Monica DelgadoBio 520

Winter 2009

www.nature.com/.../n6/fig_tab/nm0606-612_F1.html

Estrogens are a group of steroid compounds

Like all steroid hormones, estrogens readily diffuse across the cell membrane; inside the cell, where they interact with estrogen receptors (ERs)

Produced mainly by developing follicles in the ovaries

Follicle-stimulating hormone (FSH) and luteinizing hormone (LH) stimulate the production of estrogen in the ovaries

Present in both men and women

Usually present higher in women

Estrogen receptor (ER) are activated by 17-β-estradiol (estrogen)

Two types of ERs exist: ER (intracellular receptor) and the estrogen G protein coupled receptor (GPR30)

Barnard, A., et al Neuroimmunomodulation (2008)

Estrogens have a degenerative effect on the primary lymphoid organs, affecting the development, maturation and function of the immune system

Nearly 79% of autoimmune disease patients in the USA are women ERs are present on bone marrow (BM) stromal cells and resident

multipotential stem cells Today’s focus looks at the one of the primary target s of estrogen; the thymus

(which undergoes phenotypic alterations when exposed to estrogen) However, normal thymic development has also been shown to be dependent

on the estradiol/ERα signaling pathway Estrogen treatment has been shown to activate phagocytic activity and

enhance Fc-γR expression on splenic-macrophages Estradiol drastically reduces the lineage-negative Flt3+ Sca-1+ cKit+

population in the BM (a population that contains thymic homing progenators) Lin- Sca-1+ cKit+ (LSK) cells are the principal circulating progenitors with T

lineage potential

Barnard, A., et al Neuroimmunomodulation (2008)Kletcha, A,, et al Journal of Endocrinology (2006)

Barnard, A., et al Neuroimmunomodulation (2008)

In adults, T-cell precursors enter the thymus at the cortico-medullary junction

Uncommited progenators include CD4- CD8- double negative 1 (DN1) cells, in region 2 cells differentiate into the DN2 stage and undergo a proliferative clonal expansion (loss of NK cell potential)

T-cell lineage commitment and the onset of T-cell receptor β-chain rearrangement occurs in DN3 stage (region 3)

Transition from DN to CD4+ CD8+ DP occurs in region 4

DP cells then migrate back through the cortex and into the medulla

Nature Reviews Immunology 4, 278-289 (April 2004)

High levels of estrogens contribute to the development of autoimmunity as a result of degenerative effects on the primary lymphoid organs

Significant thymic atrophy began 48h after infection of estradiol

(A) Thymus cellularity declines over the period of 6 days where it reached a minimun of 19% (compared to control)

(B) Preferential reduction of DP thymocytes due to estrodiol

Age-matched pairs of 6-12wk old male C57BL/6 mice (strain used to show Th1 responses) were injected daily with vehicle only or 17-β-estradiol for 1-6 days

(A) While total BM cellularity was not affected, after 24h there was a greater than 60% reduction in the percentage of Flt3+ LSK cells in estradiol treated mice

(B) Further reduction seen at 48h with decrease in lineage-negative c-Kit+ cells, LSK cells, and Flt3+ LSK cells

Loss of cells in the BM is not likely to contribute significantly to the depletion of DP thymocytes

CD117 also calledC-kit is a cytokine receptor found on Hematopoietic stem cells

% of total bone marrow

% of Lin-, c-Kit cells

% of bone marrow LSK cells

Pregnancy constitutes a major challenge to the immune system

Although localized mechanisms contribute to fetal evasion from immune attack, maternal alloreactive lymphocytes persist

In addition to their function in suppressing autoimmune responses, maternal regulatory T cells suppress aggressive allogeneic response directed at the fetus

Positive effects of pregnancy (counter hypothesis)

(A) Regulatory T cells prevent rejection of the fetal allograft

In an allogeneic context, regulatory T cells are necessary for the prevention of a maternal immune response against the fetus

Allogeneic = sourced from a genetically non-identical member of the same speciesSyngeneic = Genetically identical or closely related, immunologically compatible

Filled circles = females fertilized by C57BL/6 males (Th1 response)Open circles= females fertilzed by BALB/c males (Th2 response)Filled boxes= non pregnant females

Exposure to pregnancy levels of E2 reduced thymic size and cellularity (A)

Thymic atrophy was attenuated in AERKO mice but not obliterated (B)

E2 induced thymic atrophy was fractionally seen in both ER-α and GPR30-but not in ERβ-deficient mice (B)

Suggest that ERα and GPR30, but not Erβ, played partial roles in E2-induced thymic atrophy

Note: Erα- and Erβ-knockout (AERKO and BERKO) mice

Critique: Column B thymic size irregularities were not addressed

Note: DERKO = mice with inactivated ERα and ERβ

The estrogenic response to thymic atrophy were absent or largely reduced in DERKO mice

WT – DERKO / WT X 100

100 – Dependent effect

E2 Treatment inhibits TNF-α producing inflammatory cells in CNS of mice with Experimental autoimmune encephalomyelitis (EAE)

Myelin oligodendrocyte glycoprotein (MOG)-35-55-stimulated CNS mononuclear cells from E2-treated C57BL/6J mice showed a large reduction in intracellular staining TNF-α

Critique: Increase in T cell was not really addressed

Estrogen treatment increases nitric oxide levels in the supernatants from Con-A-stimulated splenocytes when compared to that from placebo treated mice

Blocking with Con-A + CTLA-41g fusion protein leads to a decrease in nitric oxide levels

Previous studies by Karpuzoglu’s group suggest that estrogen-induced up-regulation of iNOS/nitric oxide in activated splenocytes is likely to be mediated through IFN-γ

Note:Con-A = T cell mitogenCTLA4 (Cytotoxic T-Lymphocyte Antigen 4)

Article Conclusions made Support Hypothesis?

Zoller and Kersh (2006) Estradiol drastically reduces the thymic homing progenators in the BM (thymic atrophy)

Y

Aluvihare, Kallikourdis, and Betz (2004)

Regulatory T Cells Mediate Maternal Tolerance to the Fetus

N (counter aspect)

Wang et al (2008) GPR30 Contributes to Estrogen-Induced Thymic Atrophy

Y

Lindberg, M. K., et al (2002) Estrogen effects such as thymic atrophy are mainly EFα mediated

Y

Offner, H., and Polanczyk, M. (2006)

E2 treatment inhibits TNF-α producing inflammatory cells in CNS of mice with EAT

N

Karpuzoglu, E., Ahmed, S. A. (2006)

Estrogens are not only involved in regulation of normal immune responses, but also are implicated in many autoimmune diseases

Y

Elevated estradiol induces loss of T lineage progenators in the BM (Zoller and Kersh 2006)

Regulatory T cells are required for the maternal immune system to tolerate the fetal allograft (Aluvihare, Kallikourdis, and Betz 2004)

Wang et al showed that E2-induced thymic atrophy involves both ERα and GPR30, but not ERβ

Karpuzoglu’s group showed that estrogen treatment in mice markedly upregulates the levels of iNOS mRNA through IFN-γ; Also estrogens are not only involved in regulation of normal immune responses, but also are implicated in many autoimmune diseases

The exact pathway of estrogen-induced thymic degeneration is yet to be elucidated/established

Further research should focus on the different and frequently opposite effects exerted by physiologic and pharmacologic doses of estrogens (dose-related effects)

Further studies must be directed to the inflammatory mediators (i.e., cytokines) that seem to alter the peripheral metabolism of sex hormones and complicate the effects of sex hormones on susceptibility to autoimmunity

There are no available data on cytokine measurements on non-immune hyperthyroidsm

Questions?