The induction of tolerance to thyroglobulin significantly reduces the severity of thyroiditis in obese strain chickens

Obese strain chickens develop severe spontaneous autoimmune thyroiditis several weeks after hatching, characterized by mononuclear cell infiltration and antibodies to thyroglobulin (Tg). The presence of antibodies to Tg suggests that Tg is an important antigen in this disease, but it does not provide definitive evidence. To clarify this point, Obese strain chicks were tolerized at hatching with Tg and then examined up to 6 wk later for antibodies to Tg, thyroid pathology, and function. Various tolerance regimens were tested. The optimal conditions were i.v. injection of Tg within 24 hr of hatching, and injection of at least 1 mg. Tg isolated from normal thyroid glands was satisfactory, and it did not have to be deaggregated. Tolerance induced by the above procedure significantly retarded all parameters of autoimmunity, although by 6 wk of age some of the tolerized chicks had severe thyroiditis. Multiple weekly injections of Tg were no more effective than a single injection at hatching. Interestingly, a single injection at hatching was very effective, yet it was cleared from the circulation within 24 hr. In summary, tolerance induced with Tg had a profound effect on the disease and thus provides good evidence for the role of Tg in this disease.     

Experimental autoimmune thyroiditis in mice chronically treated from birth with anti-IgM antibodies

The role of T lymphocytes in the pathogenesis of experimental autoimmune thyroiditis in mice is well established while the role of B lymphocytes is unclear. Mice with thyroid lesions have thyroglobulin antibodies whereas these antibodies can occur in mice immunized with Tg that do not develop thyroid lesions. To determine whether thyroglobulin antibodies are necessary for the development of the thyroid infiltrates with mononuclear cells, which are characteristic for experimental autoimmune thyroiditis, AKR mice chronically treated from birth with goat anti-mouse IgM antibodies were immunized with mouse thyroglobulin in Freund's complete adjuvant when they were 7 weeks old. Control mice, similarly immunized, were chronically injected from birth with normal goat gamma-globulin. Three weeks after immunization, all mice were sacrificed, thyroglobulin antibodies in the serum were measured by hemagglutination assay and enzyme-linked immunosorbent assay, and thyroid pathology was assessed. The serum concentration of IgG and IgM, the percentage of B and T lymphocytes in the spleen (flow cytometry), and the in vitro proliferative response of spleen lymphocytes to stimulation by PHA, LPS, and Tg were also measured. All mice treated with anti-IgM antibodies did not have detectable thyroglobulin antibodies but 63% of these mice and 88% of control mice (all of which had thyroglobulin antibodies) had thyroid lesions. Mice treated with anti-IgM antibodies that did not have thyroid lesions had a more pronounced depression of B lymphocytes than similarly treated mice that had thyroid lesions. These experiments suggest that thyroglobulin antibodies are not necessary for the development of thyroid infiltrates with mononuclear cells. B lymphocytes could still participate in the production of experimental autoimmune thyroiditis by presenting thyroglobulin to helper T lymphocytes.     

cDNA immunization of mice with human thyroglobulin generates both humoral and T cell responses: a novel model of thyroid autoimmunity

Thyroglobulin (Tg) represents one of the largest known self-antigens involved in autoimmunity. Numerous studies have implicated it in triggering and perpetuating the autoimmune response in autoimmune thyroid diseases (AITD). Indeed, traditional models of autoimmune thyroid disease, experimental autoimmune thyroiditis (EAT), are generated by immunizing mice with thyroglobulin protein in conjunction with an adjuvant, or by high repeated doses of Tg alone, without adjuvant. These extant models are limited in their experimental flexibility, i.e. the ability to make modifications to the Tg used in immunizations. In this study, we have immunized mice with a plasmid cDNA encoding the full-length human Tg (hTG) protein, in order to generate a model of Hashimoto's thyroiditis which is closer to the human disease and does not require adjuvants to breakdown tolerance. Human thyroglobulin cDNA was injected and subsequently electroporated into skeletal muscle using a square wave generator. Following hTg cDNA immunizations, the mice developed both B and T cell responses to Tg, albeit with no evidence of lymphocytic infiltration of the thyroid. This novel model will afford investigators the means to test various hypotheses which were unavailable with the previous EAT models, specifically the effects of hTg sequence variations on the induction of thyroiditis.     

Thymosin alpha 1-induced modulation of cellular responses and functional T-cell subsets in mice with experimental autoimmune thyroiditis

The effects of Ta-1, a peptide constituent of thymosin fraction 5, were studied on murine autoimmune thyroiditis using two congenic strains of mice, B10.Br (Br) and B10.D2 (D2), which are sensitive and resistant to experimental autoimmune thyroiditis (EAT) induction, respectively. EAT was induced by either 2 weekly iv injections of mouse thyroglobulin with adjuvant lipopolysaccharide (LPS) or intradermal injection of thyroglobulin mixed with complete Freund's adjuvant (CFA). The criteria for induction and intensity of thyroiditis were the level of lymphoid infiltration in the thyroid gland and the titer of anti-thyroglobulin antibodies. Ta-1 was given in 5 or 10 daily sc injections in doses ranging from 0.0001 to 0.1 microgram/injection. The injections were commenced at varying intervals from the 1st to the 4th week after immunization. T-Cell subsets in the spleens were determined 2 weeks after the first antigen injection and thyroid infiltration was determined 3 weeks later. Treatment with Ta-1 between the two antigen injections increased the level of thyroiditis in resistant mice, but had no effect in sensitive mice. Treatment for the first 2 weeks had similar effects in resistant mice, but also suppressed thyroiditis in the sensitive strain. Later treatments, during the 3rd and 4th weeks after immunization also revealed immunomodulating properties of Ta-1, with a suppressing effect on thyroiditis in sensitive mice and an enhancing effect in the resistant strain. Both effects of Ta-1 were dose dependent. The effects of Ta-1 on the individual phenotypes were also dose dependent. The dose of 0.01 microgram greatly lowered the percentages of Lyt-2+3+ cells in D2 mice and mildly increased the percentages in Br mice, but did not change the Lyt-1+ cell level in either strain. On the other hand, the dose of 0.001 microgram greatly increased the percentage of Lyt-1+ cells in D2 mice and mildly decreased it in the Br strain, but did not alter the Lyt-2+3+ cell subset in either strain. Thus, both doses of Ta-1 modulated Lyt-1+/2+3+ ratios, with each dose affecting a different T-cell subset. The changes in the response to thyroglobulin are apparently exerted through the regulation of the functional T-cell subset balance.     

Inhibition of experimental autoimmune thyroiditis in mice by anti-I-A antibodies

Experimental autoimmune thyroiditis is induced in mice by immunization with thyroglobulin emulsified in Freund's complete adjuvant. The disease is characterized both by thyroid infiltration with mononuclear cells and by circulating thyroglobulin antibodies. The magnitude of the thyroid infiltration and the titer of thyroglobulin antibodies are controlled by genes in the I-A subregion of the major histocompatibility complex (H-2). We investigated the in vivo effect of monoclonal anti-Ia antibodies on experimental autoimmune thyroiditis in susceptible mice. Antibodies were given around the time of immunization, later after immunization, and to mice with established disease. Monoclonal antibody produced by the hybridoma line 10-3.6 (anti-I-Ak, s, u, v, z, f) completely prevented both production of thyroglobulin antibodies and thyroid infiltrates, when given shortly before or at the time of antigen administration. This effect was dose-dependent and this monoclonal antibody decreased the severity of the disease when given after the antigen challenge but did not fully suppress established thyroiditis. The same antibody markedly decreased the number of B lymphocytes in the spleen and decreased the thyroglobulin-induced spleen cell proliferation when either given in vivo or added in vitro to cell cultures. Antibodies produced by the hybridoma line 11.2.12 (anti-I-Ak) did not show an inhibitory effect on the disease. These experiments suggest that in this model of murine thyroiditis anti-Ia antibodies act on antigen-presenting cells. Furthermore, only one monoclonal antibody, anti-Ia, suppressed the immune response to thyroglobulin, suggesting a possible role for the isotype and specificity of anti-Ia antibody.     

Modulation of lymphocyte functions by group A streptococcal membrane

Protoplast membranes isolated from group A streptococci suppress functions of mouse B cells in vivo and in vitro. Intraperitoneal injection 24 or 72 hr (but not 12 hr) before collection of lymphoid cells results in a selective decrease in the mitogenic response of bone marrow cells to dextran sulfate (DS). The response of bone marrow cells to lipopolysaccharide (LPS), and spleen cells to both DS and LPS, is unaltered. In vitro exposure of lymphocytes to membranes concomitantly with mitogen reduces the response to both DS and LPS, however, the DS response is more susceptible to low doses of membrane. Suppression of the response to DS in vitro is not mediated by cells bearing Thy 1.2 antigen. Neither the phytohemagglutinin (PHA)-responsive cells nor the adherent cells participate in suppression of the LPS response in vitro. In contrast to the suppression of B-cell functions neither the PHA nor concanavalin A (Con A) response of mouse bone marrow, spleen, or thymus cells is altered by streptococcal protoplast membranes injected 24 hr before collection of cells. In vitro exposure of spleen cells to a limited range of concentrations of membrane results in an enhanced proliferative response of spleen cells stimulated by suboptimal doses of PHA. This synergism is not mediated by the adherent cells. Addition of membranes to spleen cell cultures in vitro has no effect upon the response of spleen cells to suboptimal doses of Con A or to optimal doses of either Con A or PHA. Higher concentrations of membranes reduce the proliferative response of both control and mitogen-stimulated cells. This nonselective suppression by high doses of membranes is not due to toxicity. Delayed hypersensitivity to sheep erythrocytes is potentiated by injection of membranes. These studies suggest that streptococcal membranes preferentially suppress the immature B cells and enhance certain T-cell functions.     

Lack of effect of propylthiouracil and methylmercaptoimidazole on thyroglobulin biosynthesis

Experiments were performed both in vivo and in vitro to test a previous proposal that part of the antithyroid action of the thioureylene drugs, propylthiouracil (PTU) and methylmercaptoimidazole, can be attributed to inhibition of thyroglobulin (Tg) biosynthesis. Rat thyroid lobes were incubated in leucine-free Eagle's medium containing bovine thyroid-stimulating hormone and 0, 0.1-0.2, or 1 mM drug. After a 30-min preincubation, 5 mu Ci of [14C]leucine were added and the incubation was continued for 4 hr. The soluble fraction was analyzed by sucrose density gradient centrifugation, and the fractions corresponding to the 19S Tg peak were pooled and assayed for 14C. No inhibition of 14C incorporation into 19S Tg was observed, even in thyroid lobes incubated in the presence of 1 mM methylmercaptoimidazole or 2 mM PTU. At the same time, 14C incorporation into 19S Tg was completely inhibited when lobes were incubated in the presence of 0.1 mM puromycin. In vivo, rats received an injection of PTU (1 mumol/100 g body wt), followed 60 min later by an injection of 25 mu Ci of [14C]leucine. Blood samples and thyroids were taken 5 hr after the [14C]leucine injection. Serum thyroid-stimulating hormone was not significantly affected by the PTU injection. The thyroid-soluble fraction was analyzed by sucrose density gradient centrifugation. No significant differences between saline and PTU-injected groups were observed in [14C]leucine incorporation into 19S Tg. We conclude from both our in vitro and our in vivo studies that PTU and methylmercaptoimidazole have no inhibitory effect on thyroglobulin synthesis in rat thyroids and that such inhibition does not play a significant role in the antithyroid action of these drugs.     

Suppression in murine experimental autoimmune thyroiditis: in vivo inhibition of CD4+ T cell-mediated resistance by a nondepleting rat CD4 monoclonal antibody.

Genetically susceptible mice become resistant to experimental autoimmune thyroiditis (EAT) induction with mouse thyroglobulin (MTg) and lipopolysaccharide after pretreatment with deaggregated MTg (dMTg). Recent work showed this suppression to be mediated by CD4+ suppressor T cells (Ts). To study Ts action in vivo, we used a rat IgG2a monoclonal antibody (mAb), YTS 177.9, which modulates CD4 antigen in vivo without depleting CD4+ cells. Initial studies showed that after two 1-mg doses of mAb 7 days apart, extensive CD4 antigen modulation of peripheral blood leukocytes occurred within 4 days. Mice given CD4 mAb 24 hr before dMTg (2 doses, 7 days apart) were resistant to EAT induction when immunized with MTg and LPS 20 days later. Also, anti-rat IgG2a titers were reduced following challenge with heat-aggregated rat IgG2a compared to controls. Subsequent analysis of serum in CD4 mAb-treated animals revealed that mAb was present in the circulation for 14 days. Moreover, mice given CD4 mAb and dMTg, then challenged after only 10 days, when CD4 mAb was still circulating, developed a significantly higher incidence of thyroid damage than controls. These findings suggest that modulation of CD4 antigen does not interfere with Ts activation, but the presence of CD4 mAb, at the time of autoantigenic challenge, can interfere with tolerance to EAT induction. Thus, the direct relationship between the presence of CD4 mAb and inhibition of EAT suppression implicates a role for CD4 molecules in the mediation of suppression.     

Peptides of human thyroglobulin reactive with sera of patients with autoimmune thyroid disease

Autoantibodies to thyroglobulin (Tg) are a prominent feature of the two autoimmune thyroid diseases, chronic lymphocytic (Hashimoto's) thyroiditis and Graves' disease. Similar autoantibodies are found in the serum of many normal individuals without evidence of thyroid disease. Previous studies have indicated that patients with autoimmune thyroid disease recognize epitopes of Tg which are not usually recognized by normal individuals. The goal of this investigation was to identify peptide fragments of Tg bearing these disease-associated epitopes. For this purpose, we utilized a panel of mAbs that bind to different epitopes of the Tg molecule. One of these mAbs (137C1) reacted with an epitope that was also recognized by the sera of patients with autoimmune thyroiditis. In the present study, we show that two peptides (15 and 23 kDa) that reacted with mAb 137C1 are located in different parts of the Tg molecule. Each peptide inhibited the binding of mAb 137C1 to the other peptide and to the intact Tg, indicating that the same epitope was represented on the two peptides. Loops and helices of the secondary structure of the two peptides might be involved in the conformational epitope recognized by mAb 137C1. A striking finding of this study is that two apparently unrelated fragments of the Tg molecule bind to the same mAb. These findings may have important ramifications with regard to epitope spread and the progression of the autoimmune response to disease.     

Beneficial effect of testosterone in the treatment of chronic autoimmune thyroiditis in rats

Early thymectomy and sublethal irradiation of normal rats consistently induces a sex-dependent chronic autoimmune thyroiditis. Females are much more susceptible to this autoimmune disorder than are males. The possible therapeutic effects of testosterone (Te) on established autoimmune thyroiditis has been investigated in this model. The pathologic condition of the gland before treatment was monitored by a thyroid grafting and extirpation techniques. Te administration by either parenteral injection or implantation caused significant regression of established thyroiditis. Repeated doses of Te ester in oil were found to be more effective than powdered free-Te given by implantation, and frequently produced complete resolution of chronic lesions involving the entire gland. In these thyroids, there was reappearance of normal thyroid architecture and complete absence of mononuclear cellular infiltration. However, no inhibitory effect on serum autoantibody production to thyroglobulin was noted with any form of Te treatment. These observations strengthen the concept that cellular rather than humoral mechanisms are involved in the pathogenesis of thyroiditis.     

A specific IgG in Graves' ophthalmopathy and its relation to retro-orbital and thyroid autoimmunity.

An IgG (ophthalmopathic immunoglobulin) that binds to retro-orbital antigen was identified in serum from patients with active Graves'' ophthalmopathy, and its nature and specificity were investigated. Dose related binding of this immunoglobulin to retro-orbital antigens prepared from guinea pig harderian gland or porcine eye muscle was found, which could be abolished by prior incubation with antigen. The immunoglobulin did not bind to thyroid membranes, thyroid microsomes, or thyroglobulin or interact with liver, skeletal muscle, or fat membranes. Serum with high activity of thyrotrophin binding inhibiting immunoglobulin did not react with retro-orbital antigen, and this activity was not affected by preincubation of the serum with retro-orbital antigen. Thyroid stimulating hormone was also without effect on retro-orbital antigen. It is concluded that Graves'' ophthalmopathy is associated with a specific ophthalmopathic immunoglobulin that reacts with retro-orbital antigen as distinct from thyroid antigens, and that the autoimmune response is directed towards retro-orbital antigens. This suggests that the ophthalmopathy is an entity distinct from autoimmune thyroid disease.     

Characterization of a human follicular thyroid carcinoma cell line (UCLA RO 82 W-1)

A thyroid tumor cell line has been established from the metastases of a follicular carcinoma in a female patient. Although the primary tumor released thyroglobulin (Tg) into the circulation (greater than 10,000 ng/ml), the uptake of I131 was less than 2%. After 37 replications the doubling time was 4 days and confluency was reached after 7 days from inoculation of 3 x 10(7) cells. This human thyroid tumor cell line has now been growing in culture for several years. An aneuploid chromosomal pattern was observed (62-82 chromosomes). A pair of X chromosomes was present but no Y chromosome was found which is compatible with the female origin of the cell line. EM studies revealed the presence of microvilli. Immunoperoxidase staining using specific anti-human Tg antisera indicated the presence of Tg within the cells. Nude mice developed solid-cystic tumors within 6 months after injection of the cells. The basal release of immunodetectable Tg, as measured in a perifusion system, increased in response to thyroid stimulating hormone (TSH) (P less than 0.025) or TSH combined with theophylline (P less than 0.001). Unusual isoenzyme patterns for galactose-1-phosphate-uridyltransferase (GALT) and phosphoglucomutase1 (PGM1) were detected in the tumor, compared with normal human fibroblasts and blood cells and isoenzyme patterns from the patient's lymphocytes. Because this malignant human thyroid follicular cell line has retained the ability to synthesize Tg it represents a valuable model for the study of human follicular carcinomas.     

Immunoregulatory abnormalities in autoimmune thyroid diseases

We have investigated the ability of lymphocytes from normal subjects and patients with autoimmune thyroid diseases to respond to a thyroidal antigen (human thyroglobulin, hTG) and a non-thyroidal antigen (Keyhole limpet hemocyanin, KLH) in vitro, using a hapten (trinitrophenol, TNP)-carrier system. This system is based on the concept that the T helper cells which respond to hTG or KLH should stimulate anti-TNP antibody producing B cells in the presence of TNP conjugated hTG (TNP-hTG) or KLH (TNP-KLH). After 5 or 6 days of culture of peripheral blood mononuclear cells with pokeweed mitogen (PWM), PWM and TNP-hTG, or PWM and TNP-KLH, IgM anti-TNP and IgM anti-sheep red blood cell (SRBC) plaque forming cells (PFC) were enumerated. The results showed that (1) in normal controls, hTG caused only suppression in both TNP and SRBC response, and KLH caused dose-related enhancement and suppression in TNP response without a change in SRBC response, and (2) in patients, both hTG and KLH resulted in dose-related enhancement in TNP response without a change in SRBC response. These data suggest that patients with autoimmune thyroid diseases have regulatory cell abnormalities confined to a thyroid antigen.     

FURTHER STUDIES ON MOBILIZATION OF CFUs

Mobilization of CFUs from haemopoietic tissues into circulation was studied after injection of different bacterial lipopolysaccharides (LPS), zymosan, phytohaemagglutinin (PHA), concanavalin A (Con A), trypsin and di-isopropyl-fluorophosphate-inhibited trypsin. All bacterial LPS used gave an increase of CFUs in the peripheral blood at 1 h after i.v. injection. Some variation in activity could not be excluded. As with Salmonella typhosa LPS, zymosan gave an increase in circulating CFUs during the first few hr and a second peak a few days later. After injection of zymosan as well as S. typhosa LPS the second peak in the blood was accompanied by a large increase in CFUs numbers in the spleen. PHA gave an immediate mobilization of CFUs, but the mobilization after injection of Con A during the first few hr occurred more slowly. After injection of S. typhosa LPS, zymosan and PHA the blood C3 level was found to be depressed considerably. This might indicate that the complement system is involved in the early mobilization of CFUs. Dexamethasone, a synthetic hormone which has been reported to give sequestration of several cell types in the bone marrow, did not inhibit the early and late mobilization of CFUs which normally occurs after injection of S. typhosa LPS.     

Analyses of the serum concentrations and antigenic determinants of thyroglobulin in chickens susceptible to autoimmune thyroiditis

These studies revealed an abnormal elevation in serum thyroglobulin (Tg) concentrations of the thyroiditis-prone OS chicks. Their serum levels, measured by a sensitive and specific RIA, remained high for the first 2 wk of age, a time preceding significant lymphoid infiltration of their thyroid glands. The elevated serum Tg levels probably resulted from, or were related to, hyperactivity of the OS thyroid gland. Reducing the activity of the OS thyroid gland by exogenous administration of T4 caused a temporary but significant reduction in thyroidal infiltration and the synthesis of Tg antibodies. In addition to the analysis of serum concentrations of Tg, a competitive binding RIA was developed to determine whether unique antigenic determinants exist on Tg isolated from OS thyroid glands. This was proved to be unlikely because OS chicks produced autoantibodies that fully cross-reacted with Tg isolated from normal chicken thyroid glands. The relationships between intrinsic thyroid hyperactivity, high serum Tg levels, the sensitization of OS thymocytes, and thyroiditis are discussed.     

Critical role of iodination for T cell recognition of thyroglobulin in experimental murine thyroid autoimmunity

We have used two clonotypically distinct thyroglobulin (Tg)-specific, I-Ak restricted monoclonal T cell populations to investigate the role of thyroid peroxidase-catalyzed iodination in Tg recognition by autoreactive T cells. The results showed that these T cells could recognize Tg only it it was sufficiently iodinated. Unlike normal mouse Tg, noniodinated mouse Tg was unable to induce significant thyroid lesions but could trigger the production of Tg autoantibodies. In these experiments, the importance of T cell recognition of iodination-related epitopes was emphasized by the inability of serum antibodies to distinguish Tg on the basis of iodine content, whether they were induced with normal or noniodinated Tg. Therefore, thyroid peroxidase-dependent modification of Tg would appear to be central to its recognition by autoreactive T cells and hence its capacity to induce autoimmune thyroid lesions.     

Effect of bacterial lipopolysaccharide on the in vitro secondary antibody response in mice. I. Description of the suppressive capacity of lipopolysaccharide.

Bacterial lipopolysaccharide (LPS) suppressed the in vitro secondary antibody response in mice to the protein antigens human gamma globulin (HGG) and turkey gamma globulin (TGG). This is the first report of LPS inhibiting a secondary antibody response. Consistent suppression was dependent on the time of LPS addition; LPS added at culture initiation was less effective than LPS added 12 to 48 hr later. The mitogenic moiety of LPS was the inhibitory principle, as shown by the lack of suppression of spleen cells from C3H/HeJ mice, and the inability of the polysaccharide component, but not the lipid component of LPS, to suppress A/J spleen cells. The mechanism of suppression by LPS was not due to large numbers of B cells proliferating in response to LPS, since removal of B cells not bearing specificity for the priming antigen did not reduce suppression by LPS. However, the possibility exists that LPS may act through B cells specific for the priming antigen.     

The thyroglobulin gene as the first thyroid-specific susceptibility gene for autoimmune thyroid disease

Recent linkage and association studies provide evidence for thyroglobulin (Tg) being an autoimmune thyroid disease (AITD) susceptibility gene. The Tg locus has been reported to be linked with AITD in two independent studies, and further analysis demonstrated that markers within the Tg gene were associated with AITD. Furthermore, missense single-nucleotide polymorphisms (SNPs) in the Tg gene were shown to be associated with autoimmune thyroiditis in both mice and humans. If Tg is confirmed as a susceptibility gene for AITD, it could provide a novel therapeutic target.     

Further studies on mobilization of CFUs.

Mobilization of CFUs from haemopoietic tissues into circulation was studied after injection of different bacterial lipopolysaccharides (LPS), zymosan, phytohaemagglutinin (PHA), concanavalin A (Con A), trypsin and di-isopropyl-fluorophosphate-inhibited trypsin. All bacterial LPS used gave an increase of CFUs in the peripheral blood at 1 h after i.v. injection. Some variation in activity could not be excluded. As with Salmonella typhosa LPS, zymosan gave an increase in circulating CFUs during the first few hr and a second peak a few days later. After injection of zymosan as well as S. typhosa LPS the second peak in the blood was accompanied by a large increase in CFUs numbers in the spleen. PHA gave an immediate mobilization of CFUs, but the mobilization after injection of Con A during the first few hr occurred more slowly. After injection of S. typhosa LPS, zymosan and PHA the blood C3 level was found to be depressed considerably. This might indicate that the complement system is involved in the early mobilization of CFUs. Dexamethasone, a synthetic hormone which has been reported to give sequestration of several cell types in the bone marrow, did not inhibit the early and late mobilization of CFUs which normally occurs after injection of S. typhosa LPS.     

Is there a double pathway for the secretion of thyroglobulin: a "short circuit" weakly glycosyl-iodinated and a "long circuit" strongly glycosyl-iodinated?

Intact rat thyroid lobes incubated in vitro release recently synthesized thyroglobulin (Tg) into the media at a faster rate than they release thyroglobulin stored in follicular structures. Differential release of this Tg fraction cannot be explained by morphological alterations in thyroid architecture during incubation. This rapidly excreted fraction exhibits a low density on rubidium chloride gradients characteristic of poorly sialylated and poorly iodinated thyroglobulin, comigrating on rubidium chloride gradients with thyroglobulin isolated from tunicamycin treated glands. This poorly sialylated and poorly iodinated thyroglobulin is itself unaffected in its density or release into the media by tunicamycin treatment. Tg isolated from the media of tunicamycin treated glands has nearly the same low iodine and low sialic acid content as rat serum thyroglobulin and does not incorporate radiolabelled glucosamine. This fraction thus appear to duplicate properties of low glycosylated-low iodinated thyroglobulin released from thyroid cells in organisms that have no follicular structures and no follicular storage process as well as from thyroid tissue in patients with thyroid disease states, particularly thyroid tumors. Thus it is proposed a "short loop" pathway of low-glycosylated low-iodinated thyroglobulin directly into circulation, that bypasses and is not stored in the follicular lumen, the "long loop".