Nutrient-gene interactions in mitochondrial function: vitamin A needs are increased in BHE/Cdb rats

The BHE/Cdb rat has a maternally inherited mutation in the ATPase 6 mitochondrial gene that associates with impaired oxidative phosphorylation (OXPHOS) and glucose intolerance. A longevity study revealed that feeding an egg-rich (vitamin A-rich) diet delayed the onset of impaired glucose tolerance. Two experiments were conducted to test the hypothesis that BHE/Cdb rats require more dietary vitamin A than normal rats. Experiment 1 was a dose-response study examining OXPHOS in BHE/Cdb rats fed one of six levels of vitamin A. In experiment 2 BHE/Cdb and Sprague-Dawley rats were used. The rats were depleted of retinol stores, then repleted with 4 or 12 IU vitamin A/g diet. Vitamin A status was assessed in depleted, never depleted, and depleted/repleted rats. OXPHOS was optimized at 4 IU/g diet for the Sprague-Dawley rats and 12 IU/g diet for the BHE/Cdb rats. These results suggested that the criteria for vitamin intake adequacy in the BHE/Cdb rats is the optimization of mitochondrial OXPHOS. Using this criteria, we conclude that diabetes-prone BHE/Cdb rats require more dietary vitamin A than normal rats.     

Attenuation of circadian rhythms of food intake and respiration in aging diabetes-prone BHE/Cdb rats

The hypothesis that BHE/Cdb rats with mutations in their mitochondrial genome might accommodate this mutation by changing their food intake patterns was tested. Four experiments were conducted. Experiments 1 and 2 examined food intake patterns of BHE/Cdb rats fed a stock diet or BHE/Cdb and Sprague-Dawley rats fed a high-fat diet from weaning. Experiment 3 examined the daily rhythms of respiration and heat production in these rats at 200 days of age. Experiment 4 examined the effects of diet composition on these measurements at 50-day intervals. The Sprague-Dawley rats, regardless of diet, had the typical day-night rhythms of feeding and respiration. In contrast, the BHE/Cdb rats fed the high-fat diet showed normal rhythms initially, but with age, these rhythms were attenuated. The changes in rhythms preceded the development of glucose intolerance.     

Further studies of diabetes-prone BHE/Cdb rats: increased sensitivity to calcium ion suppression of oxidative phosphorylation

The responsiveness of hepatic mitochondria isolated from hyperthyroid and control Sprague-Dawley (SD) and diabetes-prone BHE/Cdb rats was studied. Hyperthyroidism was induced through the addition of thyroxine (T(4)) to the diet (2 mg/kg of diet). Oxidative phosphorylation (OXPHOS) with the addition of adenosine diphosphate (ADP) or an 8:1 mixture of adenosine monophosphate (AMP):ADP was studied. Dose response curves of state 3 and state 4 respiration, respiratory control (RC) ratio, and ADP:O ratio to calcium levels (0-7.5 microm) were generated. Mitochondria from BHE/Cdb rats were more sensitive to the addition of calcium than mitochondria from SD rats, as judged by losses in OXPHOS. T(4) treatment potentiated this strain difference and we conclude that the diabetes phenotype in the BHE/Cdb rat is probably related not only to the previously described mutation in the F(O)ATPase but also to a defect in the efflux of the calcium ion that, in turn, affected the regulation of OXPHOS.     

Comparative studies of the responses of two strains of rats to an essential fatty acid deficient diet

A comparative study of two strains of rats to an EFA deficient diet was conducted. Parameters of insulin status in BHE and Sprague-Dawley rats were measured. No differences in growth were observed. The strains differed in their hepatic and adipose tissue response to insulin stimulation of glucose oxidation and conversion to fatty acids. Hepatic tissue from EFA deficient BHE rats converted more glucose to fatty acid under the influence of insulin than their controls while diet had no effect on glucose oxidation. Hepatic tissue from EFA deficient Sprague-Dawley rats oxidized more glucose than their controls but diet did not affect fatty acid synthesis. A reverse of these strain and diet differences was observed in adipose tissue. These results suggest that the genetic heritage of the rat may determine the type of response to EFA deficiency.     

Comparative studies on mitochondrial respiration in four strains of rats (Rattus norvegicus)

Mitochondrial respiration and ATP synthesis were examined in young rats of the Sprague-Dawley, Wistar, BHE and Zucker strains. Both lean and obese Zucker rats were studied. Pyruvate-supported state 3 respiration was highest in mitochondria from Sprague-Dawley rats and least in mitochondria from obese Zucker rats. Succinate-supported state 3 respiration was highest in the Wistar group and least in the Sprague-Dawley rats. There appeared to be no relationship between oxygen consumption and the genetic tendency for hepatic hyperlipogenesis. ATP synthesis was greatest in the obese Zucker rats and least in the Sprague-Dawley rats. Differences in liver weights and mitochondrial yields may explain, in part, these observed strain differences in mitochondrial activity.     

Ultraviolet irradiation of human skin causes functional vitamin A deficiency, preventable by all-trans retinoic acid pre-treatment

We report here that ultraviolet irradiation substantially reduced the mRNA and protein of the two major nuclear retinoid receptors, RAR-gamma and RXR-alpha, in human skin in vivo. Pre-treatment with retinoic acid mitigated this loss of nuclear retinoid receptors. Ultraviolet irradiation caused a near-total loss of retinoic acid induction of two RAR/RXR target genes, cellular retinoic acid binding protein-II and RA 4-hydroxylase, but did not affect 1,25-dihydroxyvitamin D3 induction of the vitamin D receptor/RXR-regulated gene vitamin D 24-hydroxylase. In effect, ultraviolet irradiation causes a functional vitamin A deficiency that may have deleterious effects on skin function, contributing to skin photo-aging and carcinogenesis.     

Studies of 5'-deiodinase activity in rats differing in hepatic lipogenic activity

Studies of the relationship of hepatic 5'-deiodinase activity to hepatic lipogenic capacity were conducted. Rats of the Zucker, BHE, and Sprague-Dawley strains were used. BHE and Sprague-Dawley rats were starved and refed a 65% glucose diet, whereas lean and obese Zucker rats were fed a stock diet; the rats were thus different in hepatic lipogenic capacity. After hepatic 5'-deiodinase activity was determined, we found that rats genetically predisposed to increased hepatic lipogenesis had less deiodinase activity than rats without this genetic feature. The role of the interaction between the thyroid hormones and glucocorticoid in the activity of hepatic deiodinase was also studied. Adrenalectomized (ADX) or intact BHE and Sprague-Dawley rats were injected with saline, thyroxine, or triiodothyronine and either saline or glucocorticoid. The normal Sprague-Dawley rats made predictable adjustments to their deiodinase activity when their hormonal status was manipulated, whereas the BHE rats responded as though these manipulations were corrections rather than additions or deletions.     

Endothelial cell specific molecule-1--a newly identified protein in adipocytes.

Expression of the endothelial cell-specific molecule (ESM)-1 was originally identified in lung and kidney endothelial cells, where its expression is regulated by cytokines. In vitro, ESM-1 interferes with the molecular mechanisms of immune cell migration by binding to adhesion molecules. In this study, we have explored the expression of ESM-1 in isolated human adipocytes and in rat adipose tissue depots. Human primary adipocytes were cultivated after collagenase digestion and used for in vitro incubation studies. Adipocytes were also isolated from different fat depots of Sprague-Dawley rats. Gene expression was quantified by TaqMan RT-PCR using specific human and rat ESM-1 primers. The cellular localisation of ESM-1 was determined by confocal microscopy using a specific antibody. ESM-1 expression in human adipocytes was stimulated by phorbol ester, an activator of protein kinase C, and by retinoic acid, an activator of nuclear receptors. The maximum increase in gene expression was 3.2-fold after 72 h treatment with phorbol ester and 4.6-fold after 72 h treatment with retinoic acid. The highest expression was found in subcutaneous rat adipose tissue - two-fold compared to epididymal and six-fold compared to intrascapular brown adipose tissue. As obesity is related to systemic inflammation (examplified by increased circulating levels of C-reactive protein and interleukin-6), the formation of ESM-1 in adipocytes and its activation by protein kinase C may play a role in the regulation of inflammatory processes.     

Histological evaluation of rat mammary tumours after treatment with retinoic acid analogues — phytol, TTNPB and vitamin D3 analogue seocalcitol

The effects of retinoic acid analogue — phytol (precursor of retinoic X receptors ligand), TTNPB (retinoic acid receptor agonist) and seocalcitol (EB1089, analogue of vitamin D₃) in mammary tumours of Sprague-Dawley rats induced with 1-methyl-1-nitrosourea (MNU) were investigated. Treatment with phytol, TTNPB and seocalcitol may have some protective and therapeutical effects on malignant processes. Treatment with these components in combination of TTNPB and phytol or seocalcitol and phytol inhibited progression of MNU-induced tumours of the rat mammary gland, and also induced decrease of tumour burden and volume in comparison with treated control group. Treatment of rats with the above compounds had no effect on malignity and invasiveness of carcinomas.     

Effects of retinoic acid on the concentrations of radioactive metabolites of retinol in tissues of rats maintained on a retinol-deficient diet

The effects of feeding retinoic acid for 2 and 6 days on the metabolism of labeled retinol in tissues of rats maintained on a vitamin A deficient diet was studied. The metabolites of retinol were analyzed by high performance liquid chromatography. Feeding retinoic acid for 2 days significantly reduced the blood retinol and retinyl ester levels without affecting the vitamin A content of the liver. In intestine and testis the content of labeled retinoic acid was decreased significantly by dietary retinoic acid. Addition of retinoic acid to the diet for 6 days resulted, in addition to decreased blood retinol and retinyl ester values, in an increase in the retinyl ester values in the liver. The accumulation of retinyl ester in the retinoic acid fed rat liver was accompanied by an absence of labeled retinoic acid. Kidney tissue was found to contain the highest levels of labeled retinoic acid, retinol, and retinyl esters; dietary retinoic acid did not alter the concentrations of these retinoids in the kidney during the experimental period. Since kidney retained more vitamin A when the liver vitamin A was low and also dietary retinoic acid did not affect the concentrations of radioactive retinoic acid in the kidney, it is suggested that the kidney may play a major role in the production of retinoic acid from retinol in the body.