Rat Pituitary Gland Research Articles

Nicotine inhibits expression of Prrx1 in pituitary stem/progenitor cells through epigenetic regulation, leading to a delayed supply of growth-hormone-producing cells

ObjectiveNicotine, a toxic component of smoking, adversely affects animal growth and reproduction by decreasing secretion of anterior pituitary hormones. However, it has not been clarified whether nicotine inhibits the supply of endocrine cells in the pituitary gland. The present study investigated short- and long-term effects of persistent nicotine exposure on the pituitary glands of young animals. DesignThree-week-old male Wistar rats were exposed to nicotine (1 mg/kg body weight/day) for 7 days, and gene expression, cell numbers, and DNA methylation status were analyzed on the following day and 4 weeks after final treatments. ResultsThe expression level of the stem cell marker Sox2 was not changed by nicotine exposure throughout the experiment. On the other hand, nicotine inhibited expression of a progenitor cell marker, Prrx1, and growth hormone (Gh). Immunohistochemical analysis showed that the SOX2-positive cells positive for PRRX1 in nicotine-treated groups decreased to 61% (4-week-old) and 70% (8-week-old) of the saline-treated controls. In addition, the proportion of GH-positive cells in nicotine-treated group was 14% lower than that of saline-treated controls. Furthermore, first intron hypermethylation of Prrx1 was detected by a bisulfite sequence of genomic DNA from the anterior lobe of the rat pituitary gland. ConclusionsWe show that persistent nicotine exposure in young animals inhibits expression of Prrx1 in pituitary stem/progenitor cells through epigenetic regulation, leading to a delayed supply of GH-producing cells.

Antiprolactinoma Effect of Hordenine by Inhibiting MAPK Signaling Pathway Activation in Rats.

Prolactinomas are harmful to human health, and the clinical first-line treatment drug is bromocriptine. However, 20% prolactinomas patients did not respond to bromocriptine. Hordenine is an alkaloid separated from Fructus Hordei Germinatus, which showed significant antihyperprolactinemia activity in rats. The aim of this study was to explore the effect and mechanism of hordenine on prolactinomas in rats. The study used estradiol to induce prolactinomas, which caused the activation of the pituitary mitogen-activated protein kinase (MAPK) pathway in rats significantly. The treatment of hordenine restored estradiol, induced the overgrowth of pituitary gland, and reduced the prolactin (PRL) accumulation in the serum and pituitary gland of rats by blocking the MAPK (p38, ERK1/2, and JNK) activation and production of inflammatory cytokines, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). The antiprolactinoma effect of hordenine was mediated by inhibiting the MAPK signaling pathway activation in rats.

Age-Dependent Mapping of Somatotropic Hormone in Rat Pituitary Gland; Histological, Histochemical and Immunohistochemical Studies

The current study was performed to investigate age-dependent immunoexpression and localization of somatotropin in the pituitary gland. Twenty adult Wistar rats (fifteen females and five males) were housed; four animals per cage with observation of pregnancy. Samples from pituitary gland were collected from different ages of male offsprings. Tissue specimens were processed, and stained with Hematoxylin and Eosin (HE but basophils were difficult to be seen then appeared at stage II. Special stain results showed increasing in somatotrophs number from stage I to stage III then decreased at the last stage. Immunoexpression of somatotropin in pars distalis showed that somatotrophs were numerous and uniformly distributed at the first three stages then they were restricted in clusters at the last stage.

Combined irradiation by gamma rays and carbon nuclei increases the C/EBP-β LIP-isoform content in the pituitary gland of rats

C/EBP-β, a basic leucine zipper transcription factor, has important roles in the regulation of organism’s immune and inflammatory responses. Wistar rats subjected to the combined irradiation were characterized by an increase in the content of C/EBP-β LIP-isoform in the pituitary gland. The obtained data indicate that moderate doses of ionizing radiation to initiate the endoplasmic reticulum stress response and are likely to initiate C/EBP-β-mediated cell death according to the apoptotic scenario. This study also confirms the earlier hypothesis about the alterations of the hypothalamic-pituitary-adrenocortical axis in response to moderate doses of ionizing radiation.

Combined Irradiation by Gamma Rays and Carbon Nuclei Increases the C/EBP-β LIP Isoform Content in the Pituitary Gland of Rats.

C/EBP-β, a basic leucine zipper transcription factor, has important roles in the regulation of the body immune and inflammatory responses. Wistar rats subjected to combined irradiation were characterized by an increase in the content of the C/EBP-β LIP isoform in the pituitary gland. The obtained data indicate that moderate doses of ionizing radiation to initiate the endoplasmic reticulum stress response and are likely to initiate C/EBP-β-mediated cell death according to the apoptotic scenario. This study also confirms the earlier hypothesis about the alterations of the hypothalamic-pituitary-adrenocortical axis in response to moderate doses of ionizing radiation.

Functional survival of rat pituitary gland in hypothermic storage for pituitary transplantation.

Deteriorated pituitary function can lead to serious complications that might need lifelong hormone replacement therapy. However, long-term hormone administration can have significant adverse effects. Thus, it would be more desirable to restore pituitary function by pituitary transplantation. In this study, we investigated functional preservation of extracted pituitary gland in special preservation solution under hypothermic condition for pituitary transplantation. We obtained nineteen pituitary glands from 250-300g male Sprague-Dawley rats via parapharyngeal approach. These extracted glands were divided into three pieces and stored in histidine-tryptophan-ketoglutarate (HTK) solution at 4°C and compared to their corresponding glands stored in phosphate buffer saline (PBS). Light and electron microscopic examinations were performed to identify morphological changes of pituitary gland at 0,3, and 7days after storage. TUNEL assay to confirm cell viability, and adenosine-triphosphate (ATP) concentration were also serially examined. Tissue architecture and cellular viability of specimens preserved in HTK solution for 3days were considerably maintained and similar to those in normal pituitary gland (0day specimen). In contrast, specimens stored in PBS were markedly destroyed after 3days of storage. After 7days of storage, significant degeneration occurred in tissues stored in both HTK and PBS. However, tissue architecture was preserved more in specimens stored in HTK solution than those stored in PBS. ATP concentration decreased more rapidly in specimens stored in PBS solution, but there was no statistical significance (p= 0.055). Extracted rat pituitary gland supplemented with special preservation solution could be preserved for 3days under hypothermic condition.

Role of GPER in the anterior pituitary gland focusing on lactotroph function.

Ovarian steroids control a variety of physiological functions. They exert actions through classical nuclear steroid receptors, but rapid non-genomic actions through specific membrane steroid receptors have been also described. In this study, we demonstrate that the G-protein-coupled estrogen receptor (GPER) is expressed in the rat pituitary gland and, at a high level, in the lactotroph population. Our results revealed that ~40% of the anterior pituitary cells are GPER positive and ~35% of the lactotrophs are GPER positive. By immunocytochemical and immuno-electron-microscopy studies, we demonstrated that GPER is localized in the plasmatic membrane but is also associated to the endoplasmic reticulum in rat lactotrophs. Moreover, we found that local Gper expression is regulated negatively by 17β-estradiol (E2) and progesterone (P4) and fluctuates during the estrus cycle, being minimal in proestrus. Interestingly, lack of ovarian steroids after an ovariectomy (OVX) significantly increased pituitary GPER expression specifically in the three morphologically different subtypes of lactotrophs. We found a rapid estradiol stimulatory effect on PRL secretion mediated by GPER, both in vitro and ex vivo, using a GPER agonist G1, and this effect was prevented by the GPER antagonist G36, demonstrating a novel role for this receptor. Then, the increased pituitary GPER expression after OVX could lead to alterations in the pituitary function as all three lactotroph subtypes are target of GPER ligand and could be involved in the PRL secretion mediated by GPER. Therefore, it should be taken into consideration in the response of the gland to an eventual hormone replacement therapy.

Effects of Aqueous Leaf Extract of Lawsonia inermis on Aluminum-induced Oxidative Stress and Adult Wistar Rat Pituitary Gland Histology.

Objectives:The aim of this study was to investigate the antioxidant effect of aqueous Lawsonia inermis leaf extract on aluminum-induced oxidative stress and the histology of the pituitary gland of adult Wistar rats.Methods:Thirty-five adult male Wistar rats weighing between 100-196g and 15 mice of the same weight range were included in the study. Lawsonia inermis extracts and aluminum chloride (AlCl3) were administered for a period of three weeks to five rats per group. The subjects in Group 1 (control) were given pellets and distilled water. Group 2 received 60mg/kg/d of aqueous extract of Lawsonia inermis. Group 3 was given 0.5mg/kg/d of AlCl3. Group 4 was administered 0.5mg/kg/d of AlCl3 and 60mg/kg/d of aqueous Lawsonia inermis extract orally. Group 5 received 0.5mg/kg/d of AlCl3 and 75mg/kg/d of aqueous Lawsonia inermis extract orally. Group 6 was given 0.5mg/kg/d of AlCl3 and 100mg/kg/d of aqueous Lawsonia inermis extract orally. Group 7 was administered 0.5mg/k/d of AlCl3 and 5mg/Kg/d ascorbic acid in distilled water orally. Twenty-four hours after the last administration, the animals were weighed, sedated with chloroform, and had their pituitary glands located, removed, and weighed on an electronic analytical balance.Results:Decreased cell counts were observed in the pituitary gland micrographs of the Wistar rats given 0.5mg of aluminum chloride, whereas the Wistar rats given 0.5mg of aluminum chloride and varying doses of Lawsonia inermis had increased dose-dependent cell counts.Conclusion:Aqeuous Lawsonia Inermis leaf extract increased the cell counts of the pituitary glands of adult male Wistar rats, in addition to alleviating aluminum-induced oxidative stress.

Gonadotropin-releasing hormone and kisseptin-10 regulate nuclear receptor subfamily 5 group a member 1/catenin beta 1/ nuclear receptor subfamily 0 group B member 1 activity in female rat anterior pituitary gland.

In this study, we tested the hypothesis that modulation of endogenous gonadotropin-releasing hormone (Gnrh) neuronal network activity alters the mRNA expression of nuclear receptor subfamily 5 group A member 1 (Nr5a1), through one of the component of Wnt pathway signaling - catenin beta 1 (Ctnnb1) (its co-activator), and its co-repressor nuclear receptor subfamily 0, group B member 1 (Nr0b1) in the female rat pituitary gland in vivo. Adult ovariectomized rats were given a serial infusion of Gnrh, kisspeptin-10, Gnrh + Gnrh antagonist (Antide), or kisspeptin-10 + kisspeptin antagonist (kisspeptin-234) into the third ventricle of the brain. The anterior pituitary and blood was used to mRNA and protein expression analysis. We demonstrated that Gnrh up-regulates Nr5a1 mRNA expression in the anterior pituitary and induces NR5A1 depletion in gonadotropes. Gnrh administration increased both Ctnnb1 mRNA expression and protein synthesis, and induced activation of cellular Ctnnb1 via translocation from the gonadotropes cytoplasm to nucleus. After kisspeptin-10 treatment, up-regulation of Nr0b1 mRNA and protein expression in the anterior pituitary was observed. These data indicate that Gnrh-neuron-mediated network activity alters Nr5a1 gene transcription and translation in gonadotrope cells and this effect may result from the changes induced in the Ctnnb1 and Nr0b1 gene/protein expression balance.

Immunohistochemical Study of the Laminin α5 Chain and Its Specific Receptor, Basal Cell Adhesion Molecule (BCAM), in both Fetal and Adult Rat Pituitary Glands.

Laminin, a major basement membrane protein, comprises three subunit chains: α, β, and γ chains. Among these chains, only the laminin α chain is capable of signaling via laminin receptors. Although laminin isoforms containing the α5 chain were reported to be the first laminin produced during rat anterior pituitary gland development, the functions of these isoforms are unknown. We used immunohistochemical techniques to localize the laminin α5 chain and its specific receptor, basal cell adhesion molecule (BCAM), in fetal and adult pituitary gland. Laminin α5 chain immunoreactivity was observed in the basement membrane of the primordial adenohypophysis at embryonic days 12.5 to 19.5. Double immunostaining showed that BCAM was present and co-localized with the laminin α5 chain in the tissue. Quantitative analysis showed that the laminin α5 chain and BCAM were expressed in the anterior pituitary gland during postnatal development and in adulthood (postnatal day 60). In the adult gland, co-localization of the laminin α5 chain and BCAM was observed, and BCAM was detected in both the folliculo-stellate cells and endothelial cells. These results suggest that laminin α5 chain signaling via BCAM occurs in both the fetal adenohypophysis and adult anterior pituitary gland.

Molecular basis of growth hormone daily mRNA and protein synthesis in rats

AimsDaily and seasonal rhythms coordinate the endocrine and metabolic functions. The pituitary gland is the master regulator of several endocrine activities, and its function is classically regulated by endocrine signals from its target glands as well as from the hypothalamus. The growth hormone (GH) produced and secreted by the anterior pituitary presents a pulsatile secretion throughout the 24-hour cycle. However, the molecular mechanisms regulating the daily pattern of GH secretion are still unclear. Herein we investigated whether circadian GH mRNA and protein synthesis is modulated by acute adjustments in the stability and expression of GH mRNA. Main methodsGH mRNA and protein content were evaluated by real-time PCR and Western blotting, respectively, in pituitary gland of rats euthanized every 3 h during a 24-h period at the Zeitgeber times (ZT3 to ZT24). The GH mRNA poly(A) tail length was determined by RACE-PAT assay. Key findingsWe identified two main peaks of GH mRNA level in the pituitary gland of rats; one in the middle of the light-cycle and another in the middle of the dark-cycle. The latter was associated with an increase in pituitary GH protein content. Interestingly, an increment in the poly(A) tail length of the GH transcript was observed in association to reduced migration rate of the GH transcript and increased mRNA content in the dark-cycle period. SignificanceOur findings provide evidence that changes in the GH mRNA poly(A) length may underlie the circadian pattern of GH mRNA and protein levels in the pituitary gland of rats.

Ameliorating effect of edible bird’s nest against lead acetate toxicity on the rat hypothalamic–pituitary–ovarian axis and expressions of epidermal growth factor and vascular endothelial growth factor in ovaries

Lead acetate (LA) alters ovarian histology, size and function, as well as decreases oestrogen production. LA is also toxic to rat ovary and pituitary gland. Edible bird’s nest (EBN) can reportedly alleviate these damages. This study aimed to evaluate the protective effect of EBN supplement on the ovaries and pituitary glands of rats exposed to LA toxicity. Five treatment groups were established as follows: group 1 (C), which was given distilled water; group 2 (T0), which was administered with LA (10 mg/kg body weight (BW); and groups 3 (T1), 4 (T2) and 5 (T3), which were given LA (10 mg/kg BW) plus graded concentrations of 30, 60 and 120 mg/kg BW of EBN, respectively. Rats were euthanised at day 35 to collect ovary and pituitary gland for histomorphological study and the expression analysis of epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF). Plasma was used to determine concentrations of E2, progesterone (P4), growth hormone (GH) and prolactin (P). E2, P4, GH and P levels were lower (p < 0.05) in T0, whereas T3 had the highest E2 concentration. All EBN-treated groups had increased number of survival follicles, as well as significant proliferation of gonadotrophic cells in the pituitary gland and decreased degenerative changes. All immunohistochemistry results showed significantly higher EGF and VEGF expression levels (p < 0.05) in T3. Overall, results showed the detrimental effects of LA on the ovary and pituitary gland and revealed the ameliorating effect of oral EBN supplementation against LA toxicity.

Correction to: SOX10-positive cells emerge in the rat pituitary gland during late embryogenesis and start to express S100β.

The published online version contains mistake in Table 1, Table 2, and some data in Materials and Methods.

Plasma ACTH concentration and pituitary gland histo-pathology in rats infected with Trypanosoma brucei brucei.

Human African trypanosomiasis is one of the neglected and re-emerging infectious diseases in Africa with over 60 million people being at risk of contracting the disease. To investigate the effects of Trypanosoma brucei brucei infection on secretion of adrenocorticotropic hormone (ACTH) and histology of the pituitary gland and paraventricular nucleus in rats. Rats were randomly divided into two groups, control and experimental. Experimental rats were injected intraperitonially with 0.2ml of blood containing 1.0 × 104 live T.b.brucei parasites. Tail blood samples were collected weekly for the determination of plasma concentration of ACTH. The pituitary gland and coronal section of brain were processed histologically and observed microscopically. There was a significant difference (p = 0.0190) in plasma ACTH concentration between the control and experimental rats. Histological alterations were observed in both the pituitary and paraventricular nucleus of experimental rats. T.b.brucei infection causes histological changes in both the paraventricular nucleus and pituitary gland in rats. These histological changes could account for the decrease in corticotropin releasing hormone (CRH) and ACTH production in the infected rats.

SOX10-positive cells emerge in the rat pituitary gland during late embryogenesis and start to express S100β.

In the pituitary gland, S100β-positive cells localize in the neurohypophysis and adenohypophysis but the lineage of the two groups remains obscure. S100β is often observed in many neural crest-derived cell types. Therefore, in this study, we investigate the origin of pituitary S100β-positive cells by immunohistochemistry for SOX10, a potent neural crest cell marker, using S100β-green fluorescence protein-transgenic rats. On embryonic day 21.5, a SOX10-positive cell population, which was also positive for the stem/progenitor cell marker SOX2, emerged in the pituitary stalk and posterior lobe and subsequently expanded to create a rostral-caudal gradient on postnatal day 3 (P3). Thereafter, SOX10-positive cells appeared in the intermediate lobe by P15, localizing to the boundary facing the posterior lobe, the gap between the lobule structures and the marginal cell layer, a pituitary stem/progenitor cell niche. Subsequently, there was an increase in SOX10/S100β double-positive cells; some of these cells in the gap between the lobule structures showed extended cytoplasm containing F-actin, indicating a feature of migration activity. The proportion of SOX10-positive cells in the postnatal anterior lobe was lower than 0.025% but about half of them co-localized with the pituitary-specific progenitor cell marker PROP1. Collectively, the present study identified that one of the lineages of S100β-positive cells is a SOX10-positive one and that SOX10-positive cells express pituitary stem/progenitor cell marker genes.

Two types of rat pituitary somatotrophs secrete growth hormone with different biological and immunological profiles

ObjectiveTwo stable subpopulations of somatotrophs reside in the rat pituitary gland. We tested the hypothesis that one produced growth hormone (GH) with greater activity when tested in the tibial line bioassay (BGH) than the other, while differences in the activities between the two groups would be less dramatic when measured by immunoassay (IGH). DesignA series of studies using hypophysectomized rats, hollow fibers, treatments and culture models were used to differentiate differences in Type I and Type II anterior pituitary somatotrophs in both function and production of immunoactive and bioactive growth hormone. ResultsWe found that dense, Type II somatotrophs (>1.070g·cm−3) differed markedly in their secretion patterns of IGH vs BGH in different In vitro and in vivo tests. In culture, Type II cells secreted five times as much BGH, and three fourths as much IGH as the less dense Type I cells. Production (storage and secretion) of BGH was 7-fold greater by Type II cells whereas IGH production was identical for the two cell types. Implantation of Type II cells into hypophysectomized rats significantly increased body weight, epiphyseal cartilage thickness, and muscle weight of the recipients; in contrast, Type I cells elicited only a small increase in body weight. Type I somatotrophs isolated from rats which had been previously fasted or insulin-treated subsequently showed only small, inconsistent changes in release relative to that from cells in the unfractionated cell population. However, release of BGH from the Type II cells was markedly decreased. ConclusionBoth IGH and BGH should be considered in the elucidation of GH physiology.

Sequencing and Identification of Endogenous Neuropeptides with Matrix-Enhanced Secondary Ion Mass Spectrometry Tandem Mass Spectrometry.

Matrix-enhanced secondary ion mass spectrometry (ME-SIMS) has overcome one of the biggest disadvantages of SIMS analysis by providing the ability to detect intact biomolecules at high spatial resolution. By increasing ionization efficiency and minimizing primary ion beam-induced fragmentation of analytes, ME-SIMS has proven useful for detection of numerous biorelevant species, now including peptides. We report here the first demonstration of tandem ME-SIMS for de novo sequencing of endogenous neuropeptides from tissue in situ (i.e., rat pituitary gland). The peptide ions were isolated for tandem MS analysis using a 1 Da mass isolation window, followed by collision-induced dissociation (CID) at 1.5 keV in a collision cell filled with argon gas, for confident identification of the detected peptide. Using this method, neuropeptides up to m/z 2000 were detected and sequenced from the posterior lobe of the rat pituitary gland. These results demonstrate the potential for ME-SIMS tandem MS development in bottom-up proteomics imaging at high-spatial resolution.

Molecular-level effects of acupuncture on depression: a genome-wide transcriptome analysis of pituitary gland in rats exposed to chronic restraint stress

ObjectiveTo investigate the antidepressant mechanism of acupuncture at the molecular level in the pituitary gland of rats exposed to chronic restraint stress (CRS). MethodsForty-eight rats were randomly divided into a control group (C), model group (M), fluoxetine group (F), and acupuncture group (A). CRS was induced in the model group by restraining rats for 28 days. Groups F and A were given fluoxetine administration and acupuncture intervention, respectively, 1 h before the CRS procedures were implemented. On the 29th day, rat pituitary glands were harvested for RNA extraction and RNA-sequencing analysis. ResultsDifferentially expressed gene analysis identified 148, 87, and 179 response genes differentially expressed in group C vs group M, group F vs group M, and group A vs group M, respectively. Using Gene Ontology term enrichment analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis, we identified the up-regulation of gene sets involved in extracellular space, receptor binding, and monoamine neurotransmitter metabolism, and the down-regulation of gene sets involved in immune response and inflammatory response as the prevailing transcriptomic signatures in the pituitary gland of rats treated with CRS, fluoxetine, or acupuncture. ConclusionOur findings suggest that acupuncture has a multitarget antidepressant effect at the molecular and behavioral levels.

Cell type-specific localization of Ephs pairing with ephrin-B2 in the rat postnatal pituitary gland.

Sox2-expressing stem/progenitor cells in the anterior lobe of the pituitary gland form two types of micro-environments (niches): the marginal cell layer and dense cell clusters in the parenchyma. In relation to the mechanism of regulation of niches, juxtacrine signaling via ephrin and its receptor Eph is known to play important roles in various niches. The ephrin and Eph families are divided into two subclasses to create ephrin/Eph signaling in co-operation with confined partners. Recently, we reported that ephrin-B2 localizes specifically to both pituitary niches. However, the Ephs interacting with ephrin-B2 in these pituitary niches have not yet been identified. Therefore, the present study aims to identify the Ephs interacting with ephrin-B2 and the cells that produce them in the rat pituitary gland. In situ hybridization and immunohistochemistry demonstrated cell type-specific localization of candidate interacting partners for ephrin-B2, including EphA4 in cells located in the posterior lobe, EphB1 in gonadotropes, EphB2 in corticotropes, EphB3 in stem/progenitor cells and EphB4 in endothelial cells in the adult pituitary gland. In particular, double-immunohistochemistry showed cis-interactions between EphB3 and ephrin-B2 in the apical cell membranes of stem/progenitor cell niches throughout life and trans-interactions between EphB2 produced by corticotropes and ephrin-B2 located in the basolateral cell membranes of stem/progenitor cells in the early postnatal pituitary gland. These data indicate that ephrin-B2 plays a role in pituitary stem/progenitor cell niches by selective interaction with EphB3 in cis and EphB2 in trans.

Image analysis for TSH mRNA in situ hybridization in pituitary glands from rats with thyroid follicular cell hypertrophy after treatment with three different test compounds

The goal of this in situ hybridization and image analysis technique is to study the effects of new pharmacological/chemical entities on the thyroid and pituitary gland in rats, reveal the pathogenesis of thyroid follicular cell hypertrophy and to retrospectively exclude the risk of thyroid tumor development in humans.In the present study, we describe the increase of thyroid-stimulating hormone- (TSH-) beta subunit mRNA in the pars distalis of the pituitary gland and the quantitative measurement of TSH mRNA positive cells from rats of three 4-week toxicity studies treated with three different test compounds inducing thyroid follicular cell and hepatocellular hypertrophy in rats. Compared to immunohistochemistry (IHC), in situ hybridization (ISH) for TSH was found to be more sensitive.With this technique we are able to exclude a direct effect of the test compound on the thyroid gland by showing the activation of thyrotrope cells from the pituitary gland and therefore this technique retrospectively enables us to exclude a possible risk for humans at an early stage of drug development. Also in case blood serum samples for evaluation of TSH are not available anymore or hepatocellular hypertrophy is not present (close metabolic relationship between thyroid gland and liver in rodents), the described method allows retrospective investigations on thyroid follicular cell hypertrophy or hyperplasia. This can be of high relevance in human safety assessment for certain drugs in order to exclude a primary effect on the thyroid gland especially when it comes to thyroid neoplasia in rodents as previously described.