P. Govindarajulu

Testosterone and estradiol up-regulate androgen and estrogen receptors in immature and adult rat thyroid glands in vivo.

Thyroid gland is one of the non-classical target organs for sex steroids. Presence of androgen and estrogen receptors in the neoplastic and non-neoplastic thyroid glands of mammalian species is well documented. The aim of the present study is to elucidate the changes in serum and thyroidal sex steroids, and their receptors in the thyroid gland of rats from immature to adult age under gonadectomized (GDX) and sex steroids replaced conditions. Normal Wistar male and female rats from immature to adult age (day 21, 30, 45, 60 and 160 post-partum (pp)) were used in the present study. One group (I) of rats was GDX at an early age (day 10 pp) and the other group (II) at the adult age (day 120 pp). Group I rats were sacrificed at different experimental periods such as 21, 30, 45 and 60 days pp, and group II rats were sacrificed at day 160 pp. Another group of GDX rats from group I and II were replaced with physiological doses of testosterone or estradiol. Serum and thyroidal concentrations of sex steroids were estimated by RIA method and the concentrations of receptors by radioreceptor assay. Gonadectomy significantly decreased serum and thyroidal testosterone and estradiol and concentrations of androgen receptor (AR) and estrogen receptor (ER) in the thyroid. Replacement of sex steroids to GDX rats restored the normal level of sex steroids, AR and ER. Therefore, it is suggested from the present study that (i). sex steroids up-regulate their own receptors in the thyroid, (ii). sex steroids may influence thyroid growth and the proliferation of thyrocytes by modulating their receptor concentrations in the thyroid.

Developmental profiles of TSH, sex steroids, and their receptors in the thyroid and their relevance to thyroid growth in immature rats

Sex steroids are reported to influence thyroid pathogenesis in human and experimental animals. However, there is no report on this phenomenon during the early developmental period. The mitotic activity of thyrocytes in rats reaches its peak by day 10 postpartum. Thyrocytes actively proliferate in immature rats during the first three postnatal weeks, during which the pre-pubertal rise in serum titers of testosterone and estradiol has been recorded. The aim of the present study was to analyze whether there is a physiological relevance between thyroid growth and sex steroids during the postnatal period. Serum and thyroid tissue hormones (TSH, testosterone, and estradiol) were assayed by liquid phase RIA, and receptors for these hormones were also quantified. The peak rate of thyrocyte proliferation was observed during the second postnatal week in rats. Since the concentrations of sex steroids and their receptors also reached a peak around this period, it is suggested that elevated sex steroids and their receptors in the thyroid might enhance thyrocyte proliferation. A positive correlation between thyroid growth indices and sex steroids and their receptors further strengthens this suggestion. This is a preliminary study, and further experimental study may strengthen this proposal. This is the first report to show the availability of sex steroids and their receptors in the thyroid glands of immature rats under normal conditions.

Testosterone and estradiol differentially regulate TSH-induced thyrocyte proliferation in immature and adult rats.

Though sex steroids are found to influence thyroid pathogenesis in human and in animals, their role in normal thyroid growth and thyrocyte proliferation is not yet understood fully. The present study is addressed to know the effect of testosterone and estradiol on the basal and TSH-induced thyrocyte proliferation in immature and adult rats in vitro. The male and female Wistar rats were gonadectomized (GDX) and one group of GDX rats were supplemented with either testosterone or estradiol. After the experimental period, the rats were sacrificed by decapitation and thyroid glands were removed, washed in Hanks Balanced Salt Solution (HBSS), pH 7.4 and digested with the enzyme mixture containing 0.08% collagenase and 0.12% dispase in HBSS. The isolated follicles were washed thrice with Dulbeccos modified Eagles medium (DMEM) containing 0.5% fetal bovine serum (FBS), and were cultured in Falcons tissue culture flasks containing 5 ml DMEM with FBS (5%) transferrin (5 microg/ml), hydrocortisone (10(-8) M), somatostatin (10 microg/ml), insulin (10 microg/ml) and glycyl-L-histidyl-L-lysine acetate (10 microg/ml). The cells (2.5 x 10(4)) were exposed to various exponential doses of TSH or testosterone (6.25-800 ng/ml) or estradiol (6.25-800 pg/ml). It is suggested from the present study that both TSH and sex steroids enhance thyrocyte proliferation. The mitogenic effect of TSH is greater than that of sex steroids. Sex steroids modulate TSH-induced cell proliferation in a gender-specific manner.

TESTOSTERONE AND ESTRADIOL DIFFERENTIALLY REGULATE THYROID GROWTH IN WISTAR RATS FROM IMMATURE TO ADULT AGE

Experimental and clinical data from cancerous thyroid tissue suggest gender as one of the major factors influencing thyroid growth. However, there is not much information on the effect of sex steroids on the normal development and growth of the thyroid. The aim of the present study was to examine the effect of sex steroids on thyroid growth, serum TSH and TSH receptors in the thyroid, at various stages of sexual maturation. Normal Wistar male and female rats from immature to adult age (day 21, 30, 45, 60 and 160 post partum (PP)) were used in the present study. One group of rats was gonadectomized (GDX) (Group II); another 2 groups of GDX rats (Groups III and IV) were treated with replacement doses (i.e. physiological amounts) of either testosterone or estradiol. Thyroid growth indices (thyroid weight, concentration of DNA, mitotic index and numerical density of thyrocytes), serum TSH and concentration of TSH receptors in the thyroid were assessed. It is concluded from the present study that (i) sex steroids influence thyroid growth by interfering with the secretion of TSH and regulating its receptor number in the thyroid (ii) while testosterone facilitates the mitogenic activity of the thyroid gland of rats irrespective of sex and age, estradiol plays a gender-specific stimulatory role on thyroid growth.

Testosterone and estradiol modulate TSH-binding in the thyrocytes of Wistar rats: influence of age and sex

Age and sex are important factors that influence thyroid pathophysiology. Though sex steroids are known to enhance thyrotropin (TSH) mRNA expression and incidence of thyroid tumours, there is no report on their effects on TSH action under normal physiological conditions. In the present study, the effects of testosterone (T) and estradiol (E2) on thyroidal TSH-receptor (TSH-R) concentration, and TSH-binding to thyrocytes (in vitro) were elucidated in immature and mature Wistar rats. Immature (10 days old) and adult (120 days old) rats of either sex were gonadectomized (GDX) and one group of GDX rats was treated with physiological doses of T and another with E2. Immature GDX rats were supplemented with the steroids for 10 days and adults were supplemented with the steroids for 30 days. While supplementation of steroids to immature rats was begun immediately after surgery, for adult rats it was started 10 days after gonadectomy. The rats were killed at the end of the experimental period. Gonadectomy significantly decreased serum TSH, and TSH-R concentration under in vivo condition and [125I]-TSH binding to thyrocytes under in vitro conditions. Supplementation of T to male and E2 to female GDX rats restored normality of the parameters. Thyrocytes of immature male rats challenged with linearly increasing doses of TSH or T (6.25-800 ng/ml) showed a dose-dependent increase in TSH-binding. However, thyrocytes of immature female rats challenged with T showed a gender-specific response. While there was a linear increase in TSH-binding in thyrocytes of males, a biphasic response was evident in thyrocytes of females. In the case of thyrocytes from adult rats, T induced a dose-dependent change in TSH-binding in males, which reached the peak in response to 12.5 ng T, and diminished thereafter. In contrast, E2 was inhibitory to TSH-binding to thyrocytes of adult male rats. On the other hand, E2 showed a clear gender-specific stimulation of TSH-binding in thyrocytes of females and an inhibition of the same in males. TSH and sex steroids upregulated TSH receptors in immature rats, whereas the effect was biphasic in adult rat thyrocytes. It is concluded from the present study that sex steroids modulate TSH-binding in rat thyrocytes, which may vary according to the age and sex of the animals.

Testosterone and estradiol have specific differential modulatory effect on the proliferation of human thyroid papillary and follicular carcinoma cell lines independent of TSH action.

Differential effects of testosterone and estradiol on the proliferation of human thyroid papillary (NPA-87-1) and follicular (WRO-82-1) carcinoma cell lines were assessed by [3H]-thymidine incorporation and the cell number. Cells (2.5 × 105) plated in 24-well culture plates in 400 µL RPMI-1640 medium/well, under 5% CO2 and 95% air; at 37°C were exposed to linearly increasing concentrations of human thyroid-stimulating hormone (hTSH) (1.25–640 ng/mL), testosterone (1.25–640 ng/mL), or estradiol (1.25–640 pg/mL) for 24 h. Testosterone and estradiol increased the proliferation of NPA cell line in a dose-dependent manner; flutamide (an anti-androgen) and tamoxifen (an anti-estrogen) (10−8, 10−7, 10−6, and 10−5 mol/L) effectively inhibited the testosterone and estradiol-induced cell proliferation, respectively. While flutamide inhibited the stimulatory effect of testosterone on the WRO cell line, tamoxifen augmented the inhibitory effect of estradiol. TSH did not have any effect on the proliferation of NPA or WRO cell lines, and testosterone-estradiol had no impact on TSH binding to these cells. N-ethylmalemide (5α-reductase inhibitor) (10−8–10−5 mol/L) did not modulate basal and testosterone-induced cell proliferation, indicating the direct effect of testosterone without getting converted into dihydrotestosterone (DHT). Both the cell lines tested positive for androgen and estrogen receptors and were up-regulated by the respective ligands. It is concluded that testosterone and estradiol modify the proliferation of thyroid cancer cells through homologous up-regulation of their own receptors, which is independent of TSH, and their effects may vary according to the cell type.

Modulatory effect of estradiol and testosterone on the development of N-nitrosodiisopropanolamine induced thyroid tumors in female rats.

Both experimental and clinical research support the conclusion that thyroid tumors are sex dependent. Also, several studies have pointed out that the use of oral contraceptives is associated with a higher risk of thyroid tumor. Most of the existing reports suggest indirect effects of sex steroids on thyroid tumor growth in women. In this work, we present data to support the direct promoting effect of estradiol and testosterone on carcinogen‐induced thyroid tumorigenesis. Thyroid tumors were induced in rats by a combination of N‐nitrosodiisopropanolamine (DHPN) and phenobarbital (PB) treatment. Serum thyroid hormones, thyroid stimulating hormone (TSH), steroid hormones, thyroidal steroid concentration, androgen and estrogen receptors were quantified. Serum thyroid hormones and TSH suggested euthyroid status of the all experimental animals. Ovariectomy decreased the incidence of DHPN + PB induced thyroid tumor when compared with ovary intact rats and estradiol/testosterone supplementation increased the same. Thyroidal estradiol level and its nuclear receptors increased in the tumor tissue specifically. Testosterone supplementation to DHPN‐treated ovariectomized rats specifically induced the development of malignant thyroid tumors. Addition of estradiol in vitro to thyrocytes induced a higher proliferation rate. Our data proves a direct promoting role of estrogen on carcinogen‐induced thyroid tumor development.

Effects of piperine on the lipid composition and enzymes of the pyruvate-malate cycle in the testis of the rat in vivo.

Effects of piperine at two oral doses (5 and 10 mg/kg body weight for 30 days) on the lipid composition and some lipogenic enzymes of the rat testis were studied. Piperine treatment depleted the total lipid content which was mainly due to the diminution of the total phospholipid concentration. All the classes of phospholipids were decreased markedly following high dose piperine treatment. In contrast, a marked increase in total cholesterol and cholesterol ester was evident with a concomitant fall in free cholesterol. A similar trend was found for the total glyceride glycerol and its fractions. Total glyceride glycerol and triacyl glycerol showed a significant increase at the expense of diacyl glcyerol in rats treated with the high dose of pipeline. Lipogenic enzymes, malate dehydrogenase (MDH), malic enzyme (ME) and isocitrate dehydrogenase (ICDH) were inhibited by the high dose and only MDH and ME activities were inhibited by the low dose treatment.

Impact of altered thyroid hormone status on prostatic glycosidases

The impact of hyper- and hypothyroidism on prostatic glycosidases was investigated. Hyper-thyroidism was induced by administering L-thyroxine (25 micrograms/100 g body weight/day) for 60 days and hypothyroidism was induced by total thyroidectomy. To test the direct influence of thyroid hormones, prostatic lobes were incubated with different concentrations (10, 25 and 50 ng/mL) of T3 and beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase were assayed. Serum levels of thyroid hormones, oestradiol and testosterone increased in hyperthyroid, and decreased in hypothyroid rats. TSH decreased in hyperthyroid, and an opposite trend was seen in thyroidectomized, rats. Prostatic [anterior (coagulating glands), dorsolateral and ventral prostates] beta-glucosidase, beta-galactosidase, beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase activities increased uniformly in hyperthyroid, and decreased in thyroidectomized, rats. In vitro studies showed a dose-dependent stimulatory effect of T3 on beta-N-acetylglucosaminidase and beta-N-acetylgalactosaminidase in all three lobes of the prostate. From the present study, it is concluded that hyperthyroidism augments and hypothyroidism inhibits prostatic glycosidases and T3 has a direct stimulatory effect on these enzymes.

Prostate-thyroid axis: Stimulatory effects of ventral prostate secretions on thyroid function

Endocrine cells of the prostate secrete thyrotropin (TSH)‐releasing hormone (TRH), TRH‐ and TSH‐like peptides, and growth factors. Propylthiouracil‐ and methimazole‐induced hypothyroidism increased prostatic levels of TRH in rats as in hypothalamus, whereas thyroxine (T4) replacement decreased TRH. From these reports, we inferred the existence of a prostate‐thyroid axis.