Treatment with a tissue-selective oestrogen complex does not affect disease pathology but reduces pre-BI cells in lupus-prone mice.

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.

Roles of transactivating functions 1 and 2 of estrogen receptor-alpha in bone.

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.

The role of estrogen receptor-α and its activation function-1 for growth plate closure in female mice.

High estradiol levels in late puberty induce growth plate closure and thereby cessation of growth in humans. In mice, the growth plates do not fuse after sexual maturation, but old mice display reduced longitudinal bone growth and high-dose estradiol treatment induces growth plate closure. Estrogen receptor (ER)-α stimulates gene transcription via two activation functions (AFs), AF-1 and AF-2. To evaluate the role of ERα and its AF-1 for age-dependent reduction in longitudinal bone growth and growth plate closure, female mice with inactivation of ERα (ERα(-/-)) or ERαAF-1 (ERαAF-1(0)) were evaluated. Old (16- to 19-mo-old) female ERα(-/-) mice showed continued substantial longitudinal bone growth, resulting in longer bones (tibia: +8.3%, P < 0.01) associated with increased growth plate height (+18%, P < 0.05) compared with wild-type (WT) mice. In contrast, the longitudinal bone growth ceased in old ERαAF-1(0) mice (tibia: -4.9%, P < 0.01). Importantly, the proximal tibial growth plates were closed in all old ERαAF-1(0) mice while they were open in all WT mice. Growth plate closure was associated with a significantly altered balance between chondrocyte proliferation and apoptosis in the growth plate. In conclusion, old female ERα(-/-) mice display a prolonged and enhanced longitudinal bone growth associated with increased growth plate height, resembling the growth phenotype of patients with inactivating mutations in ERα or aromatase. In contrast, ERαAF-1 deletion results in a hyperactive ERα, altering the chondrocyte proliferation/apoptosis balance, leading to growth plate closure. This suggests that growth plate closure is induced by functions of ERα that do not require AF-1 and that ERαAF-1 opposes growth plate closure.

Effects of oestradiol and raloxifene on the induction and effector phases of experimental postmenopausal arthritis and secondary osteoporosis.

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.

Interleukin 17A: a Janus-faced regulator of osteoporosis.

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.