TSH-producing Cells Research Articles

Characterization and co-localization of thyroid stimulating hormone and melatonin receptors in the pars tuberalis of mares during the breeding and non-breeding seasons

Control of seasonal reproduction in horses has yet to be fully elucidated; however, duration of melatonin secretion is a seasonal regulator of the hypothalamic-pituitary-gonadal (HPG) axis. The hypophyseal pars tuberalis (PT) appears to play a key role in the transduction of melatonin signal through melatonin responsive, PT-specific cells that produce thyroid stimulating hormone (TSH). The objectives of these experiments were to characterize and colocalize TSH-producing cells and melatonin receptors (MT1r) in the PT during breeding and non-breeding seasons. Pituitaries from nine light-horse type mares, aged 6 to 26 years, were obtained following euthanasia during the breeding and non-breeding seasons. Mares were determined to be either cycling (n = 5) or seasonally anovulatory (n = 4) based on season, gross inspection of ovarian structures, and plasma progesterone concentrations. Immediately following euthanasia, the carotid artery was perfused with sodium nitrite and 4% paraformaldehyde before decapitation and dissection of PT from the hypophysis. Immunofluorescent detection of TSH was carried out on fixed-frozen PT sections and number of immunopositive cells per 1 mm2 were counted and averaged for three replications per horse to compare TSH distribution between breeding and non-breeding season using Mann Whitney U test. TSH-immunoreactive cells were present in PT and were greater (P < 0.05) in PT during the breeding season comparedto non-breeding season; few to no immunoreactive cells were seen in PT from non-breeding season. In situ hybridization (ISH) was carried out using RNAscope HD Red Detection Kit and a custom MT1r probe (Ec-MTNR1A) to confirm MT1r localization and compare MT1r distribution between breeding and non-breeding seasons. Mean percentage of MT1r immunopositive cells, determined as a percentage of total number of cells, and expression of MT1r mRNA per cell, determined as mean number of dots (representing mRNA) per cell, were analyzed using Mann Whitney U test. Melatonin receptor mRNA was densely expressed in glandular cells of the PT and no differences were found in either percent immunopositive cells or mean number of mRNA dots per cell between breeding and non-breeding seasons. To determine if MT1r colocalize with TSH-producing cells, ISH was carried out alongside immunofluorescent detection of TSH as previously described. In cycling mares, MT1r mRNA colocalized with TSH-producing cells in the PT; however, not all TSH-producing cells colocalized with MT1r and vice versa. Lack of TSH-producing cells accounted for absence of colocalization in seasonally anovulatory mares. The PT is considered a major target for melatonin, and colocalization with TSH supports a role for TSH as a modulator of seasonal reproduction in the mare. This is further evidenced by increased TSH immunosignal during the breeding season.

A new insight into GH regulation and its disturbance from nutrition and autoimmune perspectives.

GH activates GH receptors, which activates IGF-1 in the liver through a cascade of processes. The GH/IGF-1 axis plays an important role in the regulation of metabolism. Insufficient GH secretion results in short stature in childhood, while adult GH deficiency (AGHD) is observed in adulthood. The early diagnosis of AGHD is important for early initiation of GH replacement therapy. This review described the regulatory mechanisms of GH signaling based on nutritional status and a novel disease concept pathogenesis that causes AGHD. GH-dependent IGF-1 production in the liver is regulated by a complex interplay between nutritional status, hormones, and growth factors. GH resistance is an adaptive response that enhances survival during starvation and malnutrition. Sirtuin 1 (SIRT1) negatively regulates GH-induced IGF-I production in the liver by directly inhibiting STAT5 activation, which causes GH resistance under starvation and malnutrition. The presence of autoantibodies is strongly associated with the disruption of immune tolerance in pituitary cells. Pituitary-specific transcription factors (PIT-1) are essential for the development, differentiation, and maintenance of GH, PRL, and TSH producing cells. However, the underlying mechanism that causes immune intolerance to PIT-1 remain unclear. The GH-IGF-1 system plays a pivotal role in growth, and the involvement of SIRT1 in this regulatory mechanism presents an intriguing perspective on the interplay between nutrient metabolism and lifespan. The discovery of the anti-PIT-1 pituitary antibody, a novel disease concept associated with AGHD, has provided valuable insights, which serves as a significant milestone towards unraveling the complete pathogenesis of the disease.

Fluctuation of CD9/SOX2-positive cell populations during the turnover of GH- and TSH-producing cells in the adult anterior pituitary gland.

The adenohypophysis is comprised of the anterior and intermediate lobes (AL and IL, respectively). Cluster of differentiation 9 (CD9)- and sex-determining region Y-box 2 (SOX2)-positive cells are stem/progenitor hormone-producing cells in the AL. They are located in the marginal cell layer (MCL) facing Rathke's cleft between the AL and IL (primary niche) and the parenchyma of the AL (secondary niche). We previously showed that, in rats, CD9/SOX2-positive cells in the IL side of the MCL (IL-side MCL) migrate to the AL side (AL-side MCL) and differentiate into prolactin-producing cells (PRL cells) in the AL parenchyma during pregnancy, lactation, and diethylstilbestrol treatment, all of which increase PRL cell turnover. This study examined the changes in CD9/SOX2-positive stem/progenitor cell niches and their proportions by manipulating the turnover of growth hormone (GH)- and thyroid-stimulating hormone (TSH)-producing cells (GH and TSH cells, respectively), which are Pit1 lineage cells, as well as PRL cells. After induction, the isolated CD9/SOX2-positive cells from the IL-side MCL formed spheres and differentiated into GH and TSH cells. We also observed an increased GH cell proportion upon treatment with GH-releasing hormone and recovery from continuous stress and an increased TSH cell proportion upon propylthiouracil treatment, concomitant with alterations in the proportion of CD9/SOX2-positive cells in the primary and secondary niches. These findings suggest that CD9/SOX2-positive cells have the potential to supply GH and TSH when an increase in GH and TSH cell populations is required in the adult pituitary gland.

A novel thymoma-associated autoimmune disease: Anti-PIT-1 antibody syndrome

Anti-PIT-1 antibody syndrome has recently been reported and characterized by acquired growth hormone (GH), prolactin (PRL), and thyroid-stimulating hormone (TSH) deficiencies associated with autoimmunity to a pituitary specific transcription factor PIT-1, which plays an essential role in GH-, PRL-, and TSH-producing cells. Although circulating anti-PIT-1 antibody and PIT-1-reactive cytotoxic T cells (CTLs) were detected in the patients, the pathophysiology and precise mechanisms for the autoimmunity remain unclarified. During the follow up, thymoma was diagnosed in all 3 cases with anti-PIT-1 antibody syndrome. Immunohistochemical analysis revealed that PIT-1 was strongly expressed in neoplastic cortical thymic epithelial cells. Importantly, after thymectomy, the titer of anti-PIT-1 antibody decreased and reactivity of CTLs toward PIT-1 diminished. These data strongly suggest that the aberrant expression of PIT-1 in the thymoma plays a causal role in the development of this syndrome. Thus, we define that this syndrome is a novel thymoma-associated autoimmune disease.

Impact of isolated TSH levels in and out of normal range on different tissues

Routine treatment of thyroid cancer (TC) includes long-term suppression of TSH. The necessity of this treatment in low- and intermediate-risk patients as well as the extent of TSH suppression is currently under discussion. A literature search was performed to illustrate the role of TSH in extrathyroidal cells and to identify potential reasons for different effects of exogenously suppressed and endogenously low TSH levels. Although adverse effects of subnormal and supranormal TSH blood levels on heart and brain have not been consistently found, studies show a clear negative effect of suppressed TSH levels on bone mineral density. Experimental data also support an important role of TSH in the immune system. The ability of levothyroxine (L-T4) to regulate TSH levels and triiodothyronine levels in a physiological manner is limited. Reduction of circadian changes in TSH levels, decrease of thyroid hormone-binding proteins, prevention of potential compensatory increases of TSH levels (e.g., in old age), and unresponsiveness of TSH-producing cells to TRH on L-T4 treatment might cause adverse effects of suppressed TSH levels. In view of the adverse effects of aggressive TSH suppression, achieving the suggested levels of TSH between 0.9 and 1 mU/l in the treatment of low-to-intermediate risk TC patients appears justified.

MECHANISMS IN ENDOCRINOLOGY: Impact of isolated TSH levels in and out of normal range on different tissues.

Routine treatment of thyroid cancer (TC) includes long-term suppression of TSH. The necessity of this treatment in low- and intermediate-risk patients as well as the extent of TSH suppression is currently under discussion. A literature search was performed to illustrate the role of TSH in extrathyroidal cells and to identify potential reasons for different effects of exogenously suppressed and endogenously low TSH levels. Although adverse effects of subnormal and supranormal TSH blood levels on heart and brain have not been consistently found, studies show a clear negative effect of suppressed TSH levels on bone mineral density. Experimental data also support an important role of TSH in the immune system. The ability of levothyroxine (l-T4) to regulate TSH levels and triiodothyronine levels in a physiological manner is limited. Reduction of circadian changes in TSH levels, decrease of thyroid hormone-binding proteins, prevention of potential compensatory increases of TSH levels (e.g., in old age), and unresponsiveness of TSH-producing cells to TRH on l-T4 treatment might cause adverse effects of suppressed TSH levels. In view of the adverse effects of aggressive TSH suppression, achieving the suggested levels of TSH between 0.9 and 1 mU/l in the treatment of low-to-intermediate risk TC patients appears justified.

Anterior Pituitary Morphology and Hormone Production During Sustained Critical Illness in a Rabbit Model

Prolonged critical illness is hallmarked by striking alterations in the somatrope, thyrotrope, and lactotrope axes, the severity of which is associated with the risk of morbidity and mortality. The exact role of the pituitary gland in these alterations is unknown. We studied the impact of sustained critical illness on pituitary morphology and hormone production in a standardized rabbit model of prolonged (7 days) burn injury-induced critical illness. In healthy and prolonged critically ill rabbits, we determined pituitary weight, size, morphology and orientation of the somatrope, lactotrope and thyrotrope cells and the pituitary expression of GH, PRL, and TSH at gene and protein level. The weight of the pituitary gland was unaltered by 7 days of critical illness. Also, spatial orientation and morphology of the GH, PRL, and TSH producing cells remained normal. In prolonged critically ill rabbits GH mRNA levels were higher and PRL mRNA levels were lower than in healthy controls, whereas TSH mRNA was not affected. The sizes of GH, PRL, or TSH producing cells and the pituitary content of GH, PRL, and TSH proteins were unaltered. In conclusion, in this rabbit model of prolonged critical illness, the morphology of the pituitary gland and the pituitary GH, PRL, and TSH content was normal. The alterations in pituitary hormone mRNA levels with sustained critical illness are compatible with altered hypothalamic and peripheral regulation of pituitary hormone release as previously suggested indirectly by responses to exogenous releasing factors.

Effects of the anti-thyroidal compound potassium-perchlorate on the thyroid system of the zebrafish

The increasing pollution of aquatic habitats with anthropogenic compounds has led to various test strategies to detect hazardous chemicals. However, information on effects of pollutants in the thyroid system in fish, which is essential for growth, development and parts of reproduction, is still scarce. Other vertebrate groups such as amphibians or mammals are well-studied; so the need for further knowledge especially in fish as a favored vertebrate model test organism is evident. Modified early life-stage tests were carried out with zebrafish exposed to the known thyroid inhibitor potassium perchlorate (0, 62.5, 125, 250, 500 and 5000μg/L) to identify adverse effects on the hypothalamic-pituitary–thyroid axis. Especially higher perchlorate concentrations led to conspicuous alterations in thyroidal tissue architecture and to effects in the pituitary. In the thyroid, severe hyperplasia at concentrations ≥500μg/L together with an increase in follicle number could be detected. The most sensitive endpoint was the colloid, which showed alterations at ≥250μg/L. The tinctorial properties and the texture of the colloid changed dramatically. Interestingly, effects on epithelial cell height were minor. The pituitary revealed significant proliferations of TSH-producing cells resulting in alterations in the ratio of adeno- to neurohypophysis. The liver as the main site of T4 deiodination showed severe glycogen depletion at concentrations ≥250μg/L. In summary, the thyroid system in zebrafish showed effects by perchlorate from concentrations ≥250μg/L, thus documenting a high sensitivity of the zebrafish thyroid gland for goitrogens. In the future, such distinct alterations could lead to a better understanding and identification of potential thyroid-disrupting chemicals.

Chapter Nine - Constitutively Active Thyrotropin and Thyrotropin-Releasing Hormone Receptors and Their Inverse Agonists

Receptors for thyrotropin-releasing hormone (TRH) and thyrotropin (thyroid-stimulating hormone-TSH) are important regulators of the function of the TSH-producing cells of the anterior pituitary gland and the thyroid gland, respectively, and thereby play a central role in thyroid hormone homeostasis. Although the roles of TRH- and TSH-stimulated signaling in these endocrine glands are well understood, these receptors are expressed in other sites and their roles in these extraglandular tissues are less well known. Moreover, one of the two subtypes of TRH receptors (TRH-R2) and the single TSH receptor (TSHR) exhibit constitutive signaling activity and the roles of constitutive signaling by these receptors are poorly understood. One approach to studying constitutive signaling is to use inverse agonists. In this chapter, we will describe the experimental procedures used to measure constitutive signaling by TRH-R2 and TSHR and the effects of their specific inverse agonists.

Expression of Adiponectin and Adiponectin Receptors in Human Pituitary Gland and Brain

Background/Aims: Adiponectin and its receptors, AdipoR1 and AdipoR2, constitute integral components of energy homeostatic mechanism in peripheral tissues. Recent studies have implicated adiponectin in central neural networks regulating food intake and energy expenditure. The present study aimed at investigating the possible expression and distribution of adiponectin and its receptors in human pituitary gland, hypothalamus and different brain areas. Methods: Sections of the pituitary gland, hypothalamus and adjacent basal forebrain area, cerebrum and cerebellum from 35 autopsy cases, were examined using HE, PAS-Orange G, luxol fast blue/cresyl violet stains and single and double immunohistochemistry using adiponectin, AdipoR1, AdipoR2, choline acetyltransferase, FSH, LH, TSH, GH, ACTH and prolactin-specific antibodies. Age and BMI mean values ± SD of the autopsy cases were 56 ± 18 years and 27 ± 5 kg/m², respectively. Results: Strong adiponectin expression was observed in pituitary gland. In pars distalis (PD), adiponectin localized in GH, FSH, LH and TSH-producing cells and in pars tuberalis (PT) in FSH, LH and TSH-producing cells. Strong to moderate expression of AdipoR1 and AdipoR2 was observed in PD by the same cell types as adiponectin. No immunoreactivity for adiponectin receptors was noted in cells of PT. Intense AdipoR1 immunostaining was observed in neurons of lateral hypothalamic area and of nucleus basalis of Meynert (NBM). Conclusions: Adiponectin and its receptors expression in human pituitary might indicate the existence of a local system, modulating endocrine axes. Furthermore, the presence of AdipoR1 in hypothalamus and NBM suggests that adiponectin may participate in central neural signaling pathways controlling energy homeostasis and higher brain functions.

Expression of MSX1 in Human Normal Pituitaries and Pituitary Adenomas

Transcription factors play specific roles in the development and differentiation of normal pituitary tissues and pituitary adenoma. The transcription factor, muscle segment homeobox 1 (MSX1), which belongs to the homeobox gene family, binds the promoter region of the glycoprotein hormone alpha-subunit (SU) in TSH-producing cells in the mouse pituitary and regulates alpha-SU expression. The present study investigated MSX1 expression in the normal human pituitary. In addition, 50 pituitary adenomas were examined using immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR) to clarify the role of MSX1 in the development and functional differentiation of pituitary adenoma cells. In the normal pituitary, MSX1 was predominantly expressed in the cytoplasm of GH-producing cells. Furthermore, MSX1 immunoreactivity was observed in the cytoplasm of some alpha-SU-producing cells. It is interesting to note that, in the pituitary adenoma, MSX1 was expressed in the nucleus of GH- and TSH-producing adenomas. RT-PCR using RNA extracted and purified from formalin-fixed paraffin-embedded pituitary adenoma specimens revealed MSX1 mRNA expressed in GH- and TSH-producing adenomas. Immunoelectron microscopy demonstrated MSX1 localized at intranuclear heterochromatin and euchromatin, which suggests transcriptional activity. These results suggest that MSX1 plays a specific role in human pituitary adenoma.

Immunoreactive TSH cells in juvenile and peripubertal rats after estradiol and human chorionic gonadotropin treatment

Different hormones and growth factors control the homeostasis of the anterior pituitary gland. We examined the morphological features of pituitary thyroid stimulating hormone (TSH)-producing cells in juvenile and peripubertal female rats after treatments with estradiol-dipropionate (EDP), human chorionic gonadotropin (hCG) and a combination of both hormones (EDP+hCG). TSH-producing cells were labelled using a peroxidase-antiperoxidase immunohistochemical procedure for binding of a rabbit anti-rat beta-thyrotropic polyclonal antisera. Morphometric differences in the cytoplasmic and nuclear volume densities in thyrotropes after the treatments were determined using a stereological method. The relative weights of the pituitary glands were significantly higher in the EDP- and EDP+hCG-treated juvenile and peripubertal rats than in untreated age-matched controls. Treatment with EDP promoted a decrease, and treatment with hCG an increase, of the cellular and nuclear volumes in TSH cells in both juvenile and peripubertal females in comparison with the respective controls. Treatment with a combination of EDP+hCG did not induce any significant changes. The cytoplasmic and nuclear volume densities in TSH cells in the EDP+hCG-treated group were significantly higher than in the EDP-treated, and significantly lower than in hCG-treated rats at both growth stages. These findings suggest that estradiol and hCG exerted opposite effects on pituitary TSH-immunoreactive cells. The observed effects on thyrotrope morphology were apparently independent of the stage of development.

High Level Expression of Prop-1 Gene in Gonadotropic Cell Lines

Prop-1 acts as an upstream regulator for the Pit-1 gene to induce development of Pit-1 lineage pituitary cell lines, GH-, PRL-, and TSH-producing cells, in the early stage of pituitary organogenesis. Furthermore, Prop-1 is presumed to be involved in the function of FSH/LH-producing cells, gonadotropes, since the defective Prop-1 gene shows hypogonadism. Recently, we reported evidence that Prop-1 directly regulates expression of the porcine FSHbeta gene, thus providing a novel advance in understanding the function of Prop-1 in FSH/LH production and hypogonadism. This study was intended to demonstrate the expressions of Prop-1 gene in pituitary tumor-derived cell lines. RT-PCR analyses were conducted of Pit-1, glycoprotein alpha subunit (alphaGSU), GnRH receptor, and cyclophilin A (a ubiquitously expressing gene). We observed expression of the Pit-1 gene in alphaT1-1, TalphaT1, MtT/S, GH3, and TtT/GF cells, expression of the alphaGSU gene in alphaT1-1, alphaT3-1, LbetaT2, LbetaT4, TalphaT1, and GH3 cells, and expression of GnRH receptor gene in alphaT3-1, LbetaT2, LbetaT4, and GH3 cells, respectively. These expression profiles were in accord with their cell lineages, with only a few exceptions. To accurately measure the expression level of the Prop-1 gene, a quantitative analysis was performed using the real-time PCR method. This analysis demonstrated that the LbetaT2 and LbetaT4 gonadotrope cell lines, which express the FSHbeta gene, express the Prop-1 gene. Taken together with our previous observation that Prop-1 is present in the adult porcine pituitary gonadotropes, Prop-1 might also be involved in development of gonadotropes and hormone production.

Immunocytochemical detection of effects of TRH and T 4 on prolactin- and TSH-producing cells in the pituitary gland of Rana ridibunda

Effects of synthetic thyrotropin-releasing hormone (TRH) and various doses of thyroxin (T 4) on prolactin (PRL)-producing cells and thyrotropic cells in the pituitary were investigated in adult male and female Rana ridibunda frogs. Animals were given 200 μg TRH once a week for 4 weeks and 0.2–0.5 mg T 4 during 3 days per week for a period of 2 weeks by injections in the groin. PRL-producing cells and thyrotropic cells were identified with light microscopical and electron microscopical immunocytochemical methods, using rabbit anti-PRL and rabbit anti-thyroid stimulating hormone (TSH) as primary antibodies. TRH caused cytological changes in both cell types, which were consistent with increased synthesis and release of both PRL and TSH. Treatment with 0.5 mg T 4 activated both cell types less than TRH treatment did, whereas 0.2 and 0.4 mg T 4 caused inactivation of both cell types. In conclusion, mammalian TRH is effective on both types of frog pituitary cells. Our study suggests that T 4 has a positive rather than a negative effect when concentrations above a certain threshold are given.

Hipotireoidismo primário simulando volumoso macroadenoma hipofisário

A 21-year-old woman complaining of 8-month amenorrhea associated to weight gain, galactorrhea and frequent headaches, presented for clinical evaluation; her laboratory tests were: TSH: 1192 mUI/ml (0.27-4.2); TT4: 1.0 microg/dl (4.4-11.4 l); TT3: 0.41 ng/ml (0.7-2.1); prolactin: 69.2 ng/ml (3-20) and a diagnosis of myxedema associated to galactorrhea was made. A hypothalamic-pituitary magnetic resonance imaging (MRI) showed a suprasellar and intrasellar mass lesion of 1.9 x 1.4 x 1.9 cm, determining compression and deviation of the optic chiasm. Due to the possibility of hyperplasia of the TSH-producing cells, treatment of hypothyroidism was initiated with levothyroxine. Two months later, upon normalization of thyroid hormones and TSH levels, a second MRI showed an anatomically normal pituitary gland. Regression of the pituitary mass after treatment with levothyroxine confirmed the hypothesis of pituitary hyperplasia secondary to primary hypothyroidism. Our findings support the importance of determining thyroid function tests during the investigation of pituitary masses and thus avoiding an unnecessary surgery.

Physiological Relevance of Thyroid Stimulating Hormone and Thyroid Stimulating Hormone Receptor in Tissues other than the Thyroid

Decades of research have provided strong evidence for a reciprocal relationship between the immune system and hormones of the hypothalamus–pituitary–thyroid (HPT) axis. Thyroid stimulating hormone (TSH), in particular, has been shown to have a variety of immune-regulating cytokine-like activities that can influence the outcome of T cell development in the thymus and intestine, and can affect the magnitude of antibody and cell-mediated responses of peripheral lymphocytes. Production of TSH and the expression of the TSH receptor are widely but selectively distributed across many different types of hematopoietic cells in the bone marrow, as well as among subsets of dendritic cells, monocytes and lymphocytes in the spleen and lymph nodes. In addition to their role in immunity, the involvement of TSH-producing hematopoietic cells in the microregulation of thyroid hormone activity represents a novel and potentially important aspect of the TSH-mediated immune-endocrine circuit.

Effect of centrally administered somatostatin on pituitary thyrotropes in male rats.

The effects of intracerebroventricularly administered somatostatin (SRIH-14 or -28) on growth and function of pituitary thyrotropes (TSH-cells) were examined in adult male Wistar rats. The animals were implanted with an intracerebroventricular cannula and after a recovery period, administered three 1 microg doses of SRIH-14 or -28 dissolved in 5 microl saline every second day. Controls were treated in the same way with the same volume of saline only. TSH-producing cells were studied using the peroxidase-antiperoxidase immunohistochemical procedure. Blood samples were collected for hormone (TSH) analyses 5 days after the last injection. Both SRIH-treatments significantly decreased (p < 0.05) all morphometric parameters obtained for TSH-cells in comparison with controls. The volume of TSH-cells decreased by 27%, nuclei by 44% and volume density by 33% in animals treated with SRIH-14. In animals treated with SRIH-28, these parameters were also significantly decreased (p < 0.05) (22%, 31%, and 25% respectively) compared to control rats. Serum concentrations of TSH were significantly decreased (p < 0.05) by 15% in SRIH-14- and by 12% in SRIH-28-treated animals in comparison with the controls. These observations suggest that centrally administered SRIH- 14 or -28 is specifically involved in the control of growth and secretory activity of TSH cells.

Time course observation of thyroid proliferative lesions and serum levels of related hormones in rats treated with kojic acid after DHPN initiation.

Time course changes in thyroid proliferative lesions as well as related hormone levels in the blood of male F344 rats given N-bis(2-hydroxypropyl)nitrosamine (DHPN: 2800 mg/kg body weight, single s.c. injection) as an initiation treatment followed by pulverized basal diet containing 0% (Group 2), 2% (Group 3) or 4% (Group 4) kojic acid (KA) were examined at Weeks 1, 2, 4, 8 and 12. As an untreated control group (Group 1), rats were given basal diet for 13 weeks and examined in the same manner. Serum T3/T4 levels in the DHPN + 2% KA and DHPN + 4% KA groups were significantly reduced as compared with the DHPN-alone group at each time point. Serum TSH levels in both DHPN + KA groups were significantly increased at each time point in a treatment period-dependent manner from Weeks 1 to 12, and the extent of elevation was more remarkable in the DHPN + 4% KA group. At Week 2, there were no statistically significant intergroup differences in liver T4-UDP-GT activities on a milligram microsomal protein basis. Histopathologically, no thyroid proliferative lesions were observed in the untreated control group or the DHPN-alone group. However, diffuse follicular cell hypertrophy and decreased colloid in the thyroid were apparent in all rats of the DHPN + KA groups at each time point. In addition, focal follicular cell hyperplasias and adenomas of the thyroid were observed at high incidence in the DHPN + 2% KA group from Week 4 and in the DHPN + 4% KA group from Week 8. Multiplicities of focal follicular cell hyperplasias and adenomas of the thyroid in the DHPN + 2% KA group were significantly greater than those in the DHPN + 4% KA group at Week 8. In the pituitary, an increase in the number of TSH producing cells with expanded cytoplasm was apparent from Weeks 4 to 12 in both DHPN + KA groups. These results strongly suggest that thyroid proliferative lesions were induced by KA administration due to continuous serum TSH stimulation through the negative feedback mechanism of the pituitary-thyroid axis, resulting from depression of serum T3 and T4.

Immunoreactive TSH cells in the pituitary of female middle-aged rats after treatment with estradiol or calcium

The pituitary TSH cell structure of middle-aged (14-month-old) female Wistar rats chronically treated with estradiol dipropionate (EDP), calcium glucoheptonate (Ca) or with the combination of both was studied. TSH-producing cells were examined in the pituitary pars distalis using rabbit anti-rat beta-thyrotropin (TSH) serum and peroxidase-antiperoxidase immunohistochemistry. A stereological method for the determination of morphometric changes of the volume of TSH cells and nuclei as well as of their number and relative volume densities was used. All examined morphometric parameters in treated animals showed a significant decrease in comparison with immunoreactive TSH cells of the controls; the most significant decrease was observed in EDP-treated rats. These results together with structural features of immunoreactive TSH cells in the pituitary of middle-aged rats after chronic application of EDP or Ca indicate that both compounds inhibit these cells.

Immunohistochemical studies of TSH-producing cells in the pituitary and expression of growth factors in thyroidal proliferative lesions in rats treated with thiourea and excess vitamin A.

Changes of TSH-producing cells in the pituitary and thyroid expression of the growth factors, transforming growth factor alpha (TGF alpha) and epidermal growth factor receptor (EGFR), as well as cyclin D1, were investigated immunohistochemically in order to clarify their contribution to the enhancing effects of excess vitamin A (VA) on thyroidal carcinogenesis induced by thiourea (TU). Male rats were allocated to 4 groups, control, TU, VA, and TU + VA, respectively, receiving no treatment, water containing 0.2% TU, diet containing 0.1% VA, and both for 10 or 19 weeks after a single s.c. injection of DHPN (2800 mg/kg) for initiation. Immunohistochemistry using antibodies against TSH demonstrated enlargement of TSH-producing cells in the TU + VA group as compared to the TU group, supporting our conclusion that enhanced TSH stimulation is mainly responsible for promoting the effects of excess VA. Since the expression of TGF alpha, EGFR, and cyclin D1 in thyroid proliferative lesions did not exhibit any differences between the TU and TU + VA groups in the present study, these factors are unlikely to participate in VA enhancement of carcinogenesis.