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PubMed2002

Cellular mechanisms of nociception in the frog.

Cellular mechanisms underlying defense reactions induced by noxious heat and acids were studied in frogs (Rana pipiens) by measuring whole cell membrane currents in cultured dorsal root ganglion (DRG) neurons. Seventy-eight of 82 DRG neurons exposed to 3-s ramps of increasing temperature to 48 degrees C exhibited an inward current (I(HEAT)) of 490 +/- 70 pA at -70 mV. I(HEAT) exhibited reversal at approximately 10 mV with a pronounced outward rectification, suggesting opening of nonselective cation channels. In frogs, in contrast to mammals, I(HEAT) was not influenced by capsaicin (5 microM), capsazepine (10 microM), or ruthenium red (10 microM). In a large proportion (approximately 80%) of heat-sensitive DRG neurons, acids produced a large slowly inactivating sodium carried current (I(ACID)) with average pH(50) 5.7. I(ACID) was blocked by 1 mM amiloride (to 22%) and was absent if extracellular Na(+) was substituted by Cs(+). Elevating temperature to 38 degrees C increased I(ACID), whereas temperatures >40 degrees C profoundly inhibited it (by 82 +/- 2%; n = 42). The inhibition was long-lasting (>30 s) but fully reversible. Phorbol ester myristate acetate (PMA, 1 microM) and forskolin (1 microM) inhibited I(ACID) to 37 +/- 5% (n = 5) and 78 +/- 8% (n = 4), respectively. It is suggested that I(HEAT) in frog DRG neurons is carried through capsaicin-insensitive nonselective cation channels distinct from vanilloid receptor in mammals, whereas I(ACID) is carried through amiloride-sensitive sodium channels that are strongly inhibited by noxious heat, possibly due to activation of the intracellular messenger systems.

Acids↗

PubMed2004

Cerebral vasospasm: a consideration of the various cellular mechanisms involved in the pathophysiology.

The cellular mechanisms responsible for cerebral vasospasm (CVS) occurring after subarachnoid hemorrhage (SAH) have been of major interest over the past 50 years. The present review describes how each of the discrete anatomic components that comprise the cerebral artery may contribute to the pathology of CVS. The blood extravasated after SAH is hemolyzed and undergoes degradation with resultant production of free radicals, known to be powerful initiators of vascular damage. An inflammatory response is generated activating both leukocytes and platelets with subsequent release of inflammatory agents. The cerebral artery affected by CVS undergoes phenotypic change involving both the endothelial and smooth muscle cells. In the endothelium the production of nitric oxide and prostacyclin is affected. In the smooth muscle cells signal transduction pathways that enhance the function of the contractile proteins and induce the upregulation of contractile receptors are activated. In parallel, there is evidence that nervous reflex pathways involving the trigeminal ganglion and the hypothalamus are activated. However, the relative contributions of each of the systems are speculative. Therapy may be directed at disrupting the cascade leading from the SAH insult to CVS or at overcoming the dysfunction incurred by CVS; possible therapeutical interventions are considered.

Blood Cells↗

PubMed2005

The cellular mechanisms involved in the vasodilator effect of nebivolol on the renal artery.

Nebivolol is known as a highly selective beta1-adrenoceptor antagonist. Based on the reported vasodilator effect of nebivolol, we examined the cellular mechanisms by which the drug induces renal artery vasodilation, an issue of potential relevance for condition associated with high blood pressure. To this purpose, myograph and patch-clamp techniques were used. Small mouse renal arteries were placed in the myograph chamber, and after the optimal concentration for the vasodilator effect of nebivolol was established (50 microM), the arteries were further investigated to assess the potential contribution of nitric oxide (NO) and of Ca2+ ions to the nebivolol-induced effect, by exposing the arteries to the specific inhibitors such as N(G)-nitro-L-arginine methylester (L-NAME, 100 microM), ethylenglycol-bis-(beta-amino-ethylen ester) N,N'-tetraacetic acid (EGTA, 4 microM) and thapsigargin (1 microM). The expression of NO synthase was evaluated by the Western-blot technique. Using myograph and patch-clamp techniques applied on intact renal artery, we investigated the role of beta2-adrenoceptors, of myoendothelial junctions and of Ca(2+)-activated K+ channels in the vasodilatory effects of nebivolol, using 100 microM butoxamine, 40 microM 18 beta-glycyrrhetinic acid, 1 mM tetraethylammonium, and 100 nM iberiotoxin, respectively. The results showed that the cellular mechanisms of the vasodilator effect of nebivolol on the renal artery entail (i) activation of the endothelial beta2-adrenoceptor, (ii) participation of [Ca2+]i, (iii) increase in NO and eNOS, and (iv) activation of Ca(2+)-activated K+ channels. The cellular mechanisms underlying vasodilator effect of nebivolol on the artery explain the favorable effect of this drug in hypertension.

Acetylcholine↗

PubMed1998

Cellular mechanisms of adrenaline-stimulated anion secretion by the mouse endometrial epithelium.

The uterine fluid composition is largely determined by the absorptive and secretory activities of the endometrial epithelium. The present study explored the cellular mechanisms involved in adrenaline-stimulated anion secretion across the cultured mouse endometrial epithelium using the short-circuit current (ISC) technique in conjunction with transporter inhibitors and channel blockers. Cultured endometrial epithelial monolayers responded to basolateral application of adrenaline with an increase in ISC, which was attributable to both Cl- and HCO3- secretion. When extracellular Cl- or HCO3- was removed, the adrenaline-induced response, as measured by the total charge transfer per unit area, was reduced to 53% and 46%, respectively. When both Cl- and HCO3- were absent from the bathing solutions, the adrenaline-induced response was reduced to only 2% of the response when both ions were present, indicating substantial contribution of Cl- and HCO3- secretion to the adrenaline-stimulated response. Cellular mechanisms, e.g., transporters and ion channels, involved in Cl- or HCO3- secretion were investigated separately. Cl- secretion was found to depend on the activities of basolaterally located Na+-K+-ATPase, Na+-K+-2Cl- cotransporter, and K+ channels, while evidence suggested that HCO3- secretion depends substantially on basolaterally situated Na+-HCO3- cotransporter and Na+-H+ exchanger. Similar to what was seen for Cl- exit, a large portion of HCO3- appeared to exit apically through anion channels. The results indicate that the uterine fluid composition in the mouse may be regulated by adrenaline through stimulation of both Cl- and HCO3- secretion and may be fine-tuned through an elaborate operation of different cellular mechanisms.

Animals↗

PubMed2006

Cellular mechanism for spontaneous calcium oscillations in astrocytes.

AIM: To determine the Ca2+ source and cellular mechanisms of spontaneous Ca2+ oscillations in hippocampal astrocytes. METHODS: The cultured cells were loaded with Fluo-4 AM, the indicator of intracellular Ca2+, and the dynamic Ca2+ transients were visualized with confocal laser-scanning microscopy. RESULTS: The spontaneous Ca2+ oscillations in astrocytes were observed first in co-cultured hippocampal neurons and astrocytes. These oscillations were not affected by tetrodotoxin (TTX) treatment and kept up in purity cultured astrocytes. The spontaneous Ca2+ oscillations were not impacted after blocking the voltage-gated Ca2+ channels or ethylenediamine tetraacetic acid (EDTA) bathing, indicating that intracellular Ca2+ elevation was not the result of extracellular Ca2+ influx. Furthermore, the correlation between the spontaneous Ca2+ oscillations and the Ca2+ store in endoplasmic reticulum (ER) were investigated with pharmacological experiments. The oscillations were: 1) enhanced when cells were exposed to both low Na+ (70 mmol/L) and high Ca2+ (5 mmol/L) solution, and eliminated completely by 2 micromol/L thapsigargin, a blocker of sarcoplasmic reticulum Ca2+-ATPase; and 2) still robust after the application with either 50 micromol/L ryanodine or 400 micromol/L tetracaine, two specific antagonists of ryanodine receptors, but depressed in a dose-dependent manner by 2-APB, an InsP3 receptors (InsP3R) blocker. CONCLUSION: InsP3R-induced ER Ca2+ release is an important cellular mechanism for the initiation of spontaneous Ca2+ oscillation in hippocampal astrocytes.

Animals↗

PubMed2001

Molecular and cellular mechanisms of allergic disease.

The molecular and cellular mechanisms mediating the allergic inflammatory cascade involve multiple mediators, cell types, and pathways. Of particular interest are the pathways regulated by the T(H)2 lymphocyte, which result in release of IL-4 (important to IgE synthesis) and IL-5 (important to eosinophil proliferation). IL-4 regulates differentiation of naïve T(H)0 cells to develop a T(H)2 phenotype and stimulates B cells to produce IgE. Cross-linking by allergen of IgE affixed to high-affinity receptors on mast cells and basophils triggers degranulation and the release of preformed inflammatory mediators (important to the early phase response), and subsequently initiates synthesis and the release of lipid mediators and cytokines (which may contribute to the late phase response). Eosinophils may also play a prominent role in the development of bronchial hyperreactivity. IL-5, which is a lineage-specific eosinophil growth factor, increases the formation of eosinophils from progenitor cells and, in concert with CCR3 active chemokines, increases their trafficking to sites of allergic inflammation. An improved understanding of the basic mechanisms of allergic inflammation has led to the discovery of molecular targets involved in the initial events of the inflammatory cascade. Potential targets for the development of novel therapies for allergic disease include IgE, the T(H)2 lymphocyte, and T(H)2-derived cytokines, IL-4 and IL-5.

Eosinophils↗

PubMed1995

Molecular and cellular mechanisms of atherosclerosis.

At least three distinct cellular mechanisms are currently thought to be responsible for the initiation of atherosclerotic lesions in humans: (1) accumulation of lipids and plasma-derived lipoproteins in the arterial intima; (2) smooth muscle cell migration from the media into the intima, and smooth muscle cell proliferation or accumulation, or both, within the intima; and (3) accumulation of platelet and/or fibrin deposits in the intima. Independent of the triggering factor, it appears that the first step in atherogenesis involves activation of repair mechanisms in the blood vessel in an attempt to restore vascular homeostasis, which involves a delicate balance of growth promoting and growth inhibitory activities of the vascular wall cells. The healing process involves a series of specific and temporally coordinated events, such as platelet aggregation, monocyte adhesion and migration across endothelial cells, and migration and proliferation of vascular smooth muscle cells, which are normally orchestrated by a variety of growth factors, cytokines, adhesion molecules, and extracellular matrix proteins in a controlled, although not yet fully understood, manner. Alteration in any of these steps during the healing process can lead to changes in the patterns of protein synthesis, processing, and secretion in these cells. These in turn result in connective tissue deposition, lipid accumulation, cell death, mineralization, and finally the development of advanced, occluding atherosclerotic plaques.

Arteriosclerosis↗

PubMed1996

Food allergy, oral tolerance and immunomodulation--their molecular and cellular mechanisms.

This paper describes the molecular and cellular mechanisms of food allergy and oral tolerance including immunomodulation. Food allergy is triggered by an aberrant immune response elicited by oral administration of dietary antigens. Oral tolerance is a state of immunological unresponsiveness induced by oral administration of dietary antigens and is thought to serve to suppress food allergy. This review first describes the characteristic properties of T and B cells relating to milk allergy and also the location of binding sites to T and B cells on allergen molecules. The immunogenicity of allergens is shown to be reduced by the modulations of the T cell binding site, using sophisticated methods such as site-specific mutagenesis. Furthermore, this review focuses on oral tolerance with special reference to the identification of lymphocyte compartment subsets and the immunological mechanism relating to oral tolerance. Finally, the application of oral tolerance for the treatment of allergy is speculated on.

Animals↗

PubMed2006

Adaptation of cellular mechanical behavior to mechanical loading for osteoblastic cells.

Numerous cellular biochemical responses to mechanical loading are transient, indicating a cell's ability to adapt its behavior to a new mechanical environment. Since load-induced cellular deformation can initiate these biochemical responses, the overall goal of this study was to investigate the adaptation of global, or whole-cell, mechanical behavior, i.e., cellular deformability, in response to mechanical loading for osteoblastic cells. Confluent cell cultures were subjected to 1 or 2 Pa flow-induced shear stress for 2 h. Whole-cell mechanical behavior was then measured for individual cells using an atomic force microscope. Compared to cells maintained under static conditions, whole-cell stiffness was 1.36-fold (p=0.006) and 1.70-fold (p<0.001) greater for cells exposed to 1 and 2 Pa shear loading, respectively. The increase in shear stress magnitude from 1 to 2 Pa also caused a statistically significant, 1.25-fold increase in cell stiffness (p=0.02). Increases in cell stiffness were not altered in either flow group for 70 min after flow was terminated (p=0.15). Flow-induced rearrangement of the actin cytoskeleton was also maintained for at least 90 min after flow was terminated. Taken together, these findings support the hypothesis that cells become mechanically adapted to their mechanical environment via cytoskeletal modifications. Accordingly, cellular mechanical adaptation may play a key role in regulation of cellular mechanosensitivity and the related effects on tissue structure and function.

3T3 Cells↗

PubMed1989

Cellular mechanisms underlying modulation of breathing pattern in mammals.

Understanding the generation and modulation of respiratory pattern requires knowledge of the cellular and network properties of the central nervous system controller. Although considerable efforts have focused on network properties, recent efforts in several laboratories have emphasized the importance of cellular mechanisms. Using a novel experimental in vitro system, we have been able to investigate several classes of cellular mechanisms difficult or impossible to study in vivo or in tissue slices. The conclusions and hypotheses that we have made include the following: 1. Respiratory rhythm and spatiotemporal patterns of (pre)motoneuronal activity are separately generated. 2. Cl- -dependent inhibition is critical in burst pattern formation. 3. Cl- -dependent and at least one type of K+-dependent (GABAB) inhibition is not necessary for rhythm generation. 4. Inhibitory neurotransmitters, including those acting through second messenger systems, can modulate respiratory rhythm and pattern. 5. Pacemakers underlie rhythm generation. 6. Excitatory inspiratory drive to respiratory motoneurons is mediated by an excitatory amino acid, which acts on both post- and presynaptic receptors. The effort to verify these conclusions and test these hypotheses should greatly enhance our understanding of the nervous control of respiration.

Animals↗

PubMed2004

Comparison of cellular mechanisms underlying histamine release from rat mast cells induced by ionic and nonionic radiographic contrast media.

PURPOSE: To determine the cellular mechanisms underlying mast cell histamine release induced by ionic and nonionic radiographic contrast media. MATERIALS AND METHODS: Histamine release from rat pulmonary mast cells was measured after incubation with various radiographic contrast media. The cellular cAMP content was determined by an enzymatic immunoassay. RESULTS: Both ionic and nonionic contrast media stimulated the histamine release, although the former was more potent than the latter. Dibutyryl cAMP suppressed histamine release evoked by ionic but not nonionic contrast media in a manner dependent on A kinase. The cellular cAMP content was lowered only by ionic contrast media. However, a secretory phospholipase A2 inhibitor p-bromophenacyl bromide inhibited both ionic and nonionic contrast media-evoked histamine releases. CONCLUSION: We demonstrated for the first time the difference and similarity in the cellular mechanisms underlying histamine release induced by ionic and nonionic contrast media, in which the reduction in cAMP was specific for ionic materials and the activation of secretory phospholipase A2 may be common to both agents.

Acetophenones↗

PubMed1984

Age-related structural changes in trabecular and cortical bone: cellular mechanisms and biomechanical consequences.

It is proposed that there are two structurally different forms of bone loss with different rates, cellular mechanisms, and biomechanical effects. Rapid bone loss is the result of excessive depth of osteoclastic resorption cavities. This leads in trabecular bone to perforation of structural elements, increased size of marrow cavities, and discontinuity of the bone structure, and in cortical bone to subendosteal cavitation and conversion of the inner third of the cortex to a trabecularlike structure, which then undergoes the same changes as the trabecular bone originally present. These structural characteristics reduce the strength of the bones to a greater extent than the reduction in the amount of bone by itself would suggest. Slow bone loss results from incomplete refilling by osteoblasts of resorption cavities of normal or reduced size. This leads to simple thinning of residual structural elements in both trabecular and cortical bone, and reduces the strength of the bones in proportion to the reduction in the amount of bone. This concept, although derived mainly from an examination of postmenopausal bone loss, may be applicable to other osteopenic states. At the same time as bone loss is occurring on the endosteal surface, rapidly or slowly, bone is being added to the periosteal surface, but much more slowly than during growth. The cellular mechanism is the converse of that causing slow bone loss, consisting of slight overfilling of shallow resorption cavities. Slow periosteal gain serves to partly offset the structural weakness resulting from endosteal loss, but is not directly compensatory.(ABSTRACT TRUNCATED AT 250 WORDS)

Aging↗

PubMed1989

Cellular mechanisms of nitrate action.

It is now generally accepted that organic nitrates generate their vasodilator action via production of nitric oxide. However, the cellular location of the metabolic enzyme(s) responsible for such conversion has not been defined. We examined the production of nitric oxide, via chemiluminescence detection, by various cellular fractions of the bovine coronary artery. We were able to show that the highest activity resides in the plasma membrane. Future isolation and characterization of such metabolic systems will greatly assist our understanding of nitrate action and tolerance. Several cellular mechanisms for nitrate tolerance have been proposed. Among the most popular theories is the "intracellular sulfhydryl depletion hypothesis" originally proposed by Needleman et al. The primary supportive data for this mechanism are that exogeneously added thiols (such as N-acetylcysteine) can potentiate the in vivo activity of nitroglycerin and can partially reverse nitrate tolerance. We showed that a cellular-impermeant thiol, viz: glutathione, can also potentiate the hemodynamic effect of nitroglycerin in rats. We subsequently showed that exogenously administered thiols can promote the formation of vasoactive S-nitrosothiols in blood. Thus, the beneficial effects of thiols on nitrate action might be mediated through an extracellular pathway. Another cellular mechanism for nitrate tolerance suggested that tolerance is caused by an alteration of the enzyme, guanylate cyclase. We showed, however, that blood vessels made tolerant to nitroglycerin remain fully responsive (in terms of in vitro relaxation) toward nitric oxide and S-nitrosothiols. These data showed that, as far as relaxation is concerned, nitrate tolerance did not cause a significant alteration of guanylate cyclase activity toward nitric oxide and S-nitrosothiols.

Animals↗

PubMed1985

Cellular mechanisms in the actions of antiglaucoma drugs.

There are several classes of drugs currently in use for the therapeutic management of the glaucomas. Although the ocular hypotensive effects of these agents have been well characterized and described, little is known of their site of action and cellular mechanism. This review attempts to describe those cellular mechanisms that may be linked to the actions of several classes of antiglaucoma drugs. Special emphasis was placed on drug actions and 1) the adenylate cyclase system; 2) receptor-coupled phosphoinositide turnover; 3) prostaglandins and 4) ion transport processes. Models are presented depicting proposed cellular sites of the interaction of the antiglaucoma drugs with these cellular processes.

Adenylyl Cyclases↗

PubMed2001

Cellular mechanisms of bone resorption in breast carcinoma.

The cellular mechanisms that account for the increase in osteoclast numbers and bone resorption in skeletal breast cancer metastasis are unclear. Osteoclasts are marrow-derived cells which form by fusion of mononuclear phagocyte precursors that circulate in the monocyte fraction. In this study we have determined whether circulating osteoclast precursors are increased in number or have an increased sensitivity to humoral factors for osteoclastogenesis in breast cancer patients with skeletal metastases (+/- hypercalcaemia) compared to patients with primary breast cancer and age-matched normal controls. Monocytes were isolated and cocultured with UMR 106 osteoblastic cells in the presence of 1,25 dihydroxyvitamin D3[1,25(OH)2D3] and human macrophage colony stimulating factor (M-CSF) on coverslips and dentine slices. Limiting dilution experiments showed that there was no increase in the number of circulating osteoclast precursors in breast cancer patients with skeletal metastases (+/- hypercalcaemia) compared to controls. Osteoclast precursors in these patients also did not exhibit increased sensitivity to 1,25(OH)2D3or M-CSF in terms of osteoclast formation. The addition of parathyroid hormone-related protein and interleukin-6 did not increase osteoclast formation. The addition of the supernatant of cultured breast cancer cell lines (MCF-7 and MDA-MB-435), however, significantly increased monocyte-osteoclast formation in a dose-dependent fashion. These results indicate that the increase in osteoclast formation in breast cancer is not due to an increase in the number/nature of circulating osteoclast precursors. They also suggest that tumour cells promote osteoclast formation in the bone microenvironment by secreting soluble osteoclastogenic factor(s).

Aged↗

PubMed2002

Cellular mechanisms of orexin actions on paraventricular nucleus neurones in rat hypothalamus.

Using whole-cell patch clamp techniques we have examined the cellular mechanisms underlying the effects of orexin A (OX-A) on electrophysiologically identified magnocellular and parvocellular neurones in the rat hypothalamic paraventricular nucleus (PVN). The majority of magnocellular neurones (67 %) showed concentration-dependent, reversible depolarizations in response to OX-A. These effects were abolished in tetrodotoxin (TTX), suggesting them to be indirect effects on this population of neurones. OX-A also caused increases in excitatory postsynaptic current (EPSC) frequency and amplitude in magnocellular neurones. The former effects were again blocked in TTX while increases in mini-EPSC amplitude remained. Depolarizing effects of OX-A on magnocellular neurones were also found to be abolished by kynurenic acid, supporting the conclusion that these effects were the result of activation of a glutamate interneurone. Parvocellular neurones (73 % of those tested) also showed concentration-dependent, reversible depolarizations in response to OX-A. In contrast to magnocellular neurones, these effects were maintained in TTX, indicating direct effects of OX-A on this population of neurones. Voltage clamp analysis using slow voltage ramps demonstrated that OX-A enhanced a non-selective cationic conductance with a reversal potential of -40 mV in parvocellular neurones, effects which probably explain the depolarizing effects of this peptide in this subpopulation of PVN neurones. These studies have identified separate cellular mechanisms through which OX-A influences the excitability of magnocellular and parvocellular PVN neurones.

Animals↗

PubMed1992

Cellular mechanism of the insulin-like effect of growth hormone in adipocytes. Rapid translocation of the HepG2-type and adipocyte/muscle glucose transporters.

The cellular mechanism whereby growth hormone (GH) acutely stimulates adipocyte glucose uptake was studied in cultures of primary rat adipocytes differentiated in vitro. Preadipocytes were isolated by collagenase digestion of inguinal fat-pads from young rats and were differentiated in the presence of 3-isobutyl-1-methylxanthine, insulin and dexamethasone. The development of an adipocyte morphology (i.e. lipid inclusions) was observed over 6 days after initiation of differentiation. Coincident with this phenotypic change was an increase in glyceraldehyde-3-phosphate dehydrogenase (GPDH) activity and in cellular content of the HepG2-type (Glut1) and adipocyte/muscle (Glut4) glucose transporter isoforms as determined by Western immunoblotting of total cellular protein. Age-matched undifferentiated cells expressed the Glut1 transporter and low levels of GPDH, but neither accumulated lipid nor exhibited measurable expression of the Glut4 protein. On day 6 after the initiation of differentiation, GH and insulin stimulated 2-deoxy[14C]glucose uptake in a dose- and time-dependent fashion in adipocytes cultured under serum-free conditions for at least 15 h. Western-blot analysis of subcellular fractions revealed that both GH and insulin rapidly (within 20 min) stimulated translocation of the Glut1 and Glut4 proteins from a low-density microsomal fraction to the plasma membrane. Confirmatory evidence was provided in immunocytochemical experiments utilizing antisera directed against the C-terminal region of the Glut4 protein and a fluorescein isothiocyanate-labelled second antibody. Observation of the cells via confocal laser microscopic imaging was consistent with glucose transporter redistribution from an intracellular region to the plasma membrane after treatment with GH or insulin. On the basis of these data, we suggest that the insulin-like effect of GH on adipocyte glucose transport involves translocation of the Glut1 and Glut4 proteins to the plasma membrane. Furthermore, stimulation of glucose-transporter translocation by both GH and insulin may indicate a common cell signalling element between the adipocyte GH and insulin receptors or, alternatively, the existence of multiple cellular mechanisms for stimulating glucose-transporter translocation.

1-Methyl-3-isobutylxanthine↗

PubMed2006

Cellular mechanism of estrogen-induced thymic involution in wall lizard: caspase-dependent action.

The present study, for the first time in an ectothermic vertebrate, demonstrates the cellular mechanism of estrogen-induced thymic involution. Ovariectomy in lizards during the preparatory phase of the reproductive cycle resulted in distinct differentiation of cortico-medullary regions and increase in cellularity, especially in the cortical region. The ovariectomy-induced changes were reversed following administration of 17-estradiol (E2), suggesting a primary role of E2 in causing thymic atrophy. To understand the cellular mechanism of E2-induced thymic atrophy, in vitro effect of E2 was investigated on thymocyte proliferation and apoptosis. E2 decreased the uptake of tritiated thymidine (3H-TdR) by thymocytes in a dose-dependent manner, suggesting that estrogen directly inhibits the thymocyte proliferation. Unlike proliferation, E2 did not have any direct effect on thymocyte apoptosis, as evident by DNA gel electrophoretic, flow cytometric or fluorescence microscopic studies. However, in the presence of thymic epithelial cell-rich stromal components (TEC), E2 treatment at low or high concentrations resulted in depolarization of plasma membrane, DNA fragmentation and decrease in DNA content. This suggests that E2 indirectly, through TEC-secreted factors, controls thymocyte apoptosis. Similar result was observed following fluorescence microscopy. The indirect effect of E2 was further ascertained with the findings that E2-pretreated TEC-conditioned medium accelerated the thymocyte apoptosis. Nevertheless, exposure of thymocytes to E2 was seen to be inevitable for the apoptotic action of TEC-secreted paracrine factors. In the presence of TEC, a positive reaction for caspase-3, -7 and -9 and enzyme substrate, poly(ADP-ribose) polymerase (PARP) in response to E2 suggests the caspase-dependent thymocyte apoptosis in the wall lizard Hemidactylus flaviviridis. Further, E2 was shown to act through genomic pathway, since the receptor antagonist tamoxifen and transcription/translation inhibitors blocked its apoptotic action. Interestingly, the apoptotic effect of E2 was effectively decreased by progesterone.

Animals↗