Giuseppe Caso

Control of muscle protein kinetics by acid-base balance.

Purpose of reviewAbnormalities of acid-base balance accompany many pathological conditions. Acidosis is associated with several diseases such as chronic renal failure, diabetic ketosis, severe trauma and sepsis, and chronic obstructive respiratory disease, which are often associated with muscle wasting. There is evidence that acidosis can induce muscle protein catabolism and it could therefore be an important factor contributing to loss of muscle protein in these conditions. This review aims at outlining the effects of acid-base balance abnormalities on muscle protein metabolism, and will in particular summarize and evaluate the most recent studies on the impact of pH on control of muscle protein metabolism. Recent findingsAcidosis has been shown to promote muscle protein catabolism by stimulating protein degradation and amino acid oxidation. This effect is achieved through up-regulation of the ubiquitin-proteasome pathway - one of the major enzyme systems for protein degradation. Recent studies in animals and humans have also shown that acidosis inhibits muscle protein synthesis. Little is known about the mechanisms by which acidosis depresses protein synthesis, or of the impact of alkalosis on protein metabolism. SummaryIncreasing evidence suggests that acidosis promotes muscle protein wasting by both increasing protein degradation and inhibiting protein synthesis. Correction of acidosis may therefore help to preserve muscle mass and improve the health of patients with pathological conditions associated with acidosis.

Tumour cell growth in culture: dependence on arginine.

The amino acid arginine has been shown to affect the growth of several tumours, although the mechanisms of its action are not clear. In the present study, using a human breast tumour cell line (MCF-7), we investigated the arginine requirements of tumour cells for optimal protein synthesis and growth, and the metabolic pathway responsible for the arginine-dependent growth. The results showed that MCF-7 cells are highly dependent on arginine for growth and that the requirement for arginine is much higher than for an indispensable amino acid, leucine, indicating that arginine is needed for pathways other than protein synthesis. In arginine-free cultures, growth could be completely restored by the urea cycle intermediate citrulline. However, arginine could not be replaced by the urea cycle intermediate and the direct precursor for polyamine synthesis, ornithine, or by the polyamine putrescine, suggesting that the high dependence on arginine is not due to a requirement for polyamine synthesis. Moreover, inhibition of NOS [NO (nitric oxide) synthase] did not affect cell protein synthesis and growth, and the arginine analogue and substrate for NOS, homoarginine, could not replace arginine, implying that the conversion of arginine into NO is not involved in the growth-promoting effects of arginine. The major determinant for the high dependence of MCF-7 cells for arginine was found to be the irreversible conversion of this amino acid into ornithine by the intracellular enzyme arginase. The conversion into ornithine caused a progressive depletion of arginine from the culture medium, which ultimately inhibited cell protein synthesis and halted growth. Intracellular arginase activity may be the major factor determining the requirement for arginine of all cells in culture.

The prediction of basal metabolic rate in female patients with anorexia nervosa

OBJECTIVE: To evaluate in female patients with anorexia nervosa the accuracy of a specific predictive formula for basal metabolic rate (BMR) already proposed in the literature and to derive a new disease-specific equation with the same purpose.DESIGN: Cross-sectional study.SUBJECTS: One-hundred and twenty adolescent girls (<18 y) and young-adult women (18–30 y) with anorexia nervosa.MEASUREMENTS: BMR was determined by indirect calorimetry or predicted according to the Schebendach formula, which was specifically derived for anorexia nervosa.RESULTS: On average the Schebendach formula performed well in the adolescent group but not in the young-adult group. The range including 95% of the predicted–measured differences was in both cases wider than 2000 kJ/day. In the young-adult patients the accuracy of the prediction was also related to age and body mass index. Weight and age (but not height or body mass index) emerged as predictors of BMR in the sample as a whole, and only weight when the two age groups were considered separately, thus leading to three different equations. The intercepts of these regression lines were very close and not significantly different from zero while their standard error of estimate was 500–550 kJ/day.CONCLUSION: The Schebendach formula is not very accurate in estimating the BMR of female anorectic patients. Moreover, in this group the relationship between BMR and weight was altered. The predictive formulas proposed by the present study have a reasonable prediction power.

Peripheral fat loss and decline in adipogenesis in older humans

OBJECTIVE Aging is associated with a redistribution of body fat including a relative loss of subcutaneous peripheral fat. These changes in body fat can have important clinical consequences since they are linked to increased risk of metabolic complications. The causes and mechanisms of loss of peripheral fat associated with aging are not clear. The aim of this study was to assess whether defects in adipogenesis contribute to fat loss in aging humans, as suggested from animal studies, and to evaluate the role of inflammation on pathogenesis of fat loss. MATERIALS/METHODS Preadipocytes isolated from subcutaneous peripheral fat of healthy young and elderly subjects were compared in their ability to replicate and differentiate. RESULTS The results show that both the rate of replication and differentiation of preadipocytes are reduced in older subjects. The reduction in adipogenesis is accompanied by a higher plasma level of the inflammatory marker, soluble tumor necrosis factor receptor 2, and greater release of tumor necrosis factor α from fat tissue. CONCLUSIONS Thus, the gradual relative loss of peripheral fat in aging humans may in part result from a defect in adipogenesis, which may be linked to inflammation and increased release of proinflammatory cytokines from fat tissue.

Role of Phospholipase C-β in RNA interference

Phospholipase C-β (PLCβ) enzymes are activated by G proteins in response to agents such as hormones and neurotransmitters, and have been implicated in leukemias and neurological disorders. PLCβ activity causes an increase in intracellular calcium which ultimately leads to profound changes in the cell. PLCβ localizes to three cellular compartments: the plasma membrane, the cytosol and the nucleus. Under most cell conditions, the majority of PLCβ localizes to the plasma membrane where it interacts with G proteins. In trying to determine the factors that localize PLCβ to the cytosol and nucleus, we have recently identified the binding partner, TRAX. TRAX is a nuclease and part of the machinery involved in RNA interference. This review discusses the interaction between PLCβ and TRAX, and its repercussions in G protein signaling and RNA silencing.

Effect of ritonavir and atazanavir on human subcutaneous preadipocyte proliferation and differentiation.

Protease inhibitors (PIs) have been implicated in the development of HIV-associated lipodystrophy through a reduction in the differentiation of preadipocytes. While atazanavir (ATV) is associated with fewer clinical metabolic abnormalities in the short-term, the effects of long-term exposure are not known. ATV effects on preadipocyte replication or differentiation would indicate the potential for long-term problems. This study compared ritonavir (RTV) and ATV effects on preadipocyte replication and differentiation in human primary cultures. Preadipocytes from subcutaneous fat were studied in the presence of therapeutic concentrations of RTV and ATV for replication, differentiation, and adipokine secretion. The effects of the drugs on the expression of PPARgamma and related genes during differentiation were also assessed by real-time quantitative PCR. RTV induced a significant inhibition of preadipocyte proliferation, differentiation and adiponectin secretion. ATV at concentrations within the range of therapeutic levels did not affect differentiation or adiponectin secretion, but did have inhibitory effects on preadipocyte proliferation. Inhibition of differentiation by PIs was associated with decreased expression of PPARgamma, C/EBPalpha, and aP2 genes. In summary, although ATV at therapeutic levels has a smaller impact on adipogenesis, alterations in preadipocyte proliferation suggest the potential for adverse effects with long-term use.

Altered protein metabolism following coronary artery bypass graft (CABG) surgery

The aim of the present study was to investigate the acute effect of CABG (coronary artery bypass graft) surgery on the rates of synthesis of muscle protein, the positive acute-phase protein fibrinogen and the negative acute-phase protein albumin. Synthesis rates of muscle protein, fibrinogen and albumin were measured simultaneously before and 4 h after the end of surgery from the incorporation of L-[(2)H(5)]phenylalanine (given at 43 mg/kg of body weight) in 12 patients undergoing CABG surgery. Surgery was performed either with the use of extracorporeal circulation with cardiopulmonary bypass (on-pump; n=5) or with the beating heart procedure without cardiopulmonary bypass (off-pump; n=7). Post-surgical muscle protein fractional synthesis rates were decreased by 36+/-6.5% compared with pre-surgical values (1.59+/-0.10 compared with 0.97+/-0.08%/day respectively; P<0.001). In contrast, the synthesis rates of both fibrinogen (36+/-4 compared with 100+/-11 mg.day(-1).kg(-1) of body weight; P<0.0001) and albumin (123+/-12 compared with 178+/-19 mg.day(-1).kg(-1) of body weight; P<0.001) were both significantly increased after surgery. No significant differences were found between surgery performed with or without cardiopulmonary bypass. In conclusion, the results demonstrate that CABG surgery has a profound effect on protein metabolism, with a differential response of protein synthesis in muscle and liver.

Effect of insulin on whole body protein metabolism in children with type 1 diabetes

Purpose of reviewUntreated type 1 diabetes (T1D) is associated with abnormalities in protein metabolism, leading to protein loss. These alterations can be particularly detrimental in children, affecting both normal growth and development. A better understanding of the effects of insulin on protein metabolism in children with T1D is essential for optimizing therapy and minimizing consequences of the disease.The aim of the present review is to outline the effects of insulin on whole body protein metabolism in T1D, focusing particularly on studies in children with T1D. Recent findingsWhole body protein degradation and amino acid oxidation are enhanced in children with T1D. Insulin reduces the rates at which body proteins are degraded. Whole body protein synthesis is either unaffected or reduced by insulin, even when insulin is administered together with amino acids to prevent insulin-dependent hypoaminoacidemia. Provision of insulin with oral nutrients improves protein balance by inhibiting whole body protein degradation, but does not affect protein synthesis. SummaryIn children with T1D the anticatabolic effects of insulin on whole body protein metabolism are mainly exerted through a reduction in rates at which body proteins are degraded. Nutritional factors enhancing the anabolic effect of insulin need to be further elucidated.

Dysregulation of CXCL9 and reduced tumor growth in Egr-1 deficient mice

BackgroundEarly growth response-1 (Egr-1) is an immediate-early transcription factor inducible in the vasculature in response to injury, shear stress, and other stimuli. Mice lacking Egr-1 have a profound deficit in the ability to recover from femoral artery ligation, suggesting a role in neovascularization. Previous studies have shown that manipulating Egr-1 expression can have either positive or negative effects on tumor growth. We hypothesized that Egr-1 knockout mice might exhibit reduced tumor growth, possibly due to a reduced capacity to respond to angiogenic signals from a growing tumor.ResultsWe injected 106 Lewis lung carcinoma (LLC1) cells subcutaneously in the flank of wild type and Egr-1 knockout mice. The average mass of tumors from wild type mice at 12 days after implantation was 413 +/- 128 mg, while those from Egr-1-/- mice was 219 +/- 81 mg (p = 0.001, mean +/- SD). However, sectioning the tumors and staining with anti-CD31 antibodies revealed no difference in the vascularity of the tumors and there was no difference in angiogenic growth factor expression. Expression of the chemokine Mig (CXCL9) was increased 2.8-fold in tumors from knockout mice, but no increase was found in serum levels of Mig. Natural killer cells have a 1.7-fold greater prevalence in the CD45+ cells found in tumors from Egr-1-/- mice compared to those from wild type mice. Immunohistochemical staining suggests that Mig expression in the tumors comes from invading macrophages.ConclusionMice deficient in Egr-1 exhibit reduced growth of LLC1 tumors, and this phenomenon is associated with overexpression of Mig locally within the tumor. There are no obvious differences in tumor vascularity in the knockout mice. Natural killer cells accumulate in the tumors grown in Egr-1-/- mice, providing a potential mechanism for the reduction in growth.

Does Increasing Blood pH Stimulate Protein Synthesis in Dialysis Patients

Background: Although the mechanism of muscle wasting in end-stage renal disease is not fully understood, there is increasing evidence that acidosis induces muscle protein degradation and could therefore contribute to the loss of muscle protein stores of patients on hemodialysis, a prototypical state of chronic metabolic acidosis (CMA). Because body protein mass is controlled by the balance between synthesis and degradation, protein loss can occur as result of either increased breakdown, impaired synthesis, or both. Correction of acidosis may therefore help to maintain muscle mass and improve the health of patients with CMA. We evaluated whether alkalizing patients on hemodialysis might have a positive effect on protein synthesis and on nutritional parameters. Methods: Eight chronic hemodialysis patients were treated daily with oral sodium bicarbonate (NaHCO3) supplementation for 10–14 days, yielding a pre-dialytic plasma bicarbonate concentration of 28.6 ±1.6 mmol/l. The fractional synthesis rates (FSR) of muscle protein and albumin were obtained by the L-[2H5ring]phenylalanine flooding technique. Results: Oral NaHCO3 supplementation induced a significant increase in serum bicarbonate (21.5 ± 3.4 vs. 28.6 ± 1.6 mmol/l; p = 0.018) and blood pH (7.41 vs. 7.46; p = 0.041). The FSR of muscle protein and the FSR of albumin did not change significantly (muscle protein: 2.1 ± 0.2 vs. 2.0 ± 0.5% per day, p = 0.39; albumin: 8.3 ± 2.2 vs. 8.6 ± 2.5% per day, p = 0.31). Plasma concentrations of insulin-like growth factor 1 decreased significantly (33.4 ± 21.3 vs. 25.4 ± 12.3 nmol/l; p = 0.028), whereas thyroid-stimulating hormone, free thyroxin and free triiodothyronine did not change significantly and nutritional parameters showed no improvement. Conclusion: In contrast to other findings, raising the blood pH of dialysis patients was not associated with a positive effect on albumin and muscle protein synthesis, or nutritional and endocrinal parameters.