Stress-induced Levels Research Articles

Effects of prolactin level on burn-induced aberrations in myelopoiesis.

In this study, we sought to determine if prolactin (PRL) had any influence on burn-induced alterations in myelopoiesis and serum IL-6, IL-10, IL-12, IFN-gamma, TNF-alpha, and MCP-1 levels. To do this, we used mice that were PRL normal, PRL deficient, or hyperprolactinemic and had received a 15% total body surface area burn, sham treatment, or no treatment. We performed clonogenic assays of bone marrow cells, and we found that sham treatment significantly decreased monocyte/macrophage (M) colony formation relative to the control group in the PRL-deficient and PRL-normal mice (P < 0.01). Hyperprolactinemia attenuated the sham-induced decrease in M colony formation. Burn injury significantly increased M colony formation relative to the sham group with an equal significance in the PRL-deficient and PRL-normal mice (P < 0.05). We also showed that burn led to a significant increase in GM colony formation relative to the sham group. This burn-induced increase was significant in the PRL-normal (P < 0.05) and the PRL-deficient (P < 0.01) mice. In the PRL-normal mice, burn injury caused a 2.1-fold increase in the GM colony number, whereas in the PRL-deficient mice burn led to a 2.6-fold increase in GM colony number. When comparing the effects of burn injury on colony formation to the control groups, there were no significant differences seen, irrespective of the PRL level. We observed that all of the cytokines studied, with the exception of IL-10, were influenced by either sham treatment, burn injury, or both forms of stress. This stress-induced response occurred most often in animals that were either hypo- or hyperprolactinemic. We conclude that the PRL level was able to influence the sham-induced and burn-induced alterations in GM and M colony formation. Under euprolactinemic conditions, mice exhibited less often with stress-induced serum cytokine level alterations. We did not find any significant correlations with any of the serum cytokine levels and the ability to form colonies. Importantly, the sham treatment led to immune alterations independent of, and sometimes opposite of burn-induced effects.

Thymus-derived glucocorticoids are insufficient for normal thymus homeostasis in the adult mouse

BackgroundIt is unclear if thymus-derived glucocorticoids reach sufficient local concentrations to support normal thymus homeostasis, or if adrenal-derived glucocorticoids from the circulation are required. Modern approaches to this issue (transgenic mice that under or over express glucocorticoid receptor in the thymus) have yielded irreconcilably contradictory results, suggesting fundamental problems with one or more the transgenic mouse strains used. In the present study, a more direct approach was used, in which mice were adrenalectomized with or without restoration of circulating corticosterone using timed release pellets. Reversal of the increased number of thymocytes caused by adrenalectomy following restoration of physiological corticosterone concentrations would indicate that corticosterone is the major adrenal product involved in thymic homeostasis.ResultsA clear relationship was observed between systemic corticosterone concentration, thymus cell number, and percentage of apoptotic thymocytes. Physiological concentrations of corticosterone in adrenalectomized mice restored thymus cell number to normal values and revealed differential sensitivity of thymocyte subpopulations to physiological and stress-inducible corticosterone concentrations.ConclusionThis indicates that thymus-derived glucocorticoids are not sufficient to maintain normal levels of death by neglect in the thymus, but that apoptosis and possibly other mechanisms induced by physiological, non stress-induced levels of adrenal-derived corticosterone are responsible for keeping the total number of thymocytes within the normal range.

PHYSIOLOGICAL CONDITION IN MAGELLANIC PENGUINS: DOES IT MATTER IF YOU HAVE TO WALK A LONG WAY TO YOUR NEST?

Abstract Colony edges, as opposed to interiors, are often considered less advantageous nesting places in colonial species. For temperate-breeding penguins, inland colony edges should be less desirable than other edges, as there are added costs of walking farther inland, and ambient temperatures are higher. During settlement and incubation, we compared body condition and baseline and stress-induced levels of the hormone corticosterone in male Magellanic Penguins (Spheniscus magellanicus) nesting on the sea edge of a colony with those nesting on the inland edge, >800 m from shore. Body condition in both groups was significantly lower during settlement than during incubation, but was similar in both groups within breeding stages. Corticosterone levels were similar between breeding stages and for groups within each breeding stage. While body condition can vary over time, penguins appear to be well buffered to physiological extremes, as they do not show modification of corticosterone levels with variations in ...

Reconstituted High-Density Lipoprotein Exhibits Neuroprotection in Two Rat Models of Stroke

Background: Reconstituted high-density lipoprotein (rHDL) is prepared from apolipoprotein A-I, isolated from human plasma, and soybean-derived phosphatidylcholine and exhibits biochemical and functional characteristics similar to endogenous nascent high-density lipoprotein (HDL). This study tested the hypothesis that pretreatment with rHDL may reduce neuronal damage in 2 experimental rat models of stroke. Methods: In the first model, an excitotoxic lesion was induced by unilateral injection of N-methyl-D-aspartate (NMDA) in the right striatum (excitotoxic lesion model). In the second model, temporary occlusion of the middle cerebral artery (MCA) was attained by inserting a nylon thread through the carotid artery and blood flow was restored 30 min later (MCAo model). In both models, either rHDL (120 mg/kg) or saline (control) were infused over 4 h, starting 2 h before the injection of NMDA or the induction of ischemia, respectively. 24 h after the interventions, the rats were sacrificed and the brains removed for histochemical preparation. The necrotic area was delimited using an image analysis system. In addition, the levels of reactive oxygen species (ROS) in human endothelial (ECV 304) and neuroblastoma (SK-N-BE) cell lines were measured fluorometrically as 2′,7′-dichlorofluorescein fluorescence in the presence and absence of rHDL and under basal and stress-induced conditions. Results: In the excitotoxic lesion and MCAo models, pretreatment with rHDL significantly reduced the brain necrotic area by 61 and 76%, respectively (p < 0.01). Overnight incubation of ECV 304 and SK-N-BE cells with 0.5 mg/ml rHDL decreased basal and stress-induced ROS levels by 73 and 72% (ECV 304) and by 76 and 43% (SK-N-BE), respectively (p < 0.01). Conclusion: These results suggest that rHDL reduces neuronal damage after onset of ischemic stroke, possibly by involving an anti-oxidative mechanism. Thus, rHDL may be a powerful neuroprotective tool for the treatment of cerebrovascular diseases.

Piezospectroscopic studies of the re-orientation process of defect complexes

The stress-induced electronic level splitting pattern can provide information regarding the symmetry of a defect complex. If the components of the piezospectroscopic tensor are known it is also possible to deduce information about the lattice relaxation in the vicinity of the defect. Based on a complete piezospectroscopic analysis of the vacancy–oxygen complex in silicon we confirm that this defect has an orthorhombic symmetry in the stable configuration while in the unstable configuration of the saddle point on the reconfiguration trajectory we demonstrate that it has trigonal symmetry. The piezospectroscopic description of static defects is, in some cases, inadequate to deal with some defect complexes when they are observed at relatively high temperatures. This is often the case for the high-resolution Laplace deep level transient spectroscopy (DLTS) measurements. At very low temperatures the vacancy–oxygen–hydrogen complex has monoclinic symmetry while at the temperatures used by us for Laplace DLTS measurements the observed symmetry is orthorhombic due to fast hydrogen jumps. A similar re-orientation (position averaging) process occurs for the divacancy in silicon. However, this is true only for the divacancy in the single negative charge state while the symmetry of the complex in the double negative state is stable trigonal. We discuss how these re-orientation processes can be identified from the observed piezospectroscopic parameters for both cases.

The inhibitory effect of ginseng saponins on the stress-induced plasma interleukin-6 level in mice

The effect of ginseng saponins on plasma interleukin-6 (IL-6) in non-stressed and immobilization-stressed mice were investigated. Ginseng total saponins, ginsenosides Rb2, Rg1 and Rd administered intraperitoneally attenuated the immobilization stress-induced increase in plasma IL-6 level. But, intracerebroventricular injection of each ginsenoside did not affect plasma IL-6 level induced by immobilization stress. Ginsenosides Rb2, Rd and Rg1 significantly decreased norepinephrine and/or epinephrine-induced increase of IL-6 level in macrophage cell line (RAW 264.7). Thus, it can be suggested that the inhibitory action of ginseng saponins against the immobilization stress-induced increase of plasma IL-6 level would be in periphery; at least in part, mediated by blocking norepinephrine- and/or epinephrine-induced increase of IL-6 level in macrophage rather than in the brain. Ginseng saponins might be proposed as a possible candidate in the research or therapeutic modulation of stress-related disorders.

The inhibitory effect of ginseng saponins on the stress-induced plasma interleukin-6 level in mice

The effect of ginseng saponins on plasma interleukin-6 (IL-6) in non-stressed and immobilization-stressed mice were investigated. Ginseng total saponins, ginsenosides Rb2, Rg1 and Rd administered intraperitoneally attenuated the immobilization stress-induced increase in plasma IL-6 level. But, intracerebroventricular injection of each ginsenoside did not affect plasma IL-6 level induced by immobilization stress. Ginsenosides Rb2, Rd and Rg1 significantly decreased norepinephrine and/or epinephrine-induced increase of IL-6 level in macrophage cell line (RAW 264.7). Thus, it can be suggested that the inhibitory action of ginseng saponins against the immobilization stress-induced increase of plasma IL-6 level would be in periphery; at least in part, mediated by blocking norepinephrine- and/or epinephrine-induced increase of IL-6 level in macrophage rather than in the brain. Ginseng saponins might be proposed as a possible candidate in the research or therapeutic modulation of stress-related disorders.

Effects of corticosterone on muscle mitochondria identifying different sensitivity to glucocorticoids in Lewis and Fischer rats.

Previous studies in rat have demonstrated decreased number of mitochondria and uncoupling of oxidative phosphorylation after administration of glucocorticoids but at supraphysiological doses and using synthetic glucocorticoids. To analyze the relationships between corticosterone levels (the natural glucocorticoid in rat) and muscle mitochondrial metabolism, Lewis and Fischer 344 rats were bilaterally adrenalectomized and implanted with different corticosterone pellets (0, 12, 50, 100, and 200 mg of corticosterone). Rats bearing a corticosterone pellet delivering corticosterone at concentrations in the range of chronic stress-induced levels presented a lower amount of functional muscle mitochondria with a decrease in cytochrome c oxidase and citrate synthase activities and a depletion of mitochondrial DNA. Moreover, a strain difference in tissue sensitivity to corticosterone was depicted both in end-organ sensitive to glucocorticoids (body, thymus, and adrenal weights) and in muscle mitochondrial metabolism (Lewis > Fischer). Interestingly, this strain difference was also observed in the absence of corticosterone, with a deleterious effect on muscle mitochondrial metabolism in Fischer rats, whereas no effects were observed in Lewis rats. We therefore postulate that corticosterone is necessary for muscle mitochondrial metabolism exerting its effects in Fischer rats with an inverted U curve, whereby too little (only Fischer) or too much (Fischer and Lewis) corticosterone is deleterious to muscle mitochondrial metabolism. In conclusion, we propose a general model of coordinate regulation of mitochondrial energetic metabolism by glucocorticoids.

Differential mechanisms for regulation of the stress response across latitudinal gradients.

We examined plasticity of the stress response among three populations of the white-crowned sparrow (Zonotrichia leucophrys). These populations breed at different elevations and latitudes and thus have breeding seasons that differ markedly in length. We hypothesize that in populations where birds raise only one or rarely two broods in a season, the fitness costs of abandoning a nest are substantially larger than in closely related populations that raise up to three broods per season. Thus individuals with short breeding seasons should be less responsive to stressors and therefore less likely to abandon their young. In our study, baseline and handling-induced corticosterone levels were similar among populations, but corticosteroid-binding globulins differed, leading to a direct relationship between stress-induced free corticosteroid levels and length of breeding season. There were also population-specific differences in intracellular low-affinity (glucocorticoid-like) receptors in both liver and brain tissue. Although investigations of population-based differences in glucocorticoid secretion are common, this is the first study to demonstrate population-level differences in binding globulins. These differences could lead to dramatically different physiological and behavioral responses to stress.

11beta-hydroxysteroid dehydrogenase complementary deoxyribonucleic acid in rainbow trout: cloning, sites of expression, and seasonal changes in gonads.

11beta-Hydroxysteroid dehydrogenases (11beta-HSDs) are important steroidogenic enzymes for catalyzing the interconversion of active glucocorticoid (cortisol and corticosterone) and inert 11-keto forms (cortisone and 11-dehydrocorticosterone) in mammals. In teleosts, 11beta-HSD also plays a role in the production of the predominant androgen, 11-ketotestosterone, in male fish. In this study we cloned cDNAs encoding rainbow trout 11beta-HSD (rt11beta-HSD) from testes and head kidney. The predicted amino acid sequence, hydrophobicity analysis, and transient transfection assays with rt11beta-HSD in HEK293 cells showed that rt11beta-HSD is a homolog of mammalian 11beta-HSD type 2. rt11beta-HSD transcripts are present in steroidogenic tissues and in a number of other tissues. Strong in situ hybridization signals for rt11beta-HSD transcripts were found in Leydig cells of testes, in thecal cells of the early vitellogenic ovarian follicles, and in thecal and granulosa cells of the midvitellogenic and postovulatory follicles. Weaker signals were also found in head kidney interrenal cells from juvenile rainbow trout. Seasonal changes in rt11beta-HSD transcripts in testes showed a pattern similar to that of stress-induced serum cortisol levels, but not to serum androgen levels. High levels of rt11beta-HSD transcripts were found in ovarian follicles from late vitellogenesis through ovulation. These results raise the possibility of a role for rt11beta-HSD in the protection of developing gonads from the inhibitory effects of stress-induced cortisol.

Interactions between ecology, demography, capture stress, and profiles of corticosterone and glucose in a free-living population of Australian freshwater crocodiles

In this study we examined three aspects pertaining to adrenocortical responsiveness in free-ranging Australian freshwater crocodiles ( Crocodylus johnstoni). First, we examined the ability of freshwater crocodiles to produce corticosterone in response to a typical capture-stress protocol. A second objective addressed the relationship between capture stress, plasma glucose and corticosterone. Next we examined if variation in basal and capture- stress-induced levels of plasma corticosterone was linked to ecological or demographic factors for individuals in this free-ranging population. Blood samples obtained on three field trips were taken from a cross-sectional sample of the population. Crocodiles were bled once during four time categories at 0, 0.5, 6, and 10 h post-capture. Plasma corticosterone increased significantly with time post-capture. Plasma glucose also significantly increased with duration of capture-stress and exhibited a positive and significant relationship with plasma corticosterone. Significant variation in basal or stress induced levels of corticosterone in crocodiles was not associated with any ecological or demographic factors including sex, age class or the year of capture that the crocodiles were sampled from. However, three immature males had basal levels of plasma corticosterone greater than 2 standard deviations above the mean. While crocodiles exhibited a pronounced adrenocortical and hyperglycaemic response to capture stress, limited variation in adrenocortical responsiveness due to ecological and demographic factors was not evident. This feature could arise in part because this population was sampled during a period of environmental benigness.

Plasma corticosterone in American kestrel siblings: effects of age, hatching order, and hatching asynchrony

Although it is well documented that hatching asynchrony in birds can lead to competitive and developmental hierarchies, potentially greatly affecting growth and survival of nestlings, hatching asynchrony may also precipitate modulations in neuroendocrine development or function. Here we examine sibling variation in adrenocortical function in postnatally developing, asynchronously hatching American kestrels ( Falco sparverius) by measurements of baseline and stress-induced levels of corticosterone at ages 10, 16, 22, and 28 days posthatching. There was a significant effect of hatching order on both baseline and stress-induced corticosterone levels during development and these effects grew stronger through development. First-hatched chicks exhibited higher baseline levels than later-hatched chicks throughout development and higher stress-induced levels during the latter half of development. Furthermore, there was significant hatching span (difference in days between first- and last-hatched chicks) × hatching order interaction on both baseline and stress-induced corticosterone levels during development. Hatching span was also positively correlated with both measures of corticosterone and body mass in first-hatched chicks, but was negatively correlated with these factors through most of the development in last-hatched chicks. It is known that hatching asynchrony creates mass and size hierarchies within kestrel broods and we suggest that hierarchies in adrenocortical function among siblings may be one physiological mechanism by which these competitive hierarchies are maintained.

Systemic apomorphine alters HPA axis responses to interleukin-1β adminstration but not sound stress

Apomorphine is a dopamine receptor agonist that was recently licensed for the treatment of erectile dysfunction. However, although sexual activity can be stressful, there has been little investigation into whether treatments for erectile dysfunction affect stress responses. We have examined whether a single dose of apomorphine, sufficient to produce penile erections (50 μg/kg, i.a.), can alter basal or stress-induced plasma ACTH levels, or activity of central pathways thought to control the hypothalamic-pituitary-adrenal axis in rats. An immune challenge (interleukin-1β, 1 μg/kg, i.a.) was used as a physical stressor while sound stress (100 dB white noise, 30 min) was used as a psychological stressor. Intravascular administration of apomorphine had no effect on basal ACTH levels but did substantially increase the number of Fos-positive amygdala and nucleus tractus solitarius catecholamine cells. Administration of apomorphine prior to immune challenge augmented the normal ACTH response to this stressor at 90 min and there was a corresponding increase in the number of Fos-positive paraventricular nucleus corticotropin-releasing factor cells, paraventricular nucleus oxytocin cells and nucleus tractus solitarius catecholamine cells. However, apomorphine treatment did not alter ACTH or Fos responses to sound stress. These data suggest that erection-inducing levels of apomorphine interfere with hypothalamic-pituitary-adrenal axis inhibitory feedback mechanisms in response to a physical stressor, but have no effect on the response to a psychological stressor. Consequently, it is likely that apomorphine acts on a hypothalamic-pituitary-adrenal axis control pathway that is unique to physical stressors. A candidate for this site of action is the nucleus tractus solitarius catecholamine cell population and, in particular, A2 noradrenergic neurons.

A small nuclear RNA, hdm365, is the major processing product of the human mdm2 gene.

mdm2 encodes for an E3 ubiquitin ligase targeting constitutively expressed p53 for proteasomal degradation. Several protein isoforms have been described for human MDM2 (HDM2), some of which may correspond to splicing variants detectable by RT-PCR in many tumors. Upon cellular stress, p53 becomes resistant to MDM2 and, in a feedback loop, up-regulates mdm2 transcription. The physiological relevance of stress-induced mdm2 gene activity is not well understood. We describe a small nuclear RNA of 365 bases comprised of the first five hdm2 exons and lacking polyadenylation. hdm365 precedes full-length hdm2 RNA expression after induction by p53 and accumulates to significant levels in the nucleus, detectable at the site of hdm2 transcription and processing only. Considering a 10-fold lower stability and high steady-state levels of the novel RNA species, hdm365 appears to be the major processing product of hdm2 transcripts. hdm365 induction was observed after ectopic expression of p53 and after DNA damaging treatment of tumor cell lines, primary fibroblasts and lymphocytes, and was not related to apoptosis. Corresponding truncated transcripts were observed in hdm2 amplified cells. High stress-inducible expression levels, absence of a corresponding protein, and nuclear localisation of hdm365 suggest a novel RNA-based function for hdm2.

Gender specific effect of neonatal handling on stress reactivity of adolescent rats.

Early neonatal handling of rat pups produces dampened hypothalamic-pituitary-adrenal axis reactivity to stress in adult male offspring. However, less is known about whether there is a similar effect for females. Although, most studies of neonatal handling have examined subsequent effects during adulthood, adolescence is an important developmental stage for stress responsivity. To address these issues, the effect of neonatal handling on the endocrine stress response and brain activity of male and female rats was determined in response to acute restraint stress during adolescence. Consistent with previous findings in adult males, neonatal handling reduced restraint stress-induced hormone levels in adolescent males. However, in contrast, we found elevated plasma hormone concentrations in handled females. A gender-specific handling effect on brain activity was also evident, with significantly increased stress-induced activation of the posterior cingulate cortex of handled females, as measured by c-fos mRNA expression. The striking gender difference in the effect of early neonatal handling provides evidence that this must be considered as an important variable in subsequent stress responsivity induced by early manipulations.

Prolactin enhances production of interferon-gamma, interleukin-12, and interleukin-10, but not of tumor necrosis factor-alpha, in a stimulus-specific manner.

Prolactin, an anterior pituitary hormone, has been shown to have a role in immunomodulation. Some reports have shown the importance of prolactin in activating lymphocytes and macrophages, while in hyperprolactinemia patients, prolactin was found to decrease lymphocyte activation and natural killer function. In the present work, at physiological (15ng/ml) and stress-induced levels (30ng/ml) of prolactin, interferon-gamma (IFN-gamma) and interleukin (IL)-12 p70 levels, but not of IL-10 and tumor necrosis factor-alpha (TNF-alpha), increased significantly (p<0.05-0.006) in phytohemeagglutinin (PHA)+lipopolysaccharide (LPS)-stimulated whole blood. However, no such effect was observed at high concentrations of prolactin (100-300ng/ml). In addition, 15ng/ml of prolactin reversed hydrocortisone suppressive effect on IFN-gamma, IL-12 p70, and IL-10 production in PHA+LPS-stimulated whole blood. On the other hand, in LPS-stimulated whole blood, prolactin enhanced significantly (p=0.027) the production levels of IL-10, but not of IFN-gamma, IL-12 p70, and TNF-alpha, in non-concentration-dependent manner. These results suggest that prolactin modulates cytokine response during antigenic response, and this modulation is stimulus specific.

Effects of corticosterone on the proportion of breeding females, reproductive output and yolk precursor levels

In this study we investigated the role of corticosterone (B) in regulating the proportion of laying females, timing of breeding, reproductive output (egg size and number), and yolk precursor levels in chronically B-treated female zebra finches ( Taeniopygia guttata). Corticosterone treatment via silastic implant elevated plasma B to high physiological (stress-induced) levels (24.1±5.3 ng/ml at 7-days post-implantation). B-treated females had high plasma levels of very-low density lipoprotein (VLDL) but low levels of plasma vitellogenin 7-days post-implantation, suggesting that corticosterone inhibited yolk precusor production and perhaps shifted lipid metabolism away from production of yolk VLDL and towards production of generic (non-yolk) VLDL. Only 56% of B-treated females ( n=32) initiated laying, compared with 100% of sham-implanted females ( n=18). In females that did breed, corticosterone administration delayed the onset of egg laying: B-treated females initiated laying on average 14.5±0.5 days after pairing compared to 6.4±0.5 days in sham-implanted females. B-treated females that laid eggs had significantly higher plasma B levels at the 1st-egg stage (45.9±9.0 ng/ml) than did sham-implanted females (7.9±6.8 ng/ml). Despite this there was no difference in mean egg mass, clutch size, or egg composition in B-treated and sham-implanted females. These results are consistent with the idea that elevated corticosterone levels inhibit reproduction, but contrast with studies of other oviparous vertebrates (e.g., lizards) in relation to the role of corticosterone in regulating egg and clutch size.

Corticosterone levels during post-natal development in captive American kestrels ( Falco sparverius)

We investigated post-natal development of the adrenocortical stress-response system in captive American kestrels ( Falco sparverius) by measurements of baseline and stress-induced levels of corticosterone at ages 10, 16, 22, and 28 days post-hatching. Baseline levels of corticosterone increased significantly during post-natal development and although chicks aged 10- and 16-days old exhibited comparable baseline corticosterone levels, those of 22-day-old chicks were significantly higher and those of 28-day-old chicks close to fledging were higher than all younger groups. Chicks in this study exhibited low levels of stress-induced corticosterone early in development and did not exhibit adult-type stress-induced levels of corticosterone until 22 days of age post-hatching. Finally, although baseline and stress-induced levels of 28-day-old birds were significantly higher than one-year-old adults, there was no relationship between baseline corticosterone concentrations and time to nest departure. The fact that baseline levels of corticosterone are low during early development and then increase during later development may be an adaptation to the negative effects of chronically elevated corticosterone levels and as previously noted in other studies may minimize these negative effects on rapid growth and development in young birds, potentially maximizing normal growth. The ability of even young kestrel chicks to elevate corticosterone levels in response to stress suggests that they may be able to physiologically cope with food shortages associated with unpredictable food resources which wild kestrels often face.

The influence of dexamethasone on Hsp70 level and association with glucocorticoid receptor in the liver of unstressed and heat-stressed rats

The aim of the present study was to examine the influence of dexamethasone on the levels of heat shock protein Hsp70 and glucocorticoid hormones receptor as well as on the interaction of these two proteins in the liver cytosol and nuclei of unstressed and rats exposed to whole body hyperthermic stress. The results, obtained by quantitative immunoblotting, have shown that dexamethasone provoked a reduction of Hsp70 basal level and an increase in its stress-induced level in the nuclei, supporting the idea that this hormone may be a factor included in the regulation of Hsp70 level both under normal and stress conditions. The cytosolic reduction and nuclear elevation of the glucocorticoid hormones receptor level by dexamethasone were also observed. Co-immunopurification of Hsp70 and glucocorticoid hormones receptor has revealed that the changes of cytosolic and nuclear levels of the two examined proteins resulted in the changes of their interaction within the respective cellular compartments. Thus, 41 ?C heat stress was shown to cause at least two-fold elevation of Hsp70/GR ratio within the glucocorticoid hormones receptor heterocomplexes both in the presence and in the absence of dexamethasone. The results support the view that glucocorticoid hormones signaling pathway and heat shock system are interrelated.

Tensile stress-dependent collagen XII and fibronectin production by fibroblasts requires separate pathways

The intracellular mechanisms controlling mechano-dependent production of the two extracellular matrix proteins collagen XII and fibronectin were analyzed. Fibroblasts were cultured on either tensed (attached) or released (floating) collagen type-I gels, respectively. Collagen XII and fibronectin production was three- to fivefold higher under tensed than under released conditions. The general inhibitor of tyrosine phosphorylation, genistein (50 μM), and the MAP kinase inhibitor PD98059 (20 μM) selectively reduced collagen XII accumulation by tensed cultures. Addition of PD98059, but not genistein, downregulated tensile stress-induced tyrosine phosphorylation levels of ERK1/2 and focal adhesion kinase. Staurosporine as well as pretreatment with phorbol ester, which constitute means to downregulate classical and novel PKC activity, specifically blocked collagen XII but not fibronectin accumulation in tensed fibroblasts. ERK1/2 phosphorylation levels were not affected by staurosporine treatment. Chronic exposure to the protein kinase C inhibitors bisindolylmaleimide and calphostin C blocked increased production of both fibronectin and collagen XII from cells under tension. The data manifest that the mechano-dependent production of collagen XII and fibronectin requires separate pathways. The FAK-ERK1/2 pathway, a genistein-sensitive tyrosine kinase, and a distinct classical/novel PKC appear selectively required for increased production of collagen XII in cells under tensile stress, whereas fibronectin induction is regulated by a different PKC-dependent pathway.