Low-amplitude, low-frequency electric field-stimulated bone cell proliferation may in part be mediated by increased IGF-II release.

We have developed an in vitro model incorporating a low-amplitude (10(-7) V/cm), low frequency (f less than 100 Hz), capacitively coupled electric field in order to study the mechanism through which an electric field may increase bone cell proliferation. Utilizing this model we have previously shown that electric field-stimulated bone cell proliferation was dependent on release of mitogen activity into the culture medium from exposed cells. The current studies were intended to characterize this mitogen activity. In these studies we found that electric field-stimulated human bone cell proliferation was associated with increased IGF-II mRNA accumulation and IGF-II secretion suggesting that IGF-II may in part mediate the increase in bone cell proliferation following electric field exposure.     

Orchidectomy attenuates impaired endothelial effects of a high-salt diet in Sprague-Dawley rats

The effect of sex hormones on vascular reactivity is considered one of the underlying factors contributing to gender differences in cardiovascular functions and diseases. Experiments were designed to investigate the role of androgens in salt-induced hypertension by assessing the relaxation response of isolated aortic rings to acetylcholine and sodium nitroprusside in the presence or absence of l-nitroarginine methyl ester in Sprague-Dawley rats. The rats were either orchidectomized or sham-operated, with or without testosterone replacement, and were placed on a normal or high-salt diet for 6?weeks. The results indicate a significant increase (p?< 0.001) in the mean arterial blood pressure of rats on the high-salt diet, when compared with control or orchidectomized rats. Orchidectomy elicited a reduction in mean arterial blood pressure (p?< 0.01), while testosterone replacement normalized mean arterial blood pressure to values seen in intact rats on the high-salt diet. The high-salt diet reduced the relaxation response to acetylcholine both in the presence and absence of inhibition of endothelial nitric oxide synthase with l-nitroarginine methyl ester. Bilateral orchidectomy attenuated the impaired endothelial function induced by the high-salt diet in rats, but this was reversed by concomitant administration of testosterone, suggesting a role for androgens in enhancing long-term vascular smooth muscle tone and hence maintenance of high blood pressure in salt-induced hypertension.     

Neuronal-glial interactions in rats fed a ketogenic diet

Glucose is the preferred energy substrate for the adult brain. However, during periods of fasting and consumption of a high fat, low carbohydrate (ketogenic) diet, ketone bodies become major brain fuels. The present study was conducted to investigate how the ketogenic diet influences neuronal-glial interactions in amino acid neurotransmitter metabolism. Rats were kept on a standard or ketogenic diet. After 21 days all animals received an injection of [1-(13)C]glucose plus [1,2-(13)C]acetate, the preferential substrates of neurons and astrocytes, respectively. Extracts from cerebral cortex and plasma were analyzed by (13)C and (1)H nuclear magnetic resonance spectroscopy and HPLC. Increased amounts of valine, leucine and isoleucine and a decreased amount of glutamate were found in the brains of rats receiving the ketogenic diet. Glycolysis was decreased in ketotic rats compared with controls, evidenced by the reduced amounts of [3-(13)C]alanine and [3-(13)C]lactate. Additionally, neuronal oxidative metabolism of [1-(13)C]glucose was decreased in ketotic rats compared with controls, since amounts of [4-(13)C]glutamate and [4-(13)C]glutamine were lower than those of controls. Although the amount of glutamate from [1-(13)C]glucose was decreased, this was not the case for GABA, indicating that relatively more [4-(13)C]glutamate is converted to GABA. Astrocytic metabolism was increased in response to ketosis, shown by increased amounts of [4,5-(13)C]glutamine, [4,5-(13)C]glutamate, [1,2-(13)C]GABA and [3,4-(13)C]-/[1,2-(13)C]aspartate derived from [1,2-(13)C]acetate. The pyruvate carboxylation over dehydrogenation ratio for glutamine was increased in the ketotic animals compared to controls, giving further indication of increased astrocytic metabolism. Interestingly, pyruvate recycling was higher in glutamine than in glutamate in both groups of animals. An increase in this pathway was detected in glutamate in response to ketosis. The decreased glycolysis and oxidative metabolism of glucose as well as the increased astrocytic metabolism, may reflect adaptation of the brain to ketone bodies as major source of fuel.     

Fatty acid utilization by young Wistar rats fed a cafeteria diet

The content and accretion of fatty acids in 30, 45 and 60-day old Wistar rats fed either reference chow or a cafeteria diet has been studied, together with their actual fatty acid intake during that period. Diet had a small overall effect on the pattern of deposition of fatty acids, but the deposition of fat was much higher in cafeteria rats. The fat-rich cafeteria diet allowed the direct incorporation of most fatty acids into lipid storage, whilst chow-feeding activated lipogenesis and the deposition of a shorter chain and more saturated type of fatty acids. During the second month of the rat's life, the elongation pathway as well as ⁹-desaturase became functional, thus helping to shape the pattern of fatty acids actually accrued. The 60-day rats showed a relative impairment in the operation of ⁵-desaturase, since their lipids had a higher C20:4/C20:3 ratio than those of the diet ingested. Cafeteria-diet feeding minimized this effect since the large supply of dietary polyunsaturated fatty acids made the operation of the elongation-desaturase pathways practically unnecessary.     

Cell division in kidneys of rats fed a potassium-deficient diet

Mitoses is stimulated in the kidneys of adult rats fed a potassium-deficient diet. A statistically significant increase in mitotic figures appears first in the collecting ducts of the inner strip of the outer medulla after two days on the K+ deficient diet (P less than 0.05). After six days, mitosis also increases in the collecting ducts in the outer stripe and in the inner medulla (P less than 0.01). After eight days there is a significant rise in mitotic activity in the cells of the proximal convoluted tubule (P less than 0.01). There is a questionable increase in mitosis found only on the fifth day in the distal convoluted tubule. In all other cell types there is no statistically significant increase in cell division over the normal low levels that are observed in the cells of the controls rats.     

Effect of a perinatal high-salt diet on blood pressure control mechanisms in young Sprague-Dawley rats

In the present investigation we sought to determine if a perinatal high-salt treatment affects blood pressure at an early age (30 days), and if so, to determine the mechanisms responsible for the hypertension. Pregnant dams were given an 8% NaCl diet [high-salt (HS) rats] during the final one-third of gestation and throughout the suckling period. After weaning, the pups continued to receive the high-salt diet until testing at age 30 days. Control groups received a normal-salt diet (NS rats). In HS rats, mean arterial pressure (MAP) was significantly increased (110 +/- 5 vs. 96 +/- 3 mmHg) compared with NS rats. Blockade of brain AT(1) receptors with intracerebroventricular losartan decreased MAP in HS but not NS rats. Blockade of alpha-adrenergic receptors with intravenous phentolamine or ganglionic transmission with intravenous chlorisondamine produced a greater decrease in MAP in HS rats. Baroreflex control of heart rate was assessed using a four-parameter logistics function. The mid-range MAP (p3) was significantly increased in the HS rats. No other baroreflex parameters were affected. Specific binding of (125)I-[Sa (1),Ile(8)]ANG II to AT(1) receptors was increased in the subfornical organ (SFO) of the HS rats. Expression of AT(1a) receptor mRNA was greater in both SFO and PVN of the HS rats. These data suggest that even at an early age, Sprague-Dawley rats treated with a perinatal high-salt diet are hypertensive. The elevated blood pressure appears to be caused by increased sympathetic nervous activity, resulting, in part, from increased brain AT(1) receptor activation.     

Neuro-inflammation induced in the hippocampus of 'binge drinking' rats may be mediated by elevated extracellular glutamate content

The neuropathological and immune changes induced in the brain by 'binge drinking' have been investigated in a rat model. Evidence of neuro-inflammation was identified in the 'binge drinking' rat model of alcohol abuse after 3 weeks of administration of 2 or 3 g/kg ethanol (EtOH), three times per day for two consecutive days, followed by 5 days of abstinence: Firstly, alveolar macrophages, isolated from these animals, showed significant increases in inducible nitric oxide synthase, as assayed by nitrite release, both before and after lipopolysaccaharide stimulation. Secondly, significant numbers of activated microglia were present in the dentate gyrus region of the hippocampus of the 'binge drinking' model, after major histocompatibility complex class II staining, by comparison with the control. Microdialysis studies in the ventral hippocampus identified a significant increase in the basal extracellular concentration of glutamate, in both the 2 and 3 g/kg administered 'binge drinking' rats. In contrast, no changes in the hippocampal extracellular concentrations, of GABA and taurine, or the dopamine and serotonin metabolites were observed under basal conditions. A further dose of EtOH induced a significant decrease in the concentrations of both 3,4-dihydroxyphenylacetic acid and 5-hydroxyindoleacetic acid, whereas glutamate, taurine and GABA levels were unaffected. There was no evidence that EtOH preference was initiated by the 'binge drinking' regimen. Our results suggest that the possible toxicity associated with 'binge drinking' maybe directed by the elevated glutamate levels, which in turn, activate phagocytic cells to release their inflammatory cytokines and chemokines, ultimately leading to neuro-inflammation.     

Regulation of muscle protein degradation,not synthesis,by dietary leucine in rats fed a protein-deficient diet

The aim of this study was to elucidate the effects of long-term intake of leucine in dietary protein malnutrition on muscle protein synthesis and degradation. A reduction in muscle mass was suppressed by leucine-supplementation (1.5% leucine) in rats fed protein-free diet for 7 days. Furthermore, the rate of muscle protein degradation was decreased without an increase in muscle protein synthesis. In addition, to elucidate the mechanism involved in the suppressive effect of leucine, we measured the activities of degradation systems in muscle. Proteinase activity (calpain and proteasome) and ubiquitin ligase mRNA (Atrogin-1 and MuRF1) expression were not suppressed in animals fed a leucine-supplemented diet, whereas the autophagy marker, protein light chain 3 active form (LC3-II), expression was significantly decreased. These results suggest that the protein-free diet supplemented with leucine suppresses muscle protein degradation through inhibition of autophagy rather than protein synthesis.     

Iron status in rats fed a purified diet without vitamin A

The effect of vitamin A deficiency on iron status was investigated in rats. After 28 d of feeding either low or high vitamin A diets (0 vs 4000 IU of vitamin A per kg feed), the final body weight was slightly but significantly lowered by the low vitamin A diet. Plasma retinol concentrations were decreased in rats fed diets low in vitamin A. Marginal vitamin A deficiency produced slightly, but significantly lower blood hemoglobin concentrations; it did not clearly affect hematocrit. The concentration of iron in liver was significantly higher when diets low in vitamin A were fed while significantly lower levels were observed in femur.     

Excessive hepatic copper accumulation in jaundiced rats fed a high-copper diet

The response of copper metabolism to dietary copper challenge was investigated in jaundiced rats with elevated plasma concentrations of conjugated bilirubin as a result of impaired canicular transport of bilirubin glucuronides. Control and jaundiced rats were fed purified diets with either normal (64 μmol Cu/kg) or high (640 μmol Cu/kg) concentration of added copper. Copper loading produced a greater increase in hepatic copper concentrations in the jaundiced than in control rats. The greater dietary-copper-induced increase in hepatic copper in the jaundiced rats can be explained by the observed smaller rise in biliary copper excretion and a greater efficiency of dietary copper absorption. In individual rats, there was a positive relationship between hepatic copper concentrations and biliary copper concentrations. It is suggested that not the transport of copper from liver cells to bile but that from plasma to bile is diminished in the jaundiced rats. The elevated plasma copper concentrations in the jaundiced rats may support this suggestion.     

The effects of ox-LDL in human atherosclerosis may be mediated in part via the toll-like receptor 4 pathway

Toll-like receptor 4 (TLR4) may provide a potential pathophysiological link between lipids and infection/inflammation and atherosclerosis. Oxidized low-density lipoprotein (ox-LDL) makes it more atherogenic than its native form. The purpose of the study was to investigate the relationship between the effects of ox-LDL in human atherosclerosis and the expression of TLR4. We studied the relationship between TLR4 and ox-LDL, pro and con, using both real-time quantitative RT-PCR and RNA interference technology through in vitro cell culture. Nuclear factor kappa B (NF-κ B) activity and the concentrations of monocyte chemoattractant protein-1(MCP-1) and interleukin-8 (IL-8) were detected by ELISA. The results showed that the expression of TLR4 increased in response to ox-LDL. Simultaneously, NF-κ B relative activity and the concentrations of MCP-1 and IL-8 in cell supernatant were upregulated by ox-LDL in a dose-dependent manner. TLR4 expression was inhibited by small interference RNA(siRNA) plasmid expression vectors; NF-κ B activity and the secretions of MCP-1 and IL-8 in response to ox-LDL were significantly lower in the group whereinTLR4 expression has been inhibited than that in the group wherein TLR4 expression has not been inhibited. We suggest that the atherogenic effects of ox-LDL could be mediated in part via the TLR4 pathway. Furthermore, inhibition of TLR4 expression may downregulate the NF-κ B activity and secretions of MCP-1 and IL-8 in monocytes due to oxidized LDL, resulting in the alleviation of the progress of atherosclerosis.     

Differential sympathetic and angiotensinergic responses in rats submitted to low- or high-salt diet

The present study was designed to evaluate, in Wistar rats, the effect of high- or low-salt diet on the hemodynamic parameters and on the renal and lumbar sympathetic nerve activity. The renal gene expression of the renin angiotensin system components was also evaluated, aiming to find some correlation between salt intake, sodium homeostasis and blood pressure increase. Male Wistar rats received low (0.06% Na, TD 92141-Harlan Teklad), a normal (0.5% Na, TD 92140), or a high-salt diet (3.12% Na, TD 92142) from weaning to adulthood. Hemodynamic parameters such as cardiac output and total peripheral resistance, and the renal and lumbar sympathetic nerve activity were determined (n=45). Plasma renin activity, plasma and renal content of angiotensin (ANG) I and II, and the renal mRNA expression of angiotensinogen, renin, AT1 and AT2 receptors were also measured (n=24). Compared to normal- and low-salt diet-, high-salt-treated rats were hypertensive and developed an increase (P<0.05) in total peripheral resistance and lumbar sympathetic nerve activity. A decrease in renal renin and angiotensinogen-mRNAs and in plasma ANG II and plasma renin activity was also found in salt overloaded animals. The renal sympathetic nerve activity was higher (P<0.05) in low- compared to high-salt-treated rats, and was associated with an increase (P<0.05) in renal ANG I and II and with a decrease (P<0.05) in AT2 renal mRNA. Plasma ANG I and II and plasma renin activity were higher in low- than in normal-salt rats. Our results show that increased blood pressure is associated with increases in lumbar sympathetic nerve activity and total peripheral resistance in high-salt-treated rats. However, in low-salt-treated rats an increase in the renal sympathetic nerve was correlated with an increase in the renal content of ANG I and II and with a decrease in AT2 renal mRNA. These changes are probably in favor of the antinatriuretic response and the sodium homeostasis in the low-salt group.