Iodine distribution in the thyroid follicles of the hagfish,Eptatretus burgeri and lamprey,Lampetra japonica: Electron-probe X-ray microanalysis

Summary In the thyroid follicles of species of cyclostomes, a hagfish and a lamprey, the distribution of stable iodine was examined by electron-probe X-ray microanalysis. A high concentration of stable iodine, heterogeneously distributed, was observed in the follicular cells of hagfish thyroid follicles. In the lamprey a low concentration of iodine was seen in the follicular lumina. The relative values for stable iodine determined in this way corresponded to values obtained by a chemical analytical method.     

Different concentrations of thyroid peroxidase and thyroglobulin in the nuclear envelope and the endoplasmic reticulum throughout the cytoplasm.

Thyroid peroxidase (TPO) and thyroglobulin (TG) represent two major glycoproteins of thyroid follicular cells performing biological functions such as iodination, transcytosis of thyroglobulin, and formation of thyroid hormones. They are involved in thyroid autoimmunity and thyroid inborn metabolic disorders. Studying these processes at a molecular level includes the determination of their precise intracellular distribution. An evaluation of the relative concentrations of TG and TPO in different subcellular compartments was carried out in stimulated human follicular cells using thin-frozen sections and the immunogold technique. It is documented that TG is transported from the endoplasmic reticulum and the Golgi apparatus to the follicular lumen by transport vesicles; most of it being present in the expanded endoplasmic reticulum throughout the cytoplasm. On the other hand, gold particles indicating TPO are adjacent to the membranes of the exocytotic pathway. They do not label the basolateral membrane but show the strongest density in the nuclear envelope and the apical membrane. The labeling density of TPO is about four times higher in the nuclear envelope than in the endoplasmic reticulum throughout the cytoplasm. In contrast, TG is concentrated three times higher in the rough endoplasmic reticulum throughout the cytoplasm than in the nuclear cisternae. Our results give the first quantitative evidence that TPO and TG are concentrated in different subcompartments of the endoplasmic reticulum. Because previous studies demonstrated the nuclear envelope as the site where the synthesis of endogenous peroxidase (Br?kelmann, J., D. W. Fawcett, Biol. Reprod. 1, 59-71 (1969)) begins, we suggest that synthesis of these functionally related proteins happens in specialized parts of the endoplasmic reticulum.(ABSTRACT TRUNCATED AT 250 WORDS)     

Inhibition of thyroid-restricted genes by follicular thyroglobulin involves iodinated degree

Follicular thyroglobulin (TG) reflects the storage of both iodine and thyroid hormone. This is because it is a macromolecular precursor of thyroid hormone and organic iodinated compound in follicular lumen. Thus, it may have an important feedback role in thyroid function. In this study, monolayer cells were cultured and follicles were reconstituted with primary pig thyroid cells in vitro. Reconstituted follicles were treated with iodine and methimazole (MMI), a drug that blocks iodine organification and reduces the degree of TG iodination in follicular lumen. The high degree of iodinated TG in follicular lumen was observed to inhibit thyroid-restricted gene expression. To confirm this finding, monolayer thyroid cells were treated with a different degree of TG iodination at the same concentration. These iodinated TG were extracted from reconstituted follicles of different groups. In this manner, this study provides firsthand evidence suggesting that follicular TG inhibits the expressions of thyroid-restricted genes NIS, TPO, TG, and TSHr.     

Theory on the evolutionary history of lamprey metamorphosis: role of reproductive and thyroid axes

Metamorphosis is a developmental strategy used by only a small number of extant fishes and little is known about its phylogenetic development during the evolution history of this large group of vertebrates. The present report provides a putative evolutionary history of metamorphosis in the lamprey, an extant agnathan with direct descendancy from some of the oldest known vertebrates. The study reviews recent data on the role of the thyroid gland and its hormones in metamorphosis, summarizes some recent views on the evolution of the endostyle/follicular thyroid in lampreys, and provides new data on the content of two gonadotropin-releasing hormones (GnRH-I and -III) in brain during goitrogen-stimulated, precocious metamorphosis. These new data support an earlier viewpoint of a relationship between thyroid and reproductive axes during metamorphosis. It is proposed that the earliest lampreys were paedomorphic larvae and they lived in a marine environment; as such, they resembled in many ways the larvae from which the ancient protochordates, Larvacea, are derived. The iodide-concentrating efficiency of the endostyle was a critical factor in the evolution of metamorphosis and this gland was replaced by a follicular thyroid, for postmetamorphic animals needed to store iodine following their invasion of freshwater. Larval growth and postmetamorphic reproduction in freshwater became fixtures in the lamprey life cycle; a non-parasitic adult life-history type appeared later. The presence among extant lampreys of two different adult life-history types, and examples of the lability of the timing of sexual maturation in some species, imply that there has been a complex interplay between the thyroid and reproductive axes during the evolution of metamorphosis in lampreys. This proposal is consistent with what we know of interplay of these axes in extant adult lampreys and with the long-held viewpoint that thyroid function and sexual maturation are an association with an ancient history.     

Ascidian homologs of mammalian thyroid peroxidase genes are expressed in the thyroid-equivalent region of the endostyle.

The endostyle is a pharyngeal organ for the internal filter feeding of urochordates, cephalochordates, and larval lamprey. This organ is also considered to be homologous to the follicular thyroid gland of higher vertebrates. Thyroglobulin (Tg) and thyroid peroxidase (TPO) are specifically expressed in the thyroid gland of higher vertebrates, and they play an important role in iodine metabolism for the synthesis of thyroid hormones. Previous histochemical observations showed that iodine-concentrating and peroxidase activities were detected in zones 7, 8, and 9 of the ascidian endostyle, suggesting that these zones contains cells that are equivalent to those in the vertebrate follicular thyroid. In order to investigate the molecular developmental mechanisms involved in the formation and function of the endostyle, with special reference to the evolution of the thyroid gland, in the present study, we isolated and characterized cDNA clones for TPO genes, CiTPO from Ciona intestinalis and HrTPO from Halocynthia roretzi. Northern blot and in situ hybridization analyses revealed that the expression of the ascidian TPO genes was restricted to zone 7, one of the elements equivalent to the thyroid. These results provide the first evidence at the gene expression level for shared function between a part of the ascidian endostyle and the vertebrate follicular thyroid gland. J. Exp. Zool. ( Mol. Dev. Evol. ) 285:158-169, 1999.     

Molecular Analysis of Structural Abnormalities in Papillary Thyroid Carcinoma Genome

Rearrangements of the protooncogene RET (RET/PTC) and somatic mutations of the gene BRAF are the most common events in the etiopathogenesis of papillary thyroid carcinoma (PTC). The rates of RET/PTC rearrangements and BRAF mutations in different nodular formations of the thyroid gland (TG) have been estimated. Comparative expression analysis of the extracellular (EC) and tyrosine kinase (TK) domains of RET has shown that 14% (12 out of 85) of PTC cases are RET/PTC-positive, including one RFP/RET-, two RET/PTC3-, and seven RET/PTC1-positive tumors, as well as two unidentified chimeric RET/PTC oncogenes. The standard T1796A transversion in the gene BRAF has been found in 60% (55 out of 91) PTC cases with the use of amplification refractory mutation system–polymerase chain reaction (ARMS–PCR). Somatic mutation G1753A and deletion del1800_1811 have been identified in PTC for the first time. The absence of the BRAF mutations in RET/PTC-positive tumors allows these two genetic defects to be regarded as alternative mechanisms of the RAS–RAF–MEK–ERK mitogen-activated protein (MAP) kinase cascade activation in PTCs. In none of the three follicular thyroid carcinomas (FTCs), 11 follicular thyroid adenomas (FTAs), and 13 nodular goiters have either BRAF mutations or RET/PTC rearrangements been found. Thus, the RET/PTC chimeric oncogenes and BRAF somatic mutations are specific markers of PTC and can be used for the preoperative diagnosis of these tumors.     

X-ray microanalysis on the thyroid follicle of the hagfish, Eptatretus burgeri and lamprey, Lampetra japonica.

Intracellular dense bodies, cytoplasmic matrices, and luminal colloids in the thyroid follicle of cyclostomes, hagfish and lamprey were examined to identify the distribution of iodine using an energy dispersive X-ray microanalyzer attached to a scanning transmission electron microscope. High level of iodine was detected only in the large dense body of the hagfishes, while it was too slight in quantity to detect by this method in the cytoplasmic matrix as well as in the luminal colloid. In the adult lamprey thyroid, an appreciable amount of iodine was detected in a few large dense bodies. In mice and rats, it is very hard to detect iodine in the luminal colloid, intracellular colloid droplet, and in the lysosomal dense granule by this method, though these structures have been well known to contain a fairly large amount of iodine which is combined with thyroglobulin. These facts mean that intracellular dense bodies in the thyroid follicular epithelium of the cytoclostome, especially of the hagfish have an extremely larger amount of iodine. These bodies are considered to be secondary lysosomes or residual bodies containing reabsorbed colloid materials which are highly condensed.     

Morphological changes in the ultrastructure of the thyroid epithelium of Notophthalmus viridescens after hypophysectomy and TSH- and prolactin stimulation

Summary In the newt, Notophthalmus viridescens, hypophysectomy results in progressive atrophy of the thyroid cells to the point of irreversible degeneration. After exclusive TSH-stimulation of hypophysectomized newts, increased endocytotic activity of the follicular epithelium is observed. Prolactin stimulation under the same conditions prevents atrophy but does not result in increased cell activity, as expressed by the reduced amount of microvilli and the lack of endocytotic activity. Combined TSH- and prolactin stimulation also results in cell activation, but the activation level of exclusively TSH-stimulated cells is not reached. Although prolactin prevents cellular atrophy and degeneration of the follicular epithelium, it reduces the TSH-induced activation of the thyroid epithelium.     

Movement of radio-tagged anadromous sea lamprey during the spawning migration in the River Mondego (Portugal)

The available spawning habitat for the anadromous sea lamprey, Petromyzon marinus L., population that enters the River Mondego has been drastically reduced in the last 20 years. The installation of a fish passage in the first impassable dam, the Açude-Ponte, would enable sea lamprey to recolonise the 34.6-km river stretch between the Açude-Ponte and Raiva dams. In order to assess the suitability of the upstream river stretches for this species, 10 radio-tagged sea lamprey were released upstream of the Açude-Ponte dam and tracked continuously throughout the entire migratory path. Lamprey were unable to pass over intact weirs that had been built for recreational purposes. Sea lamprey movements were more frequent during dusk and night than other periods. Increased river discharge at night, resulting from the operation of the Raiva power station, stimulated lamprey movements but reduced ground speeds recorded. After reaching a certain location, some of the animals maintained their position for several weeks, before undergoing a new movement. Two of the main tributaries of this river stretch were used by some sea lamprey, indicating that the animals were able to find these historical spawning grounds.     

Volume and pH regulation in agnathan erythrocytes: comparisons between the hagfish,Myxine glutinosa,and the lampreys,Petromyzon marinus and Lampetra fluviatilis

The responses of hagfish (Myxine glutinosa) and lamprey (Lampetra fluviatilis and Petromyzon marinus) erythrocytes to osmotic swelling in hypoosmotic medium and to acid-base disturbances induced by ammonium chloride prepulse were studied. The erythrocytes of hagfish regulated neither cell volume after osmotic swelling nor intracellular pH after acidification. In contrast, the erythrocytes of lamprey lost potassium and chloride after osmotic swelling, whereby their volume recovered. Furthermore, the red cell pH of lamprey recovered from experimental acidification in a nominally bicarbonate-free medium in the presence of sodium, confirming that the pathway involved is sodium/proton exchange.Abbreviation DMO 5,5-dimethyloxazolidine-2,4-dione     

Variability in migration timing of adult Pacific lamprey (<Emphasis Type="Italic">Lampetra tridentata</Emphasis>) in the Columbia River,U.S.A.

We examined the effects of river environment on the timing of spawning migrations by anadromous Pacific lamprey, Lampetra tridentata, in the Columbia River (U.S.A.). In a 41-year time series of adult lamprey counts, migration timing was earliest in warm, low-discharge years and latest in cold, high-flow years. Threshold temperatures associated with run timing were similar throughout the dataset despite significant impoundment-related warming, suggesting that temperature-dependent migration cues have been temporally stable. Within each year, migration rates of PIT-tagged lampreys were positively correlated with temperature and negatively correlated with discharge through multiple river reaches, offering additional evidence for environmental control of upstream movement. Both visual count and PIT-tag data indicated that there may be population-based differences in migration timing within the aggregate Columbia River lamprey run. These life history and behavioral results have potentially far-reaching implications for management of lamprey species.     

Distribution of lamprey gonadotropin-releasing hormone-III (GnRH-III) in brains of larval lampreys (Petromyzon marinus)

Two immunoreactive forms of gonadotropinreleasing hormone (GnRH), lamprey GnRH-I and lamprey GnRH-III, were found in neurons in larval sea lampreys (Petromyzon marinus). Using antisera preferentially directed against either lamprey GnRH-I or-III, dense reaction product was seen in cell bodies in the rostral hypothalamus and preoptic area. Reaction product was also dense in fibers to and within the neurohypophysis, in addition to numerous fibers which projected caudally, beyond the neurohypophysis through the mesencephalon. The majority of immunoreactive GnRH was lamprey GnRH-III, and when lamprey GnRH-I was seen, it was in cells that appeared to contain both forms of GnRH. A small number of cells found in the caudal hypothalamus contained only immunoreactive lamprey GnRH-III, and these may constitute a functional subgroup within the population of GnRH neurons. In animals undergoing metamorphosis there was a large increase in reaction product in all GnRH-containing cells and fibers. A striking change within the distribution of GnRH cells was localized to a distinct group of GnRH-immunoreactive cells (GnRH-I and-III) in the ventral anterior hypothalamic area. These cells were minimally detectable in larvae, but during metamorphosis became densely filled with immunoreactive product in perikarya and distal processes. The results are consistent with the hypothesis that lamprey GnRH-III is an important form of GnRH during the maturation of GnRH cells and fibers, and further indicates that these cells have attained their normal positions in the preoptic area and hypothalamus before metamorphosis.     

Fungal Thyroiditis: An Overview

The authors review the epidemiology, clinical manifestations, diagnosis, and treatment of fungal thyroiditis cases previously reported in the medical literature. Aspergillus was by far the most common cause of fungal thyroiditis. Immunocompromised patients, such as those with leukemia, lymphoma, autoimmune diseases, and organ-transplant patients on pharmacological immunosuppression were particularly at risk. Fungal thyroiditis was diagnosed at autopsy as part of disseminated infection in a substantial number of patients without clinical manifestations and laboratory evidence of thyroid dysfunction. Local signs and symptoms of infection were indistinguishable from other infectious thyroiditis and included fever, anterior cervical pain, thyroid enlargement sometimes associated with dysphagia and dysphonia, and clinical and laboratory features of transient hyperthyroidism due to the release of thyroid hormone from follicular cell damage, followed by residual hypothyroidism. Antemortem diagnosis of fungal thyroiditis was made by direct microscopy and culture of a fine-needle aspirate, or/and biopsy in most cases. Since most patients with fungal thyroiditis had disseminated fungal infection with delay in diagnosis and treatment, the overall mortality was high.     

Sequential analysis of experimental autoimmune thyroiditis induced by neonatal thymectomy in the Buffalo strain rat

We have studied the evolution of thyroiditis induced by neonatal thymectomy in Buffalo strain rats, with particular emphasis on the thyroid lymphocytic infiltrate. The earliest change was increased endothelial Ia expression, and infiltration of the thyroid at 5 weeks by ED1- and ED2-positive macrophages and B and T cells. The T cells comprised equal numbers of Ox 8 (T cytotoxic/suppressor)- and W3/25 (T helper)-positive cells. Ia-positive thyroid follicular cells were seen only in the presence of a T-cell infiltrate. Thyroglobulin antibody levels, thyroid weight, thyroid follicular cell Ia expression, and lymphocytic infiltration of the thyroid were maximal between Weeks 12 and 24, and impairment of macrophage function by injection of silica at this time produced amelioration of disease. The thyroid weight returned to control levels by Week 34 and Ia expression by thyroid cells disappeared. Circulating Ox 8-positive T cells were reduced between Weeks 12 and 24 and by Week 34 had returned to control levels. Our results indicate that the mononuclear infiltrate precedes thyroid follicular cell Ia expression and macrophages play an important role in perpetuating thyroiditis. Recovery from disease is accompanied by a return to normal in circulating suppressor/cytotoxic T cells.     

Bicarbonate permeability and immunological evidence for an anion exchanger-like protein in the red blood cells of the sea lamprey,Petromyzon marinus

Physiological and immuno-blotting experiments were used to determine whether the red blood cell membrane of a primitive vertebrate, the sea lamprey Petromyzon marinus, contained a counterpart similar to the vertebrate anion exchange protein known as AE1 or band 3. Results of the physiological experiments which measured CO₂ production after adding H¹⁴CO ₃ ^- to the extracellular saline, indicated significant transmembrane bicarbonate movement in lamprey blood which unlike that in most vertebrates, was insensitive to inhibition by 4,4 diisothiocyanatostilbene-2,2 disulfonic acid. The present study also showed that lamprey red blood cells possess acetazolamide-sensitive carbonic anhydrase which is an important component of CO₂ production by vertebrate red blood cells. Polyclonal immunoglobulins against a 12 amino acid domain in the C-terminus of the mouse AE1 recognized a trout red blood cell membrane protein with a relative molecular mass of 97 kDa, but failed to immunoreact with any membrane proteins from the red blood cells of lamprey. Antibodies against trout AE1 immunoreacted with trout red blood cell membrane proteins of approximately 97 kDa, 200 kDa and >200 kDa. Interestingly, only a 200-kDa membrane protein from the red blood cells of the primitive lamprey immunoreacted with the trout anti-AE1 immunoglobulin proteins. Therefore, lamprey red blood cells appear to possess an AE1-like protein that may be physiologically different than that in most other vertebrates.     

Hyaluronidase and collagenase inhibitory activities of the herbal formulation <Emphasis Type="Italic">Triphala guggulu</Emphasis>

Myrrh (guggulu) oleoresin from the Commiphora mukul tree is an important component of antiarthritic drugs in Ayurvedic medicine. Clinical data suggest that elevated levels of hyaluronidase and collagenase type 2 enzymes contribute significantly to cartilage degradation. Triphala guggulu (TG) is a guggulu-based formulation used for the treatment of arthritis. We assessed the chondroprotective potential of TG by examining its effects on the activities of pure hyaluronidase and collagenase type 2 enzymes. Triphala shodith guggulu (TSG), an intermediate in the production of TG, was also examined. A spectrophotometric method was used to assay Hyaluronidase activity, and to detect potential Hyaluronidase inhibitors. Aqueous and hydro-alcoholic extracts of TSG showed weak but dose-dependent inhibition of hyaluronidase activity. In contrast, the TG formulation was 50 times more potent than the TSG extract with respect to hyaluronidase inhibitory activity. A validated X-ray film-based assay was used to measure the gelatinase activity of pure collagenase type 2. Hydro-alcoholic extracts of the TG formulation were 4 times more potent than TSG with respect to collagenase inhibitory activity. Components of Triphala were also evaluated for their inhibitory activities on hyaluronidase and collagenase. This is the first report to show that the T2 component of Triphala (T. chebula) is a highly potent hyaluronidase and collagenase inhibitor. Thus, the TG formulation inhibits two major enzymes that can degrade cartilage matrix. Our study provides the first in vitro preclinical evidence of the chondroprotective properties of TG.     

Dietary ethanol and lipid synthesis in Drosophila melanogaster

When cultured on a defined diet, ethanol was an efficient substrate for lipid synthesis in wild-type Drosophila melanogaster larvae. At certain dietary levels both ethanol and sucrose could displace the other as a lipid substrate. In wild-type larvae more than 90% of the flux from ethanol to lipid was metabolized via the alcohol dehydrogenase (ADH) system. The ADH and aldehyde dehydrogenase activities of ADH were modulated in tandem by dietary ethanol, suggesting that ADH provided substrate for lipogenesis by degrading ethanol to acetaldehyde and then to acetic acid. The tissue activity of catalase was suppressed by dietary ethanol, implying that catalase was not a major factor in ethanol metabolism in larvae. The activities of lipogenic enzymes, sn-glycerol-3-phosphate dehydrogenase, fatty acid synthetase (FAS), and ADH, together with the triacylglycerol (TG) content of wild-type larvae increased in proportion to the dietary ethanol concentration to 4.5% (v/v). Dietary ethanol inhibited FAS and repressed the accumulation of TG in ADH-deficient larvae, suggesting that the levels of these factors may be subject to a complex feedback control.This research was supported by National Institutes of Health Grant GM-28779 to B.W.G. and a Monash University Research Grant to S.W.M.