Estrogen receptor (ER) ligands modulate hemopoiesis and immunity in the normal state, during autoimmunity, and after infection or trauma. Dendritic cells (DC) are critical for initiation of innate and adaptive immune responses. We demonstrate, using cytokine-driven culture models of DC differentiation, that 17-β-estradiol exerts opposing effects on differentiation mediated by GM-CSF and Flt3 ligand, the two cytokines that regulate DC differentiation in vivo. We also show that estradiol acts on the same highly purified Flt3+ myeloid progenitors (MP) to differentially regulate the DC differentiation in each model. In GM-CSF-supplemented cultures initiated from MP, physiological amounts of estradiol promoted differentiation of Langerhans-like DC. Conversely, in Flt3 ligand-supplemented cultures initiated from the same MP, estradiol inhibited cell survival in a dose-dependent manner, thereby decreasing the yield of plasmacytoid and conventional myeloid and lymphoid DC. Experiments with bone marrow cells from ER-deficient mice and the ER antagonist ICI182,780 showed that estradiol acted primarily via ERα to regulate DC differentiation. Thus, depending on the cytokine environment, pathways of ER signaling and cytokine receptor signaling can differentially interact in the same Flt3+ MP to regulate DC development. Because the Flt3 ligand-mediated differentiation pathway is important during homeostasis, and GM-CSF-mediated pathways are increased by inflammation, our data suggest that endogenous or pharmacological ER ligands may differentially affect DC development during homeostasis and disease, with consequent effects on DC-mediated immunity.

Fibromyalgia (FMS), a predominantly female (85%) syndrome, affects an estimated 2% of the US population with skeletal muscle ache, fatigue, headache, and sleep disorder. The pathogenesis of FMS is unknown and there is no laboratory test for diagnosis. In this study, plasma levels of 25 cytokines and chemokines in 92 female patients with FMS and 69 family members were measured compared to 77 controls. Trans-endothelial migration of normal leukocytes in response to FMS plasma and the cytokine profile of human myoblasts were analyzed. High levels of MCP-1 (P<0.001) and eotaxin (P<0.01) were found in patients and family members compared to controls. Patients (56/92) treated with the single agent guaifenesin (>3 months) had higher levels of eotaxin than those not treated (P<0.01). Diluted plasma from patients increased the migration of normal eosinophils and monocytes, but not neutrophils, through an endothelial/Matrigel barrier only when mast cells are included in the lower wells (P<0.05). Furthermore, myoblasts can secrete MCP-1, eotaxin, and IP-10, while treatment with MCP-1 caused secretion of IL-1beta, eotaxin and IP-10. FMS is associated with inflammatory chemokines, that MCP-1 and eotaxin may contribute to the symptoms of FMS, and that similar cytokine profiles found in family members support the idea that FMS has a genetic component. Furthermore, the chemokine profile associated with FMS has direct effects on the migration of eosinophils and monocytes in the presence of mast cells, and skeletal muscle itself may secrete.

The steroid hormone estrogen regulates the differentiation, survival, or function of diverse immune cells. Previously, we found that physiological amounts of 17β-estradiol act via estrogen receptors (ER) to promote the GM-CSF-mediated differentiation of dendritic cells (DC) from murine bone marrow progenitors in ex vivo cultures. Of the two major subsets of CD11c+ DC that develop in these cultures, estrogen is preferentially required for the differentiation of a CD11bintLy6C− population, although it also promotes increased numbers of a CD11bhighLy6C+ population. Although both DC subsets express ERα, only the CD11bhighLy6C+ DC express ERβ, perhaps providing a foundation for the differential regulation of these two DC types by estrogen. The two DC populations exhibit distinct phenotypes in terms of capacity for costimulatory molecule and MHC expression, and Ag internalization, which predict functional differences. The CD11bintLy6C− population shows the greatest increase in MHC and CD86 expression after LPS activation. Most notably, the estrogen-dependent CD11bintLy6C− DC express langerin (CD207) and contain Birbeck granules characteristic of Langerhans cells. These data show that estrogen promotes a DC population with the unique features of epidermal Langerhans cells and suggest that differentiation of Langerhans cells in vivo will be dependent upon local estrogen levels and ER-mediated signaling events in skin.

Most immune cells, including myeloid progenitors and terminally differentiated dendritic cells (DC), express estrogen receptors (ER) making these cells sensitive to estrogens. Our laboratory recently demonstrated that 17-β-estradiol (E2) promotes the GM-CSF-mediated development of CD11c+CD11bint DC from murine bone marrow precursors. We tested whether the therapeutic selective estrogen receptor modulators (SERM), raloxifene and tamoxifen, can perturb DC development and activation. SERM, used in treatment of breast cancer and osteoporosis, bind to ER and mediate tissue-specific agonistic or antagonistic effects. Raloxifene and tamoxifen inhibited the differentiation of estrogen-dependent DC from bone marrow precursors ex vivo in competition experiments with physiological levels of E2. DC differentiated in the presence of SERM were assessed for their capacity to internalize fluoresceinated Ags as well as respond to inflammatory stimuli by increasing surface expression of molecules important for APC function. Although SERM-exposed DC exhibited increased ability to internalize Ags, they were hyporesponsive to bacterial LPS: relative to control DC, they less efficiently up-regulated the expression of MHC class II, CD86, and to a lesser extent, CD80 and CD40. This phenotype indicates that these SERM act to maintain DC in an immature state by inhibiting DC responsiveness to inflammatory stimuli. Thus, raloxifene and tamoxifen impair E2-promoted DC differentiation and reduce the immunostimulatory capacity of DC. These observations suggest that SERM may depress immunity when given to healthy individuals for the prevention of osteoporosis and breast cancer and may interfere with immunotherapeutic strategies to improve antitumor immunity in breast cancer patients.

Design of dual antagonists for the chemokine receptors CCR2 and CCR5 will be greatly facilitated by knowledge of the structural differences of their binding sites. Thus, we computationally predicted the binding site of the dual CCR2/CCR5 antagonist N