Effects of Deuterium Depletion on Age-Declining Thymopoiesis In Vivo.

The thymus provides maturation and migration of T cells to peripheral organs of immunity, where they recognize diverse antigens and maintain immunological memory and self-tolerance. The thymus is known to be involved with age and in response to stress factors. Therefore, the search for approaches to the restoration of thymopoiesis is of great interest. The present investigation was aimed at evaluating how prolonged deuterium depletion affects morphogenetic processes and the physiological transition of the thymus to age-related involution. The study was performed on 60 male Wistar rats subjected to consumption of deuterium-depleted water with a 10 ppm deuterium content for 28 days. The control rats consumed distilled water with a normal deuterium content of 150 ppm. The examination found no significant differences in body weight gain or the amount of water consumed. The exposed rats exhibited similar to control dynamics of the thymus weight but significant changes in thymic cell maturation according to cytofluorimetric analysis of thymic subpopulations. Changes in T cell production were not monotonic and differentially engaged morphogenetic processes of cell proliferation, differentiation, and migration. The reactive response to deuterium depletion was a sharp increase in the number of progenitor CD4-CD8- cells and their differentiation into T cells. The compensatory reaction was inhibition of thymopoiesis with more pronounced suppression of differentiation of T-cytotoxic lymphocytes, followed by intensification of emigration of mature T cells to the bloodstream. This period lasts from 3 to 14 days, then differentiation of thymic lymphocytes is restored, later cell proliferation is activated, and finally the thymopoiesis rate exceeds the control values. The increase in the number of thymic progenitor cells after 3-4 weeks suggests consideration of deuterium elimination as a novel approach to prevent thymus involution.

Postnatal Exposure to the Endocrine Disruptor Dichlorodiphenyltrichloroethane Affects Adrenomedullary Chromaffin Cell Physiology and Alters the Balance of Mechanisms Underlying Cell Renewal.

Dichlorodiphenyltrichloroethane (DDT) is a wide-spread systemic pollutant with endocrine disrupting properties. Prenatal exposure to low doses of DDT has been shown to affect adrenal medulla growth and function. The role of postnatal exposure to DDT in developmental disorders remains unclear. The aim of the present investigation is to assess growth parameters and the expression of factors mediating the function and renewal of chromaffin cells in the adult adrenal medulla of male Wistar rats exposed to the endocrine disruptor o,p'-DDT since birth until sexual maturation. The DDT-exposed rats exhibited normal growth of the adrenal medulla but significantly decreased tyrosine hydroxylase production by chromaffin cells during postnatal period. Unlike the control, the exposed rats showed enhanced proliferation and reduced expression of nuclear ß-catenin, transcription factor Oct4, and ligand of Sonic hedgehog after termination of the adrenal growth period. No expression of pluripotency marker Sox2 and absence of Ascl 1-positive progenitors were found in the adrenal medulla during postnatal ontogeny of the exposed and the control rats. The present findings indicate that an increase in proliferative activity and inhibition of the formation of reserve for chromaffin cell renewal, two main mechanisms for cell maintenance in adrenal medulla, in the adult DDT-exposed rats may reflect a compensatory reaction aimed at the restoration of catecholamine production levels. The increased proliferation of chromaffin cells in adults suggests excessive growth of the adrenal medulla. Thus, postnatal exposure to DDT alters cell physiology and increases the risk of functional insufficiency and hyperplasia of the adrenal medulla.

Bilateral Shifts in Deuterium Supply Similarly Change Physiology of the Pituitary-Thyroid Axis, but Differentially Influence Na+/I- Symporter Production.

Deuterium, a stable isotope of hydrogen, is abundant in organisms. It is known to produce various biological effects. However, its impact in thyroid hormone synthesis and secretion is poorly studied. The aim of this investigation was to evaluate the dynamics of thyroid hormones and pituitary thyroid-stimulating hormone secretion during bilateral shifts in deuterium supply and assess a possible role of the Na+/I- symporter (NIS), the main iodide transporter, in altered thyroid function. The experiment was performed on adult male Wistar rats, which consumed deuterium-depleted ([D] = 10 ppm) and deuterium-enriched ([D] = 500,000 ppm) water for 21 days. The assessment of total thyroxine and triiodothyronine and their free fractions, as well as thyroid-stimulating hormone in blood serum, revealed the rapid response of the thyroid gland to shifts in the deuterium/protium balance. The present investigation shows that the bilateral changes in the deuterium body content similarly modulate thyroid hormone production and functional activity of the pituitary gland, but the responses of the thyroid and pituitary glands differ. The response of the thyroid cells was to increase the synthesis of the hormones and the pituitary thyrotropes, in order to reduce the production of the thyroid-stimulating hormone. The evaluation of NIS serum levels found a gradual increase in the rats that consumed deuterium-enriched water and no differences in the group exposed to deuterium depletion. NIS levels in both groups did not correlate with thyroid hormones and pituitary thyroid-stimulating hormone production. The data obtained show that thyroid gland has a higher sensitivity to shifts in the deuterium body content than the hypothalamic-pituitary complex, which responded later but similarly in the case of deuteration or deuterium depletion. It indicates a different sensitivity of the endocrine glands to alterations in deuterium content. It suggests that thyroid hormone production rate may depend on deuterium blood/tissue and cytosol/organelle gradients, which possibly disturb the secretory process independently of the NIS.

Emerging Role of Deuterium/Protium Disbalance in Cell Cycle and Apoptosis.

Deuterium, a stable isotope of hydrogen, is a component of water and organic compounds. It is the second most abundant element in the human body after sodium. Although the concentration of deuterium in an organism is much lower than that of protium, a wide variety of morphological, biochemical, and physiological changes are known to occur in deuterium-treated cells, including changes in fundamental processes such as cell division or energy metabolism. The mode and degree of changes in cells and tissues, both with an increase and a decrease in the concentration of deuterium, depends primarily on the time of exposure, as well as on the concentration. The reviewed data show that plant and animal cells are sensitive to deuterium content. Any shifts in the D/H balance outside or inside cells promote immediate responses. The review summarizes reported data on the proliferation and apoptosis of normal and neoplastic cells in different modes of deuteration and deuterium depletion in vivo and in vitro. The authors propose their own concept of the effects of changes in deuterium content in the body on cell proliferation and death. The altered rate of proliferation and apoptosis indicate a pivotal role of the hydrogen isotope content in living organisms and suggest the presence of a D/H sensor, which is yet to be detected.

Developmental Exposure to DDT Disrupts Transcriptional Regulation of Postnatal Growth and Cell Renewal of Adrenal Medulla.

Dichlorodiphenyltrichloroethane (DDT) is the most widespread persistent pollutant with endocrine-disrupting properties. DDT has been shown to disrupt secretory and morphogenetic processes in the adrenal cortex. The present investigation aimed to evaluate transcriptional regulation of postnatal growth of the adrenal medulla and formation of the pools necessary for self-renewal of medullary cells in rats that developed under low-dose exposure to DDT. The study was performed using male Wistar rats exposed to low doses of o,p'-DDT during prenatal and postnatal development. Light microscopy and histomorphometry revealed diminished medulla growth in the DDT-exposed rats. Evaluation of Ki-67 expression in chromaffin cells found later activation of proliferation indicative of retarded growth of the adrenal medulla. All DDT-exposed rats exhibited a gradual decrease in tyrosine hydroxylase production by adrenal chromaffin cells. Immunohistochemical evaluation of nuclear ß-catenin, transcription factor Oct4, and ligand of sonic hedgehog revealed increased expression of all factors after termination of growth in the control rats. The DDT-exposed rats demonstrated diminished increases in Oct4 and sonic hedgehog expression and lower levels of canonical Wnt signaling activation. Thus, developmental exposure to the endocrine disruptor o,p'-DDT alters the transcriptional regulation of morphogenetic processes in the adrenal medulla and evokes a slowdown in its growth and in the formation of a reserve pool of cells capable of dedifferentiation and proliferation that maintain cellular homeostasis in adult adrenals.

Developmental Exposure to Endocrine Disrupter DDT Interferes with Age-Related Involution of Thymus.

The impact of endocrine-disrupting chemicals on the development and involution of the immune system is a possible reason for the increased incidence of disorders associated with inappropriate immune function. The thymus is a lymphoid and also an endocrine organ, and, accordingly, its development and functioning may be impaired by endocrine disruptors. The aim was to evaluate age-related thymus involution in mature rats exposed to the endocrine disruptor DDT during prenatal and postnatal ontogeny. Methodology included in vivo experiment on male Wistar rats exposed to low doses of DDT during prenatal and postnatal development and morphological assessment of thymic involution, including the immunohistochemical detection of proliferating thymocytes. The study was carried out at the early stage of involution. Results: DDT-exposed rats exhibited a normal anatomy, and the relative weight of the thymus was within the control ranges. Histological and immunohistochemical examinations revealed increased cellularity of the cortex and the medulla, higher content of lymphoblasts, and more intensive proliferation rate of thymocytes compared to the control. Evaluation of thymic epithelial cells revealed a higher rate of thymic corpuscles formation. Conclusion: The data obtained indicate that endocrine disrupter DDT disturbs postnatal development of the thymus. Low-dose exposure to DDT during ontogeny does not suppress growth rate but violates the developmental program of the thymus by slowing down the onset of age-related involution and maintaining high cell proliferation rate. It may result in excessive formation of thymus-dependent areas in peripheral lymphoid organs and altered immune response.

Impact of Prenatal and Postnatal Exposure to Endocrine Disrupter DDT on Adrenal Medulla Function.

Epinephrine is the most abundant catecholamine hormone, produced by the nervous system and adrenal glands. Endocrine disruption of epinephrine synthesis, secretion and signaling is less studied than steroid and thyroid hormones. Dichlorodiphenyltrichloroethane (DDT) is recognized as one of the most prominent environmental contaminants with a long half-life. It is a potent endocrine disrupter affecting sex steroid, mineralocorticoid, glucocorticoid and thyroid hormone production. Exposure to low doses of DDT is universal and begins in utero. Therefore, we studied adrenal medulla growth and function in male Wistar rats exposed to low doses of DDT during prenatal and postnatal development until puberty and adulthood, as well as rats exposed to DDT since the first day of postnatal development. All the exposed rats demonstrated lowered epinephrine blood levels, gradually reducing with age. DDT was found to inhibit the synthesis of tyrosine hydroxylase and affect the mitochondrial apparatus of epinephrine-producing cells during puberty and even after maturation. Low-dose exposure to DDT from birth resulted in more pronounced changes in adrenomedullary cells and a more profound decrease (up to 50%) in epinephrine secretion in adult rats. Prenatal onset of exposure demonstrated a mild effect on epinephrine-producing function (30% reduction), but was associated with lower rate of adrenal medulla growth during maturation and 25% smaller adrenal medullar size in adult rats. All subjects exposed to low doses of DDT failed to develop adaptive changes and restore proper epinephrine production. These results indicate a dysmorphogenetic effect of prenatal exposure and disruption of secretory function of adrenal chromaffin cells by postnatal exposure to DDT.

Dichlorodiphenyltrichloroethane and the Adrenal Gland: From Toxicity to Endocrine Disruption.

Endocrine disruptors are exogenous compounds that pollute the environment and have effects similar to hormones when inside the body. One of the most widespread endocrine disruptors in the wild is the pesticide dichlorodiphenyltrichloroethane (DDT). Toxic doses of DDT are known to cause cell atrophy and degeneration in the adrenal zona fasciculata and zona reticularis. Daily exposure in a developing organism to supposedly non-toxic doses of DDT have been found to impair the morphogenesis of both the cortex and the medulla of the adrenal glands, as well as disturbing the secretion of hormones in cortical and chromaffin cells. Comparison of high and very low levels of DDT exposure revealed drastic differences in the morphological and functional changes in the adrenal cortex. Moreover, the three adrenocortical zones have different levels of sensitivity to the disruptive actions of DDT. The zona glomerulosa and zona reticularis demonstrate sensitivity to both high and very low levels of DDT in prenatal and postnatal periods. In contrast, the zona fasciculata is less damaged by low (supposedly non-toxic) exposure to DDT and its metabolites but is affected by toxic levels of exposure; thus, DDT exerts both toxic and disruptive effects on the adrenal glands, and sensitivity to these two types of action varies in adrenocortical zones. Disruptive low-dose exposure leads to more severe affection of the adrenal function.

Impaired Morphogenesis and Function of Rat Adrenal Zona Glomerulosa by Developmental Low-Dose Exposure to DDT Is Associated with Altered Oct4 Expression.

Dichlorodiphenyltrichloroethane (DDT) is a persistent organic pollutant and one of the most widespread endocrine disrupting chemicals. The impact of low-dose exposure to DDT on the morphogenesis of the adrenal gland is still poorly understood. The development and function of zona glomerulosa in rats has been found to be associated with changes in the expression of the transcription factor Oct4 (Octamer 4), which is the most important player in cell pluripotency. The aim of the study was to investigate the morphogenesis and function of rat adrenal zona glomerulosa in rats exposed to low doses of DDT during prenatal and postnatal development and to determine the possible role of Oct4 in DDT-mediated structural and functional changes. The DDT-exposed rats demonstrated slower development and lower functional activity of the zona glomerulosa during the pubertal period associated with higher expression of Oct4. Further, accelerated growth and restoration of hormone production was associated with, firstly, a decrease in Oct4 expressing cells and secondly, the loss of the inverse relationship between basal aldosterone levels and the number of Oct4 expressing cells. Thus, the transcriptional factor Oct4 exhibited an altered pattern of expression in the DDT-exposed rats during postnatal development. The results of the study uncover a novel putative mechanism by which low doses of DDT disrupt the development of adrenal zona glomerulosa.

Differential Disrupting Effects of Prolonged Low-Dose Exposure to Dichlorodiphenyltrichloroethane on Androgen and Estrogen Production in Males.

Dichlorodiphenyltrichloroethane (DDT) is the most widespread, persistent pollutant and endocrine disruptor on the planet. Although DDT has been found to block androgen receptors, the effects of its low-dose exposure in different periods of ontogeny on the male reproductive system remain unclear. We evaluate sex steroid hormone production in the pubertal period and after maturation in male Wistar rats exposed to low doses of o,p'-DDT, either during prenatal and postnatal development or postnatal development alone. Prenatally and postnatally exposed rats exhibit lower testosterone production and increased estradiol and estriol serum levels after maturation, associated with the delayed growth of gonads. Postnatally exposed rats demonstrate accelerated growth of gonads and higher testosterone production in the pubertal period. In contrast to the previous group, they do not present raised estradiol production. All of the exposed animals exhibit a reduced conversion of progesterone to 17OH-progesterone after sexual maturation, which indicates putative attenuation of sex steroid production. Thus, the study reveals age-dependent outcomes of low-dose exposure to DDT. Prenatal onset of exposure results in the later onset of androgen production and the enhanced conversion of androgens to estrogens after puberty, while postnatal exposure induces the earlier onset of androgen secretion.

Transcription factors ß-catenin and Hex in postnatal development of the rat adrenal cortex: implication in proliferation control.

Transcriptional regulation of growth, maturation, and cell turnover in adrenal cortex during postnatal development has been significantly less studied than in embryonic period, while elucidation of factors mediating its normal postnatal morphogenesis could clarify mechanisms of tumorigenesis in adrenal cortex. Expression of transcription factors Hex, ß-catenin, and Wnt signaling in the adrenal cortex of male pubertal and postpubertal Wistar rats were examined. Adrenal cortex morphology and hormone production during postnatal development were also studied. Adrenocortical zones demonstrated similar reduction of Ki-67-expressing cells, but different patterns of morphological and functional changes. Age-dependent decrease in percentage of cells with membrane localization of ß-catenin and stable rate of cells with nuclear ß-catenin, indicative of Wnt signaling activation, were revealed in each cortical zone. Nuclear ß-catenin was not observed in immature areas of zona fasciculata. No association between Wnt signaling activation and rates of proliferation as well as changes in secretion of adrenocortical hormones was observed in postnatal development of rat adrenal cortex. Hex, known as antiproliferative factor, showed up-regulation of expression after puberty. Strong inverse correlations between ratio of Hex-positive cells and proliferating cells were found in zona glomerulosa and zona fasciculata. Zona reticularis demonstrated moderate correlation. Thus, these findings suggest a role for Hex in proliferation control during postnatal development of the rat adrenal cortex and possible implication of Hex down-regulation in adrenocortical dysplasia and neoplasia, which requires further study. Evaluation of Hex expression may also be considered a potent tool in assessment of cell proliferation in rat adrenal cortex.

Developmental exposure to low doses of dichlorodiphenyltrichloroethane impairs proliferative response of thymic lymphocytes to Concanavalin A in rats.

The aim of the research was to study formation of thymic lymphocytes proliferative response to T cell mitogen Concanavalin A in 7, 42, and 70 days-old male Wistar rats developmentally exposed to low doses of endocrine disruptor dichlorodiphenyltrichloroethane (2.90 ± 0.13 µg/kg body weight). The thymus of the exposed rats did not show morphological abnormalities. Exposure to the endocrine disrupter was found to alter age-dependent changes of thymic lymphocyte proliferative activity and attenuate proliferative response to Concanavalin A in puberty and adulthood. Insufficient response to mitogen was mediated by higher content of actively proliferating Ki-67-positive lymphoblasts compared to the control values. Insufficient proliferative response to mitogen in developmentally exposed to the endocrine disruptor rats may provide higher risk of impaired cellular immune reactions.