaCl, respectively. In another study, LPL and HTGL were partially purified from human postheparin plasma. The independent effects of these enzymes on the lipoprotein association of apo E were then examined after incubation of plasma in the absence or presence of one or both lipases. Data from both types of in vitro study showed that LPL-mediated triglyceride hydrolysis in the absence of HTGL activity was accompanied by a loss of apo E from triglyceride-rich lipoproteins, a gain or no change in the apo E-containing lipoproteins the size of intermediate density lipoproteins (IDL) and inconsistent changes in the apo E mass associated with high density lipoproteins (HDL). HTGL activity, on the other hand, in the absence of LPL, resulted in a redistribution of apo E from lipoproteins the size of IDL and a gain by those of HDL size. These studies thus support previous in vivo studies which pointed toward a specific role for HTGL in the processing of apo E containing IDL.     

The effect of alcohol ingestion on hepatic aromatase activity and plasma steroid hormones in the rat

Chronic alcohol ingestion in the rat resulted in increased hepatic aromatase activity, elevation of plasma estradiol, and a decrease in plasma testosterone levels. Testicular incubation studies indicated that the source of the estrogen was not of gonadal origin but was, most likely, due to increased peripheral conversion. The failure of HCG in vitro to restore testicular secretion of testosterone to normal levels suggested a direct action of alcohol, or a metabolic product, on gonadal secretory processes, as distinct from trophic hormone effects. This study demonstrates that many of the hormonal alterations seen in cirrhosis of the liver in man may be produced directly by alcohol feeding without cirrhotic changes in the rat.     

The effect of ethanol on hepatic sodium plus potassium activated adenosine triphosphatase activity in the rat

We studied the acute an chronic effects of ethanol on hepatic Na+, K+-ATPase activity. Graded doses of ethanol given 1 h before death acutely increased Na+, K+-ATPase activity in a dose-related manner to a maximum of 138% of control with a simultaneous serum ethanol of 35.4 mM (163 mg%). Rats chronically fed 10% ethanol as their only fluid for 5 days had normal Na+, K+-ATPase activity, however, when ethanol was withdrawn 14 h before death Na+, K+-ATPase activity fell to 60% of control. The in vitro addition of ethanol to membrane preparations from untreated rats caused an increase in Na+, K+-ATPase activity with ethanol concentrations less than 81 mM with decreased activity at higher concentrations. In membranes from rats treated chronically with ethanol Na+, K+-ATPase activity did not respond to in vitro ethanol while the decreased activity noted after ethanol withdrawal was reversed by in vitro ethanol. These results suggest hepatocytes respond to the acute effects of ethanol by membrane adaptation which restores Na+, K+-ATPase activity to normal in the presence of ethanol. This adaptive response is probably due to changes in membrane lipids modulating membrane fluidity.     

Cloning of rat hepatic lipase cDNA: evidence for a lipase gene family.

Clones for rat hepatic lipase were isolated by probing a rat liver cDNA library in lambda gt11 with an oligonucleotide synthesized on the basis of a partial peptide sequence. The cloned messenger codes for a protein of 472 amino acids plus a hydrophobic leader sequence of 22 amino acids. The unglycosylated protein has a predicted molecular weight of 53,222 and contains two potential sites for N-glycosylation. The protein bears striking regions of homology with other known lipases and contains peptide sequences that have been implicated in lipid binding. The homologous mRNA is present in liver tissue but no detectable mRNA is observed in the adrenal gland, despite the reported presence of hepatic lipase in both the liver and the adrenal gland. No mRNA was seen in any of a variety of other tissues.     

实验性兔脂肪肝肝脂酶活性变化及茶多酚的作用

目的 观察肝组织中肝脂酶(HL)活性变化与肝细胞脂肪变性的关系,探讨茶多酚对HL活性变化的影响及机制。 方法 雄性新西兰白兔分为三组:脂肪肝组6只喂高脂饲料;茶多酚组7只喂高脂饲料,同时口服茶多酚200μg·g-1·d-1;正常对照组6只喂普通兔饲料。实验8周后测定血脂水平、血浆HL活性水平,肝组织做病理形态学检查同时用吐温法行HL活性染色,用硫代巴比妥酸比色法测定肝组织丙二醛(MDA)含量。 结果 脂肪肝组均产生重度混合型脂肪肝,血总胆固醇(TC)为(28.49±5.99)mmol/L,正常对照组为(1.11±0.82)mmol/L,t=21.654,P<0.05;血清低密度脂蛋白胆固醇(LDL-c)为(12.15±1.95)mmol/L,正常对照组为(0.71±1.14)mmol/L,t=12.400,P<0.05;肝组织MDA含量为(75.58±29.88)nmol/mg,正常对照组为(43.64±16.95)nmol/mg,t-2.278,P<0.05。每100μm2肝组织中HL活性点个数为1.76±0.10,正常对照组为4.14±0.05,t=3.306,P<0.05。茶多酚组:肝脏病理改变5只兔为中度混合型脂肪肝,2只兔为轻度混合型脂肪肝,明显轻于脂肪肝组。血TC为(16.87±6.58)mmol/L,明显低于脂肪肝组高于正常对照组(t值分别为5.786和3.968,P<0.05)。血LDL-c为(5.10±4.19)mmol/L,明显低于脂肪肝组(t=2.478,P<0.05)高于正常对照组(t=3.7634,P<0.05)。肝组织MDA含     

Comparison of myocardial and adipose tissue lipase activity in the rat

Effects of catecholamines and iodoacetamide on lipase activity (pH 6.8) and triglyceride mobilization were investigated in adipose tissue, myocardial slice and isolated perfused heart preparation of the rat. Epinephrine and isoproterenol did not enhance myocardial lipase activity in vitro and in perfused hearts. No detectable amount of fatty acid was released from myocardial slice by epinephrine in vitro. Epinephrine, however, induced an increase in lipase activity and in the amount of fatty acid released from the adipose tissue. The results suggest a difference in properties of lipase present in the myocardium and in the adipose tissue. Iodoacetamide decreased lipase activity in vitro both in the myocardium and in the adipose tissue. Triglyceride content in the perfused heart receiving iodoacetamide at 30 min of perfusion was higher than that of the control. However, there was no correlation between triglyceride mobilization and lipase activity in these 2 tissues.