Kisspeptin deficiency leads to abnormal adrenal glands and excess steroid hormone secretion

Abstract

Knockout mice for the kisspeptin receptor, Kiss1r (Kiss1r-/-) and its ligand kisspeptin, Kiss1 (Kiss1-/-) replicate the phenotype of isolated hypogonadotropic hypogonadism (IHH) associated with variants of these genes in humans. A recent report suggests that kisspeptin may be involved in human fetal adrenocortical development and function. Herein, we characterized the adrenal function and morphology in Kiss1-/- mice that do not go through normal puberty. Two fetal markers were expressed in eosinophilic cells potentially derived from the X-zone that should disappear at puberty in male mice and during the first pregnancy in female animals. Although the hypercorticosteronism observed in Kiss1-/- females corrected overtime, hyperaldosteronism persisted at 14 months and correlated with the overexpression of Star. To determine if KISS1 and KISS1R genes are involved in the development of primary aldosteronism (PA) and hypercortisolism [Cushing's syndrome (CS)] in humans, we sequenced these 2 genes in 65 patients with PA and/or CS. Interestingly, a patient with CS presented with a germline KISS1 variant (p.H90D, rs201073751). We also found three rare variants in the KISS1R gene in three patients with PA: p.C95W (rs141767649), p.A189T (rs73507527) and p.R229R (rs115335009). The two missense variants have been previously associated with IHH. Our findings suggest that KISS1 may play a role in adrenal function in mice and possibly adrenocortical steroid hormone secretion in humans, beyond its recently described role in human fetal adrenocortical development.

Figure 1

The X-zone does not regress in Kiss1^−/− adrenals. (A) Histology analysis showed an accumulation of cells at the border of the cortex and the medulla that may be fetal X-zone cells in the adrenal glands of female Kiss1^−/− mice that were not observed in Kiss1^+/+ animals at 5 months of age. C: Cortex, M: Medulla. A white dashed line is indicating the border between the cortex and the medulla. Arrowhead points to the fetal cells composing the X-zone. (B) 20αHSD staining confirmed the fetal-like differentiation of the cells. The white arrowhead points to the fetal cells composing the X-zone. (C) The expression of X-zone markers, Akr1c18, Pik3c, Inha and Cyp17a1 was increased in the adrenal glands of female Kiss1^−/− mice (compared with Kiss1^+/+) both at 5 and 12 months of age, by RT qPCR. Bars represent the induction fold of at least four individual adrenals ± standard deviation. P-value was calculated using Student’s t-test. (D–E) Histology by hematoxylin and eosin (D) and 20αHSD staining (E) demonstrate that fetal cells were still observed in the adrenal glands ofKiss1^+/+and Kiss1^−/− animals at 12 months.

Figure 2

Female Kiss1^−/− animals have primary hyperaldosteronism. (A) Serum corticosterone and ACTH concentration were up- and downregulated, respectively, in Kiss1^−/−female compared with Kiss1^+/+ animals at 5 months of age. Bars represent the mean of either corticosterone or ACTH level of at least six individual adrenals ± standard deviation. (B) RTqPCR for mc2r, Star, Cyp11a1, Hsd3b, Cyp21, Cyp11b1 and Cyp11b2 in Kiss1^+/+, Kiss1^+/− and Kiss1^−/− from the adrenal glands of 5-month-old female mice. Bars represent the mean of the hormone level of at least six individual glands ± standard deviation. P-value was calculated using Student’s t-test. (C) Immunofluorescence staining of Akr1b7 on Kiss1^+/+ and Kiss1^−/− adrenals at 5 months. (D) Serum aldosterone concentration was significantly increased in Kiss1^−/− females at both 5 and 12 months. Bars represent the mean of aldosterone level of at least six individual samples ± standard deviation.