Modulation in Wistar Rats of Blood Corticosterone Compartmentation by Sex and a Cafeteria Diet

Abstract

In the metabolic syndrome, glucocorticoid activity is increased, but circulating levels show little change. Most of blood glucocorticoids are bound to corticosteroid-binding globulin (CBG), which liver expression and circulating levels are higher in females than in males. Since blood hormones are also bound to blood cells, and the size of this compartment is considerable for androgens and estrogens, we analyzed whether sex or eating a cafeteria diet altered the compartmentation of corticosterone in rat blood. The main corticosterone compartment in rat blood is that specifically bound to plasma proteins, with smaller compartments bound to blood cells or free. Cafeteria diet increased the expression of liver CBG gene, binding plasma capacity and the proportion of blood cell-bound corticosterone. There were marked sex differences in blood corticosterone compartmentation in rats, which were unrelated to testosterone. The use of a monoclonal antibody ELISA and a polyclonal Western blot for plasma CBG compared with both specific plasma binding of corticosterone and CBG gene expression suggested the existence of different forms of CBG, with varying affinities for corticosterone in males and females, since ELISA data showed higher plasma CBG for males, but binding and Western blot analyses (plus liver gene expression) and higher physiological effectiveness for females. Good cross- reactivity to the antigen for polyclonal CBG antibody suggests that in all cases we were measuring CBG.The different immunoreactivity and binding affinity may help explain the marked sex-related differences in plasma hormone binding as sex-linked different proportions of CBG forms.

Modulation of SHBG binding to testosterone and estradiol by sex and morbid obesity

“…The 'estradiolprone' SHBG, structurally close to 'testosterone-prone' SHBG should be formed by differential post-translational selective cleavage (26) of the SHBG transcript (22), thus preserving the similarity and similar (albeit probably not identical) immunoreactive properties (24) of both SHBG molecular species. The protein measured in this study had immunoreactive properties obviously dependent on the affinity and specificity of the antibody used, decanted toward one or other of the isoforms postulated, a situation parallel to that described by our group in rat CBG (52). Thus, in fact, we were not measuring SHBG (under the one molecule-for-all-hormones concept) but either a composite (with unknown proportions of both isoforms) or essentially one of these isoforms, that closer to bind/regulate testosterone.…”

“…Then, densitometric measurements were done using the Total-Lab program (Nonlinear USA, Durham, NC, USA). This Western blot is an adaptation of that previously developed by our group for the measurement of rat plasma CBG (52).…”

“…The basic procedure for hormone-binding assays was the same described by us previously for CBG (52). In short, samples of just-thawed plasma (10 μL) were mixed with 1 mL of PBS (phosphate-buffered saline) pH 7.4 containing 1 g/L gelatin.…”

The Food Energy/Protein Ratio Regulates the Rat Urea Cycle but Not Total Nitrogen Losses

“…Nitrogen content was measured with a semiautomatic Kjeldahl procedure using a ProNitro S system (JP Selecta, Abrera, Spain). Initial rat nitrogen content was estimated from the percentage of N previously published (at day 0) of control animals from the same stock (and provider), age, and sex [ 18 , 19 ], and was adjusted using the initial body weight of each animal used in this experiment. Total accrued N was calculated from the values of body nitrogen obtained at the end of the experiment and discounting the estimated initial nitrogen content.…”

In rats fed high-energy diets, taste, rather than fat content, is the key factor increasing food intake: a comparison of a cafeteria and a lipid-supplemented standard diet

“…Energy expenditure was calculated as previously described ( Rothwell, Saville & Stock, 1982 ) from the difference between the ingested energy and the increase in body energy content of the animals. Energy content increase was estimated using reference data from our previous studies using rats of the same stock, age and sex ( Romero et al, 2013 ; Romero et al, 2014 ). Sodium (salt) intake was calculated from food intake and the sodium content of the different food components used ( Fernández-López et al, 1994 ).…”