Effect of peri-pubertal high caffeine exposure on the growth and function of reproductive organs in immature female rats

Abstract

Previously, it has been reported that caffeine intake was inversely associated with luteal phase free estradiol level in women as well as testosterone production in immature male rats. Chronic caffeine exerts negligible effects on reproductive organ in normal female, but is unknown whether this is also true for children and adolescents. Because reproductive organs such as the ovary and uterus undergo extensive structural and functional modifications during the puberty, children and adolescents may be more vulnerable to caffeine exposure than adults. However, research on whether the caffeine consumption alters female pubertal parameters is sparse, and impacts of caffeine on the female reproductive organ growth and ovarian steroidogenesis have not been examined during the puberty. Here, the effects of high caffeine exposure on sexual maturation and ovarian estradiol production were investigated using immature female rat. Twenty-two days old female SD rats were divided into control and caffeine groups fed with 120 and 180mg/kg/day for 4 or 8 weeks. Female pubertal parameters such as vaginal opening and estrous cycle were monitored. At the end of experiment, one of ovaries from each animal was processed for histological analysis and the other was prepared for ex vivo granulosa cells (GCs) culture. In addition, GCs was prepared from 50 days old rat and cultured for determining in vitro effect of caffeine on estradiol production. Real time RT-PCR was performed to analyze aromatase and estrogen receptor genes expression in GCs and uterus, respectively. There was a significant delay in vaginal opening and 1st diestrus in caffeine-fed groups. In addition, the proportion of irregular cycle was relatively increased in caffeine-fed animals, but histological analysis of the ovary was not different in terms of proportion by follicle growing stage compared to the control. On the other hand, serum estradiol levels were significantly increased in caffeine-fed animals after 2 and 4 weeks of exposure, but comparable between groups after 8 weeks of exposure. Simultaneously, estradiol secretion was significantly increased in ex vivo and in vitro cultured GCs with caffeine treatment. In parallel to these, aromatase mRNA and protein expressions, which is responsible steroidogenic enzyme for estradiol production, were also significantly increased in caffeine-treated cells. However, ovarian weight or uterine length was substantially reduced in caffeine-exposed animals compared to the controls. These results demonstrate that peripubertal exposure to caffeine abnormally increased serum estradiol and aromatase expression in the ovary; it may disturb coordinated regulation of hypothalamo-pituitary-ovarian axis during the puberty, thereby overall interference with sexual maturation. Therefore, caffeine might work as an endocrine imbalance inducer during the puberty and further research in human and rodent models is necessary to define the impact of caffeine on a female's endocrine and reproductive health.