Interleukin-6 and its considerable role in the pathogenesis of thyrotoxicosis-related disturbances of bone turnover in postmenopausal women

Background: Thyrotoxicosis is more frequent in postmenopausal women than in the general population, effectively accelerating bone turnover. Interleukin-6 has been shown to be involved in the pathogenesis of bone disorders. Thus, the aim of the present study was to assess the role of IL-6 and its soluble receptor in the pathogenesis of thyrotoxicosis-related disturbances of bone turnover in oestrogen-deficient women. Material and methods: The study was carried out in 40 subjects with toxic nodular goitre in three groups: Group 1 &#8212; 13 premenopausal females, mean age 36 &#177; 15 years (PremTx&#8594;PremEu); Group 2 &#8212; 12 postmenopausal females, mean age 66 &#177; 14 years (PostTx&#8594;PostEu); and Group 3 &#8212; 15 males, mean age 45 &#177; 21 years (MTx&#8594;MEu). Overt thyrotoxicosis and euthyreosis after treatment with thyrostatics were confirmed by thyrotropin, free thyroxine and free triiodothyronin concentrations. Serum levels of bone turnover markers: TRACP5b and osteocalcin as well as serum IL-6 and IL-6sR were determined using ELISA kits. Results: TRACP5b/osteocalcin quotient was significantly elevated in the PostTx females compared to the PremTx women (p < 0.02). There was a positive correlation between serum TRACP5b and osteocalcin in the studied patients (R = 0.45, p < 0.001). Levels of serum IL-6 values were significantly elevated in PostTx: 3.0 (2.14&#8211;6.40) and MTx: 2.24 (1.60&#8211;5.10), compared to PremTx females: 1.39 (0.96&#8211;2.14) (p < 0.01 and p < 0.05 respectively). There were significant positive correlations between IL-6 and IL-6sR concentrations (R = 0.22, p < 0.05) and between IL-6sR and TRACP5b serum levels (R = 0.23, p < 0.05). Conclusions: The results of our study suggest that interleukin-6 plays a considerable role in the pathogenesis of thyrotoxicosis-related disturbances of bone turnover in oestrogen-deficient women. (Pol J Endocrinol 2011; 62 (4): 299&#8211;302)Wstęp: Nadczynność tarczycy występuje częściej u kobiet po menopauzie w porównaniu z populacją ogólną, skutecznie przyspieszając obrót kostny. Wykazano, że interleukina 6 (IL-6) odgrywa istotną rolę w regulacji obrotu kostnego. Uwzględniając ten fakt, celem obecnej pracy była próba oceny roli IL-6 i jej rozpuszczalnego receptora w patogenezie zaburzeń obrotu kostnego w przebiegu tyreotoksykozy u kobiet po menopauzie. Materiał i metody: Badanie przeprowadzono u 40 osób z nadczynnym wolem guzkowym w 3 grupach: 1 &#8212; 13 kobiet przed menopauzą w wieku 36 &#177; 15 lat (PremTx&#198;PremEu), 2 &#8212; 12 kobiet po menopauzie w wieku 66 &#177; 14 lat (PostTx&#198;PostEu) i 3 &#8212; 15 mężczyzn w wieku 45 &#177; 21 lat (MTx&#198;MEu). Stan czynnościowy tarczycy potwierdzono oznaczeniem TSH, fT3 i fT4 w surowicy. Markery obrotu kostnego: TRACP5b i osteokalcyna oraz IL-6 i IL-6sR w surowicy, oznaczono zestawami ELISA. Wyniki: Iloraz TRACP5b/osteokalcyna był istotnie zwiększony u kobiet PostTx w porównaniu z grupą PremTx (p < 0,02). Stwierdzono dodatnią korelację między TRACP5b i osteokalcyną (R = 0,45, p < 0,001). Stężenie IL-6 było istotnie zwiększone w grupie PostTx: 3,0 (2,14&#8211;6,40) i MTx: 2,24 (1,60&#8211;5,10) w porównaniu z odnotowanym u kobiet z grupy PremTx: 1,39 (0,96&#8211;2,14) (odpowiednio: p < 0,01 i p < 0,05). Wykazano istotną dodatnią korelację pomiędzy IL-6 i IL-6sR (R = 0,22, p < 0,05) oraz pomiędzy IL-6sR i TRACP5b (R = 0,23, p < 0,05). Wnioski: Podsumowując, wyniki obecnej pracy wskazują, że IL-6 odgrywa ważną rolę w patogenezie zaburzeń obrotu kostnego w przebiegu tyreotoksykozy u kobiet po menopauzie. (Endokrynol Pol 2011; 62 (4): 299&#8211;302

Serum interleukin-16 and RANTES during treatment of Graves&prime; orbitopathy with corticosteroids and teleradiotherapy

Introduction: To assess the usefulness of circulating IL-16 and RANTES measurements as markers of Graves&prime; orbitopathy (GO) activity and to estimate the role of these cytokines in GO pathogenesis. Material and methods: 42 individuals were divided into four groups: Group 1 comprised 15 euthyroid patients with clinical symptoms of GO who underwent corticosteroid therapy consisting of intravenous infusions of methylprednisolone (MP) and teleradiotherapy (TR); Group 2 comprised ten patients with hyperthyroid GD (Gtx); Group 3 comprised ten patients with GD in euthyreosis (Geu); and Group 4 comprised seven healthy volunteers age- and sex-matched to Groups 1&#8211;3. Serum samples were collected 24 hours before the first dose of MP, 24 hours after the first dose of MP, before TR, and at the end of therapy. Serum IL-16 and RANTES were determined by ELISA and TSH-Rab by RIA. Results: Serum IL-16 levels in patients with GO were significantly elevated at the end of therapy: 346 pg/mL (257&#8211;538) compared to IL-16 values before treatment: 250 ng/mL (211&#8211;337) and to the control group. RANTES serum concentrations did not significantly differ between studied groups, and immunosuppressive treatment did not influence its level. A negative correlation between TSH-Rab and RANTES was found in all studied groups (R = &#8211;0.32, p < 0.01). Conclusions: Our data suggests that IL-16 may exert an immunoregulatory effect in Graves&prime; orbitopathy. Serum measurements of both IL-16 and RANTES may be clinically useful; however, establishing their place in the diagnostics and treatment monitoring of GO needs further research. (Pol J Endocrinol 2012; 63 (2): 92&#8211;96)Wstęp: Ocena przydatności oznaczania krążącej IL-16 i RANTES jako wskaźników aktywności GO oraz określenie roli tych cytokin w patogenezie GO. Materiał i metody: 42 osoby w 4 grupach: 1 &#8212; 15 pacjentów z klinicznymi objawami orbitopatii w eutyreozie (GO), którzy poddali się leczeniu kortykosteroidami przy zastosowaniu podawanego dożylnie metylprednizolonu (MP) i teleradioterapii (TR); 2 &#8212; 10 pacjentów z chorobą Gravesa w nadczynności (Gtx); 3 &#8212; 10 pacjentów z chorobą Gravesa w eutyreozie (Geu); 4 &#8212; 7 zdrowych ochotników dobranych pod względem płci i wieku do grup 1.&#8211;3. Próbki krwi pobrano 24 h przed MP, 24 h po 1. dawce MP, przed TR i po zakończeniu leczenia. Stężenia IL-16 i RANTES w surowicy oznaczono metodą ELISA, a TSH-Rab &#8212; metodą RIA. Wyniki: Stężenie IL-16 w surowicy u pacjentów z GO było istotnie wyższe po zakończeniu terapii &#8212; 346 pg/ml (257&#8211;538) w porównaniu z wartością IL-16 przed leczeniem &#8212; 250 ng/ml (211&#8211;337) i w odniesieniu do grupy kontrolnej. Stężenie RANTES w surowicy nie różniło się istotnie między badanymi grupami i leczenie immunosupresyjne nie wpłynęło na jej wartość. Wykazano ujemną korelację między TSH-Rab i RANTES we wszystkich badanych grupach (R = &#8211;0,32; p < 0,01). Wnioski: Uzyskane dane wskazują, że IL-16 może wywierać immunomodulujący wpływ na przebieg orbitopatii. Zarówno oznaczenia IL-16, jak i RANTES w surowicy mogą być przydatne klinicznie, jednak ustalenie ich miejsca w diagnostyce i monitorowaniu leczenia GO wymaga dalszych badań. (Endokrynol Pol 2012; 63 (2): 92&#8211;96

Role of interleukin-6 on RANKL-RANK/osteoprotegerin system in hypothyroid ovariectomized mice.

Postmenopausal women frequently develop hypothyroidism. Estrogen depletion is accompanied by an increase of IL-6, accelerating bone turnover. The influence of hypothyroidism on bone metabolism in postmenopausal women is poorly understood. The aim of the study was an attempt to clarify the role of interleukin-6 on RANKL-RANK/osteoprotegerin system in hypothyroid ovariectomized mice. The study was performed on 56, 12-13 weeks old, female mice: C57BL/6J (wild-type; WT) and C57BL/6JIL6-/-Kopf (IL-6 knock-out; IL6KO). The mice were randomly divided into 8 groups with 7 mice in each one: 1/ WT controls, 2/ IL6KO controls, 3/ WT hypothyroid mice, 4/ IL6KO hypothyroid mice, 5/ WT ovariectomized, 6/ IL6KO ovariectomized, 7/ WT ovariectomized hypothyroid mice and 8/ IL6KO ovariectomized hypothyroid mice. Experimental model of menopause was produced by bilateral ovariectomy carried out in 8-9 weeks old mice. Experimental model of hypothyroidism was induced by propylthiouracyl administration in driking water. The serum levels of TRACP 5b, osteocalcin, OPG and RANKL were determined by ELISA. Serum RANKL concentrations were elevated significantly in all groups of ovariectomized mice as compared to respective controls, but in a minor degree in IL6KO hypothyroid mice as compared to wild-type animals. Moreover sRANKL values were significantly lower in IL6KO as compared to WT controls and IL6KO PTU injected mice. Osteoprotegerin serum levels were decreased in all IL-6 deficient mice and in a highest degree in sham-operated hypothyroid mice. To sum up, the results of the present study suggest that estrogens deficit is a strong stimulus for RANKL-RANK/OPG pathway that breaks an inhibitory influence of hypothyroidism even in IL-6 deficient mice

Increased percentage of L-selectin+ and ICAM-1+ peripheral blood CD4+/CD8+ T cells in active Graves' ophthalmopathy.

The purpose of the study was to evaluate the percentage of CD4+/CD8+ peripheral T cells expressing CD62L+ and CD54+ in patients with Graves' disease and to assess if these estimations could be helpful as markers of active ophthalmopathy. The study was carried out in 25 patients with Graves' disease (GD) divided into 3 groups: 1/ 8 patients with active Graves' ophthalmopathy (GO) (CAS 3-6, GO complaints pound 1 year), 2/ 9 patients with hyperthyroid GD without symptoms of ophthalmopathy (GDtox) and 3/ 8 patients with euthyroid GD with no GO symptoms (GDeu). The control group consisted of 15 healthy volunteers age and sex matched to groups 1-3. The expression of lymphocyte adhesion molecules was evaluated by using three-color flow cytometry. In GO group the percentage of CD8+CD54+, CD8+CD62L+, CD4+CD54+ and CD4+CD62L+ T cells was significantly higher as compared to controls (

Interleukin-6 is not essential for bone turnover in hypothyroid mice.

Interleukin-6 (IL-6) has been shown to be involved in the pathogenesis of several bone diseases characterized by an imbalance between bone resorption and formation. The aim of the study was to estimate serum markers of bone turnover: osteoclast-derived tartrate-resistant acid phosphatase form 5a (TRACP 5b) and osteocalcin in IL-6-deficient mice to assess the role of IL-6 in bone metabolism in hypothyroidism in mice. C57BL/6J (wild-type; WT) and C57BL/6J(IL6-/-Kopf) (IL-6 knock-out; IL6KO) mice randomly divided into 4 groups with 10 in each one: 1/ WT mice in hypothyroidism (WT-ht), 2/ WT controls, 3/ IL6KO mice with hypothyroidism (IL6KO-ht) and 4/ IL6KO controls. Experimental model of hypothyroidism was induced by intraperitoneal injection of propylthiouracyl. The serum levels of TRACP 5b and osteocalcin were determined by ELISA. Serum concentrations of TRACP 5b (median and interquartile ranges) were significantly decreased in both groups of mice with hypothyroidism: WT (3.2 (2.5-4.7) U/l) and IL6KO (2.6 (1.8-3.5) U/l) as compared to the respective controls. Similarly, serum osteocalcin levels were significantly reduced in both groups of mice in experimental hypothyroidism: WT (25.8 (23.0-28.2) ng/ml) and IL6KO (21.5(19.0-24.6) ng/ml) in comparison to the respective controls. There were no significant differences in bone turnover markers between IL6KO and WT mice both in hypothyroid and control animals. The results of the present study suggest that IL-6 does not play an important role in bone turnover in both euthyroid and hypothyroid mice

Increased percentage of L-selectin+ and ICAM-1+ peripheral blood CD4+/CD8+ T cells in active Graves' ophthalmopathy.

The purpose of the study was to evaluate the percentage of CD4+/CD8+ peripheral T cells expressing CD62L+ and CD54+ in patients with Graves' disease and to assess if these estimations could be helpful as markers of active ophthalmopathy. The study was carried out in 25 patients with Graves' disease (GD) divided into 3 groups: 1/ 8 patients with active Graves' ophthalmopathy (GO) (CAS 3-6, GO complaints pound 1 year), 2/ 9 patients with hyperthyroid GD without symptoms of ophthalmopathy (GDtox) and 3/ 8 patients with euthyroid GD with no GO symptoms (GDeu). The control group consisted of 15 healthy volunteers age and sex matched to groups 1-3. The expression of lymphocyte adhesion molecules was evaluated by using three-color flow cytometry. In GO group the percentage of CD8+CD54+, CD8+CD62L+, CD4+CD54+ and CD4+CD62L+ T cells was significantly higher as compared to controls (p&lt;0.001, p&lt;0.05, p&lt;0.01, p&lt;0.001 respectively). The percentage of CD8+CD54+ T lymphocytes was also elevated in GO group in comparison to hyperthyroid GD patients (p&lt; 0.05). CD4+CD62L+ and CD8+CD54+ percentages were also increased in GDtox and GDeu as compared to controls. We found a positive correlation between the TSHRab concentration and the percentage of CD8+CD62L+ T cells in all studied groups (r= 0.39, p&lt;0.05) and between the TSHRab level and CAS (r= 0.77, p&lt;0.05). The increased percentage of CD8+CD54+ and CD8+CD62L+ T cells in patients with Graves' ophthalmopathy may be used as a marker of immune inflammation activity

Markers of Inflammation and Fibrosis in the Orbital Fat/Connective Tissue of Patients with Graves’ Orbitopathy: Clinical Implications

Purpose. To assess FGF-β, TGF-β, and COX2 expression and immunocompetent cells in the orbital tissue of patients with severe and mild Graves’ orbitopathy. Patients and Methods. Orbital tissue was taken from 27 patients with GO: (1) severe GO (n=18), the mean clinical activity score (CAS) being 8.5 (SD 2.5); and (2) mild GO (n=9), the mean CAS being 2.2 (SD 0.8), and from 10 individuals undergoing blepharoplasty. The expression of CD4+, CD8+, CD20+, and CD68 and FGF-β, TGF-β, and COX2 in the orbital tissue was evaluated by immunohistochemical methods. Results. We demonstrated predominant CD4+ T cells in severe GO. CD68 expression was observed in the fibrous connective area of mild GO and was robust in severe GO, while the prominent TGF-β expression was seen in all GO. Increased FGF-β expression was observed in the fibroblasts and adipocytes of severe GO. No expression of COX2 was found in patients with GO. Conclusions. Macrophages and CD4 T lymphocytes are both engaged in the active/severe and long stage of inflammation in the orbital tissue. FGF-β and TGF-β expression may contribute to tissue remodeling, fibrosis, and perpetuation of inflammation in the orbital tissue of GO especially in severe GO

Markers of Inflammation and Fibrosis in the Orbital Fat/Connective Tissue of Patients with Graves’ Orbitopathy: Clinical Implications

Purpose. To assess FGF-β, TGF-β, and COX2 expression and immunocompetent cells in the orbital tissue of patients with severe and mild Graves’ orbitopathy. Patients and Methods. Orbital tissue was taken from 27 patients with GO: (1) severe GO (n=18), the mean clinical activity score (CAS) being 8.5 (SD 2.5); and (2) mild GO (n=9), the mean CAS being 2.2 (SD 0.8), and from 10 individuals undergoing blepharoplasty. The expression of CD4+, CD8+, CD20+, and CD68 and FGF-β, TGF-β, and COX2 in the orbital tissue was evaluated by immunohistochemical methods. Results. We demonstrated predominant CD4+ T cells in severe GO. CD68 expression was observed in the fibrous connective area of mild GO and was robust in severe GO, while the prominent TGF-β expression was seen in all GO. Increased FGF-β expression was observed in the fibroblasts and adipocytes of severe GO. No expression of COX2 was found in patients with GO. Conclusions. Macrophages and CD4 T lymphocytes are both engaged in the active/severe and long stage of inflammation in the orbital tissue. FGF-β and TGF-β expression may contribute to tissue remodeling, fibrosis, and perpetuation of inflammation in the orbital tissue of GO especially in severe GO

Search of reference biomarkers reflecting orbital tissue remodeling in the course of Graves’ orbitopathy

Introduction. Graves’ orbitopathy (GO) is a complication in Graves’ disease (GD) that causes disfigurement and sometimes blindness. The pathogenesis of GO remains unknown, while its symptoms demonstrate dependence between the thyroid gland and the orbit. The ongoing inflammatory process in retrobulbar tissue results in its remodeling characterized by increased volume of the orbital contents involving adipose tissue, with fibrosis and adipogenesis as predominant features. This study was aimed at the immunohistochemical verification of potential contribution and correlation between orbital expressions of IGF-1R, CD34, Foxp-3, PPAR-γ and CD4, CD68, TGF-β, FGF-β in severe and mild (long-lasting) GO. Material and methods. Forty-one orbital tissue specimens — 22 patients with severe GO, 9 patients with mild GO and 10 patients undergoing blepharoplasty as a control group — were processed by routine immunohistochemistry. Results. Increased IGF-1R, CD34 and Foxp-3 expression was found in both severe and mild GO, yet a significant correlation between CD34 and CD4, CD68, TGF-β, FGF-β expressions was observed in long-lasting GO. Conclusions. CD34 expression is proposed to be the marker of orbital tissue remodeling in the course of mild GO