Conditioned nutritional deficiencies in the cardiomyopathic hamster heart.

BACKGROUND: Evidence indicates that nutritional factors may be important in the maintenance of myocyte structure and energetics. The failing myocardium has been reported to exhibit a depletion of several nutrients that are important for the maintenance of intracellular calcium homeostasis and cellular energetics, and levels of oxidative stress. This nutrient depletion may contribute to the progressive deterioration in myocardial structure and function observed in heart failure. OBJECTIVE: To examine the extent to which advanced cardiomyopathy results in a depletion of nutrients and/or metabolites and antioxidants, and whether supplementation with these nutrients may influence cellular structure or function. SUBJECTS AND METHODS: Cardiomyopathic hamsters were randomly placed to one of the three following diet groups: chow; control gelled diet; or a supplemented gelled diet that provided taurine, carnitine, coenzyme Q10, selenium, vitamins E and C, creatine, thiamine and L-cysteine. After approximately three months of supplementation, one group of hamsters underwent functional testing using a modified Langendorff technique with biopsy samples taken for electron microscopy. Myocardial nutrient concentrations were determined in a second group of diseased and nondiseased hamsters of the same age. RESULTS: Cardiomyopathy resulted in a depletion of vitamin E, creatine, carnitine, taurine and coenzyme Q10. Supplementation resulted in improved cardiac ultrastructure, function and contractility compared with nonsupplemented hamsters. CONCLUSIONS: These studies suggest that heart failure results in 'condition-related nutrient deficiencies' that, once corrected, can significantly impact on heart function and structure.

31P-nuclear magnetic resonance studies of bioenergetic changes in skeletal muscle in malnourished human adults.

In previous studies, both animals and malnourished children receiving 25% of the protein-energy intake of a control group, resulting in a 25% weight loss, had lower ratios of phosphocreatine to beta-ATP and of phosphocreatine to inorganic phosphorus, higher free ADP concentrations, and lower free energy of ATP hydrolysis than the control group. Therefore, the effect of malnutrition on muscle energetics in adult humans was examined by using 31P-nuclear magnetic resonance spectroscopy in malnourished patients with a mean body mass index (BMI; in kg/m2) of 16.4 compared with healthy control subjects with a significantly higher body mass index of 24.5 (P < 0.005). The mean (+/- SEM) ratio of phosphocreatine (PCr) to ATP in the malnourished patients was 2.28 +/- 0.27, which was significantly lower than the ratio of 3.1 +/- 0.15 in control subjects (P < 0.02). The ratio of inorganic phosphorus (Pi) to ATP in malnourished patients was 0.33 +/- 0.04, which was significantly lower than the ratio of 0.48 +/- 0.03 in control subjects (P < 0.02), but the ratio of PCr to Pi was not significantly different from that in control subjects. There was a significant correlation between BMI and the ratio of PCr to ATP (P < 0.01) and of Pi to ATP (P < 0.01). These data suggest that progressive loss of BMI is associated with a relative loss of muscle creatine and phosphorus in relation to ATP. The findings were unlikely to have been due only to atrophy of fast-twitch fibers because such atrophy would have altered the ratio of PCr to Pi.

Stability of high-energy substrates in fast- and slow-twitch muscle: comparison of enzymatic assay of biopsy with in vivo 31P nuclear magnetic resonance spectroscopy.

The purpose of this study was to investigate the stability of ATP and PCr levels in stored muscle samples and extracts. ATP and PCr levels were measured by fluorimetric analysis in freeze-clamped biopsies of soleus, extensor digitorum longus, and gastrocnemius muscles of the rat after storage in a freezer at -70 degrees C as (i) intact wet muscle, (ii) freeze-dried muscle, and (iii) an extract of freeze-dried muscle. Assays were performed within 24 h of taking the biopsy and after variable periods of storage from 1 to 4 weeks. The data for the gastrocnemius muscles were compared with those obtained, in the same rat, by in vivo 31P NMR spectroscopy. In the biopsies, the ATP levels were stable irrespective of the duration or method of storage. The PCr levels fell by 13-16% compared with the values obtained from the assay done within 24 h of taking the biopsy, irrespective of the method of storage, but could be corrected in the freeze-dried stored muscle by expressing the data in relation to the total creatine levels. The fluorimetrically measured PCr, in whole muscle extracts of the gastrocnemius, assayed within 24 h, were comparable to those obtained from 31P NMR spectroscopy. We concluded that PCr levels in muscle are not stable during storage at -70 degrees C and should be assayed within 24 h of taking a muscle biopsy to ensure that the values are the same as those obtained by 31P NMR.

Effect of fasting, hypocaloric feeding, and refeeding on the energetics of stimulated rat muscle as assessed by nuclear magnetic resonance spectroscopy.

A previous study suggested that muscles from hypocalorically fed rats were limited in their ability to rephosphorylate ADP. During muscle contraction hydrolysis of ATP results in an increase in phosphorus, free ADP, delta GATP, and a reduction in phosphocreatine levels that is reversed during rest by rephosphorylation of ADP to ATP and the resynthesis of phosphocreatine by ATP. We therefore hypothesized that these changes would be restored more slowly during postcontraction rest in hypocalorically fed rats as compared with controls. We compared controls fed ad lib to 2-d fasted and hypocalorically fed rats, losing 20% of their weight. We also compared hypocalorically fed rats that had been refed ad lib for 7 d with age-matched controls fed ad lib. The results showed that ATP, muscle pH, and total muscle creatine levels were not different in all groups. The raised phosphorus and delta GATP levels and lower phosphocreatine/phosphorus ratio at the end of contraction changed more slowly during rest in the hypocaloric rats. These abnormalities were partially corrected by refeeding. The data taken as a whole support the concept of impaired rephosphorylation of ADP in malnourished muscle that is not completely restored by refeeding in stimulated muscle.

Comparison of six methods for force normalization in muscles from malnourished rats.

We have investigated six methods for normalizing isometric force in skeletal muscles from malnourished rats. Adult male Wistar rats were anesthetized, and the maximum isometric force production of soleus and extensor digitorum longus (EDL) muscles was measured after a 2-d fast or 1 wk of hypocaloric feeding. The force was expressed per unit muscle wet weight, dry weight, cross-sectional area, total creatine, total protein, and myofibrillar protein. We found that the absolute force of the muscles (N) and the force expressed in relation to total creatine were similar in muscles from fasted, hypocaloric, and control rats. Regardless of the method of normalization, forces produced by soleus muscles from fasted rats were the same as control values. Force expressed in relation to wet weight, dry weight, cross-sectional area, and total protein was greater in muscles from malnourished animals except for the soleus muscles from fasted rats. Force per mg myofibrillar protein was greater in the EDL muscles from fasted and hypocaloric rats than in controls, but the values for the soleus muscles were similar in the three groups. The data indicate that different methods of force normalization can lead to different conclusions concerning the ability of muscles to generate force after nutritional intervention.

Levels of norepinephrine, epinephrine, dopamine, serotonin and N-acetylserotonin in aqueous humour.

Levels of norepinephrine, epinephrine and dopamine were measured in aqueous humour samples from 13 patients admitted to hospital for cataract surgery. Serotonin and N-acetylserotonin levels were measured in aqueous samples from seven sheep eyes. Norepinephrine, epinephrine and dopamine were detected in the aqueous from all patients; the mean levels were 330, 92 and 155 pg/mL respectively. Neither serotonin nor N-acetylserotonin was detected in animal aqueous. The possible relevance of the findings to aqueous production and intraocular pressure control is discussed.

Serotonin in the human infant carotid body.

By immunocytochemistry serotonin was localized in the chief cells of the carotid body in human infants. Radioenzymatic measurement of the serotonin concentration revealed that it represents a significant proportion of the total amine content of the carotid body.

An active pathway for serotonin synthesis by renal proximal tubules.

Serotonin (5HT) has significant effects on renal metabolism and glomerular function and is a potent renal vasoconstrictor. In this study we describe and localize a highly active biosynthetic pathway for serotonin in the kidney. Rat kidneys were dissected into cortical and medullary fractions; in some experiments the cortex was also separated into subfractions enriched with glomeruli or proximal tubules. Serotonin and tryptophan hydroxylase (TyOH) were measured by radioenzymatic techniques. (table; see text) Renal denervation did not alter tryptophan hydroxylase activity. In kidneys from human cadaveric donors, cortical tryptophan hydroxylase (4.13 +/- 0.68 nM/30 min/g) exceeded that in the medulla (1.96 +/- 0.86 nM/30 min/g). Aromatic L-amino acid decarboxylase, the remaining enzyme for serotonin synthesis, is present in both rat renal cortex and medulla; however, we found 15-fold greater decarboxylase activity in proximal tubular (2070 nM/30 min/g) as compared to glomerular (131 nM/30 min/g) subfractions. We were able to demonstrate that under physiological conditions, free urine serotonin reflects actual biosynthesis by the kidney. Thus, although serotonin stores retained by the kidney appear small and relatively localized to the medulla, the enzymatic activity for the synthesis of serotonin in the kidney is comparable to that in the brain, with the complete pathway localized to renal cortical proximal tubules. These data suggest that further studies of renal serotonin metabolism may contribute to our understanding of renal function in health and disease.

Radioimmunoassay for human thymus-leukemia-associated antigen.

A competitive radioimmunoprecipitation method was developed for the quantitation of human thymus-leukemia-associated antigen (HThy-L) isolated from human thymus tissue. Antisera to the antigen were raised by immunization of rabbits with purified fractions of HThy-L. The antigen was labeled with 125l by means of a modification of the lactoperoxidase technique and subjected to Sephadex G-100 gel filtration. Analysis of fractions eluted from this column by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed two labeled components with apparent molecular weights of 45,000 and 25,000 daltons, respectively. Immunoprecipitation with anti-HThy-L antiserum demonstrated directly that the 45,000-dalton component carried HThy-L antigenicity. This fraction served as the source of labeled antigen in a competitive radioimmunoassay that permitted the detection of approximately 1 ng HThy-L. With the use of this assay, we confirmed quantitatively that the highest amounts of HThy-L were found in extracts of thymocytes, normal thymus tissues, and lymphoblasts for T-cell lines.