Cat Thyroid Research Articles

A dosing algorithm for individualized radioiodine treatment of cats with hyperthyroidism.

BackgroundRadioiodine (131I) is the treatment of choice for hyperthyroidism in cats, but current 131I‐dosing protocols can induce iatrogenic hypothyroidism and expose azotemia.ObjectivesTo develop a cat‐specific algorithm to calculate the lowest 131I dose to resolve hyperthyroidism, while minimizing risk of iatrogenic hypothyroidism and subsequent azotemia.AnimalsOne thousand and four hundred hyperthyroid cats treated with 131I.MethodsProspective case series (before‐and‐after study). All cats had serum concentrations of thyroxine (T4), triiodothyronine (T3), and thyroid‐stimulating hormone (TSH) measured (off methimazole ≥1 week). Using thyroid scintigraphy, each cat's thyroid volume and percent uptake of 99mTc‐pertechnatate (TcTU) were determined. An initial 131I dose was calculated by averaging dose scores for T4/T3 concentrations, thyroid volume, and TcTU; 80% of that composite dose was administered. Twenty‐four hours later, percent 131I uptake was measured, and additional 131I administered, as needed, to deliver an adequate radiation dose to the thyroid tumor(s). Serum concentrations of T4, TSH, and creatinine were determined 6 to 12 months later.ResultsThe median calculated 131I dose was 1.9 mCi (range, 1.0‐10.6 mCi); 1380 cats required additional 131I administration on day 2. Of the cats, 1047 (74.8%) became euthyroid, 57 (4.1%) became overtly hypothyroid, 240 (17.1%) became subclinically hypothyroid, and 56 (4%) remained hyperthyroid. More overtly (71.9%) and subclinically (39.6%) hypothyroid cats developed azotemia than euthyroid cats (14.2%; P < .0001).Conclusions and Clinical ImportanceOur algorithm for calculating individual 131I doses resulted in cure rates similar to historical treatment rates, despite much lower 131I doses. This algorithm appears to lower prevalence of both 131I‐induced overt hypothyroidism and azotemia.

Follicle-forming cat thyroid cell lines synthesizing extracellular matrix and basal membrane components: a new tool for the study of thyroidal morphogenesis.

Interactions between follicular epithelial cells and extracellular matrix (ECM) are supposed to play an important role in the development and maintenance of thyroid tissue architecture. In the present study we have therefore investigated the synthesis of ECM components by a feline thyroid cell line which is able to form follicle-like structures in vitro, and also in v-ras-transfected and control-transfected sublines. Transfections were performed by lipofection with pZSR (viral Harvey ras gene; neo) and pSV2-neo (control, neo only) plasmids. We have adapted a semisolid culture system composed exclusively of polymerized alginate and therefore devoid of ECM components. Feline cells embedded in alginate gels as single cells and cultured for up to 90 days formed cell clusters within 10 days. Follicle-like structures were formed in the original cell lines and also in the v-ras- and control-transfected cells. Differences in proliferation rates were observed, the v-ras-transfected cells growing up to two to three times faster than the non-transfected cells. Immunostaining was done using rabbit first antibodies directed against mouse collagen IV, human fibronectin, laminin (tumor Engelbreth-Holm-Swarm laminin), perlecan and other ECM components. For comparison, immunostaining was also performed on cryosections of nodular goiters of six hyperthyroid cats. The cell lines and their transfected clones stained strongly positive for collagen IV and fibronectin, and positively but less strongly for laminin and perlecan. The cat goiter tissue stained positively for collagen IV, laminin, perlecan, and fibronectin, and positive staining for S-laminin (containing the beta2-chain) was seen in blood vessel walls in this tissue. In conclusion, cat cell lines grow three-dimensionally in alginate beads over several weeks, they form follicle-like structures and express the same ECM components as the native cat goiter tissue. Transfection with v-ras does increase proliferation rate, but does not fundamentally alter formation of follicle-like structures and ECM expression. Alginate gel culture is a promising new tool for the study of follicular morphogenesis, polarity, the expression pattern of ECM components and of the interaction between thyrocytes and ECM. It avoids interference caused by gels composed of ECM components.

Thyroid growth immunoglobulins in feline hyperthyroidism.

Feline hyperthyroidism bears a strong clinical and pathologic resemblance to toxic nodular goiter in humans. To evaluate whether the observed thyroid growth might be due to circulating thyroid antibodies, as has been postulated in humans, we studied the effect of purified immunoglobulin (Ig) G preparations on a rat thyroid follicular (FRTL-5) cell line. When compared with control, hyperthyroid cat IgG caused significantly increased [3H]-thymidine (Tdr) incorporation into DNA (p less than 0.02) and stimulated cellular proliferation 15-fold. Stimulation of 3H-Tdr incorporation tended to be biphasic and could be inhibited completely by a potent, specific TSH receptor blocking antibody. Hyperthyroid cat IgG also significantly inhibited 125I-bTSH binding to porcine thyroid membranes, an effect that could be reproduced using electrophoretically pure IgG and normal cat thyroid membranes. Unlike its effect on growth, hyperthyroid cat IgG did not stimulate intracellular cAMP, and there was no correlation between thyroid function in vivo and IgG growth-promoting activity in vitro. These data suggest that elevated titers of thyroid growth IgGs, probably acting through the TSH receptor, are present in feline hyperthyroidism and may play a role in goiter formation. Unlike growth, the thyroid hyperfunction observed is not IgG dependent. Further study of feline hyperthyroidism may contribute important insights into human nodular goiter and into the mediation of thyroid growth in general.

Autonomous Growth and Function of Cultured Thyroid Follicles from Cats with Spontaneous Hyperthyroidism

Spontaneous feline hyperthyroidism is a unique experimental model of toxic nodular goiter. To determine whether feline toxic goiter is caused by extrathyroidal stimulating factors or by the intrinsic autonomy of follicular cells, primary cultures of enzymatically dissociated follicles from 15 hyperthyroid cat goiters and from 3 normal cat thyroid glands were embedded in collagen gels. Growth and function in chemically defined media were assessed by autoradiography after double labeling with 3H-thymidine and 131I-Na. Iodine organification in follicles from normal glands was TSH dependent, but intense radioiodine organification occurred in follicles from hyperfunctioning goiters even in the absence of TSH. Similarly, twice as many follicular cells of hyperfunctioning thyroid tissue, maintained without TSH in the medium, were labeled after exposure to 3H-thymidine than in follicles from normal glands. The results strongly suggest that intrinsic alterations of cell function lead to autonomy of follicular growth and function and subsequently to the development of hyperplastic nodules, causing thyrotoxicosis. The reason for the focal nature of the disease remains an unresolved challenge. Further investigation using this model may further understanding of the growth of autonomous endocrine tumors.

Prenatal development of the cat thyroid: immunohistochemical demonstration of calcitonin in the "C" cells.

The presence of calcitonin in the cat thyroid was studied immunohistochemically in a series of gland development. The first positive cells are to be found on the 38th day of gestation, i.e. 1-2 days after level nine of ontogenetic development has been reached. The cytoplasm of these cells form only a narrow border round the nucleus. With advancing development the number of calcitonin-positive cells rises, the cytoplasm becomes intensely positive and its amount increases. From approximately the 50th day of prenatal development, the initially diffusely scattered, solitary calcitonin-positive cells are gradually replaced by groups of cells, which begin to occupy a characteristic position in relation to the follicular epithelium. The largest quantity of calcitonin-positive cells is found in foetuses about to be born. In non-pregnant adult cats, the incidence of immunohistochemically calcitonin-reactive cells is more sporadic and their distribution in the lobes of the thyroid is uneven.

Autoregulation of thyroid iodide transport: evidence for the mediation of protein synthesis in iodide-induced suppression of iodide transport.

The role of protein synthesis in iodide-induced suppression of subsequent iodide transport (iodide autoregulation) was studied in cat thyroid slices. Thyroid slices were pretreated for 60-120 min in the presence or absence of either excess (30 microM) sodium iodide, inhibitors of protein synthesis, or both. Tissue was then washed in the presence of 2 mM methimazole to prevent subsequent iodination reaction and remove excess iodide. Iodide transport activity was finally evaluated by measurement of the ratio of tissue to medium iodide concentrations in 90-min incubations. Addition of 0.1 mM cycloheximide during the preexposure of thyroid tissue to excess iodide had no effect on either control levels of iodide transport or iodide-induced autoregulation. However, if thyroid tissue was treated with cycloheximide alone for 1 h before preexposure to excess iodide, there was a significant reduction in the degree of iodide-induced induced autoregulation. Similar results were obtained with pretreatment of the tissue with 0.5 mM puromycin and 1 microgram/ml emetine. These findings suggest that protein synthesis is involved in the mechanism of thyroid autoregulation of iodide transport. Cycloheximide had no effect on the ability of excess iodide to reduce TSH-stimulated cAMP formation. Two-dimensional gel electrophoresis-isoelectric focusing and Sephadex G-25 column chromatography employing dual-isotope comparison of iodoprotein labeling of control and cycloheximide-treated tissue suggest that the ability of cycloheximide to suppress iodide-induced autoregulation is associated with the reduced iodination of an 8- to 10-kilodalton soluble component of the thyroid gland.

Development of regulatory mechanisms in the thyroid: failure of iodide to suppress iodide transport activity.

Fetal, neonatal, and adult cat thyroid glands were investigated for their ability to respond to excess iodide in vitro with reduction in both iodide transport activity as measured by the 1251-T/M ratio and thyrotropin-stimulated cyclic AMP formation. Fetal and neonatal cat thyroids responded to thyrotropin addition with greater increases in cyclic AMP than those of adults. Moreover, in each group, pretreatment of the thyroid tissue with NaI (30 pmol for 2 hr) resulted in a significant reduction in the subsequent stimulation of cyclic AMP formation by thyrotropin. The 1251-T/M ratio in neonatal cat thyroid tissue was similar to that in adult animals of either sex, whereas the lz5I-T/M of fetal thyroid was greater than that of the mother. Moreover, while pretreatment of the thyroid tissue of neonatal or adult animals with excess iodide resulted in a reduction in iodide transport activity, no such reduction was observed in fetal thyroid tissue. These findings suggest that the autoregula- tory effect of iodide transport develops at a later gestational time than the action on thyrotropin-stimulated cyclic AMP formation. Exposure of thyroid tissue to high con- centrations (5-50 pA4) of iodide in vitro re- sults in a reduction in subsequently deter- mined iodide transport activity (1-3), as well as an inhibition of the stimulation of cyclic AMP formation induced by thyrotro- pin (TSH) or other thyroid stimulators (2, 4-9). These two actions of iodide on thyroid function are similar in that both are prevented by simultaneous treatment of the tissue with agents that inhibit intrathyroidal oxidative reactions, and both effects appear to occur through the mediation of as yet unidentified, iodide-containing inhibitory factors formed within the thyroid cell (1,4). There is no information available to indi- cate whether these two actions of iodide are mediated by the same or different iodide- containing inhibitory factors. The present study was undertaken to (a) ascertain whether these two actions of iodide can be observed in fetal or neonatal thyroid tissue and (b) to determine whether or not there is any difference in the fetal development of the thyroid responses to excess iodide. Material and Methods. A description of the animal groups employed in this study is presented in Table I. All animals were obtained from a local supplier (Quaker Farms, Quakertown, Pa.). They were housed in environmentally controlled rooms and allowed food and water ad libitum. In those groups utilizing newborn animals, the female was allowed to deliver normally and the date of delivery recorded during routine daily observation. Date of birth was taken as day zero. Pregnant and lactating groups were composed of animals donating the fetal and neonatal groups, re- spectively. Fetal and neonatal thyroids were obtained by careful dissection, weighed to the nearest 0.1 mg on a torsion bal- ance, and sectioned into eight pieces by dividing each lobe along its vertical and horizontal axis. Four thyroid sections, two from each lobe, were preincubated for 2 hr in 2.5 ml Eagle's minimum essential medium (10) supplemented with 30 pA4 NaI. The remaining sections were preincu- bated for 2 hr in the same medium without added iodide. Following the preincubation, the sections were subjected to two 30-min washes with Eagle's medium prior to the test incubation. This wash procedure has been shown previously to effectively re- move more than 90% of the added iodide (3). Finally, two thyroid sections from each preincubation group were taken for mea- surement of iodide transport activity and the remaining sections employed for the

Autoregulation of thyroid iodide transport: possible mediation by modification in sodium cotransport.

Cat thyroid slices were employed to determine whether preincubation with excess iodide, a procedure previously reported to depress subsequently measured iodide transport activity, is associated with any alteration in the relationship between sodium and iodide transport. Kinetic analysis revealed that the inhibition of iodide-concentrating activity by iodide pretreatment was accompanied by a reduction in the apparent affinity of the iodide transport for iodide as reflected by increase in the value of KA, the concentration of iodide required to achieve half-maximal transport activity. A reduction in extracellular sodium concentration was also accompanied by an increase in the value of KA for iodide transport. Thus, the KA of iodide transport in control thyroid tissue was increased from a value of 8.6 +/- 1.2 to 16.5 +/- 2.4 microM, by a reduction in extracellular sodium from 144 to 52 mM. In contrast, in thyroid tissue subjected to a 2-h preincubation with 30 microM NaI, a similar reduction in extracellular sodium concentration was associated with only a 20% increase in KA from a value of 17.7 +/- 2.4 to 21.7 +/- 2.2 microM. Analysis of the kinetic data suggests that the autoregulatory effect of excess iodide results in a decline in the sodium dependency of iodide transport. This loss of sodium dependency is accompanied by an increase in the affinity of the process for sodium, as well as a reduction in the affinity of the transport system for iodide.

Epinephrine inhibits thyrotropin-stimulated adenosine 3',5'-monophosphate accumulation in cat thyroid tissue.

The effect of epinephrine on in vitro cat thyroid tissue cAMP accumulation was examined in both the presence and absence of added TSH. Epinephrine alone had no significant effect on cat thyroid cAMP accumulation. However, in the presence of phentolamine, an a-adrenergic antagonist, epinephrine (1 × 10−5 m), elicited an increase in thyroid cAMP levels which was evident within 2 min, declining to basal levels within 10 min. In contrast, in the presence of propranolol, a β-adrenergic antagonist, the addition of epinephrine elicited a prompt 30% reduction in thyroidal cAMP levels which was sustained throughout the incubation period. Preexposure of cat thyroid tissue slices to epinephrine resulted in a significant 20−60% reduction in the subsequent TSH stimulation of cAMP accumulation and a similar reduction in the activation of cAMP-dependent protein kinase activity. A similar inhibition of TSH action was observed with norepinephrine but not with adrenochrome or dopamine. The inhibitory action of the catechol...

Iodide Induced Suppression of Thyrotropin-Stimulated Adenosine 3&amp;acute;,5&amp;acute;-Monophosphate Production in Cat Thyroid Slices

Cat thyroid slices were studied to investigate their responsiveness to thyrotropin stimulation of cyclic AMP accumulation. Ovine and bovine thyrotropin, in the presence of 2.5 mM aminophylline, induced a dose-dependent increase in the cyclic AMP content of cat thyroid tissue. Half-maximal stimulation of cyclic AMP accumulation was obtained at a thyrotropin concentration of 1-2 mU/ml. The maximal effect of thyrotropin was observed at 10 mU/ml, and was associated with a mean 77 +/- 19-fold increase in thyroidal cyclic AMP. Preincubation of cat thyroid tissue for 2 h with 50 micron NaI resulted in an impairment in the subsequent ability of thyrotropin to enhance cyclic AMP accumulation, without altering the level of cyclic AMP in tissues not exposed to the hormone. Preincubation alone was without effect on thyrotropin stimulation of cyclic AMP, and the inhibitory effect of iodide was prevented by addition of 3 mM methimazole to the preincubation medium. In addition, the time course of thytrotropin stimulation of cyclic AMP accumulation in cat thyroid slices was not significantly altered by the preincubation with excess iodide. These studies provide additional evidence that excess iodide inhibits the adenylate cyclase-cyclic AMP system in thyroid tissue.

Bioassay of calcitonin by determinations of blood calcium-45 levels in rats and mice.

ABSTRACT The possibility of measuring calcitonin by determinations of blood 45Ca in young rats and mice was investigated. Blood samples were taken by orbital puncture on conscious animals. In both species blood 45Ca levels fell rapidly during the first 24 h after one sc injection of 45CaCl2. Thereafter, the elimination rate was slow, indicating that an equilibrium state was reached. During this period, single iv injections of highly purified porcine calcitonin induced log-dose-dependent blood 45Ca reductions in both rats and mice. Maximal responses were recorded at 30 min after injection, and the log-dose-response regressions were linear between 0.1 and 5.0 mU in young rats, and between 2.0 and 80.0 mU in young mice. The mean indices of precision were 0.18 and 0.27, respectively. Blood 45Ca reductions were also recorded on injection of extracts from rat thyroids, cat thyroids and chicken ultimobranchial glands, as well as an injection of plasma from patients with thyroid medullary carcinoma. No reduction in blood 45Ca was obtained by injections of ACTH, TSH, cortisone, thyroxine, or insulin. The responses to calcitonin were augmented if the animals had had a low calcium intake before the experiment, and/or if the animals were injected with 0.075 m NaH2PO4 immediately before injection of the hormone. Rats were more sensitive than mice to porcine calcitonin and to rat and cat thyroid extracts, while the opposite was true with chicken ultimobranchial extracts. No inter-species difference was recorded with human calcitonin. The assay appears to be useful for both clinical and experimental purposes.

Depletion of secretory granules, calcitonin, and formaldehyde-ozone-induced fluorescence from cat thyroid C cells by vitamin D2 treatment.

Using a combination of electron microscopy, fluorometry, and bioassay, the C cells of the cat thyroid were investigated with respect to their content of secretory granules, and calcitonin, and to their formaldehyde-ozone-induced fluorescence. This fluorescence is assumed to reflect the presence of a peptide with NH2-terminal tryptophan. In cats injected with large doses of vitamin D2 daily for 5 days, the C cells were degranulated, their fluorescence intensity was lowered and the calcitonin content of the thyroid was markedly reduced. It is suggested that the proposed tryptophyl peptide in the C cells is stored in the secretory granules and that it is engaged in the storage and/or release of the hormone.

Hyperosmotic effects on the cat thyroid perfused in vitro.

Tissue blocks containing the thyroid gland of cats were perfused via the carotid artery with normal and hypertonic defibrinated bovine whole blood for 2-hour intervals. Details of apparatus and methods for in vitro perfusion are described. Hypertonic perfusion resulted in reduced Na24 and I131 passage into the gland. The rate of in vivo accumulated I131 (inorganic and organic) movement from the gland to the hypertonic perfusate was increased. Tissue sodium and potassium was increased on a dry weight basis and a thyroid weight loss was observed due to loss of water to the hypertonic perfusate. Radiochromatograms of hypertonically perfused glands showed markedly reduced total I131 and synthesis of diiodotyrosine and monoiodotyrosine. It was concluded the thyroid responded to movements of iodide into and out of the cells in a manner that reflected the osmotic state of the external environment.