RESUMO
OBJECTIVE: Systemic lupus erythematosus (SLE or lupus) is an autoimmune disease characterized by B-cell dysfunction, production of autoantibodies, and immune complex formation. Lupus is overrepresented in females, indicating that sex hormones play a role in the pathophysiology. Treatment with a tissue-selective oestrogen complex (TSEC) containing conjugated oestrogens and the selective oestrogen receptor modulator bazedoxifene (BZA) protects against postmenopausal vasomotor symptoms and osteoporosis, but its impact on organ damage in lupus is not fully understood. METHOD: We used ovariectomized MRL/lpr mice, treated with two different physiological doses of 17ß-oestradiol-3-benzoate (E2), BZA, or TSEC (E2 plus BZA), to assess early and late B-cell development and to determine histological disease manifestations in the kidneys and salivary glands. RESULTS: TSEC treatment reduced the frequency of the pre-BI population in bone marrow to levels equivalent to treatment with physiological doses of E2 alone but did not affect any of the other examined B-cell populations. Our earlier studies indicated that TSEC treatment did not aggravate disease development in ovariectomized MRL/lpr mice, while protecting against trabecular bone loss. Here, we follow up on our previous study and show that neither ovariectomy alone nor TSEC treatment of ovariectomized MRL/lpr mice influenced perivascular lymphocyte infiltration to the kidneys or salivary glands. CONCLUSION: TSEC does not aggravate a mouse model of lupus, when given in doses that protect against postmenopausal lupus-associated bone loss. This indicates that further investigations into TSEC as a treatment for osteoporosis or vasomotor symptoms in postmenopausal women with SLE are warranted.
Assuntos
Lúpus Eritematoso Sistêmico , Osteoporose , Feminino , Animais , Camundongos , Humanos , Camundongos Endogâmicos MRL lpr , Estrogênios/farmacologia , Lúpus Eritematoso Sistêmico/tratamento farmacológico , RimRESUMO
Interleukin (IL)-17A is a well-described mediator of bone resorption in inflammatory diseases, and postmenopausal osteoporosis is associated with increased serum levels of IL-17A. Ovariectomy (OVX) can be used as a model to study bone loss induced by estrogen deficiency and the role of IL-17A in osteoporosis development has previously been investigated using various methods to inhibit IL-17A signaling in this model. However, the studies show opposing results. While some publications reported IL-17A as a mediator of OVX-induced osteoporosis, others found a bone-protective role for IL-17 receptor signaling. In this study, we provide an explanation for the discrepancies in previous literature and show for the first time that loss of IL-17A has differential effects on OVX-induced osteoporosis; with IL-17A being important for cortical but not trabecular bone loss. Interestingly, the decrease in trabecular bone after OVX in IL-17A knock-out mice, was accompanied by increased adipogenesis depicted by elevated leptin levels. Additionally, the bone marrow adipose tissue expanded, and the bone-turnover decreased in ovariectomized mice lacking IL-17A compared to ovariectomized WT mice. Our results increase the understanding of how IL-17A signaling influences bone remodeling in the different bone compartments, which is of importance for the development of new treatments of post-menopausal osteoporosis.
Assuntos
Interleucina-17/fisiologia , Osteoporose/fisiopatologia , Absorciometria de Fóton , Animais , Osso Esponjoso/patologia , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Feminino , Fêmur/diagnóstico por imagem , Fêmur/patologia , Humanos , Camundongos , Camundongos Knockout , Osteogênese/efeitos dos fármacos , Osteoporose/diagnóstico por imagem , Osteoporose/etiologia , Osteoporose/patologia , Osteoporose Pós-Menopausa/etiologia , Osteoporose Pós-Menopausa/fisiopatologia , Ovariectomia/efeitos adversos , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Interleucina-17/fisiologia , Microtomografia por Raio-XRESUMO
BACKGROUND/OBJECTIVE: Postmenopausal women with systemic lupus erythematosus have an increased risk of osteoporosis and associated fractures. Their increased osteoporosis risk is probably caused by a high level of inflammation, use of glucocorticoids, impaired kidney function, and early menopause as these are known risk factors for osteoporosis. Due to these risk factors and the lack of safe and effective treatments, new therapies for the treatment of osteoporosis in this group of patients are needed. Ovariectomized MRL/lpr mice constitute a well-established model for studies of postmenopausal systemic lupus erythematosus; however, it is not clear to what extent this experimental model is associated with the development of osteoporosis. Thus, the aim of this study was to characterize the skeleton of ovariectomized MRL/lpr mice to determine the suitability of this model in studies of prospective new therapies for osteoporosis in postmenopausal systemic lupus erythematosus patients. METHODS: Skeletal parameters were measured in MRL/lpr mice and MRL/++ control mice, using peripheral quantitative computed tomography, high-resolution micro-computed tomography and biomechanical analyses. mRNA expression of bone-remodeling markers was measured by quantitative polymerase chain reaction and serological markers of lupus disease were evaluated using ELISA. RESULTS: Total bone mineral density was reduced in MRL/lpr mice compared with MRL/++ mice and MRL/lpr mice had reduced cortical and trabecular bone thickness compared with MRL/++ mice. In line with the low bone mass of MRL/lpr mice, gene expression analysis of cortical bone from these mice indicated an increased osteoclast activity as well as a decreased osteoblastogenesis and osteoblast activity, compared with MRL/++ mice. CONCLUSION: Ovariectomized MRL/lpr mice constitute a valuable experimental model for studies of osteoporosis development in postmenopausal systemic lupus erythematosus and this model is thus suitable for future studies of osteoporosis treatment in systemic lupus erythematosus.
Assuntos
Densidade Óssea , Osso e Ossos/fisiopatologia , Modelos Animais de Doenças , Osteoporose/fisiopatologia , Animais , Feminino , Humanos , Lúpus Eritematoso Sistêmico/complicações , Camundongos , Camundongos Endogâmicos MRL lpr , Osteoporose/etiologia , Pós-MenopausaRESUMO
Estrogen treatment has positive effects on the skeleton, and we have shown that estrogen receptor alpha (ERα) expression in cells of hematopoietic origin contributes to a normal estrogen treatment response in bone tissue. T lymphocytes are implicated in the estrogenic regulation of bone mass, but it is not known whether T lymphocytes are direct estrogen target cells. Therefore, the aim of this study was to determine the importance of ERα expression in T lymphocytes for the estrogenic regulation of the skeleton using female mice lacking ERα expression specifically in T lymphocytes (Lck-ERα-/-) and ERαflox/flox littermate (control) mice. Deletion of ERα expression in T lymphocytes did not affect bone mineral density (BMD) in sham-operated Lck-ERα-/- compared to control mice, and ovariectomy (ovx) resulted in a similar decrease in BMD in control and Lck-ERα-/- mice compared to sham-operated mice. Furthermore, estrogen treatment of ovx Lck-ERα-/- led to an increased BMD that was indistinguishable from the increase seen after estrogen treatment of ovx control mice. Detailed analysis of both the appendicular (femur) and axial (vertebrae) skeleton showed that both trabecular and cortical bone parameters responded to a similar extent regardless of the presence of ERα in T lymphocytes. In conclusion, ERα expression in T lymphocytes is dispensable for normal estrogenic regulation of bone mass in female mice.
Assuntos
Osso e Ossos/efeitos dos fármacos , Receptor alfa de Estrogênio/genética , Estrogênios/farmacologia , Linfócitos T/metabolismo , Animais , Densidade Óssea/efeitos dos fármacos , Densidade Óssea/genética , Osso e Ossos/metabolismo , Receptor alfa de Estrogênio/metabolismo , Feminino , Expressão Gênica , Inativação Gênica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Especificidade de Órgãos/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
Oestradiol and the selective oestrogen receptor modulator (SERM) raloxifene have been shown to ameliorate collagen-induced arthritis (CIA) in rats and in mice. One aim was to investigate if raloxifene exerts its anti-arthritic and anti-osteoporotic effects during the induction or effector phase of arthritis. A second aim was to analyse if raloxifene activates the oestrogen response element (ERE) to produce its immune-modulator effects. CIA or collagen-antibody-induced arthritis (CAIA) was induced in ovariectomized DBA/1-mice. CIA was used for evaluation of treatment during the induction, and CAIA for the effector phase of arthritis and osteoporosis development. Raloxifene, oestradiol or vehicle was administered 5 days/week. The clinical disease was evaluated continuously. Bone marrow density (BMD) was analysed with peripheral quantitative computer tomography, paws were collected for histological examination, and sera were analysed for markers of bone and cartilage turnover and proinflammatory cytokines. Transgenic luciferase (Luc)-ERE mice were immunized with collagen (CII), and after 10 days injected once with raloxifene, oestradiol or vehicle before termination. Spleens were analysed for luciferase activity to measure ERE activation. Treatment with oestradiol or raloxifene during the induction phase of CIA failed to affect arthritis. Raloxifene did not hamper disease activity in CAIA, whereas oestradiol delayed the onset and ameliorated the severity. Both raloxifene and oestradiol preserved BMD in CAIA. CII-immunization increased the oestradiol-induced ERE activation in spleen, and raloxifene activated the ERE at about 25% the intensity of oestradiol. Further experiments are needed to elucidate the exact mechanisms behind this finding.
Assuntos
Artrite Experimental/tratamento farmacológico , Estradiol/administração & dosagem , Osteoporose Pós-Menopausa/tratamento farmacológico , Cloridrato de Raloxifeno/administração & dosagem , Moduladores Seletivos de Receptor Estrogênico/administração & dosagem , Animais , Anticorpos/administração & dosagem , Artrite Experimental/induzido quimicamente , Artrite Experimental/complicações , Artrite Experimental/imunologia , Biomarcadores/sangue , Medula Óssea/patologia , Colágeno/administração & dosagem , Colágeno/imunologia , Progressão da Doença , Feminino , Humanos , Imunomodulação , Camundongos , Camundongos Endogâmicos DBA , Camundongos Transgênicos , Osteoporose Pós-Menopausa/complicações , Osteoporose Pós-Menopausa/etiologia , Osteoporose Pós-Menopausa/imunologia , Ovariectomia , Elementos de Resposta/genética , Transgenes/genéticaRESUMO
The bone-sparing effect of estrogen is primarily mediated via estrogen receptor-α (ERα), which stimulates target gene transcription through two activation functions (AFs), AF-1 in the N-terminal and AF-2 in the ligand binding domain. To evaluate the role of ERα AF-1 and ERα AF-2 for the effects of estrogen in bone in vivo, we analyzed mouse models lacking the entire ERα protein (ERα(-/-)), ERα AF-1 (ERαAF-1(0)), or ERα AF-2 (ERαAF-2(0)). Estradiol (E2) treatment increased the amount of both trabecular and cortical bone in ovariectomized (OVX) WT mice. Neither the trabecular nor the cortical bone responded to E2 treatment in OVX ERα(-/-) or OVX ERαAF-2(0) mice. OVX ERαAF-1(0) mice displayed a normal E2 response in cortical bone but no E2 response in trabecular bone. Although E2 treatment increased the uterine and liver weights and reduced the thymus weight in OVX WT mice, no effect was seen on these parameters in OVX ERα(-/-) or OVX ERαAF-2(0) mice. The effect of E2 in OVX ERαAF-1(0) mice was tissue-dependent, with no or weak E2 response on thymus and uterine weights but a normal response on liver weight. In conclusion, ERα AF-2 is required for the estrogenic effects on all parameters evaluated, whereas the role of ERα AF-1 is tissue-specific, with a crucial role in trabecular bone and uterus but not cortical bone. Selective ER modulators stimulating ERα with minimal activation of ERα AF-1 could retain beneficial actions in cortical bone, constituting 80% of the skeleton, while minimizing effects on reproductive organs.
Assuntos
Osso e Ossos/fisiologia , Receptor alfa de Estrogênio/fisiologia , Estrogênios/fisiologia , Animais , Densidade Óssea , Osso e Ossos/diagnóstico por imagem , Osso e Ossos/efeitos dos fármacos , Estradiol/farmacologia , Receptor alfa de Estrogênio/genética , Estrogênios/farmacologia , Feminino , Camundongos , Camundongos Mutantes , Tamanho do Órgão , Radiografia , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Timo/anatomia & histologia , Timo/efeitos dos fármacos , Timo/fisiologia , Ativação Transcricional , Útero/anatomia & histologia , Útero/efeitos dos fármacos , Útero/fisiologiaRESUMO
Oestrogen is not only a sex hormone but also an important regulator of the immune system. Expression of the heavy chain of IgM (mu) is essential for B-cell differentiation. However, a small number of IgA-positive B cells can be found in mice lacking the mu chain (muMT-/-). The aim of this study was to investigate the effects of oestrogen on this alternative B-cell pathway in muMT-/- mice. Our results clearly demonstrate that oestrogen increases the frequency of IgA-producing B cells in muMT-/- mice in both bone marrow and spleen cells. We also show that mature IgM-producing B cells are not required for oestrogen-mediated suppression of granulocyte-mediated inflammation or thymic involution. In conclusion, we demonstrate that 17beta-estradiol benzoate increases the frequency of IgA-producing B cells in muMT-/- mice, suggesting that oestrogen can influence the alternative B-cell pathway found in muMT-/- mice.
Assuntos
Subpopulações de Linfócitos B/imunologia , Subpopulações de Linfócitos B/metabolismo , Proliferação de Células , Estradiol/fisiologia , Imunoglobulina A/biossíntese , Cadeias mu de Imunoglobulina/genética , Animais , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos MutantesRESUMO
The isoflavone genistein (Gen) is a naturally occurring phytoestrogen found in high concentrations in soy. The biological effects of Gen have been extensively studied. The immunomodulating properties of Gen are, however, less well investigated and the results are contradictory. Our aim was to study possible estrogen agonistic and antagonistic properties of Gen in uterus, bone, lymphopoiesis and B-cell function by comparing effects in castrated and intact female mice, respectively. Oophorectomized (OVX) and sham-operated mice were treated with s.c. doses of 17beta-estradiol (E2) (0.16 mg/kg), Gen (50 mg/kg), or vehicle (olive oil) as control. Effects on bone mineral density (BMD) were studied using peripheral quantitative computerized tomography, uterine and thymus weights were examined, lymphopoiesis in thymus and bone marrow was analyzed using flow cytometry, and the frequency of immunoglobulin-producing B cells in bone marrow and spleen was studied using an ELISPOT assay. Gen was clearly antagonizing endogenous estrogen in sham-operated female mice as shown by inhibiting the uterine weight and by increasing the frequency of B lymphopoietic cells in bone marrow. The only agonistic effect of Gen was shown by increased BMD in OVX mice. Our results are discussed in the context of estrogen receptor biology.
Assuntos
Osso e Ossos/efeitos dos fármacos , Antagonistas de Estrogênios/farmacologia , Estrogênios/agonistas , Genisteína/farmacologia , Linfopoese/efeitos dos fármacos , Útero/efeitos dos fármacos , Animais , Linfócitos B/imunologia , Densidade Óssea , Medula Óssea/efeitos dos fármacos , Estradiol/agonistas , Feminino , Imunoglobulinas/metabolismo , Camundongos , Receptores de Estrogênio/agonistas , Receptores de Estrogênio/antagonistas & inibidores , Timo/efeitos dos fármacosRESUMO
Estrogen has extensive effects on the immune system. The aim of the present experiments was to compare the effects of 17beta-estradiol (E2) and 4-estren-3alpha,17beta-diol (estren) on T lymphopoiesis and T cell-dependent inflammation. In order to investigate the role of estrogen receptors (ER) in the effects of E2 and estren on the immune system, ER knock-out mice lacking both ERalpha and ERbeta (DERKO) were used. T lymphopoiesis and T cell-dependent inflammation were studied by investigating thymus cellularity, the delayed-type hypersensitivity (DTH) reaction, CD4(+) T cells in spleen and serum levels of interleukin (IL)-6. As expected, the presence of ERs was mandatory for all the effects of E2. In contrast, treatment with estren reduced thymus cellularity in ER knock-out mice, indicating an effect through ER-independent pathways. Interestingly, estren suppressed only DTH, the frequency of CD4(+) T cells in spleen and serum levels of IL-6 in wild-type (WT) mice, but not in mice lacking ERs. Thus, our study is the first to show that estren inhibits T lymphopoiesis via ER-independent pathways, whereas its suppressive effects on inflammation are ER-dependent.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Estrenos/farmacologia , Hipersensibilidade Tardia/imunologia , Linfopoese/efeitos dos fármacos , Receptores de Estrogênio/imunologia , Animais , Linfócitos T CD4-Positivos/efeitos dos fármacos , Depressão Química , Estradiol/sangue , Estradiol/imunologia , Feminino , Citometria de Fluxo , Hipersensibilidade Tardia/tratamento farmacológico , Interleucina-6/sangue , Interleucina-6/imunologia , Masculino , Camundongos , Camundongos KnockoutRESUMO
Oestrogen has a dichotomous effect on the immune system. T and B lymphopoiesis in thymus and bone marrow is suppressed, whereas antibody production is stimulated by oestrogen. In this study the importance of the oestrogen receptors (ER) ER-alpha and ER-beta in the aged immune system was investigated in 18 months old-wild type (WT), ER-alpha (ERKO), ER-beta (BERKO) and double ER-alpha and ER-beta (DERKO) knock-out mice, and compared with 4 months old WT mice. Cell phenotypes in bone marrow, spleen and thymus, and the frequency of immunoglobulin (Ig) spot forming cells (SFC) were determined. We show here that the 17-beta-oestradiol (E2)-induced downregulation of B lymphopoietic cells in bone marrow of young ovariectomized mice can be mediated through both ER-alpha and ER-beta. However, only ER-alpha is required for the age-related increased frequency of immunoglobulin M (IgM) SFC in the bone marrow, as well as for the increased production of interleukin-10 (IL-10) from cultured splenocytes in aged mice. Furthermore, increased age in WT mice resulted in lower levels of both pro- and pre-B cells but increased frequency of IgM SFC in the bone marrow, as well as increased frequency of both IgM and IgA SFC in the spleen. Results from this study provide valuable information regarding the specific functions of ER-alpha and ER-beta in the aged immune system.
Assuntos
Envelhecimento/imunologia , Receptores de Estrogênio/imunologia , Animais , Linfócitos B/imunologia , Células da Medula Óssea/imunologia , Células Cultivadas , Estradiol/sangue , Estradiol/farmacologia , Receptor alfa de Estrogênio , Receptor beta de Estrogênio , Feminino , Imunoglobulina A/biossíntese , Imunoglobulina M/biossíntese , Imunofenotipagem , Linfopoese/efeitos dos fármacos , Linfopoese/imunologia , Camundongos , Camundongos Knockout , Ovariectomia , Receptores de Estrogênio/metabolismo , Baço/imunologia , Linfócitos T/imunologiaRESUMO
Oestrogen treatment down-regulates B lymphopoiesis in the bone marrow of mice. Meanwhile it up-regulates immunoglobulin production. To understand better the oestrogen action on bone marrow male mice lacking oestrogen receptor alpha (ERalpha; ERKO mice), lacking ERbeta (BERKO mice), lacking both receptors (DERKO mice) or wild-type (wt) littermates were castrated and treated for 2.5 weeks with 30 microg/kg 17beta-oestradiol (E2) or vehicle oil as controls. The B lymphopoiesis in the bone marrow was examined by flow cytometry and mature B-cell function was studied using an ELISPOT assay enumerating the B cells in bone marrow and spleen that were actively producing immunoglobulins. In wt mice the frequency of B-lymphopoietic (B220+) cells in the bone marrow decreased from 15% to 5% upon E2 treatment. In ERKO and BERKO mice significant reduction was seen but not of the same magnitude. In DERKO mice no reduction of B lymphopoiesis was seen. In addition, our results show that E2 mediated reduction of different steps in B lymphopoiesis require only ERalpha or both receptors. In wt and BERKO mice E2 treatment resulted in significantly increased levels of B cells actively producing immunoglobulin, while in ERKO and DERKO mice no such change was seen. Similar results were found in both bone marrow and spleen. In conclusion our results clearly show that both ERalpha and ERbeta are required for complete down-regulation of B lymphopoiesis while only ERalpha is needed to up-regulate immunoglobulin production in both bone marrow and spleen.