Superiority of thyroid peroxidase DNA over protein immunization in replicating human thyroid autoimmunity in HLA-DRB1*0301 (DR3) transgenic mice.

Murine experimental autoimmune thyroiditis (EAT), characterized by thyroid destruction after immunization with thyroglobulin (Tg), has long been a useful model of organ-specific autoimmune disease. More recently, porcine thyroid peroxidase (pTPO) has also been shown to induce thyroiditis, but these results have not been confirmed. When (C57BL/6 x CBA)F(1) mice, recently shown to be susceptible to mouse TPO-induced EAT, were immunized with plasmid DNA to human TPO (hTPO) and cytokines IL-12 or GM-CSF, significant antibody (Ab) titres were generated, but minimal thyroiditis was detected in one mouse only from the TPO + GM-CSF immunized group. However, after TPO DNA immunization of HLA-DR3 transgenic class II-deficient NOD mice, thyroiditis was present in 23% of mice injected with TPO + IL-12 or GM-CSF. We also used another marker for assessing the closeness of the model to human thyroid autoimmunity by examining the epitope profile of the anti-TPO Abs to immunodominant determinants on TPO. Remarkably, the majority of the anti-TPO Abs was directed to immunodominant regions A and B, demonstrating the close replication of the model to human autoimmunity. TPO protein immunizations of HLA-DR3 transgenic mice with recombinant hTPO did not result in thyroiditis, nor did immunization of other mice expressing HLA class II transgenes HLA-DR4 or HLA-DQ8, with differential susceptibility to Tg-induced EAT. Moreover, our efforts to duplicate exactly the experimental procedures used with pTPO also failed to induce thyroiditis. The success of hTPO plasmid DNA immunization of DR3(+) mice, similar to our reports on Tg-induced thyroiditis and thyrotropin receptor DNA-induced Graves' hyperthyroidism, underscores the importance of DR3 genes for all three major thyroid antigens, and provides another humanized model to study autoimmune thyroid disease.

Graves' hyperthyroidism and thyroiditis in HLA-DRB1*0301 (DR3) transgenic mice after immunization with thyrotropin receptor DNA.

Familial and twin studies in Caucasians have established that the MHC class II allele HLA-DRB1*0301 (DR3) is a strong susceptibility gene in Graves' hyperthyroid disease (GD). To determine if a DR3 transgene could help establish an animal model for GD, we expressed DR3 molecules in class II-knockout NOD mice (H2Ag7-). DR3+g7- mice were given cardiotoxin prior to immunization on weeks 0, 3 and 6 with plasmid DNA encoding human thyrotropin receptor (TSHR). Two groups of mice were also coimmunized with plasmid DNA for IL-4 or GM-CSF. Serial bleeds on weeks 8, 11 and 14 showed that approximately 20% of mice produced thyroid-stimulating antibodies (Abs), and approximately 25% had elevated T4 levels. In particular, a subset displayed both signs of hyperthyroidism, resulting in approximately 30% with some aspect of GD syndrome. Additional mice had thyroid-stimulating blocking Abs and/or TSH-binding inhibitory immunoglobulins, while most mice showed strong labelling of TSHR+ cells by flow cytometry. Interestingly, lymphocytic infiltration with thyroid damage and Abs to mouse thyroglobulin were also noted. Vector controls were uniformly negative. Thus, DR3 transgenic mice can serve as a model for GD, similar to our earlier reports that this allele is permissive for the Hashimoto's thyroiditis model induced with human thyroglobulin.

Characterization of a novel H2A(-)E+ transgenic model susceptible to heterologous but not self thyroglobulin in autoimmune thyroiditis: thyroiditis transfer with Vbeta8+ T cells.

Recently we reported on a novel H2E transgenic, IA-negative model of experimental autoimmune thyroiditis (EAT) that excludes reactivity to self in its susceptibility pattern to heterologous thyroglobulin (Tg). In conventional, susceptible mouse strains, EAT is inducible with both homologous and heterologous Tg; e.g., human (h)Tg shares conserved thyroiditogenic epitopes with mouse (m)Tg. However, when an H2Ea(k) transgene is introduced into class II-negative B10.Ab(0) mice, which express neither surface IA (mutant Abeta-chain) nor surface IE (nonfunctional Ea gene), the resultant H2E(b) molecules are permissive for EAT induction by hTg, but not self mTg. Also, the hTg-primed cells do not cross-react with mTg. To explore this unique capacity of E+B10.Ab(0) mice to distinguish self from nonself Tg, we have developed T cell lines to examine the T cell receptor repertoire and observed a consistent Vbeta8+ component after repeated hTg stimulation. Enrichment and activation of Vbeta8+ T cells by either superantigen staphylococcal entertoxin B or anti-Vbeta8 in vitro enabled thyroiditis transfer to untreated A-E+ recipients, similar to hTg activation. Vbeta8+ T cells isolated by FACS from hTg-immunized mice also proliferated to hTg in vitro. These studies support the contribution of Vbeta8 genes to the pathogenicity of hTg in this H2A-E+ transgenic model.

Flexibility of TCR repertoire and permissiveness of HLA-DR3 molecules in experimental autoimmune thyroiditis in nonobese diabetic mice.

Experimental autoimmune thyroiditis (EAT) is inducible in genetically susceptible mice by immunization with mouse thyroglobulin (mTg). With susceptibility linked to MHC class II, EAT is useful in studying human leukocyte antigen (HLA) associations with Hashimoto's thyroiditis. In non-obese diabetic (NOD) mice, approximately 10% thyroiditis incidence occurs with aging. This potential was exploited to examine the T cell repertoire and HLA association in EAT. Similar to B10.K-Vbeta(c)mice with TCRBV genes reduced by approximately 70%, mTg-immunized NOD-Vbeta(c)mice developed thyroiditis comparable to controls, indicating plasticity of the TCR repertoire for pathogenic epitopes. HLA association was evaluated by introducing HLA-DRA/DRB1*0301 (DR3) transgene into class II-negative NOD mice (Ab(0)/NOD). Previously, this HLA-DR3 transgene rendered EAT-resistant B10.M and Ab(0)mice susceptible to both mTg- and hTg-induced EAT. These results are now confirmed. mTg-induced thyroiditis in DR3+ Ab(0)/NOD mice was comparable to that in NOD and DR3- NOD mice, and the proliferative response was stronger. By comparison, NOD mice were only moderately susceptible to hTg-induced EAT. However, thyroiditis was more severe in DR3+ Ab(0)/NOD than in DR3- NOD mice, with no difference in proliferative response to hTg harbouring heterologous epitopes. The confirmed permissiveness of HLA-DR3 molecules on an NOD background for EAT induction by both mTg and hTg supports the importance of this class II gene implicated in some patient studies.

Histologic and histochemical characterization of seminal vesicle intraluminal secretions.

CONTEXT: We have observed intraluminal crystalloid morphology in seminal vesicles that is superficially similar to that seen in prostate neoplasia, but found little information on such morphology in the literature. DESIGN: Two hundred fifty-three prostate specimens (163 needle biopsies, 75 radical prostatectomies with prostate carcinoma, 11 prostates from autopsy, and 4 cystoprostatectomies without prostate carcinoma) were examined for seminal vesicle secretions, which were categorized as (a) dense platelike inspissated, (b) fluidlike, (c) crystalloid morphology, and (d) absent. Histochemical stains (periodic acid-Schiff with and without diastase, Alcian blue at pH 2.5, and mucicarmine) were performed to characterize the nature of secretions. RESULTS: Proteinaceous secretions were identified in 82% of seminal vesicles examined. Of these, 61% had predominantly dense, platelike, inspissated secretions, 15% had predominantly fluidlike secretions, and 24% had predominantly crystalloid morphology. Although in some cases the crystalloid morphology resembled that of prostatic intraluminal crystalloids, the seminal vesicle crystalloids differed in that they were invariably multiple, had curved edges, and had varied forms (elliptical, cylindrical, rodlike, and rectangular). Seventy-one percent of seminal vesicle crystalloids were associated with dense, platelike, inspissated secretions and appeared to be created by fracturing within platelike secretions. There was no relationship between seminal vesicle crystalloid morphology and associated malignancy in the prostate gland, as it was seen in 24% of cases with prostate carcinoma and 25% of cases without prostate carcinoma (P = 1.0000). Fluidlike secretions were positive for Alcian blue (pH 2.5) and mucicarmine, whereas dense platelike secretions and crystalloid morphology were negative for Alcian blue (pH 2.5) and mucicarmine. CONCLUSIONS: Seminal vesicle secretions are fairly common and, when fluidlike, are composed of acid mucopolysaccharides. Inspissation of secretions appears to be associated with loss of acidity, presumably resulting in dense platelike secretions and crystallization. Awareness of both the crystalloid morphology in seminal vesicle tissue and the distinguishing features from prostatic crystalloids may be important while interpreting prostate needle biopsies in which seminal vesicle epithelium may be confused for prostate carcinoma because of a small acinar morphology with accompanying cytologic atypia and crystalloid morphology.

Participation of Vbeta13(+) and Vbeta1(+) T cells in transfer thyroiditis after activation of mouse thyroglobulin-primed T cells by superantigen staphylococcal enterotoxin A.

Murine experimental autoimmune thyroiditis (EAT) is a T-cell-mediated disease, but the T cell receptor (TCR) Vbeta gene usage in pathogenesis has not been well delineated. One approach is to utilize bacterial superantigens, such as staphylococcal enterotoxin (SE) A and B, to stimulate known sets of TCR Vbeta families in mouse thyroglobulin (mTg)-primed cells for thyroiditis transfer. Our previous use of SEB to activate mTg-primed cells led to no thyroiditis transfer, despite a major increase in Vbeta8(+) T cells. Unlike SEB, SEA activation did transfer thyroiditis. To determine which thyroiditogenic Vbeta(+) T cells were involved, SEA-activated T cells have now been analyzed. After repeated SEA activation in vitro, both mTg-reactive and thyroiditogenic cells persisted. FACS analysis indicated that most Vbeta13(+) cells were "large" cells (IL-2R(+)) and expressed the activation marker, transferrin receptor (CD71). RT-PCR analysis also showed the presence of both Vbeta13(+) and SEA-reactive Vbeta1(+) cells. Since our previous analyses by RT-PCR of the thyroid infiltrate after either induction or adoptive transfer have implicated both Vbeta13(+) and Vbeta1(+) cells, their activation by SEA to transfer thyroiditis further supports their role.

H2-E transgenic class II-negative mice can distinguish self from nonself in susceptibility to heterologous thyroglobulins in autoimmune thyroiditis.

Susceptibility to experimental autoimmune thyroiditis (EAT) is linked to H2-A class II genes; k and s haplotypes are susceptible, while b and f are resistant. EAT is inducible with thyroglobulins (Tgs) from several mammalian species which share portions of identical sequences. But cross-activation and cross-tolerance studies with mouse (m), human (h), and porcine (p) Tg have indicated mTg-unique T-cell epitope(s), in addition to conserved, in EAT induction. The recent introduction of the HLA-DRB1*0301 (DR3) transgene rendered major histocompatibility complex (MHC) class II-negative (Ab(0)) mice susceptible to EAT induction by both hTg and mTg, suggesting usage of conserved epitopes. Here, we introduced the H2-Ea(k) transgene into resistant B10 (H2(b)) or Ab(0) mice with a defective Ea gene to provide functional surface H2E (b haplotype) expression. Surprisingly, both transgenic strains showed severe inflammation only after hTg, but not mTg, immunization, although the moderating influence of the A(b) gene in B10 was evident. In proliferative assays, hTg-primed cells did not respond to mTg, nor to conserved 12mer peptides from three primary hormonogenic sites, two of which can activate T cells for thyroiditis transfer and cytotoxicity. The vigorous response to hTg stimulation was reduced only by Ebeta(b)-specific monoclonal antibody. EAT induction with bovine and pTg showed responses similar to hTg, suggesting thyroiditogenic epitopes shared with hTg, but not mTg. This is the first demonstration of: (1) nonpermissiveness for EAT induction with mTg, normally the most thyroiditogenic Tg and the one with unique epitopes for susceptible mice, and (2) the separation of hTg from mTg in EAT induction in H2-E-transgenic mice.

Flexibility of the thyroiditogenic T cell repertoire for murine autoimmune thyroiditis in CD8-deficient (beta2m -/-) and T cell receptor Vbeta(c) congenic mice.

In murine experimental autoimmune thyroiditis (EAT), previous studies have revealed a highly adaptable thyroiditogenic T cell repertoire which involves both CD4+ and CD8+ T cells in the susceptible H2k strain. To further test this flexibility, congenic B10.K mice lacking CD8+ T cells (B2m -/-) or harboring 70% T cell receptor (TCR) Vbeta gene deletions (Vbeta(c)) were immunized with mouse thyroglobulin (MTg) and evaluated for EAT 28 days later. All B2m -/- mice developed moderate antibodies to MTg, and thyroidal inflammation was comparable to B10.K mice, averaging 35-40%. Spleen cells (SC) from MTg-immunized mice were then injected into syngeneic recipients after stimulation in vitro with MTg or with conserved, thyroxine (T4)- or thyronine (T0)- containing 12mer peptides, hT4(5), hT0(2553), or hT4(2553), derived from the primary hormonogenic sites at position 5 or 2553 of human Tg. As previously shown in another H2k strain (CBA/J), all three peptides activated MTg-primed SC to transfer EAT in B10.K mice. hT4(5) and hT4(2553) were further tested in B10.K-Vbeta(c) and beta2m- B10.K mice. Both peptides expanded thyroiditogenic T cells in either strain, resulting in severe thyroiditis in syngeneic recipients. That EAT can develop in the absence of CD8+ T cells or in the presence of a severely restricted TCR repertoire underscores the remarkable flexibility of the thyroiditogenic T cell profile in the susceptible k haplotype.

Primary hormonogenic sites as conserved autoepitopes on thyroglobulin in murine autoimmune thyroiditis: role of MHC class II.

A few synthetic peptides corresponding to amino acid sequences on human thyroglobulin (Tg) have been reported to induce moderate thyroiditis or activate mouse Tg (MTg)-primed T cells to transfer thyroiditis in mice susceptible to experimental autoimmune thyroiditis. Using three pairs of 12-mer peptides (1-12, 2549-2560, 2559-2570), with thyroxine (T4) or noniodinated thyronine (T0) at the conserved, hormonogenic site 5, 2553, or 2567 respectively, we reported that iodination was not required for a Tg hormonogenic site to be a thyroiditogenic autoepitope. To determine the relative importance of MHC class II and T cell receptor (TCR) repertoire, we compared two EAT-susceptible k and s (CBA and A.SW) haplotypes and their respective MHC-identical strain (C57BR and SJL) with approximately 50% genomic deletion of TCR Vbeta genes. Whereas k and s strains develop MTg-induced EAT, vigorous immunization with peptides containing T4 or T0 at either 5 or 2553, but not at 2567, led to mild (10-20%) thyroiditis only in some mice of either k strain. TCR Vbeta gene differences played a minor role. T cell responses to all peptide pairs were quite similar in CBA and C57BR mice, and both hT0(2553) and hT4(2553) reciprocally primed and stimulated their T cells. In adoptive transfer, SJL mice were somewhat more responsive to peptide activation than A.SW but much weaker than k strains. By comparing T4- and T0-containing peptides in different haplotypes, we show further that antigenicity of conserved hormonogenic sites is intrinsic, dependent more on amino acid sequence and binding to appropriate class II molecules and less on TCR repertoire or iodination of T0.

Role of mouse and human class II transgenes in susceptibility to and protection against mouse autoimmune thyroiditis.

Mouse experimental autoimmune thyroiditis (EAT), a model for Hashimoto's thyroiditis, is induced by immunizing with mouse thyroglobulin (MTg). To study the extent of H2A involvement in EAT, we introduced AaAb genes from susceptible k mice into resistant or intermediately susceptible strains which do not express H2E molecules. Thyroiditis was severe in resistant B10.M (H2(f)) mice carrying the double transgene AakAbk. Likewise, thyroid infiltration was significantly extended in intermediate B10.Q (H2(q)) mice with the same transgene. To examine the effect of H2E molecules in the presence of H2A-mediated susceptibility, we introduced an Eak transgene into E- B10.S mice to express the Ebetas molecule and observed significant reduction in EAT severity in B10.S(E+) mice. On the other hand, the presence of an Ebd transgene in B10.RQB3 (H2Aq) mice resulting in the expression of H2Ebetad molecules did not alter EAT susceptibility, suggesting a role for Eb gene polymorphism in protection against EAT. We have shown recently that the HLA-DRB1(*)0301 (DR3) transgene conferred EAT susceptibility to B10. M as well as class II-negative B10.Ab0 mice. However, we report here that the HLA-DQB1(*)0601 (DQ6b) transgene in B10.M or HLA-DQA1(*)0301/DQB1(*)0302 (DQ8) transgene in class II-negative Ab0 mice did not. These studies show the differential effects of class II molecules on EAT induction. Susceptibility can be determined when class II molecules from a single locus, H2A or HLA-DQ, are examined in transgenic mice, but the overall effect may depend upon the presence of both class II molecules H2A and H2E in mice and HLA-DQ and HLA-DR in humans.

HLA-DRB1 polymorphism determines susceptibility to autoimmune thyroiditis in transgenic mice: definitive association with HLA-DRB1*0301 (DR3) gene.

Familial clustering of autoimmune thyroid diseases has led to studies of their association with human major histocompatibility complex (MHC) class II genes. One such gene implicated in Hashimoto's thyroiditis (HT) is HLA-DR3, but the association is weak and is contradicted by other reports. On the other hand, murine experimental autoimmune thyroiditis (EAT), a model for HT, presents a clear linkage with MHC class II. Moreover, it is inducible with thyroglobulin (Tg), the common autoantigen in either species. Immunization of HLA-DRB1* 0301 (DR3) transgenic mice with mouse or human Tg resulted in severe thyroiditis. In contrast, transgenic mice expressing the HLA-DRB1*1502 (DR2) gene were resistant to EAT. Our studies show that HLA-DRB1 polymorphism determines susceptibility to autoimmune thyroiditis and implicate Tg as an important autoantigen.

V beta 8.2 transgene expression interferes with development of experimental autoimmune thyroiditis in CBA k/q but not k/k mice.

The thyroiditogenic T cell receptor (TCR) repertoire is not yet well defined in murine experimental autoimmune thyroiditis (EAT). Our recent work has shown that, while V beta 8+ T cells have no major role in EAT induction with mouse thyroglobulin (MTg), V beta 13 may be involved. To examine the effect of skewing the TCR repertoire on EAT development, CBA (H2k) mice were mated with B10.Q mice harboring an ovalbumin-specific V beta 8.2 TCR transgene (trg), and the trg+ mice were backcrossed to CBA. FACS analysis showed that peripheral blood T cells from trg+ mice had about 76 and 90% V beta 8.2+ cells in the CD4+ and CD8+ subsets, respectively, compared with about 15 and 11% in trg- sibs. The transgenic CBA k/k and k/q mice were immunized with MTg and sacrificed 28 days later. In all trg+ mice, anti-MTg titers and T cell proliferative responses to MTg were significantly lowered. However, thyroid infiltration was distinctly different in the two strains of transgenic mice; a significant decrease was seen primarily in k/q, but not k/k, trg+ mice. Thus, skewing the TCR repertoire to overexpress an irrelevant TCR revealed the possession of a less flexible thyroiditogenic TCR repertoire in k/q, but not k/k, mice.

Primary hormonogenic sites as conserved autoepitopes on thyroglobulin in murine autoimmune thyroiditis. Secondary role of iodination.

We hypothesized earlier that conserved T cell epitopes and those unique to mouse thyroglobulin (MTg) contributed to its total thyroiditogenicity in murine autoimmune thyroiditis. Recent studies of synthetic peptides from human Tg (HTg) revealed no immunodominant epitopes. The role of iodine residues, considered by some to render Tg immunogenic, became unclear, since only one 12-mer peptide contained thyroxine (T4) positioned at hormonogenic site 2553. It primed T cells for thyroiditis transfer, but noniodinated peptide containing thyronine (T0) was not compared. To determine 1) whether other primary hormonogenic sites were likewise immunogenic and 2) whether iodination was requisite for this and other sites to be an autoepitope, we derivatized three pairs of 12-mer peptides, 1-12, 2549-2560, 2559-2570, containing T0 or T4 at positions 5, 2553, or 2567, respectively. The six peptides were used to stimulate MTg-primed cells in vitro and to immunize mice. None directly induced thyroiditis; peptide Abs were the lowest in mice given hT0(2567) or hT4(2567). Of the three T4-containing peptides, hT4(5) and hT4(2553), but not hT4(2567), stimulated MTg-primed or HTg-primed T cells in vitro, with hT4(2553) being the stronger. Comparing hT0(2553) with hT4(2553), both activated MTg-primed, or peptide-primed, T cells to transfer thyroiditis. The marked immunogenicity of noniodinated hT0(2553) and the poor antigenicity of hT4(5) and hT4(2567) demonstrate that immunogenicity of a conserved hormonogenic site is dependent more on its amino acid sequence than on T4 substitution. Iodination may enhance antigenicity and/or binding affinity, but it is not required for a Tg hormonogenic site to be an autoepitope.

Noninvolvement of V beta 8+ T cells in murine thyroglobulin-induced experimental autoimmune thyroiditis.

In experimental autoimmune thyroiditis (EAT) induced with mouse thyroglobulin (MTg), T cell receptor (TCR) V beta gene usage in the pathogenesis of disease is unknown. We report here studies evaluating V beta 8 gene usage in EAT, as V beta 8+ T cells are reportedly involved in some experimental autoimmune diseases. Spleen cells (SC) from MTg-immunized CBA/J (H-2k) mice were activated in vitro for adoptive transfer into syngeneic recipients. Elimination of V beta 8+ T cells by treating recipients with V beta 8 monoclonal antibody (mAb) following transfer of MTg-activated SC did not reduce disease severity. Conversely. MTg-primed SC were stimulated in vitro with V beta 8 mAb or staphylococcal enterotoxin B, which activates V beta 8+ T cells in CBA/J mice. Neither activated population transferred disease, in contrast to cells activated with MTg. Thus, in MTg-induced EAT, V beta 8+ T cells do not play a major role in pathogenesis.

Depletion of CD4+ and CD8+ cells eliminates immunologic memory of thyroiditogenicity in murine experimental autoimmune thyroiditis.

Experimental autoimmune thyroiditis (EAT) develops in genetically susceptible mice after immunization with mouse thyroglobulin (MTg), and is mediated by T cells, both CD4+ and CD8+, infiltrating the thyroid. Previous work showed that depletion of CD4+, but not CD8+, cells with rat monoclonal antibodies (mAbs) interfered with EAT induction. To test if concomitant CD4+ cell depletion and immunization led to EAT resistance, mice were reimmunized at an interval of 15 or 43 days after injection of CD4 mAbs. No resistance had been established; disease severity and anti-MTg titers were comparable to mice with primary immunization. Previous work also showed that treatment during advancing EAT with only CD4 mAbs on days 21, 25 led to long-lasting, reduced severity in EAT, whereas administration of CD8 mAbs alone reduced the smaller CD8+ subset only. However, therapy with both mAbs was most efficacious; > 50% of thyroids were purged of all cellular infiltrate after only two doses. Moreover, T cells emerging subsequent to depletion were not retained in the thyroid, despite ongoing antibody production. To test if nondepleting CD4 and CD8 mAbs were similarly effective for therapy, mAbs of the IgG2a isotype were administered during advancing EAT. No effect on thyroidal infiltration was observed, indicating that modulation of the CD4 and CD8 antigen without depletion was insufficient for efficacious therapy. To determine if combined therapy with depleting mAbs reestablished self tolerance, treated mice were reimmunized on days 70, 77, when T cell recovery was nearly complete. Thyroiditis was comparable to controls given primary immunization, despite high antibody levels.(ABSTRACT TRUNCATED AT 250 WORDS)

Coronary vessel alterations following chronic carbon monoxide exposure in the adult rat.

Adult male rats were exposed to 500 ppm CO continuously for 30 days, while litter-mate controls remained in room air (AIR). Heart weight-to-body weight ratio and hematocrit were increased significantly. Right ventricle (RV) free wall thickness was increased significantly as was right to left heart diameter and average heart diameter. Cross-sectional areas of the left ventricle (LV) free wall, interventricular septum (S) and RV midway between the apex and base were increased significantly. Morphometric analysis of the CO-exposed and AIR hearts revealed no significant differences in the number of small (27-114 microns) or larger (> 114 microns) blood vessels in any region; however, there was a trend towards an increased number of the smaller vessels, both arterioles and venules, in the CO-exposed rats. The larger arteries in the S and RV were significantly larger in the CO-exposed rats. There was a significant overall effect of CO on larger artery diameter across all heart regions, consistent with the appearance of heart radiographs taken. There were no differences in the diameter of the small vessels in any region of the heart between the CO-exposed and AIR rats. The vessel cross-sectional area of the larger vessels tended to be increased in all regions of the heart. The cross-sectional area of the large arteries in the LV was increased significantly. Arterial wall thickness was decreased significantly in RV and there was a significant overall effect of CO in decreasing wall thickness and the ratio of wall thickness-to-vessel luminal diameter in these vessels. No vascular pathology was observed. The results of this study suggest changes in coronary vessel architecture during chronic CO-induced cardiac hypertrophy and are consistent with earlier hemodynamic and morphometric studies of CO-exposed hearts.

Characterization of resistance to murine experimental autoimmune thyroiditis: duration and afferent action of thyroglobulin- and TSH-induced suppression.

Genetically susceptible mice develop experimental autoimmune thyroiditis (EAT) after immunization with mouse thyroglobulin (MTg). Earlier studies have shown that resistance to EAT induction can be activated by two regimens, pretreatment with deaggregated MTg (dMTg) or with thyroid-stimulating hormone (TSH). With both protocols, suppression is linked to a > or = 2-3 day increase in circulatory MTg level and is mediated by CD4+ suppressor T cells (Ts). To assess the duration of suppression, CBA (H-2k) mice were injected with dMTg or infused with TSH via an osmotic pump for 3-4 days and then challenged with MTg + adjuvant at intervals up to 73 days for dMTg-pretreated mice or up to 94 days for TSH-pretreated mice. Suppression was measurable for at least 73 days after injected dMTg. TSH-induced suppression was also long-lasting; resistance was strong 38 days after TSH infusion and was still measurable on Day 66. The Thy-1+, CD4+ Ts which transfer MTg-induced suppression were further characterized by treatment with I-J antibodies plus complement prior to transfer. This treatment abolished the transfer of suppression which acts in the afferent phase to interfere with EAT induction. The capacity of Ts to suppress the efferent phase of EAT was assessed in vitro and in vivo. Cells from dMTg-pretreated mice did not block the in vitro proliferative response of MTg-primed cells to MTg, nor did these cells, when left intact in situ, reduce the severity of disease produced by the adoptive transfer of thyroiditogenic cells. Similarly, TSH-induced suppression was ineffective in preventing adoptively transferred EAT. Since suppression, which correlates with a temporary increase of circulatory MTg, occurs only at the afferent phase of active immunization, these findings lend support to our earlier hypotheses that circulatory MTg serves a physiologic role in maintaining normal self-tolerance by sustaining low levels of Ts activation and that additional rise above baseline increases and prolongs resistance to EAT induction.

Chronic carbon monoxide exposure in young rats alters coronary vessel growth.

The goal of this study was to determine whether chronic monoxide exposure in the developing heart produces long-lasting coronary vasculature alterations. One-day-old male rat pups were exposed to 500 ppm CO continuously for 30 d, while littermate controls remained in room air (AIR). At 61 and 110 d of age hearts were removed, perfusion fixed, x-rayed, and processed for analysis of coronary vessel architecture. Body weight (BW) and heart weight (HW) increased with age; the ratio of HW/BW decreased. There were no differences in HW and ventricular dimensions at either age due to prior CO exposure. Morphometric analysis of the fixed hearts from CO-exposed and AIR rats revealed no significant individual group differences in the number of small (27-114 microns) or larger (> 114 microns) vessels in any heart region. The septum (S) in CO rats was an exception: There were more small veins at 61 d of age and more larger veins at 110 d of age. There was a significant increase in the number of small arteries at both ages in the CO rats across all heart regions, and in the smaller veins at 61 d of age. The large vessels in the S at 61 d of age had a significantly greater diameter in CO compared to AIR rats. This was also true for the large arteries in the S and right ventricle (RV) of the 110-d-old rats. Taking all heart regions together, the large arteries in CO rats were larger than in AIR rats. Previous CO exposure significantly increased large artery and total cross-sectional area in the S and RV at 61 d of age, and in RV at 110 d of age. Total cross-sectional area of veins in the S was also increased. Taking all heart regions together, CO significantly increased small artery cross-sectional area at 61 d of age, and small, large, and total artery cross-sectional area at 110 d of age. With one exception (small veins, 110 d of age), there was no effect of CO on vein cross-sectional area. These changes resulted in the percentage of total cross-sectional area contributed by the larger vessels being increased. Pathological examination showed nothing abnormal. The results suggest profound and persistent changes in coronary vessel architecture following chronic neonatal CO exposure.

Resistance to experimental autoimmune thyroiditis is correlated with the duration of raised thyroglobulin levels.

We have used the mouse model of experimental autoimmune thyroiditis (EAT) to examine the hypothesis that the strengthening of self-tolerance to thyroglobulin by exogenous mouse thyroglobulin (MTg) or stimulation of endogenous MTg secretion by thyroid-stimulating hormone (TSH) is correlated with the length of time MTg rises above the normal range. Bacterial lipopolysaccharide (LPS) treatment increases the initial half-life of MTg from about 3 hr to about 5 hr, probably interfering with its clearance by the mononuclear phagocytic (reticuloendothelial) system. By pretreating mice with LPS, a subtolerogenic MTg dose is rendered tolerogenic. Similarly the effect of TSH infusion by osmotic minipumps, which stimulates MTg secretion and also strengthens tolerance to MTg, can be enhanced by injecting LPS shortly after pump implantation. The resulting increase in MTg level (due to delayed clearance of MTg) is greater than that from TSH alone and suppresses further the animals' susceptibility to disease induction by MTg and adjuvant. Moreover, resistance following pretreatment with LPS and subtolerogenic MTg is mediated by CD4+ suppressor T cells, as shown recently for the suppression in mice given high doses of tolerogenic MTg. These experiments are in full agreement with the hypothesis and confirm that small increases in circulating MTg concentrations, which could occur physiologically, can be effective in protecting against EAT induction.