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The initiation of atherosclerosis involves up-regulation of molecules such as E-selectin, VCAM-1, and ICAM-1. The progression of atherosclerosis is linked to proliferation and migration of vascular smooth muscle cell via MMP-2 and MMP-9 activities. However, the etiology of atherosclerosis concerning plasticizers is unknown. We evaluated β-thujaplicin in preventing the development of atherosclerosis in a model induced by pro-inflammatory cytokines. Moreover, we established a new atherosclerosis model in vascular smooth muscle cells (VSMC) exposed to a common contact plasticizer, di(2-ethylhexyl)phthalate (DEHP). SEVC4-10 endothelial cells were treated with 50% RAW conditioned medium and A7r5 VSMC was treated with the plasticizer, with/without β-thujaplicin (4 or 12 μM). Production of E-selectin, ICAM-1, and VCAM-1 in SEVC4-10 cells as well as MMP-2/MMP-9 (both expression and activity) in VSMC were monitored. Results showed that the conditioned medium induced E-selectin and ICAM were significantly prevented by β-thujaplicin. However, inhibition on the production of VCAM by β-thujaplicin was only seen in a concentration of 12 μM. Both concentrations of β-thujaplicin also significantly prevented DEHP-induced MMP-2 and MMP-9 expression and activities. Evidence uncovers that β-thujaplicin has additional factors in amelioration of atherosclerosis and corroborates that β-thujaplicin is a strong candidate in preventing the initiation and progression of atherosclerosis.

Proliferation and migration of vascular smooth muscle cells (VSMC) are important in the development and/or progression of many cardiovascular diseases, including atherosclerosis. Evidence shows that matrix metalloproteinase (MMP)-2 and MMP-9 are related to the pathogenesis of atherosclerosis. The expressions of MMP-2 and MMP-9 in atherosclerosis are regulated via various pathways, such as p38 mitogen activated protein kinase (MAPK), extracellular signal regulated kinase 1 and 2 (ERK1/2), Akt, and nuclear factor kappa (NF-κB). Di(2-ethylhexyl) phthalate (DEHP) has been shown to induce atherosclerosis by increasing tumor necrosis factor (TNF)-α, interleukin (IL)-6, and intercellular adhesion molecule (ICAM) productions. However, whether DEHP poses any effects on MMP-2 or MMP-9 expression in VSMC has not yet been answered. In our studies, rat aorta VSMC was treated with DEHP (between 2 and 17.5 ppm) and p38 MAPK, ERK1/2, Akt, NF-κB, and MMP-2 and MMP-9 proteins and activities were measured. Results showed that the presence of DEHP can induce higher MMP-2 and MMP-9 expression than the controls. Similar results on MMP-regulating proteins, i.e., p38 MAPK, ERK1/2, Akt, and NF-κB, were also observed. In summary, our current results have showed that DEHP can be a potent inducer of atherosclerosis by increasing MMP-2 and MMP-9 expression at least through the regulations of p38 MAPK, ERK1/2, Akt, and NF-κB.

Chlorella, a type of unicellular fresh water algae, has been a popular foodstuff in Japan and Taiwan. Studies have shown the hypoglycemic effects of Chlorella in alloxan-induced and Streptozocin (STZ)-induced diabetic animals. However, the mechanisms by which Chlorella treatment affects blood glucose homeostasis have not been studied. Diabetes in ICR mice was induced by injection of STZ. Lipogenesis in vivo was measured by incorporating 3H-H2O into lipids in brown and white adipose tissues. Glucose uptake in the liver and soleus muscles was measured by assaying 2-deoxy-D-[1,2-3H] glucose levels. The effects of Chlorella on serum non-esterified fatty acids (NEFA) were measured with commercial assay kits. Insulin-stimulated lipogenic rates in brown and white adipose tissues were unaffected by Chlorella. However, Chlorella increased 2-deoxyglucose uptake in the livers and soleus muscles in normal and STZ mice compared to that in their respective controls (p < 0.01). In addition, fasting NEFA levels were lower in Chlorella-treated STZ mice compared to H2O-treated STZ mice (p < 0.005). The current results suggest that the hypoglycemic effects of Chlorella are due to an enhancement of glucose uptake in the liver and in soleus muscles. The improved insulin sensitivity after Chlorella treatment could be also due to lower NEFA levels, since insulin sensitivity is usually blunted by elevated NEFA in diabetes.

The effects of Chlorella pyrenoidosa on serum lipid profiles, after concomitant long-term treatment of high-fat diet (HFD) in rats and hamsters was studied. Wistar rats and Syrian hamsters were fed with or without various concentrations of Chlorella pyrenoidosa contained high-fat diet (CHFD) for 2, 4 and 8 weeks prior to assay of serum lipids. Fasting triglycerides, total cholesterol, and LDL cholesterol as well as HDL cholesterol levels in high-fat diet treated rats and hamster were determined. Results showed that triglycerides, total cholesterol and LDL cholesterol levels in HFD treated rats and hamsters were increased from the normal rodent diet (NRD) treated controls after 2, 4, and 8-week treatments. However, the presence of Chlorella pyrenoidosa in high-fat diets significantly decreased the levels of triglycerides, total cholesterol and LDL cholesterol with comparison to HFD group in rats and hamsters. The total cholesterol/HDL ratios, an indication of occurrence of coronary heart disease, were decreased in all CHFD treated grouped rats and hamsters which suggests administration of Chlorella pyrenoidosa could lower the occurring risk of heart diseases. In conclusion, Chlorella pyrenoidosa has the ability to prevent dyslipidemia in chronic high-fat fed animals and could be potential in use to prevent intestinal absorption of redundant lipid from our daily intake and subsequently to prevent hyperlipidemia as well as atherosclerosis.

Dexamethasone (DXM) is known as an immunosuppressive drug used for inflammation control. In the present study, we attempted to examine whether DXM administration could attenuate the hypercoagulable state and the overproduction of pro-inflammatory cytokines, improve arterial hypotension, cerebral ischemia and damage, and vital organ failure in a rat model of heat stroke. The results indicated that all the rats suffering from heat stroke showed high serum levels of tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β), accompanied with increased prothrombin time, activated partial thromboplastin time and D-D dimer, and decreased protein C. During the induction period of heat stroke, plasma levels of blood urea nitrogen (BUN), creatinine, glutamic oxaloacetic transaminase (SGOT), glutamic pyruvic transaminase (SGPT), and alkaline phosphatase (ALP), were consistently increased. High striatal levels of glycerol, glutamate, and lactate/pyruvate were simultaneously detected. On the contrary, the mean arterial pressure, plasma levels of interleukin-10 (IL-10), and local cerebral blood flow at the striatum were all decreased. Importantly, intravenous administration of DXM substantially ameliorated the circulatory dysfunction, systematic inflammation, hypercoagulable state, cerebral ischemia and damage during the induction period of heat stroke. These findings demonstrated that DXM may be an alternative therapy that can ameliorate heat stroke victims by attenuating activated coagulation, systemic inflammation, and vital organ ischemia/injury during heat stroke.

Untreated (control) obese CBA mice had lower hormone-sensitive lipase (HSL) activity and cAMP levels in brown adipose tissue than normal lean mice, but white adipose tissue HSL activity and cAMP were similar in obese and lean mice. In the obese mice, chronic ethanol treatment increased HSL activity and cAMP levels in both brown and white adipose tissue to above the levels in lean mice. In the lean mice, chronic ethanol only stimulated white adipose tissue. UK 14304 [5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline: 2 mg/kg] inhibited HSL activity in both brown and white adipose tissues in lean mice, but a higher dose (3 mg/kg) was required to produce the same inhibition in obese mice. After chronic ethanol adipose tissues were more sensitive to UK 14304; only half the dose being required to produce the same level of lipase inhibition. We propose that, although chronic ethanol consumption increases cAMP levels in adipose tissue, particularly in obese mice, it also sensitizes the tissues to alpha(2)-adrenoceptor stimulation. These effects may explain the increased sympathetic nervous system activity observed in alcohol withdrawal.

Acute S 22068 (24 mg/kg po) improved glucose tolerance and increased insulin sensitivity, assessed as the acute blood glucose response to exogenous insulin. The same acute dose did not stimulate insulin secretion or induce hypoglycemia in fed animals. Comparison of acute S 22068 to equipotent doses (with respect to effect on glucose tolerance) of gliclazide (2 mg/kg) and metformin (60 mg/kg) found S 22068 to be similar to metformin with respect to its effects on basal glucose levels (BGL) and insulin sensitivity. This also suggests that S 22068 acts by a mechanism which does not involve insulin release. Acute or sub-chronic S 22068 (14 days at 25 mg/day) had no effect on brown adipose tissue (BAT) or white adipose tissue (WAT) lipogenesis, an insulin-sensitive metabolic pathway. Sub-chronic treatment with S 22068 did not alter body weight (BW) or food intake, and resulted in tolerance to its effects on glucose metabolism and insulin sensitivity. These findings suggest that S 22068 is similar in effect to metformin, and is not insulinogenic, in contrast to the sulfonylureas or putative I 3 imidazoline site ligands.

Acute SR 58611A (0.25 mg kg−1), was effective in reducing the blood glucose response to a glucose tolerance test (GTT) in normal lean (control) and spontaneously obese/diabetic CBA/Ca mice and to be equipotent to 1.25 mg kg−1 glibenclamide in lean mice. Neither brown (BAT) nor white (WAT) adipose tissue lipogenesis was altered by acute SR 58611A (2–8 mg kg−1) in lean mice, but both increased significantly at the higher doses in the obese mice. Acute SR 58611A produced a hypoglycaemia 40 min after dosing in lean and obese animals, the duration and potency of which was less than that of glibenclamide. Plasma insulin levels increased 20 min after acute SR 58611A and glibenclamide in lean and obese mice. Chronic treatment (0.25 mg kg−1, 15 days) with SR 58611A increased its effectiveness in improving glucose tolerance, but did not affect the body weight (BW) or food intake of either lean or obese mice. Acute and chronic SR 58611A prolonged the hypoglycaemic effect of exogenous insulin in lean but not obese mice. In fed and fasted lean mice and in fasted obese mice chronic SR 58611A produced an acute hypoglycaemia 30 min post administration which was greater than after a single dose. SR 58611A maintained its effectiveness in improving glucose tolerance in lean and obese mice over a dosing period of 15 days. The improvement in glucose tolerance was achieved at a dose less than that required to stimulate adipose tissue lipogenesis and which did not affect food intake or body weight. British Journal of Pharmacology (1999) 128, 1586–1592; doi:10.1038/sj.bjp.0702946

Diurnal variation in blood and plasma ethanol levels (BACs) has been observed in animals undergoing chronic ethanol treatment, but the information available is insufficient to determine whether the different patterns seen are due to differences in ethanol administration schedules or to strain of the animal. In this study, we have compared plasma ethanol levels in males of two mouse strains with no innate preference for ethanol, TO and CBA, during two commonly employed chronic ethanol treatment schedules. Ethanol was administered in solution as sole drink (CED) (10% or 20% w/v ethanol) for 4 weeks, or in liquid diet form (ELD), (3.5% w/v ethanol for 2 days, then 7% for 5 days). Mice were housed eight per cage on a 12-h light cycle (0900-2100 hours). Plasma ethanol concentration was monitored over the 24-h period. Activity of liver alcohol dehydrogenase (ADH) was measured between 0900 and 1100 hours. CBA mice showed greater variability in body weights than TO mice, which weighed more throughout the period of study and had significantly higher total energy intakes. TO mice consumed more ELD than CBA mice. Following an initial 2-day period of 3.5% ELD, both strains decreased their diet intake when ethanol content of the diet was increased to 7% w/v, which resulted in weight loss. Mice on the CED schedules decreased their fluid intake with increasing concentration of ethanol in the drinking solution. Highest daily ethanol intakes were observed in mice on ELD (19.1 +/- 1.7 and 22.2 +/- 0.6 g/kg body weight in CBA and TO mice, respectively). Marked diurnal variation in plasma ethanol levels was observed, which was dependent on the treatment schedule, strain and method of ethanol administration. Highest levels were found in mice on the ELD schedule (104.8 +/- 7.7 mM in CBA mice, 113.5 +/- 14.5 mM in TO mice), peaking at 1900 and at 0900 hours in CBA and TO mice, respectively. Lower plasma ethanol concentrations were reached in mice on the CED schedules, peaking at midnight (34.6 +/- 8.1 mM and 35.4 +/- 8.8 mM in CBA and TO mice on 20% CED, respectively, and 3.7 +/- 1.2 mM and 6.6 +/- 2.1 mM in CBA and TO mice on 10% CED). Naive CBA mice had slightly higher liver ADH activity as compared to their TO counterparts. No effect of 10% CED on liver ADH activity was found in either mouse strain. In conclusion, we have confirmed the importance of monitoring plasma ethanol levels during chronic treatment, as there is marked diurnal variation, dependent on the light/dark cycle. Factors such as strain of the animal and the method of delivery of ethanol are also important, whereas liver ADH plays a minor role. Monitoring the daily ethanol consumption is insufficient to predict the resulting plasma levels of the drug.

Acute injection of either noradrenaline or isoprenaline in mice activated both brown (BAT) and white (WAT) adipose tissue hormone-sensitive lipase activity (HSL). Dose-response studies indicated that isoprenaline (0.05-0.15 mg/kg) produced a dose-dependent activation of HSL in both BAT and WAT, whereas SR 58611A produced no change in HSL in WAT over a dose range (1-5 mg/kg) which, at the same time, dose-dependently increased HSL activity in BAT. The other beta 3-adrenoceptor agonists, ZD 7114 (10 mg/kg) and BRL 35135 A (5 mg/kg) also selectively increased HSL activity in BAT, these doses having previously been shown to stimulate lipogenesis in vivo. Higher doses of ZD 7114 and BRL 35135 produced no further increase in HSL activity and, in the case of BRL 35135, provoked symptoms of non-selective beta-adrenoceptor activation. The increase in HSL activity could be prevented by pretreating the mice with propranolol, 10 mg/kg, i.p., 30 min prior to the agonist. The activation of HSL activity by the beta 3-adrenoceptor agonists was associated with an increase in tissue cAMP production which was also prevented by pretreatment with propranolol. The degree of cAMP accumulation was least with BRL 35135 and greatest with ZD 7114. We conclude that, in the mouse adipocyte, the atypical beta-adrenoceptor (beta 3) is present in BAT, but is not present or functional in WAT.

The time course of the effects of ethanol withdrawal on brown and white adipose tissue hormone-sensitive lipase, cAMP production, and phosphodiesterase have been investigated after chronic drinking or liquid diet schedules. Chronic drinking significantly reduced brown adipose tissue hormone-sensitive lipase activity and cAMP levels from control. During withdrawal, there was a rebound increase to 200% control, peaking 9 h into withdrawal. White adipose tissue hormone-sensitive lipase activity and cAMP accumulation were significantly raised by both treatment schedules. Ethanol liquid diet produced a significant fall in adipose tissue hormone-sensitive lipase activity and cAMP accumulation. In brown fat, there was a rebound increase in hormone-sensitive lipase activity and cAMP; in white fat, no rebound was observed. In brown fat, the reductions in hormone-sensitive lipase activity and cAMP accumulation after chronic drinking coincided with an increase in phosphodiesterase activity. In white fat, the rise in cAMP and hormone-sensitive lipase activation coincided with a decrease in phosphodiesterase activity. We conclude that the effects of chronic ethanol on hormone-sensitive lipase activity are cAMP-dependent and mediated via alterations in phosphodiesterase activity.

1. The effects of two chronic ethanol treatment schedules, which produce different plasma ethanol concentrations, on the specific activities of adipose tissue lipoprotein lipase (LPL) and hormone-sensitive lipase (HSL) have been investigated in brown and white fat. 2. Mice provided with 20% ethanol solution as sole drinking fluid for 28 days consumed between 13 and 15 g ethanol kg-1 body weight day-1 over days 22-28. The mean plasma ethanol concentration was 4.94 +/- 1.4 mM (n = 8) at 09 h 00 min on day 28 when the lipase assays were performed. Mice given ethanol in a liquid diet for 7 days consumed between 15 and 18 g ethanol day-1 over days 3-7. The mean plasma ethanol concentration was 15.9 +/- 4.7 mM (n = 8) at 09 h 00 min on day 7. These concentrations of ethanol had no effect on the activity of either LPL or HSL in vitro. 3. LPL activity in white and brown fat (expressed as nmol fatty acids released h-1 mg-1 acetone powder) was unaltered 60 min following an acute injection of ethanol (2.5 g kg-1, i.p.) which produced a mean blood ethanol level of 37.5 +/- 6.7 mM. HSL activity in white fat (expressed as nmol fatty acid released h-1 mg-1 protein) was also unaffected by this acute dose of ethanol, but the activity in brown fat was significantly reduced: 3.07 +/- 0.30 (n = 8) after ethanol compared to 4.36 +/- 0.25 (n = 12) in controls (P0.01). 4. LPL activity in white fat was little altered by either of the chronic ethanol treatment schedules whilst LPL activity in the brown fat from the same animals was significantly increased compared to the respective control values: 0.27 +/- 0.03 (ethanol drinking), control: 0.16 +/- 0.01; 0.79 +/- 0.14 (ethanol liquid diet), control: 0.39 +/- 0.05. 5. HSL activity in white fat was significantly increased by the chronic drinking treatment (7.7 +/- 0.5; control: 3.78 +/- 0.17, n = 8) at the same time that the activity in brown fat was reduced (3.76 +/- 0.2; control: 4.74 +/- 0.16). The ethanol liquid diet also reduced HSL activity in brown fat but had negligible effect in white fat. 6. The effects of the two chronic ethanol treatments on adenosine 3':5'-cyclic monophosphate (cyclic AMP) accumulation in brown and white fat were very similar, both qualitatively and quantitatively, to the effects on HSL. 7. It has been shown that brown and white adipose tissues respond differently to the presence of chronic ethanol and that the response is dependent both upon the concentration of ethanol and the nature of the diet with which the ethanol is administered. The effects of ethanol on adipose tissue HSL activity appear to be mediated via changes in the tissue cyclic AMP level and, in this respect, brown fat is more sensitive to ethanol than white fat.

The effects of a chronic ethanol drinking schedule (20% solution for 6 weeks) on energy balance and carbohydrate and lipid metabolism have been investigated in lean (32-36 g) and obese-diabetic (40-44 g) CBA/Ca mice. The untreated obese-diabetic mice exhibited hyperglycaemia, hypertriglyceridaemia, hyper-insulinaemia and insulin resistance. The chronic ethanol treatment, which yielded plasma ethanol levels of between 1 and 11 mM, lowered the blood glucose, plasma insulin and triacylglycerol levels towards normal in the obese mice, but did not affect these parameters in the lean mice. The body weight of the obese mice tended to return to normal during the 6-week drinking period, although their total energy intake (9.2-10.0 kJ/g/week, food plus ethanol-derived calories) was almost double that of the lean mice (4.8-5.4 kJ/g/week). The blood glucose response to acute insulin injection, which was significantly reduced in the obese mice, became indistinguishable from the response of normal mice after chronic ethanol treatment. Soleus muscle glycogen synthesis in both lean and obese mice was not significantly altered by ethanol drinking, but brown adipose tissue lipogenesis was significantly increased (by 50%) in the obese mice. It is proposed that ethanol is acting chronically to restore insulin sensitivity in the obese diabetic mice at doses which have little or no effect in normal lean animals. This action is exerted, at least in part, at the level of brown adipose tissue lipogenesis.