The effect of dietary salt on bone in a genetically-defined rat underloading/overloading model

Abstract

One the most serious health hazards of aging and of long term space flight is the loss of bone. The most important determinant of the debilitation due to bone loss is the peak bone mass achieved during late adolescence, which itself may be influenced by gender and environmental factors such as mineral balance. Sodium intake is considered a risk factor for both hypertension and osteoporosis. It is estimated that 30-60% of the population is hypertensive and 30-40% of the population is salt sensitive. The primary purpose of the present studies was to both delineate and combine the effects between salt intake and salt sensitive hypertensive genotype on bone. Our hypothesis was that hypertensive rats would have more severely affected bone than normotensive rats due to salt supplementation and/or genotype. In addition, how these effects might be altered by immobilization/overloading stress was examined as this further burdens NASA space pioneers. This study investigated the possible effects of an ad libidum 1% or 2% saline instead of water on the normotensive (W) and salt sensitive hypertensive (SS) young female rats. A total of 46 weight-matched female rats (7 weeks old) were used. Treated rats in the 1% study drank 1% saline ad libitum for a 42 day salt supplementation period, beginning at day 7 (after arrival) to day 49. Treated rats in the 2% study drank 2% saline ad libitum for a 42 day salt supplementation period, beginning at day 7 (after arrival) to day 49. The right hindlimb of each animal was immobilized by binding to the abdomen with 4 layers of elastic bandage tape, the hip joint in flexion and the knee and ankle joint in extension for the 42 day salt experimental period. Body weight and urine volume was measured biweekly. Food and fluid intake was monitored daily. After sacrifice, three sites (both the underloaded and the overloaded tibiae, as well as the L-2 vertebrae) were processed for histomorphometric analysis. The wet weight (g) and length (mm) of the excised right immobilized and left overloaded femur and the ulna were measured. A 3-point bending test was applied to femurs only. Immediately after the femur breaking strength measurements, bone was cut transversely, one mm from breaking point (fracture location), and a 1.0 mm cross-section was cut for morphological measurement. In addition, a 5 mm high cylinder section from each femur was cut and used for bone composition measurements along with the right ulna bone. A number of elements were analyzed at one time with Induced Coupled Plasma (ICP) spectrometry. The systolic blood pressure and heart rate were measured in the 6th week of study by the tail-cuff sphygmomanometer method. A more robust result was seen with increased concentration of saline treatment from a 1% saline threshold level, to the 2% saline level. Using two-way ANOVA, both hypertensive genotype and 2% saline treatment significantly increased blood pressure and heart rate, and decreased femur magnesium. The SS rat had significant reductions in bone mass, femur cross-sectional area and zinc concentrations with simultaneous elevations in femur stiffness, strength and calcium concentrations. Two percent saline treatment markedly increased both blood pressure and heart rate and decreased both femurs magnesium and cancellous bone in the weight-bearing tibia bone. After 6 weeks of immobilization (to simulate space weightlessness), reductions in cancellous tibia bone volume, with elevations in femur bone stiffness, mineral concentration (calcium and phosphorus) and in trace elements (zinc and manganese) were found in the underloaded femur. Our findings suggest genotype, and saline treatment, and immobilization adversely affect bone in adolescent female rats. In addition, the deleterious bone effects are site specific, affecting each site differently.