Dr. Ron’s Research Review – October 17, 2012

This week’s research review focuses on Aromatase and Calcium.

The calcium ion has an essential role in the aromatization of 4-androstene-3,17-dione to estradiol. The calcium-calmodulin complex is required for activation of aromatase by cyclic AMP. (Hochberg, Bick et al. 1986)

A calcium-deficient diet caused decreased bone mineral density and secondary elevation of estrogen in aged male rats. Testicular aromatase cytochrome P450 activity was significantly increased by 2.4-fold in the calcium-deficient diet group. The change in testicular aromatase expression might be, in part, a compensatory mechanism for the bone mineral deficiency induced by the calcium-deficient diet in aged male rats. (Kato, Mano et al. 2002)

Vitamin D receptor null mutant mice showed gonadal insufficiencies and low aromatase activity. Calcium supplementation increased aromatase activity. When the serum calcium concentration was kept in the normal range by supplementation, the aromatase activity in the ovary increased to 60%. (Kinuta, Tanaka et al. 2000)

Aromatase deficiency causes altered expression of molecules critical for calcium reabsorption in the kidneys of female mice. (Oz, Hajibeigi et al. 2007)

Dr. Ron


Articles

The dual effect of calcium on aromatization by cultured human trophoblast

         (Hochberg, Bick et al. 1986) Download

To study the effect of calcium ion on aromatization of an androgenic precursor to estradiol by the placenta, cultured term trophoblasts were used as a model system. Secretion of estradiol into the culture medium was regarded as indicating aromatization, since cells cultured with no androgenic precursors produced only insignificant amounts of estradiol. EGTA, verapamil and ionophore A23187 inhibited aromatization, while trifluoperazine, an inhibitor of the calcium-calmodulin complex, interfered with the stimulatory effect of cyclic AMP on aromatization. We conclude that calcium ion has an essential role in the aromatization of 4-androstene-3,17-dione to estradiol. The calcium-calmodulin complex is required for activation of aromatase by cyclic AMP. However, when flooded with calcium by ionophore A23187, the trophoblast is unable to effectively buffer calcium, and aromatization is inhibited.

A calcium-deficient diet caused decreased bone mineral density and secondary elevation of estrogen in aged male rats-effect of menatetrenone and elcatonin

         (Kato, Mano et al. 2002) Download

In view of the fact that a deficient calcium (Ca) intake results in osteoporosis in elderly males, we conducted an animal experiment on aged male Wistar rats given a Ca-deficient diet. The rats were divided into 2 groups according to diet: a Ca-deficient diet group (Ca content, 0.08% to 0.1%) and a regular diet group (Ca content, 0.8% to 1.2%). The Ca-deficient diet reduced bone mineral density (BMD) by approximately 12%. Administration of menatetrenone or elcatonin was able to reverse the reduction in BMD induced by Ca deficiency. The mean estradiol level in sera of rats fed the Ca-deficient diet was significantly increased to 4.3 times that in the regular diet group. However, the increased estradiol concentration was reduced after the administration of menatetrenone or elcatonin. The estrone concentrations in sera of menatetrenone- or elcatonin-treated rats fed the Ca-deficient diet decreased to a level lower than that of animals fed the regular diet. Testicular aromatase cytochrome P450 (P450(arom); estrogen synthetase) activity was significantly increased by 2.4-fold in the Ca-deficient diet group compared to that in the regular diet group, and the aromatase mRNA level was also significantly increased 1.45-fold. Testicular aromatase activity was strongly correlated with aromatase mRNA level and serum estradiol level. These data suggest that the change in testicular aromatase expression might be, in part, a compensatory mechanism for the bone mineral deficiency induced by the Ca-deficient diet in aged male rats.


Vitamin D is an important factor in estrogen biosynthesis of both female and male gonads

         (Kinuta, Tanaka et al. 2000) Download

In the present study, the role of vitamin D in the regulation of estrogen synthesis in gonads was investigated. Vitamin D receptor null mutant mice showed gonadal insufficiencies. Uterine hypoplasia and impaired folliculogenesis were observed in the female, and decreased sperm count and decreased motility with histological abnormality of the testis were observed in the male. The aromatase activities in these mice were low in the ovary, testis, and epididymis at 24%, 58%, and 35% of the wild-type values, respectively. The gene expression of aromatase was also reduced in these organs. Elevated serum levels of LH and FSH revealed hypergonadotropic hypogonadism in these mice. The gene expressions of estrogen receptor alpha and beta were normal in gonads in these mice. Supplementation of estradiol normalized histological abnormality in the male gonads as well as in the female. Calcium supplementation increased aromatase activity and partially corrected the hypogonadism. When the serum calcium concentration was kept in the normal range by supplementation, the aromatase activity in the ovary increased to 60% of the wild-type level, but LH and FSH levels were still elevated. These results indicated that vitamin D is essential for full gonadal function in both sexes. The action of vitamin D on estrogen biosynthesis was partially explained by maintaining calcium homeostasis; however, direct regulation of the expression of the aromatase gene should not be neglected.

Aromatase deficiency causes altered expression of molecules critical for calcium reabsorption in the kidneys of female mice *

            (Oz, Hajibeigi et al. 2007) Download

Kidney stones increase after menopause, suggesting a role for estrogen deficiency. ArKO mice have hypercalciuria and lower levels of calcium transport proteins, whereas levels of the klotho protein are elevated. Thus, estrogen deficiency is sufficient to cause altered renal calcium handling. INTRODUCTION: The incidence of renal stones increases in women after menopause, implicating a possible role for estrogen deficiency. We used the aromatase deficient (ArKO) mouse, a model of estrogen deficiency, to test the hypothesis that estrogen deficiency would increase urinary calcium excretion and alter the expression of molecular regulators of renal calcium reabsorption. MATERIALS AND METHODS: Adult female wildtype (WT), ArKO, and estradiol-treated ArKO mice (n = 5-12/group) were used to measure urinary calcium in the fed and fasting states, relative expression level of some genes involved in calcium reabsorption in the distal convoluted tubule by real-time PCR, and protein expression by Western blotting or immunohistochemistry. Plasma membrane calcium ATPase (PMCA) activity was measured in kidney membrane preparations. ANOVA was used to test for differences between groups followed by posthoc analysis with Dunnett's test. RESULTS: Compared with WT, urinary Ca:Cr ratios were elevated in ArKO mice, renal mRNA levels of transient receptor potential cation channel vallinoid subfamily member 5 (TRPV5), TRPV6, calbindin-D28k, the Na+/Ca+ exchanger (NCX1), and the PMCA1b were significantly decreased, and klotho mRNA and protein levels were elevated. Estradiol treatment of ArKO mice normalized urinary calcium excretion, renal mRNA levels of TRPV5, calbindin-D(28k), PMCA1b, and klotho, as well as protein levels of calbindin-D28k and Klotho. ArKO mice treated with estradiol had significantly greater PMCA activity than either untreated ArKO mice or WT mice. CONCLUSIONS: Estrogen deficiency caused by aromatase inactivation is sufficient for renal calcium loss. Changes in estradiol levels are associated with coordinated changes in expression of many proteins involved in distal tubule calcium reabsorption. Estradiol seems to act at the genomic level by increasing or decreasing (klotho) protein expression and nongenomically by increasing PMCA activity. PMCA, not NCX1, is likely responsible for extruding calcium in response to in vivo estradiol hormonal challenge. These data provide potential mechanisms for regulation of renal calcium handling in response to changes in serum estrogen levels.

References

Hochberg, Z., T. Bick, et al. (1986). "The dual effect of calcium on aromatization by cultured human trophoblast." J Steroid Biochem 24(6): 1217-9.

Kato, S., T. Mano, et al. (2002). "A calcium-deficient diet caused decreased bone mineral density and secondary elevation of estrogen in aged male rats-effect of menatetrenone and elcatonin." Metabolism 51(10): 1230-4.

Kinuta, K., H. Tanaka, et al. (2000). "Vitamin D is an important factor in estrogen biosynthesis of both female and male gonads." Endocrinology 141(4): 1317-24.

Oz, O. K., A. Hajibeigi, et al. (2007). "Aromatase deficiency causes altered expression of molecules critical for calcium reabsorption in the kidneys of female mice *." J Bone Miner Res 22(12): 1893-902.