Early initiation of anti-androgen treatment is associated with increased probability of spontaneous conception leading to childbirth in women with polycystic ovary syndrome: a population-based multiregistry cohort study in Sweden.

STUDY QUESTION: Is anti-androgen treatment during adolescence associated with an improved probability of spontaneous conception leading to childbirth in women with polycystic ovary syndrome (PCOS)? SUMMARY ANSWER: Early initiation of anti-androgen treatment is associated with an increased probability of childbirth after spontaneous conception among women with PCOS. WHAT IS KNOWN ALREADY: PCOS is the most common endocrinopathy affecting women of reproductive age. Hyperandrogenism and menstrual irregularities associated with PCOS typically emerge in early adolescence. Previous work indicates that diagnosis at an earlier age (<25 years) is associated with higher fecundity compared to a later diagnosis. STUDY DESIGN, SIZE, DURATION: This population-based study utilized five linked Swedish national registries. A total of 15 106 women with PCOS and 73 786 control women were included. Women were followed from when they turned 18 years of age until the end of 2015, leading to a maximum follow-up of 10 years. First childbirth after spontaneous conception was the main outcome, as identified from the Medical Birth Registry. PARTICIPANTS/MATERIALS, SETTING, METHODS: Participants included all women born between 1987 and 1996 with a diagnosis of PCOS in the Swedish Patient Registry and randomly selected non-PCOS controls (ratio 1:5). Information on anti-androgenic treatment was retrieved from the Swedish Prescribed Drug Registry with the use of Anatomic Therapeutic Chemical (ATC) codes. Women with PCOS who were not treated with any anti-androgenic medication were regarded as normo-androgenic, while those treated were regarded as hyperandrogenic. Women were further classified as being mildly hyperandrogenic if they received anti-androgenic combined oral contraceptive (aaCOC) monotherapy, or severely hyperandrogenic if they received other anti-androgens with or without aaCOCs. Early and late users comprised women with PCOS who started anti-androgenic treatment initiated either during adolescence (≤ 18 years of age) or after adolescence (>18 years), respectively. The probability of first childbirth after spontaneous conception was analyzed with the use of Kaplan-Meier hazard curve. The fecundity rate (FR) and 95% confidence interval for the time to first childbirth that were conceived spontaneously were calculated using Cox proportional hazards regression models, with adjustment for obesity, birth year, country of birth and education level. MAIN RESULTS AND THE ROLE OF CHANCE: The probability of childbirth after spontaneous conception in the PCOS group compared to non-PCOS controls was 11% lower among normo-androgenic (adjusted FR 0.68 (95% CI 0.64-0.72)), and 40% lower among hyperandrogenic women with PCOS (adjusted FR 0.53 (95% CI 0.50-0.57)). FR was lowest among severely hyperandrogenic women with PCOS compared to normo-androgenic women with PCOS (adjusted FR 0.60 (95% CI 0.52-0.69)), followed by mildly hyperandrogenic women with PCOS (adjusted FR 0.84 (95% CI 0.77-0.93)). Compared to early anti-androgenic treatment users, late users exhibited a lower probability of childbirth after spontaneous conception (adjusted FR 0.79 (95% CI 0.68-0.92)). LIMITATIONS, REASONS FOR CAUTION: We lacked direct information on the intention to conceive and the androgenic biochemical status of the PCOS participants, applying instead the use of anti-androgenic medications as a proxy of hyperandrogenism. The duration of anti-androgenic treatment utilized is not known, only the age at prescription. Results are not adjusted for BMI, but for obesity diagnosis. The period of follow-up (10 years) was restricted by the need to include only those women for whom data were available on the dispensing of medications during adolescence (born between 1987 and 1996). Women with PCOS who did not seek medical assistance might have been incorrectly classified as not having the disease. Such misclassification would lead to an underestimation of the true association between PCOS and outcomes. WIDER IMPLICATIONS OF THE FINDINGS: Early initiation of anti-androgen treatment is associated with better spontaneous fertility rate. These findings support the need for future interventional randomized prospective studies investigating critical windows of anti-androgen treatment. STUDY FUNDING/COMPETING INTEREST(S): This study was funded by the Health Research Council of New Zealand (18-671), the Swedish Society of Medicine and the Uppsala University Hospital. Evangelia Elenis has, over the past year, received lecture fee from Gedeon Richter outside the submitted work. Inger Sundström Poromaa has, over the past 3 years, received compensation as a consultant and lecturer for Bayer Schering Pharma, MSD, Gedeon Richter, Peptonics and Lundbeck A/S. The other authors declare no competing interests. TRIAL REGISTRATION NUMBER: N/A.

Immunohistochemical evidence for the presence of various kisspeptin isoforms in the mammalian brain.

Kisspeptins are small peptides encoded by the Kiss1 gene that have been the focus of intense neuroendocrine research during the last decade. Kisspeptin is now considered to have important roles in the regulation of puberty onset and adult oestrogen-dependent feedback mechanisms on gonadotrophin-releasing hormone secretion. Several kisspeptin antibodies have been generated that have enabled an overall view of kisspeptin peptide distribution in the brain of many mammalian species. However, it remains that the distribution of the different kisspeptin isoforms is unclear in the mammalian brain. In the present study, we report on two new N-terminal-directed kisspeptin antibodies, one against the mouse kisspeptin-52 sequence (AC053) and one against the rat kisspeptin-52 sequence (AC067), and use them to specifically map these long isoforms in the brains of mouse and rat, respectively. Kisspeptin-52 immunoreactivity was detected in the two main kisspeptin neuronal populations of the rostral periventricular area and arcuate nucleus but not in the dorsomedial hypothahamus. A large number of fibres throughout the ventral forebrain were also labelled with these two antibodies. Finally, a comparison with the most commonly used C-terminal-directed kisspeptin antibodies further suggests the presence of shorter kisspeptin fragments in the brain with specific inter- and intracellular expression patterns.

Embryonic development of kisspeptin neurones in rat.

Kisspeptins, encoded by the Kiss1 gene, play a key role in the regulation of reproductive function, although very little is known about the ontogenesis of this system. The present study aimed to determine the period of arcuate nucleus (ARC) kisspeptin cell birth and the embryonic stage and neuroanatomical sites of onset of kisspeptin immunoreactivity. Bromodeoxyuridine (BrdU) was administered to female rats at various gestational stages and double immunohistochemistry against kisspeptin and BrdU was performed on brain sections from their offspring. The period of neurogenesis of ARC kisspeptin neurones begun between embryonic day (E) 12.5 and E13.5, reached its peak at E15.5 and was not completely over at E17.5. Kiss1 mRNA was detected in mediobasal hypothalamic punches of embryos aged E14.5, E16.5, E18.5 and E22.5 by real-time reverse transcriptase-polymerase chain reaction. Accordingly, kisspeptin-immunoreactive (-IR) cells were consistently detected in the embryonic ARC from E14.5 and their number increased until E18.5 to reach approximately half the level observed in adults. Between E18.5 and E22.5, the number of kisspeptin-IR cells and hypothalamic Kiss1 expression significantly decreased, regardless of sex, and this decrease persisted until birth. Taken together, these results demonstrate that rat ARC kisspeptin neurones are born locally during an extended embryonic period and are able to synthesise kisspeptins rapidly after their birth, consistent with the hypothesis of a role during embryonic activation of the hypothalamic-hypophyseal-gonadal axis. A sex-independent decrease of kisspeptin-IR cell numbers was observed during the perinatal period, suggestive of important regulations of kisspeptin neurones around birth.

Kisspeptin-immunoreactivity changes in a sex- and hypothalamic-region-specific manner across rat postnatal development.

Kisspeptins are potent secretagogues of gonadotrophin-releasing hormone, playing a key role in puberty onset. These peptides are produced by distinct neuronal populations of the hypothalamus located in the rostral periventricular area of the third ventricle (RP3V) and arcuate nucleus (ARC). The present immunohistochemical study aimed to determine the spatiotemporal onset of kisspeptin-immunoreactivity (-IR) in the neonatal hypothalamus of male and female rats and to evaluate changes in kisspeptin-IR around puberty. Kisspeptin-IR cells and fibres could be detected from the day of birth in the ARC of both males and females. At this stage, only females displayed some kisspeptin-IR fibres in the RP3V. From postnatal day 7 to adulthood, males displayed lower levels of kisspeptin-IR than females in both regions. During infancy, kisspeptin-IR fibre density in the female decreased in the ARC, whereas it increased in the RP3V. A sex-independent decline in RP3V kisspeptin-IR fibre density was observed in the juvenile, followed by a peripubertal increase in RP3V and ARC kisspeptin-IR. These peripubertal increases in kisspeptin-IR occurred at different timings dependent on sex and region. In females specifically, the increase in kisspeptin-IR fibre density occurred first in the ARC and later in the RP3V under constant levels of circulating oestradiol. In conclusion, the present study highlights the expression of hypothalamic kisspeptins soon after birth, as well as the neonatal establishment of a strong and persisting sex difference in ARC kisspeptin-IR in rats. Moreover, a female-specific desynchronisation of the ARC and RP3V was observed with respect to the increase in kisspeptin-IR fibre density around puberty, which was not related to peripubertal variations in circulating oestradiol.

Mapping of kisspeptin fibres in the brain of the pro-oestrous rat.

Kisspeptins are a family of small peptides that play a key role in the neuroendocrine regulation of the reproductive function through neural pathways that have not yet been completely identified. The present study aimed to investigate the distribution of kisspeptin neurone fibres in the female rat brain by comparing precisely the immunoreactive pattern obtained with two antibodies: one specifically directed against kisspeptin-52 (Kp-52), the longest isoform, and the other directed against kisspeptin-10 (Kp-10), whose sequence is common to all putative mature isoforms. With both antibodies, immunoreactive cell bodies were exclusively observed in the arcuate nucleus, and immunoreactive fibres were confined to the septo-preoptico-hypothalamic continuum of the brain. Fibres were observed in the preoptic area, the diagonal band of Broca, the septohypothalamic area, the anteroventral periventricular, suprachiasmatic, supraoptic, paraventricular and periventricular nuclei, the dorsal border of the ventromedian nucleus, the dorsomedial and arcuate nuclei, and the median eminence. In the latter structure, varicose fibres were mainly distributed in the internal layer and were detected to a lesser extent throughout the external layer, including around the deeper part of the infundibular recess. Most regions of immunoreactive cells and fibres matched perfectly for the two antibodies. However, fibres in the dorsolateral septum, anterior fornix, accumbens nucleus and the lateral bed nucleus of the stria terminalis were only recognised by antibody anti-Kp-10, suggesting that anti-Kp-10 may recognise a wider range of kisspeptin isoforms than anti-Kp-52 or cross-react with molecules other than kisspeptin in rat tissue. Overall, these results illustrate the variety of projection sites of kisspeptin neurones in the rat and suggest that these peptides play a role in different functions.