ABSTRACT
The ability of the thymus gland to convert bone marrow-derived progenitor cells into single positive (SP) T-cells is well known. In this review we present evidence that the thymus, in addition to producing SP T-cells, also has a pathway for the production of double negative (DN) T-cells. The existence of this pathway was noted during our examination of relevant literature to determine the cause of sex steroid-induced thymocyte loss. In conducting this search our objective was to answer the question of whether thymocyte loss is the end product of a typical interaction between the reproductive and immune systems, or evidence that the two systems are incompatible. We can now report that "thymocyte loss" is a normal process that occurs during the production of DN T-cells. The DN T-cell pathway is unique in that it is mediated by thymic mast cells, and becomes functional following puberty. Sex steroids initiate the development of the pathway by binding to an estrogen receptor alpha located in the outer membrane of the mast cells, causing their activation. This results in their uptake of extracellular calcium, and the production and subsequent release of histamine and serotonin. Lymphatic vessels, located in the subcapsular region of the thymus, respond to the two vasodilators by undergoing a substantial and preferential uptake of gamma/delta and alpha/beta DN T- cells. These T- cells exit the thymus via efferent lymphatic vessels and enter the lymphatic system.The DN pathway is responsible for the production of three subsets of gamma/delta DN T-cells and one subset of alpha/beta DN T-cells. In postpubertal animals approximately 35 % of total thymocytes exit the thymus as DN T-cells, regardless of sex. In pregnant females, their levels undergo a dramatic increase. Gamma/delta DN T-cells produce cytokines that are essential for the maintenance of pregnancy.
Subject(s)
T-Lymphocytes/physiology , Thymocytes/physiology , Thymus Gland/physiology , Animals , Cell Differentiation , Female , Humans , Pregnancy , T-Lymphocytes/cytology , Thymocytes/cytology , Thymus Gland/cytologyABSTRACT
BACKGROUND: Female mice and rats injected with estrogen perinatally become anovulatory and develop follicular cysts. The current consensus is that this adverse response to estrogen involves the hypothalamus and occurs because of an estrogen-induced alteration in the GnRH delivery system. Whether or not this is true has yet to be firmly established. The present study examined an alternate possibility in which anovulation and cyst development occurs through an estrogen-induced disruption in the immune system, achieved through the intermediation of the thymus gland. METHODS, RESULTS AND CONCLUSION: A putative role for the thymus in estrogen-induced anovulation and follicular cyst formation (a model of PCOS) was examined in female mice by removing the gland prior to estrogen injection. Whereas all intact, female mice injected with 20 microg estrogen at 5-7 days of age had ovaries with follicular cysts, no cysts were observed in animals in which thymectomy at 3 days of age preceded estrogen injection. In fact, after restoring immune function by thymocyte replacement, the majority of thymectomized, estrogen-injected mice had ovaries with corpora lutea. Thus, when estrogen is unable to act on the thymus, ovulation occurs and follicular cysts do not develop. This implicates the thymus in the cysts' genesis and discounts the role of the hypothalamus. Subsequent research established that the disease is transferable by lymphocyte infusion. Transfer took place between 100-day-old estrogen-injected and 15-day-old naïve mice only when recipients were thymectomized at 3 days of age. Thus, a prerequisite for cyst formation is the absence of regulatory T cells. Their absence in donor mice was judged to be the result of an estrogen-induced increase in the thymus' vascular permeability, causing de facto circumvention of the final stages of regulatory T cell development. The human thymus has a similar vulnerability to steroid action during the fetal stage. We propose that in utero exposure to excessive levels of steroids such as estrogen has a long-term effect on the ability of the thymus to produce regulatory T cells. In female offspring this can lead to PCOS.
Subject(s)
Anovulation , Estrogens/toxicity , Polycystic Ovary Syndrome , Age Factors , Animals , Animals, Newborn , Anovulation/chemically induced , Anovulation/etiology , Anovulation/immunology , Autoimmunity/drug effects , Autoimmunity/immunology , Disease Models, Animal , Female , Hydrocortisone/toxicity , Hypothalamus/drug effects , Hypothalamus/physiology , Injections, Subcutaneous , Mice , Mice, Inbred A , Mice, Inbred C57BL , Polycystic Ovary Syndrome/chemically induced , Polycystic Ovary Syndrome/etiology , Polycystic Ovary Syndrome/immunology , Testosterone/toxicity , Thymectomy , Thymus Gland/drug effects , Thymus Gland/immunology , Thymus Gland/surgeryABSTRACT
In mouse ovaries, the enzyme 3 beta-hydroxysteroid dehydrogenase (HSD) is distributed between microsomes and mitochondria. Throughout the follicular phase of the estrous cycle, the HSD activity in microsomes is predominant; whereas, after LH stimulation, HSD activity during the luteal phase is highest in the mitochondria. The current study examined whether or not LH stimulation always results in an increase in mitochondrial HSD activity. This was accomplished by measuring the HSD activity in microsomal and mitochondrial fractions from ovaries of pregnant mice. These animals have two peaks of LH during gestation, and one peak of LH after parturition. It was found that mitochondrial HSD activity was highest after each peak of LH. It is proposed that mitochondrial HSD is essential for the synthesis of high levels of progesterone. The increase in HSD activity in mitochondria after LH stimulation occurs because: 1) LH initiates the simultaneous synthesis of HSD and the cholesterol side-chain cleavage enzyme (P450scc); and, 2) HSD and P450scc bind together to form a complex, which becomes inserted into the inner membrane of the mitochondria. High levels of progesterone are synthesized by mitochondrial HSD because: 1) the requisite NAD+ cofactor for progesterone synthesis is provided directly by the mitochondria, rather than indirectly via the rate limiting malate-aspartate shuttle; and, 2) the end-product inhibition of P450scc by pregnenolone is eliminated because pregnenolone is converted to progesterone.
Subject(s)
3-Hydroxysteroid Dehydrogenases/physiology , Corpus Luteum/enzymology , Mitochondria/enzymology , Mitochondrial Proteins/physiology , Progesterone/biosynthesis , 3-Hydroxysteroid Dehydrogenases/metabolism , Animals , Electron Transport Complex IV/metabolism , Female , Luteinizing Hormone/metabolism , Mice , Microsomes/enzymology , Mitochondrial Proteins/metabolism , Models, Biological , PregnancyABSTRACT
Day 3 thymectomy (D3Tx) leads to a paucity of CD4(+)CD25(+) suppressor T cells, a loss of peripheral tolerance, and the development of organ-specific autoimmune disease in adult mice. Importantly, D3Tx does not lead to autoimmune disease in all mouse strains, indicating that this process is genetically controlled. Previously, we reported linkage of D3Tx-induced autoimmune ovarian dysgenesis (AOD) and its intermediate phenotypes, antiovarian autoantibody responsiveness, oophoritis, and atrophy, to five quantitative trait loci (QTL), designated Aod1 through Aod5. We also showed interaction between these QTL and H2 as well as Gasa2, a QTL controlling susceptibility to D3Tx-induced autoimmune gastritis. To physically map Aod1, interval-specific bidirectional recombinant congenic strains of mice were generated and studied for susceptibility to D3Tx-induced AOD. Congenic mapping studies revealed that Aod1 controls susceptibility to oophoritis and comprises two linked QTL with opposing allelic effects. Aod1a resides between D16Mit211 (23.3 cM) and D16Mit51 (66.75 cM) on chromosome 16. Aod1b maps proximal of Aod1a between D16Mit89 (20.9 cM) and D16Mit211 (23.3 cM) and includes the candidate genes stefin A1, A2, and A3 (Stfa1-Stfa3), inhibitors of cathepsin S, a cysteine protease required for autoantigen presentation, and the development of autoimmune disease of the salivary and lacrimal glands following D3Tx. cDNA sequencing revealed the existence of structural polymorphisms for both Stfa1 and Stfa2. Given the roles of cathepsins in Ag processing and presentation, Stfa1 and Stfa2 alleles have the potential to control susceptibility to autoimmune disease at the level of both CD4(+)CD25(+) suppressor and CD4(+)CD25(-) effector T cells.
Subject(s)
Alleles , Autoimmune Diseases/genetics , Autoimmune Diseases/immunology , Genetic Predisposition to Disease , Oophoritis/genetics , Oophoritis/immunology , Quantitative Trait Loci/immunology , Thymectomy , Amino Acid Sequence , Animals , Animals, Newborn , Antigen Presentation/genetics , Autoantigens/immunology , Autoantigens/metabolism , Cathepsins/antagonists & inhibitors , Cathepsins/physiology , Cystatin A , Cystatins/genetics , Cystatins/isolation & purification , Cysteine Proteinase Inhibitors/genetics , Cysteine Proteinase Inhibitors/isolation & purification , Female , Genetic Linkage/immunology , Male , Mice , Mice, Congenic , Mice, Inbred A , Mice, Inbred C57BL , Molecular Sequence Data , Physical Chromosome MappingABSTRACT
Condensation of sperm chromatin occurs after spermatozoa have left the caput epididymis and are in transit to the cauda epididymis, during which time large numbers of disulfide bonds are formed. The formation of these disulfide bonds requires the repeated oxidation of the cofactor, NAD(P)H. To date, the means by which this oxidation is achieved has yet to be elucidated. Spermatozoa lose the bulk of their cytoplasm prior to leaving the testis; and, as a result, any shuttle systems for removing and transferring reducing equivalents into the mitochondria are unlikely to be operational. In an apparent preparation for the loss of cytoplasm, however, the following events occur during spermatogenesis. First, androgen-binding protein (ABP) is produced by the Sertoli cells of the testis; second, high affinity binding sites for ABP are inserted into the membrane surrounding the nucleus; and third, a nuclear location is acquired for the enzyme, 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD). We propose that after the loss of cytoplasm, the nuclear region of spermatozoa is directly accessible to constituents contained in the lumen of the caput epididymis. As a consequence, luminal ABP attaches itself to the nuclear membrane via its binding sites, and is internalized. After internalization, ABP exerts its principle function, which is to bind to luminal 5alpha-dihydrotestosterone (5alpha-DHT), thereby ensuring its availability to the enzyme, 3alpha-HSD. In the conversion of 5alpha-DHT to 3alpha-androstanediol (3alpha-Diol), NAD(P)H is oxidized. Spermatozoa that reach the cauda epididymis have fully condensed chromatin. In addition, the nuclear region retains appreciable amounts of 5alpha-DHT and 3alpha-Diol, both bound to ABP. During fertilization, the bound 3alpha-Diol is converted back to 5alpha-DHT, reducing equivalents are transferred to NAD(P)+, and disulfide bonds are broken.IVF clinics report that spermatozoa with incompletely condensed chromatin have a low percentage of fertilization. If our proposed mechanism for chromatin condensation/decondensation is borne out by further research, IVF clinics might consider preincubating spermatozoa with 5alpha-DHT in order to increase the efficiency of fertilization.
Subject(s)
Chromatin Assembly and Disassembly/physiology , Models, Biological , Rats/physiology , Spermatogenesis/physiology , 3-Hydroxysteroid Dehydrogenases/metabolism , Androgen-Binding Protein/metabolism , Androstane-3,17-diol/metabolism , Animals , Cell Nucleus/metabolism , Cell Nucleus/ultrastructure , Dihydrotestosterone/metabolism , Epididymis/cytology , Fertilization , Glutathione/metabolism , Male , NADP/metabolism , Oxidation-Reduction , Protamines/chemistryABSTRACT
PROBLEM: Female mice that are injected with estradiol-17beta (E2) and testosterone during the 7-day immune adaptive period are infertile at adulthood. To determine whether the resultant infertility can be caused by steroids other than estrogens/ androgens, this study examined the effect of injecting cortisone, alone, and in combination with E2 and testosterone, on reproductive function. METHOD OF STUDY: Neonatal (C57BL/6J x A/J)F1 B6A female mice were injected from 3 to 6 days of age with sesame oil:ethanol (9:1; v:v), alone, or containing 20 microgg cortisone acetate, 20 microg E2, or 20 microg testosterone. Two additional groups were given 20 microg cortisone acetate in combination with 20 microg E2 or 20 microg testosterone. At adulthood the animals were killed, the stage of vaginal estrus determined, the ovaries examined for the presence of corpora lutea and follicular cysts, and circulating levels of progesterone, E2, and testosterone were measured by radioimmunoassay (RIA). RESULTS: It was found that injections of cortisone seriously compromise reproductive development. For example, 11% of cortisone-injected animals had ovaries that lacked corpora lutea. In addition, 39% of cortisone-injected females had ovaries with follicular cysts. Cortisone-injected females also had low levels of circulating progesterone (18 ng/mL versus 30 ng/mL for the sesame oil-injected females). CONCLUSION: It is concluded that the deleterious effect of steroids on reproductive function, when administered during the immune adaptive period, is not restricted to estrogens and androgens. It is proposed that injections of cortisone alter T-lymphocyte subsets, which contributes to anovulation and the production of follicular cysts.
Subject(s)
Anovulation/chemically induced , Cortisone/analogs & derivatives , Cortisone/adverse effects , Immune System/drug effects , Ovarian Cysts/chemically induced , Animals , Cortisone/administration & dosage , Estradiol/administration & dosage , Female , Immune System/physiology , Mice , Mice, Inbred C57BL , Reproduction/drug effects , Reproduction/immunology , Reproduction/physiology , Testosterone/administration & dosageABSTRACT
Day 3 thymectomy (D3Tx) results in a loss of peripheral tolerance mediated by CD4(+)CD25(+) T cells and the development of autoimmune ovarian dysgenesis (AOD) in A/J and (C57BL/6J x A/J)F(1) (B6AF(1)) hybrids but not in C57BL/6J mice. Quantitative trait loci (QTL) linkage analysis using a B6AF(1) x C57BL/6J backcross population verified Aod1 and Aod2 that were previously mapped as qualitative traits. Additionally, three new QTL intervals, Aod3, Aod4, and Aod5, on chromosomes 1, 2, and 7, respectively, influencing specific subphenotypes of AOD were identified. QTL linkage analysis using the A x B and B x A recombinant inbred lines verified Aod3 and confirmed linkage to H2. Aod5 colocalized with Mater, an ovarian-specific autoantigen recognized by anti-ovarian autoantibodies in the sera of D3Tx mice. Sequence analysis of Mater identified allelic, strain-specific splice variants between A/J and C57BL/6J mice making it an attractive candidate gene for Aod5. Interaction analysis revealed significant epistatic effects between Aod1-5 and Gasa2, a locus associated with susceptibility to D3Tx-induced autoimmune gastritis, as well as with H2. These results indicate that the QTL controlling D3Tx-induced autoimmune phenomenon are both organ specific and more generalized in their effects with respect to the genesis and activity of the immunoregulatory mechanisms maintaining peripheral tolerance.