In humans, increasing dietary protein increases calcium (Ca) absorption. As measured by dual stable Ca isotopes, when healthy adults ingest a high protein diet (2.1 g/kg/day), skeletal turnover tends to be reduced and Ca absorption significantly increases and almost entirely accounts for the observed hypercalciuria. To better define this effect, rats were randomly assigned to control (20%), low (5%) or high (40%) protein diets for 7 days. All diets were isocaloric and contained 0.45% Ca and 0.35% P. Twenty-four hour urines, blood and feces were collected to assess Ca metabolism, Ca balance and bone turnover. Brush border membrane vesicles (BBMV) were prepared from duodena from rats consuming the low or high protein diet for 7 days to evaluate transcellular Ca transport.

In another group of rats consuming either the low or high protein diet, duodenal RNA was isolated and gene profiling performed. To study the direct effect of amino acids, Ca transport was evaluated across Caco-2 cell monolayers.

In rats, urinary Ca (UCa) paralleled protein intake. Rats consuming the high protein diet absorbed and retained more Ca compared to the low protein group (48.5% vs. 34.1% and 45.8% vs. 33.7% respectively, p<0.01). Ca uptake was significantly higher in the BBMV from rats consuming the high protein diet (p<0.001). Increased V_max explained the higher Ca uptake in BBMV from the high protein group (90 vs. 36 nmol Ca/mg protein/min, p<0.001- 95% CI: 46-2486 and 14-55).

No known transcellular Ca transport proteins were identified by the microarray screen but the Ca permissible tight junction protein, claudin-2, was found to be 2.5-fold higher (p<0.01) in the high protein group. Consistent with this, claudin-2 expression and paracellular Ca flux were both increased in Caco-2 cells exposed to amino acids. In conclusion, the rat model recapitulates the findings in humans in that changes in Ca absorption explain the changes in UCa. The BBMV studies indicate that increased transcellular Ca uptake contributes at least in part to the increase in Ca absorption during the high protein diet. Dietary protein/amino acids also directly improve paracellular Ca absorption likely by increasing claudin-2 expression.