地塞米松治疗小鼠实验性自身免疫性甲状腺炎的研究

本文采用细胞培养和免疫学技术研究地塞米松对自身免疫性甲状腺炎小鼠T细胞亚群、细胞因子、甲状腺抗体滴度和甲状腺组织病理形态的影响,结果显示:(1)地塞米松能明显降低EAT小鼠TGAb、TNF和IL-1水平;(2)提高Lyt-2阳性T细胞数,降低L_3T_4/Lyt-2比值;(3)明显逆转自身免疫性甲状腺炎小鼠的病理改变。结果提示地塞米松具有调整T淋巴细胞亚群,抑制细胞因子释放,抑制甲状腺过强的自身免疫反应等作用;局部注射地塞米松治疗桥本氏甲状腺炎是一种简单、有效的好方法。     

实验性自身免疫性甲状腺炎小鼠细胞免疫状态研究

作者检测了实验性自身免疫性甲状腺炎(EAT)小鼠的细胞免疫状态,结果表明:EAT小鼠脾细胞Thy-1.2和Lyt-2阳性T细胞数显著下降并伴随L3T4/Lyt-2阳性T细胞数比值升高;甲状腺球蛋白刺激的淋巴细胞增殖明显增强,但NK细胞活性无显著变化;脾细胞产生TNF和IL-1的水平明显高于正常小鼠。提示T细胞免疫功能紊乱和细胞因子的大量释放可能是引起此病的重要因素。     

Newer Insights into the Pathogenesis of Experimental Autoimmune Thyroiditis

Experimental autoimmune thyroiditis (EAT), produced in the mouse by immunization with murine thyroglobulin plus complete Freund's adjuvant, represents a valuable model for studying the pathogenesis of human chronic (Hashimoto's) thyroiditis. A major issue requiring clarification is the difference between benign autoimmunity, characterized solely by production of autoantibodies to thyroglobulin, and pathogenic autoimmunity where injury occurs to the thyroid cells. In this article, we describe the role of two key cytokines, IL12 and IFNγ, in modifying the pathogenic immune response. EAT, defined by cellular infiltration of the thyroid and the development of thyroglobulin-specific autoantibodies, is a dynamic process. Consequently, a cytokine may exert a different effect at different times during the disease process. For purposes of discussion, we propose that there are three stages in the development of EAT: priming; initiation; and progression. Administration of anti-IL12 during the priming stage and initiation dramatically decreases disease and lowers autoantibody levels. In contrast, injection of recombinant IL12 after disease was established significantly decreases the severity of disease and reduces autoantibody levels. Unlike IL-12, IFNγ was not essential for the priming of EAT. However, the severity of disease in the anti-IFNγ-treated initiation-and progression-treated animals was higher than in controls, implying a regulatory role for IFNγ. These findings emphasize that EAT involves a complex array of pathogenic mechanisms. The balance of cytokines produced during the early phase of the autoimmune reaction probably determines the progression from a harmless autoimmune response to autoimmune disease.     

Role of Late Complement Components in Experimental Autoimmune Thyroiditis

Availability of a line of rabbits deficient in the sixth complement component (C6-D) made it possible to evaluate the role of the terminal complement complex (TCC) in the development of experimental autoimmune thyroiditis (EAT) of the rabbit. Immunization with saline extract of homologous thyroid, known to be composed predominantly of thyroglobulin, led in normocomplementemic (NC) rabbits to severe thyroidiris, with cellular infiltrates occupying 50-95% of the thyroid, and to minimal or moderate thyroidiris, with 1-35% of thyroids infiltrated in C6-D rabbits. Cellular infiltrates consisted predominantly of mononuclear cells with appreciable numbers of granulocytes. Destruction of thyroid follicles was extensive and diffuse in NC rabbits, but it was only minimal and focal in C6-D rabbits. Immunohistology revealed in both groups of rabbits deposits of IgG and C3 along follicular basal laminae. In addition, NC rabbits showed deposition of C6 and MAC in thyroid follicles. These results suggested that TCC is necessary for the development of fully expressed, severe EAT; simultaneously, however, they showed that a significantly reduced EAT can develop without TCC. Administration of NC but not of C6-D rabbit serum to C6-D rabbits resulted in a significant increase in the severity of EAT. It was also shown that C6-D rabbits have "normal" T-cell activity, since they developed experimental autoallergic encephalomyelitis as readily as NC rabbits. Therefore, it is likely that development of EAT is indeed impaired by the C6 deficiency in rabbits. The requirement for TCC observed in this study may be relevant to the understanding of the pathogenesis of Hashimoto's thyroiditis, in which thyroid tissue was recently shown to contain TCC deposits.     

Effects of Anti-I-A and Anti-I-E Monoclonal Antibodies on the Induction and Expression of Experimental Autoimmune Thyroiditis in Mice

Susceptibility to experimental autoimrnune thyroiditis (EAT) in mice is linked to the I-A subregion of the major histocompatibility complex (MHC). The present study was undertaken to assess the effectiveness of anti-I-A^k monoclonal antibody (MAb) 10–2.16 in preventing or arresting the development of EAT. Spleen cells from CBA/J or (CBA/J x Balb/c) Fl mice given 10–2.16 prior to sensitization with mouse thyro-globulin (MTg) and adjuvant could not transfer EAT to normal recipients, and cells from these mice did not proliferate in vitro to MTg. Donor CBA/J mice given 10–2.16 before immunization and recipients of cells from such mice produced little MTg-specific lgGl or IgG2b antibody but did produce nearly as much IgG2a as controls. The effects of in vivo treatment with 10–2. 16 appear to be due to elimination of Ia + cells rather than to modulation of Ia or induction of suppressor T cells. When 10–2.16 was added to in vitro cultures it also prevented the proliferation and activation of sensitized CBA/J or Fl effector cell precursors. Other mAb specific for MHC class II gene products, but not associated with disease susceptibility, expressed by CBA/J (I-E^k) or FI (I-A^d) mice (14-4-4S or MK-D6 respectively), also prevented in vivo sensitization, but did not block in vitro activation. Anti-I-A^k was also effective in preventing EAT if multiple injections of mAb were given to recipients of sensitized EAT effector cells. These studies indicate that Ia + cells are required for initial sensitization and activation of EAT effector cells and also contribute to the development of severe histopathologic lesions in the thyroid during the final effector stage of EAT.     

Adoptive Transfer Murine Model of Granulomatous Experimental Autoimmune Thyroiditis

Experimental autoimmune thyroiditis (EAT) is a chronic inflammatory autoimmune disease that can be induced in genetically susceptible animals by immunization with mouse thyroglobulin (MTg) in an appropriate adjuvant or by the adoptive transfer of MTg-sensitized donor spleen cells, activated in vitro with MTg, into naive recipients. In the adoptive transfer model used in our laboratory, donor cells activated with MTg alone induce a relatively mild chronic lymphocytic form of EAT (L-EAT), in which the thyroid infiltrate consists primarily of mononuclear cells, and the thyroid inflammation persists for several months. When the same donor cells are activated with MTg together with anti-IL-2R and/or IL-12, a more severe and histologically distinct granulomatous form of EAT is induced in recipient mice. In addition to having distinct histopathologic features, granulomatous EAT (G-EAT) differs from L-EAT in that granulomatous thyroid lesions are not chronic. After reaching maximal severity 21 days after cell transfer, G-EAT thyroid lesions either resolve or the thyroids become atrophic and fibrotic by day 35. In this review, the histopathologic features of G-EAT and L-EAT are described, and our studies with the adoptive transfer G-EAT model which have focused on the mechanisms involved in induction of G-EAT in mice, and the evolution of G-EAT lesions to resolution of inflammation or fibrosis, are reviewed.     

Transfer of Experimental Autoimmune Thyroiditis by in Situ Perfusion of Thyroids with Immune Sera

The role of autoantibodies in the pathogenesis of autoimmune thyroiditis (AT) still remains poorly understood. Serum transfer experiments in experimental AT (EAT) and spontaneous AT (SAT) animal models frequently did not succeed or only resulted in minor thyroid lesions. The inconsistent results may have arisen because of technical problems inherent in serum transfer, the major of which is to obtain high enough concentrations of autoantibodies over long enough periods at the potential site of tissue injury. An attempt was made to circumvent this hurdle by repeated in situ perfusions of the rabbit thyroid via the superior thyroid artery. In situ perfusion of rabbit thyroids with high-titered homologous sera reactive with saline thyroid extract indeed led to thyroiditis characterized by granular deposits of rabbit IgG and C3 in the thyroid follicular basal laminae, cellular infiltrates consisting of mononuclear cells and granulocytes, destruction of the thyroid follicular architecture, and focal fibrosis. Perfusion with control sera lacking thyroid-reactive antibodies did not lead to thyroid lesions. These results demonstrate that humoral antibodies can induce severe AT comparable to actively induced thyroiditis.     

Humoral Autoimmune Manifestation in Subacute Thyroiditis

To study the autoimmune manifestations in subacute thyroiditis (SAT), the patterns of thyroid antibodies, thyroglobulin and circulating immune complexes were investigated in 10 patients during the course of the disease. Eight patients were thyrotoxic at diagnosis, and became euthyroid during recovery with a median observation of 8 months (4-30 months). Thyroid stimulating immunoglobulins were measured as TSH binding inhibiting immunoglobulins (TBII) and as thyroid stimulating antibodies (TSAb). TBII were present in all patients at least once during the observation period and remained detectable in six patients after recovery. TSAb were detected in three patients without relation to the hyperthyroid state. Thyroglobulin antibodies (TgAb) were present in four patients and persisted in three, while microsomal antibodies (MAb) were negative. Thyroglobulin (Tg) in the TgAb negative patients (n = 6) was high at diagnosis (median 229 micrograms/l, range 55-375) and fell rapidly during the course of SAT. Circulating immune complexes (CIC), which were found in all patients, reached maximal levels shortly after the onset of the disease and persisted after recovery. No correlation could be demonstrated between the different thyroid antibodies, and there was no clear relation between the levels of CIC and presence of the autoantibodies. However, the changes in CIC paralleled the changes in TBII, and it is suggested that immune complex formation is a major feature of the regulatory mechanisms controlling the immune responses in SAT.     

T Cell Mapping of One Epitope from Thyroglobulin Inducing Experimental Autoimmune Thyroiditis (EAT)

These data collect the advance made in the last few years in our laboratory in defining one epitope from the thyroglobulin (Tg) molecule (660 KDa) inducing Experimental Autoimmune thyroiditis (EAT) in CBA/J mice.

We achieved the characterization of one EAT-inducer Tg peptide by combining “in vitro” biochemical and immunological approaches and “in vivo” studies. Since T cells recognize degraded forms of the antigen and since endogenous antigens preferentially activate class I-restricted T cells, we hypothesized that one cytotoxic T cell hybridoma, named HTC2, which prevents further EAT induction in mice injected with Tg would be specific for one EAT inducer peptide.

In order to identify one Tg epitope inducing EAT, enzymatic treatment of the protein by trypsin, HPLC purification and sequence analysis were performed. Simultaneously, tryptic digests were used to pulse CBAJ macrophages and tested for their ability to be recognized by HTC2 cells. Lastly, when digests were recognized by HTC2 cells their capacity to induce EAT in CBA/J mice was evaluated.

To further assess the pathogenicity of the sequenced Tg peptide, one synthetic peptide was made and its capacity to induce EAT verified. By this procedure we identified for the first time one 40 amino-acid peptide from human thyroglobulin inducing EAT in CBA/J mice.     

An Experimental Model of Autoimmune Hemolytic Anemia in Mice

Autoimmune hemolytic anemia was induced in C57Bl/6J mice by multiple injections of syngeneic erythrocytes coated with Influenza A (PR8) virus. Coombs positive erythrocytes and erythrocyte-binding immunoglobulins were demonstrated by the method of antiglobulin agglutination. Viable virus was not required to induce the response, although it was more effective than inactivated preparations, and presensitization to the virus was not a prerequisite. The induced disease was mildly hemolytic. Reactive anti-erythrocyte immunoglobulins were of the IgG class, and unlike those in sera of New Zealand strain mice, they were both strain- and species-specific. These results support the concept of a viral etiology for acquired hemolytic anemia. However, the response seen in the experimental model differs from the natural response in New Zealand mice in the persistence of the anemia and the specificity of the autoantibodies. The Gross leukemia virus, being indigenous to the New Zealand strain, may persist and induce inheritable changes in host cells, thus accounting for the differences observed.     

An Experimental Model of Autoimmune Hemolytic Anemia in Mice

Autoimmune hemolytic anemia was induced in C57Bl/6J mice by multiple injections of syngeneic erythrocytes coated with Influenza A (PR8) virus. Coombs positive erythrocytes and erythrocyte-binding immunoglobulins were demonstrated by the method of antiglobulin agglutination. Viable virus was not required to induce the response, although it was more effective than inactivated preparations, and presensitization to the virus was not a prerequisite. The induced disease was mildly hemolytic. Reactive anti-erythrocyte immunoglobulins were of the IgG class, and unlike those in sera of New Zealand strain mice, they were both strain- and species-specific. These results support the concept of a viral etiology for acquired hemolytic anemia. However, the response seen in the experimental model differs from the natural response in New Zealand mice in the persistence of the anemia and the specificity of the autoantibodies. The Gross leukemia virus, being indigenous to the New Zealand strain, may persist and induce inheritable changes in host cells, thus accounting for the differences observed.     

Induction of a TC1-Mediated Experimental Autoimmune Thyroiditis by Deglycosylated Porcine Thyroglobulin

Contribution of cytotoxic T lymphocytes (CTL) to experimental autoimmune thyroiditis (EAT) was well defined (Speidel et al., Eur. J. Immunol. 1997, 27, 2391-2399, Ref. 7). The native porcine thyroglobulin (pTg) showed high sensitivity to endo-β-N-acetylglucosaminidase F (Endo F) and its molecular weights, corresponding to about 330 kDa as a monomer and 660 kDa as a dimer, were reduced to smaller molecular weight forms by Endo F and trifluoromethanesulfonic acid (TMSF). Deglycosylated porcine Tg (dgpTg) and native pTg were injected i.v. into CBA/J mice, without the aid of adjuvants. Both lymphocytic infiltrations of the thyroid glands and levels of Tg-specific CTL were similar to those found in conventional EAT induced by Tg and adjuvants. In contrast, proliferative responses in native pTg and dgpTg-injected mice could not be detected, and titers of antibodies to pTg and dgpTg were 20 times and 30 times lower than that of pTg and adjuvants, respectively. The EAT-inducer CTL belonged to the CD8⁺ cell subset and exerted their thyroiditogenic potential through release of IFN-γ. It was concluded that dgpTg-induced EAT is mediated by type 1 cytotoxic T cells (Tcl). Also, results that EAT induction of the glycosylated pTg (gpTg) was much lower than that of dgpTg, suggested that the abberant and incomplete glycosylation of the thyroglobulin is responsible for the induction of autoimmune thyroiditis.     

The Role of Iodine in the Development of Autoimmune Thyroiditis

Plasmid pLS408 includes gene,/?;'C(d) specifying Salmonella flagellin of antigenic type d with an in vitro deletion of a 48 base-pair EcoRV fragment in its central hypervariable antigenically-determinant region IV Oligonucleotides specifying peptide epitopes of antigens of unrelated pathogens inserted, in correct orientation, at the unique EcoRV site of pLS408 specify chimeric flagellins and, in many instances, cause production of functional flagella when the plasmid is placed in a flagellin-deficient AaroA live-vaccine strain of Salmonella dublin. The foreign epitope is then exposed at the surface of the flagellar filaments, as shown by the immobilizing effect of anti-epitope antibody and by immunogold electron-microscopy. The live-vaccine strain with a foreign epitope at the surface of its flagella when administered to mice by injection nearly always causes production of antibody with affinity for the foreign epitope and, sometimes, also for the source protein. Repeated injection of the live vaccine with an epitope of Streptococcus pyogenes type 5 M protein as insert caused production of opsonizing antibody and conferred partial protection against Streptococcus challenge. Injection of semi-purified chimeric flagella or flagellin, alone or with adjuvant, likewise causes antibody production, in one instance sufficient to give partial protection against influenza A virus challenge. Plasmid pLS408 with some inserts does not confer motility, either because the filaments produced are non-functional or because flagellin is made but not assembled or because little or no flagellin is produced. The features of a sequence which as insert determine production or non-production of functional flagella are not known. The effect of insertion of known T-cell epitopes and cellular immune responses to epitope inserts in flagellin are as yet little explored.     

A Thyroxine-containing Thyroglobulin Peptide Induces both Lymphocytic and Granulomatous Forms of Experimental Autoimmune Thyroiditis

Mouse thyroglobulin (MTg)-sensitized spleen cells activated in vitro with MTg can induce two histologically distinct forms of experimental autoimmune thyroiditis (EAT). MTg-sensitized cells activated with MTg alone induce a mild chronic form of EAT in which the thyroid infiltrate consists primarily of lymphocytes and other mononuclear cells (lymphocytic EAT). The same donor cells activated with MTg and anti-IL2R mAb induce a more severe and acute form of EAT with a thyroid inflammatory lesion having granulomatous histopathological features. A thyroxine-containing (T4) peptide, corresponding to positions 2549–2560 of human Tg, was shown by others to activate spleen cells of mouse thyroglobulin (MTg)-sensitized CBA/J mice to induce lymphocytic EAT. To determine if the CD4⁺effector T cells that induce granulomatous EAT can respond to the same T-cell epitope, the present study was undertaken to determine if both forms of EAT could be induced by the 2549–2560 thyroxine (T4)-containing peptide. This peptide was very effective for activation of T cells from MTg-primed CBA/J donors to induce granulomatous EAT but, in contrast to MTg, did not activate T cells from AKR/J or DBA/1 mice to induce granulomatous EAT. The T4 peptide did not apparently activate peptide-specific B cells in vivo but did activate MTg-primed B cells in vitro to produce anti-MTg autoantibody in recipient mice. These results demonstrate that a single 12-amino-acid thyroxine-containing peptide can activate T cells from CBA/J mice to induce both lymphocytic and granulo-matous EAT. However, this peptide does not activate T cells from some other EAT-susceptible strains of mice, suggesting that MTg contains multiple epitopes able to activate T cells to induce granulomatous EAT.     

CD3G Gene Defects in Familial Autoimmune Thyroiditis

The patients with CD3γ deficiency can present with different clinical findings despite having the same homozygous mutation. We report three new CD3gamma‐deficient siblings from a consanguineous family with a combined T?B+NK+ immunodeficiency and their variable clinical and cellular phenotypes despite the same homozygous mutation of the CD3G gene (c.80‐1G>C). We also re‐evaluate a previously reported non‐consanguineous family with two CD3gamma‐deficient siblings with the same mutation. The median age at diagnosis was 11 years (14 months–20 years). We found all five patients to display autoimmunity: autoimmune thyroiditis (n = 5), autoimmune haemolytic anaemia (n = 2), immune thrombocytopenia (n = 1), autoimmune hepatitis (n = 1), minimal change nephrotic syndrome (n = 1), vitiligo (n = 1) and positive antinuclear antibodies (n = 3) as well as high IgE (n = 2) and atopic eczema (n = 2). While CD3⁺TCRαβ+T cell percentages were low in all patients, only one had lymphopenia and 3 had CD3⁺T cell lymphopenia. Strikingly, we report frequent and multiple autoimmunity in tested heterozygous carriers in both families (n = 6; in 67%), and frequent autoimmunity in family members not available for testing (n = 5, in 80%). The results suggest that CD3G should be studied as a candidate gene for autoimmunity and that CD3gamma deficiency should be considered among other primary immunodeficiencies with predominantly autoimmune manifestations.     

Low Serum Vitamin D Is Associated with Anti-Thyroid Peroxidase Antibody in Autoimmune Thyroiditis

Purpose

The association between autoimmune thyroid diseases (AITDs) and vitamin D deficiency is controversial. We aimed to evaluate the relationship between serum 25-hydroxy-vitamin D₃ [25(OH)D₃] and anti-thyroid antibody levels.

Materials and Methods

25(OH)D₃, anti-thyroid antibodies, and thyroid function measured in 304 patients who visited the endocrinology clinic were analyzed. The patients were subgrouped into the AITDs or non-AITDs category according to the presence or absence of anti-thyroid antibodies. The relationship between anti-thyroid peroxidase antibody (TPOAb) and 25(OH)D₃ was evaluated.

Results

The patients with elevated anti-thyroid antibodies had lower levels of serum 25(OH)D₃ than those who did not (12.6±5.5 ng/mL vs. 14.5±7.3 ng/mL, respectively, p<0.001). Importantly, after adjusting for age, sex, and body mass index, a negative correlation (r=-0.252, p<0.001) was recognized between 25(OH)D₃ and TPOAb levels in the AITDs group, but this correlation did not exist in the non-AITDs group (r=0.117, p=0.127). 25(OH)D₃ level was confirmed as an independent factor after adjusting for co-factors that may affect the presence of TPOAb in the AITDs group.

Conclusion

25(OH)D₃ level is an independent factor affecting the presence of TPOAb in AITDs. The causal effect of 25(OH)D₃ deficiency to AITDs is to be elucidated.     

Treatment with a Novel Chemokine-Binding Protein or Eosinophil Lineage-Ablation Protects Mice from Experimental Colitis

Eosinophils are multifunctional leukocytes implicated in numerous inflammatory diseases. The present study was conducted to clarify the precise role of eosinophils in the development of colitis by using eosinophil-depleted mice and a novel chemokine-binding protein that neutralizes CCL11 action. Colitis was induced by administration of dextran sodium sulfate (DSS) to wild-type and eosinophil-deficient ΔdblGATA-1 mice. Accumulation of eosinophils in the gut of mice given DSS paralleled worsening of clinical score and weight loss. In response to DSS, ΔdblGATA-1 mice showed virtual absence of eosinophil recruitment, amelioration of clinical score, weight loss, and tissue destruction, and no lethality. There was a decrease in CXCL1 and CCL3 production and decreased neutrophil influx in the intestine of ΔdblGATA-1 mice. Transfer of bone marrow cells from wild-type mice reconstituted disease manifestation in DSS-treated ΔdblGATA-1 mice, and levels of CCL11 were increased after DSS treatment and localized to inflammatory cells. Treatment with the chemokine-binding protein evasin-4 at a dose that prevented the function of CCL11 greatly ameliorated clinical score, weight loss, overall tissue destruction, and death rates. In conclusion, the influx of eosinophils is critical for the induction of colitis by DSS. Treatment with a novel chemokine-binding protein decreased eosinophil influx and greatly ameliorated colitis, suggesting that strategies that interfere with the recruitment of eosinophils may be useful as therapy for colitis.Inflammatory bowel diseases (IBD), which include ulcerative colitis (UC) and Crohn’s disease, are chronic, relapsing, and remitting inflammatory conditions of unknown origin that affect individuals of both sexes throughout life.^1,2,3 The etiology of IBD remains unclear but presumably involves down-regulation of immunomodulatory mechanism and an uncontrolled activation of proinflammatory mediators. UC is a condition that primarily affects the superficial layer of the colon mucosa and presents clinical manifestation of disease, which include weight loss, diarrhea accompanied by blood and/or mucus, fever, and shortening of the colon. The colon of patients with UC is characterized by an increase in inflammatory infiltrate, which is composed predominantly by neutrophils, lymphocytes, mast cells, and eosinophils.^1,2,3Studies of the gut mucosa have emphasized the involvement of neutrophil granulocytes in the pathogenesis of IBD, but little attention has been given to the role of eosinophils. Recently, there has been increasing interest in the understanding of the involvement of these cells in the pathogenesis of IBD.⁴ Eosinophils are present in the healthy lamina propria of the gastrointestinal tract, where they are believed to play an important role in host defense against helminth infection.^5,6 However, eosinophils are powerful proinflammatory cells possessing the capacity to initiate or potentiate inflammatory reactions through the release of a range of inflammatory cytokines (interleukin (IL)-2, IL-4, IL-5, and IL-10), and eosinophilic granular proteins, such as major basic protein and eosinophil peroxidase (EPO).^4,5,6 Clinical investigations of bowel biopsy specimens from UC patients have demonstrated a correlation between eosinophil numbers in the mucosa, the levels of theses proteins in perfusion fluid samples, and disease severity.^7,8,9,10 Furthermore, eosinophil-active cytokines and chemokines are expressed and could be relevant for the control of colonic eosinophilia in UC.^11,12,13 However, the precise role of eosinophils and their mediators in UC remains unclear.The present study was designed to investigate the role of eosinophils in the pathogenesis of UC by using a previously described model of UC induced by dextran sodium sulfate (DSS).¹⁴ We explored the potential contribution of eosinophils to colitis pathogenesis by using mice (ΔdblGATA) in which the eosinophil lineage had been completely ablated.¹⁵ ΔdblGATA mice contain an engineered deletion of a palindromic double-enhancer binding site for GATA proteins in the 5′ region to the 1E exon of the gene encoding GATA-1 and are reported to be devoid of eosinophils both at baseline and in response to cytokine challenge, without reported effects on other hematopoietic lineages.¹⁵ Finally, we also tested the effects of treatment evasin-4, a novel chemokine-binding protein cloned from tick saliva that binds to eosinophil-active chemokines CCL11 and CCL5.^16,17