Estrogen prevents increased hepatic aquaporin-9 expression and glycerol uptake during starvation

Abstract

In starvation, glycerol is released from adipose tissue and serves as an important precursor for hepatic gluconeogenesis. By unknown sex-specific mechanisms, women suppress the endogenous glucose production better than men and respond to metabolic stress with higher plasma glycerol levels. Hepatic glycerol uptake is facilitated by aquaporin-9 (AQP9), a broad-selectivity neutral solute channel, and represents an insulin-regulated step in supplying gluconeogenesis with glycerol. In the present study, hepatic AQP9 abundance was increased 2.6-fold in starved male rats as assessed by immunoblotting and immunohistochemistry. By contrast, starvation had no significant effect on hepatic AQP9 expression in female rats. Coordinately, plasma glycerol levels remained unchanged with starvation in male rats, whereas it was increased in female rats. The different responses to starvation were paralleled by higher glycerol permeability in basolateral hepatocyte membranes from starved male rats compared with starved females. Ovariectomy led to a starvation-response pattern identical to that observed in male rats with increased hepatic AQP9 expression and unchanged plasma glycerol levels. In cultured hepatocytes, 17β-estradiol and the selective estrogen receptor α-agonist, propyl pyrazole triol, caused a decrease in AQP9 expression. Our results support that a sex-specific regulation of the hepatic glycerol channel AQP9 during starvation contributes to the higher plasma glycerol levels observed in women during fasting and possibly results in a lower cytosolic availability of glycerol. Furthermore, the sexual dimorphism in the hepatic handling of glycerol during starvation might be explained by 17β-estradiol preventing the starvation-induced increase in hepatic AQP9 abundance.

Metabolic impact of the glycerol channels AQP7 and AQP9 in adipose tissue and liver

“…Ovariectomy resulted in an increase in the hepatic abundance of AQP9 in fasted females similar to that observed in males, whereas ovariectomy did not influence the hepatic abundance of AQP9 in the fed state. This, together with results obtained by in vitro studies in hepatocytes, suggests that 17b-estradiol (E 2 ) prevents the increase in hepatic AQP9 abundance by influencing signaling pathways that are activated during metabolic stress (Lebeck et al 2012b). In support for E 2 being involved in controlling hepatic AQP9 abundance, a separate study found that neonatal exposure to diethylstilbestrol was associated with lower hepatic AQP9 abundance in 20-day-old male rats (Wellejus et al 2008).…”

“…We recently reported that the increased expression of hepatic AQP9 in response to fasting and STZ-induced diabetes mellitus does not occur in female rats (Lebeck et al 2012b). The lack of increase in hepatic AQP9 was paralleled by a lower glycerol permeability of the basolateral plasma membrane of hepatocytes and accumulation of glycerol in the blood in fasted females.…”

Hepatic AQP9 expression in male rats is reduced in response to PPARα agonist treatment

“…This discrepancy could at least partially be due to differences in the cell medium composition. Here we use 0.5% FBS in the experimental medium, which we previously have noted as a vital parameter for transcriptional regulation of AQP9 expression by estradiol in WIF-B9 hepatocytes (16). In all, our findings in HepG2 hepatocytes suggest that the results obtained in rats by pharmacological PPAR␣ activation do have human relevance.…”

“…After washing and dehydration, the tissues were embedded in paraffin wax and 2-m sections were cut with a rotary microtome (Leica). Immunoperoxidase and immunofluorescence labeling for AQP9 were performed as previously described (16,17). Microscopy of immunoperoxidase labeling was performed by using a bright-field microscope (Leica, DMIRE2) equipped with a Leica DM300 digital camera, whereas a Leica TCS SL (SP2) laser scanning confocal microscope was used for immunofluorescence detection.…”

“…Liver and cultured hepatocyte samples were immediately homogenized and prepared for storage as previously described (16,17). Proteins were separated by SDS-PAGE, and for the samples obtained in the animal experiment the initial gels were stained (Gelcode Coomassie Blue Stain Reagent, Thermo Scientific) to adjust protein loading.…”