Christian E. Lezón

Failure of Polycythemia-Induced Increase in Arterial Oxygen Content to Suppress the Anorexic Effect of Simulated High Altitude in the Adult Rat

The anorexic effect of exposure to high altitude may be related to the reduction in the arterial oxygen content (Ca(O2)) induced by hypoxemia and possibly the associated decreased convective oxygen transport (COT). This study was then performed to evaluate the effects of either transfusion-induced polycythemia or previous acclimation to hypobaria with endogenously induced polycythemia on the anorexic effect of simulated high altitude (SHA) in adult female rats. Food consumption, expressed in g/d/100 g body weight, was reduced by 40% in rats exposed to 506 mbar for 4 d, as compared to control rats maintained in room air. Transfusion polycythemia, which significantly increased hematocrit, hemoglobin concentration, Ca(O2), and COT, did not change the anorexic response to the exposure to hypobaric air. Depression of food intake during exposure to SHA also occurred in rats fasted during 31 h before exposure and allowed to eat ad libitum for 2 h during exposure. Body mass loss was similar in 48-h fasted rats that were either hypoxic or normoxic. Body mass loss was similar in normoxic and hypoxic rats, the former eating the amount of food freely eaten by the latter. Hypoxia-acclimated rats with endogenously induced polycythemia taken to SHA again had diminished food intake and lost body mass at rates that were very close to those found in nonacclimated ones. Exposure to SHA also led to a decrease in food consumption, body weight, and plasma leptin in adult female mice. Analysis of data suggest that body mass loss that accompanies SHA-induced hypoxia is due to hypophagia and that experimental manipulation of the blood oxygen transport capacity cannot ameliorate it. Leptin does not appear to be an inducer of the anorexic response to hypoxia, at least in mice and rats.

Plasma disappearance of exogenous erythropoietin in mice under different experimental conditions

Erythropoietin (EPO) is a glycoprotein hormone produced primarily in the kidneys and to a lesser extent in the liver that regulates red cell production. Most of the studies conducted in experimental animals to assess the role of EPO in the regulation of erythropoiesis were performed in mouse models. However, little is known about the in vivo metabolism of the hormone in this species. The present study was thus undertaken to measure the plasma t 1/2 of radiolabeled recombinant human EPO (rh-EPO) in normal mice as well as in mice with altered erythrocyte production rates (EPR), plasma EPO (pEPO) titer, marrow responsiveness, red cell volume, or liver function. Adult CF-1 mice of both sexes were used throughout. For the EPO life-span studies, 30 mice in each experiment were intravenously injected with 600,000 cpm of 125I-rh-EPO and bled by cardiac puncture in groups of five every hour for 6 h. Trichloroacetic acid (TCA) was added to each plasma sample and the radioactivity in the precipitate measured in a γ-counter. EPO, pEPO, marrow responsiveness, or red cell volume were altered by either injections of rh-EPO, 5-fluorouracil, or phenylhydrazine, or by bleeding, or red cell transfusion. Liver function was altered by Cl4C administration. In the normal groups of mice, the estimated t 1/2 was 182.75 ± 14.4 (SEM) min. The estimated t 1/2 of the other experimental groups was not significantly different from normal. These results, therefore, strongly suggest that the clearance rate of EPO in mice is not subjected to physiologic regulation and that pEPO titer can be really taken as the reflection of the EPO production rate, at least in the experimental conditions reported here.

Improved bone status by the β-blocker propranolol in an animal model of nutritional growth retardation

The aim of the present research was to study if the beta-blocker propranolol, which is known to increase bone mass, could reverse the adverse skeletal effects of mild chronic food restriction in weanling rats. Male Wistar rats were divided into four groups: control, control+propranolol (CP), nutritional growth retardation (NGR) and nutritional growth retardation+propranolol (NGRP). Control and CP rats were fed freely with the standard diet. NGR and NGRP rats received, for 4 weeks, 80 % of the amount of food consumed by the control and CP rats, respectively. Results were expressed as mean values and sem. Food restriction induced detrimental effects on body and femur weight and length (P < 0.05) and bone structural and geometrical properties (P < 0.001), confirming results previously shown in our laboratory. However, the beta-blocker overcame the deleterious effect of nutritional stress on load-bearing capacity, yielding load, bone stiffness, cross-sectional cortical bone area and second moment of inertia of the cross-section in relation to the horizontal axis without affecting anthropometric, histomorphometric and bone morphometric parameters. The results suggest that propranolol administration to mildly chronically undernourished rats markedly attenuates the impaired bone status in this animal model of growth retardation.

Enhanced Erythropoietin Response to Acute Hypoxemia in Mice with Pharmacological Depression of Erythropoiesis

The recent report of a depression of stimulated production of erythropoietin (EPO) in mice with enhanced erythropoiesis suggests that unknown mechanism (s) other than hypoxia may be involved in the regulation of EPO formation. The present study was thus designed to investigate EPO production during acute hypoxemia in a mouse model in which the oxygen-carrying capacity of blood, the plasma EPO level, and the plasma EPO half-life were within normal values in spite of a marked depression of the red cell production rate (RCPR) induced by cyclophosphamide (CP) administration. Injection of 100 mg/Kg of the drug into adult female CF-1 mice that previously received 0.4 ml of packed red cells depressed markedly the 24-hour RBC 59Fe uptake without affecting the plasma immunoreactive EPO level and the plasma disappearance of 1251-labeled recombinant human EPO. The EPO production rate, calculated from the change in plasma EPO levels and the estimated EPO clearance rate, after 4 h of exposure to hypobaric air was about 2.8 times higher in mice with CP-induced inhibition of the RCPR than in mice with normal RCPR. The results support the hypothesis that the EPO production rate in mammals is not only related to the oxygen supply to the tissues relative to their oxygen needs (main stimulus) but also to the erythroid activity of the marrow (modulatory action).

Hypothalamic Noradrenergic Hyperactivity and Detrimental Bone Status in an Animal Model of Nutritional Growth Retardation

We have studied hypothalamic noradrenergic activity in relation with bone status in a nutritional growth retar- dation model (ND). Control rats (C) were fed ad libitum. ND received 80% of the diet consumed by C for 4 weeks and later refed ad libitum for 8 weeks. Food restriction induced detrimental effects on body and femur weight and length (P<0.05) and bone biomechanical properties (P<0.001). Thickness of proliferative and hypertrophic zone (�m) of growth plate cartilage and bone volume (%, mean±SE) were 225.96±5.70 v. 280.70±12.52, 95.16±5.81 v. 134.60±9.30, 17.64±3.23 v. 26.80±2.03, respectively (P<0.05); anterior and posterior hypothalamus norepinephrine uptake and release and tyrosine hydroxylase activity (% of control) were 79.05±3.56, 67.00±10.00, 164.26±16.58 and 80.65±5.92, 147.00±1.00, 152.42±9.30, respectively (P<0.05). Thus, impaired biomechanical bone performance in ND could be due, in part, to the increased hypothalamic noradrenergic activity in response to restriction. Normalization of parameters with refeeding suggests no long-term side-effects in undernourished rats.

Hypophagia-induced hypometabolism during experimental anaemia in rats

Several investigators have reported a drop in oxygen (O2) consumption (VO2) and body temperature in laboratory animals during normobaric or hypobaric hypoxia. Hypophagia, with normal efficiency of protein utilisation for growth, was also observed. It has recently also been observed that hypometabolism is present during anaemic hypoxia. The present study was designed to test the experimental hypothesis that anaemic hypoxia induces hypometabolism secondary to hypophagia. Episodes of anaemia were created in adult male rats by either blood withdrawal through cardiac puncture (haemorrhagic anaemia) or phenylhydrazine administration (haemolytic anaemia). Haematrocrit, VO2, and food consumption, as indirect estimations of the level of anaemia, energy production, and appetite, respectively, were serially measured in all animals during 7 days (acute experiments) or 17 days (chronic experiments). Positive correlations were found between the three parameters during development of and recovery from anaemia during each anaemic episode. When the amount of food offered to non-anaemic rats was equalised to that freely eaten by anaemic rats, VO2 dropped in the former to almost the level found in the latter. Body composition changed during chronic anaemia because of a decrease in the lipid fraction of the body. The results confirmed the working hypothesis that hypometabolism, which has been considered as an immediate, emergency-type response to both hypoxic and anaemic hypoxia, can be considered as a response secondary to hypophagia because of depressed appetite. How appetite is adapted to the mechanisms which control O2 convection and O2 availability is not known at present.

Monounsaturated fatty acids-rich diets in hypercholesterolemic-growing rats.

Abstract The effects of replacing dietary saturated fat by different monounsaturated fatty acid (ω-9MUFA) sources on serum lipids, body fat and bone in growing hypercholesterolemic rats were studied. Rats received one of the six different diets: AIN-93G (control, C); extra virgin olive oil (OO) + C; high-oleic sunflower oil (HOSO) + C or atherogenic diet (AT) for 8 weeks; the remaining two groups received AT for 3 weeks and then, the saturated fat was replaced by an oil mixture of soybean oil added with OO or HOSO for 5 weeks. Rats consuming MUFA-rich diets showed the highest body fat, hepatic index and epididymal, intestinal and perirenal fat, and triglycerides. T-chol and non-HDL-chol were increased in HOSO rats but decreased in OO rats. Bone mineral content and density were higher in both OO and HOSO groups than in AT rats. This study casts caution to the generalization of the benefits of MUFA for the treatment of hypercholesterolemia.

mRNA of cytokines in bone marrow and bone biomarkers in response to propranolol in a nutritional growth retardation model

BACKGROUND The aim of this study was to assess mRNA of IL-6, TNFα and IL-10 cytokines in bone marrow, possible mediators involved in altered bone remodeling with detrimental consequences on bone quality in NGR (Nutritional growth retardation) rats. METHODS Weanling male Wistar rats were assigned either to control (C) or experimental group (NGR) (n=20 each). C and NGR groups were assigned to 2 groups according to receiving saline solution (SS) or propranolol hydrochloride (P): C, C+P (CP), NGR or NGR+P (NGRP). For 4 weeks, NGR and NGRP rats received 80% of the amount of food consumed by C and CP, respectively, the previous day, corrected by body weight. P (7 mg/kg/day) was injected ip 5 days/week, for 4 weeks in CP and NGRP rats. Body weight and length were recorded. After 4 weeks, blood was drawn. Femurs were dissected for RNA isolation from bone marrow and mRNA of cytokines assays. RESULTS Food restriction induced a significant negative effect on body growth in NGR and NGRP rats (p<0.001). P had no effects on zoometric parameters (p>0.05). CTX-I increased in NGR rats vs. C (p<0.001), but diminished in NGRP (p<0.01). Serum osteocalcin, PTH, calcium and phosphate levels remained unchanged between groups (p>0.05). In NGR, bone marrow IL-6 mRNA and IL-10 mRNA levels were low as compared to other groups (p<0.05). In contrast, bone marrow TNF-α mRNA levels were significantly high (p<0.05). CONCLUSIONS This study provides evidences that NGR outcomes in a bone marrow proinflammatory microenvironment leading to unbalanced bone remodeling by enhancement of bone resorption reverted by propranolol.

Efecto de diferentes dosis de propranolol sobre la eficiencia estructural y mecánica esquelética en un modelo animal de retraso del crecimiento

OBJECTIVE To assess in a growth retardation (GR) model the impact of different propranolol (P) doses on anthropomorphometric and biomechanical variables of the appendicular skeleton. MATERIALS AND METHODS Twenty-one day-old male Wistar rats were divided into the following groups: control (C), C+P3.5 (CP3.5); C+P7 (CP7); C+P10.5 (CP10.5); C+P14 (CP14); ED, ED+P3.5 (EDP3.5); ED+P7 (EDP7); ED+P10.5 (EDP10.5), and ED+P14 (EDP14). Control animals with/without P were fed a rodent diet ad libitum. GR rats with/without P were given 80% of the same diet per 100g body weight for 4 weeks (T4). Propranolol 3.5, 7, 10.5, and 14 mg/kg/day was intraperitoneally injected 5 days/week for 4 weeks to the CP3.5 and EDP3.5; CP7 and EDP7; CP10.5 and EDP10.5, and CP14 and EDP14 groups respectively. RESULTS At T4, energy restriction had negative effects upon overall growth, femur, and its mechanical competence. Propranolol improved bone rigidity in GR animals at doses of 7 and 10.5mg/kg/day, with a maximum response at 7 mg/kg/day. CONCLUSIONS Propranolol 7 mg/kg/day would be the most effective dose for modeling incorporation of bone, as shown by the increased skeletal structural and mechanic efficiency in this animal model of growth retardation. Such effect may result from maintenance of mechanosensor viability, changes in its sensitivity, the biomechanical reference point and/or effector response in GR rats.

Actividad del sistema neuronal LHRH en un modelo animal de retraso del crecimiento

OBJECTIVE Mild and chronic energy restriction results in growth retardation with puberal delay, a nutritional disease known as nutritional dwarfing (ND). The aim of the present study was to assess the profile of hypothalamic luteinizing hormone-releasing hormone (LHRH) release, at baseline and under glutamate stimulation, in ND rats to elucidate gonadotrophic dysfunction. Reproductive ability during refeeding was also studied. MATERIAL AND METHODS At weaning, 60 male rats were assigned to two groups of 30 animals each: a control and an experimental group. Control rats were fed ad libitum with a balanced rodent diet. The experimental group received 80% of the diet consumed by the control group for 4 weeks. After 4 weeks of food restriction, the ND group was fed freely for 8 weeks. Ten rats from each group were sacrificed every 4 weeks for assays. RESULTS At week 4, body weight and length were significantly diminished in the experimental group vs. the control group (p<0.001). No changes were observed in LHRH baseline release, pulse frequency or amplitude in the experimental group compared with the control group at any time. However, under glutamate stimulation, LHRH release was significantly higher in ND rats than in control rats at week 4 (p<0.05). Refeeding the ND group allowed the rats to reach overall growth and reproductive ability. CONCLUSIONS The results of the present study suggest that the response to the facilitatory effect of glutamate on LHRH release in post-restricted ND rats is probably related to a lesser central nervous system maturation in relation to their chronological age. The adequate somatic growth and normal reproductive ability attained with refeeding suggest the reversibility of the two energetically costly processes compromised by global, mild and chronic food restriction.