Lucia Kšinantová

Downregulation of metastasis suppressor genes in malignant pheochromocytoma

There is no reliable method currently available to predict malignant potential of pheochromocytoma based on conventional histology or genetic, molecular or immunohistochemical markers. Metastasis suppressor genes affect the spread of several cancers and, therefore, may provide promise as prognostic markers or therapeutic targets for malignant pheochromocytoma. We hypothesized that the downregulation of metastasis suppressor genes in malignant pheochromocytoma may play a role in malignant behavior. We applied quantitative real‐time polymerase chain reaction (QRT‐PCR) to 11 metastasis suppressor genes. These genes are known to be involved in the regulation of important cancer‐related cellular events, such as cell growth regulation and apoptosis (nm23‐H1, TIMP‐1, TIMP‐2, TIMP‐3, TIMP‐4, TXNIP and CRSP‐3), cell–cell communication (BRMS‐1), invasion (CRMP‐1) and cell adhesion (E‐Cad and KiSS1). The study included 15 benign and 10 malignant pheochromocytomas. Six metastasis suppressor genes (nm23‐H1, TIMP‐4, BRMS‐1, TXNIP, CRSP‐3 and E‐Cad) were downregulated significantly in malignant compared to benign pheochromocytoma (p < 0.05, Mann‐Whitney U‐test). We applied a non‐linear rule using median malignant value (MMV) as a threshold to use metastasis suppressor genes to distinguish malignant from benign samples. After cross‐validation, the non‐linear rule produced no errors in 10 malignant samples and 3 errors in the 15 benign samples, with an overall error rate of 12%. These results suggest that downregulation of metastasis suppressor genes reflect malignant pheochromocytoma with a high degree of sensitivity. Thus, we conclude that altered function of these metastasis suppressor gene pathways may play an important role in the malignant behavior of pheochromocytoma. Published 2004 Wiley‐Liss, Inc.

Increased thyroid volume, prevalence of thyroid antibodies and impaired fasting glucose in young adults from organochlorine cocktail polluted area: Outcome of transgenerational transmission?

In 137 females (F) and 94 males (M) aged 21-35 years from organochlorines (OCs) polluted area (POLL) increased thyroid volume (ThV), prevalence of antibodies to thyroperoxidase (TPOab), thyrotropin receptor (TRab) and of impaired fasting glucose (IFG) was found compared to 116 F and 107 M from background pollution area (BCGR). In F and M from POLL also strikingly increased level of PCBs, DDE and HCB was found. Such findings were compared to the generation of their parents aged 41-55 years consisting of 320F/213M from POLL and 406F/231M from BCGR. However, in spite of strikingly lower level of those OCs in young adults from POLL, they showed about the same prevalence of adverse health signs as the old generation. From such reason 44 young F and 40 young M with lowest PCBs level from POLL were selected to obtain nearly the same PCB level as found in all young F and M from BCGR. In such PCB adjusted groups the prevalence of TPOab, TRab, IFG and increased ThV was still significantly higher than that in all young subjects from BCGR. At the same time, also the level of DDE and HCB in such PCBs adjusted groups was considerably lower. It was concluded that such adverse effects in young adults from POLL possibly did not result from their actual OCs levels, but very likely from their exposure to high OCs levels of their mothers during their prenatal and perinatal life. Thus, the data may be compatible with present views on transgenerational transmission of endocrine disruptors action.

Endocrine Regulation of Subcutaneous Fat Metabolism during Cold Exposure in Humans

Abstract: Increased oxidation of carbohydrates and free fatty acids is a well‐known phenomenon during cold stress. Nevertheless, sources of the fuels used have not been fully clarified as yet. Thus, the aim of our study was to evaluate the effect of acute cold exposure on lipid and carbohydrate metabolism in human subcutaneous adipose tissue and to identify the possible regulatory mechanisms involved. Ten volunteers were exposed for 30 min to an ambient temperature of 4°C. Interstitial metabolism was assessed with the aid of the microdialysis technique. Lipolysis intensity was evaluated from changes of glycerol concentration in plasma and in dialysate. Cold exposure induced a significant increase of glycerol concentration both in plasma (by 199 ± 16%, p < 0.01) and in dialysate (by 308 ± 58%, p < 0.001). No changes in glucose concentration were found whether in plasma or in the dialysate. Ethanol concentration in dialysate increased (148 ± 15%, p < 0.01), indicating a slower blood flow in the subcutaneous region. Plasma concentrations of various gluco‐ and/or lipid‐regulatory hormones remained unaffected by the cold exposure, except for norepinephrine, which rose about threefold (309 ± 41%, p < 0.001). The data indicate an important role for subcutaneous adipose tissue in mobilization of free fatty acids during cold exposure. This process seems to be regulated by the sympathetic nervous system, whereas hormones involved in the regulation of lipid metabolism, such as epinephrine, insulin, cortisol, and growth hormone, may play a less significant role—at least under the conditions studied.

The response of endocrine system to stress loads during space flight in human subject

The responses of endocrine system to the exposure to stress-work load and hormonal changes during oral glucose tolerance tests were studied in the Slovak astronaut before (three weeks before flight), during (on the 4th and the 6th days of space flight), and after space flight (1-3 days and 15-17 days after space flight) on board of space station MIR. Blood samples during the tests were collected via cannula inserted into cubital vein, centrifuged in the special appliance Plasma-03, frozen in Kryogem-03, and at the end of the 8-day space flight transferred to Earth in special container for hormonal analysis. Preflight workload produced an increase of plasma norepinephrine and a moderate elevation of epinephrine levels. Plasma levels of insulin, growth hormone, prolactin and cortisol were not markedly changed immediately or 10 min after the end of work load. The higher increases of plasma growth hormone, prolactin and catecholamine levels were noted after workload during space flight as compared to preflight response. The higher plasma glucose and insulin levels were noted during the oral glucose tolerance test in space flight and also in the post flight period. Plasma epinephrine levels were slightly decreasing during glucose tolerance test; however, plasma norepinephrine levels were not changed. The similar patterns of catecholamine levels during glucose tolerance test were found when compared the preflight, in-flight and post flight values. These data demonstrate the changes of the dynamic responses of endocrine system to stress-work and metabolic loads during space flight in human subject.

Somatotropic, lactotropic and adrenocortical responses to insulin-induced hypoglycemia in patients with rheumatoid arthritis.

Abstract: Neuroendocrine mechanisms have been suggested to play an important role in the onset and progression of rheumatoid arthritis (RA). The aim of this study was to evaluate hypothalamic‐pituitary functions in RA patients by measurement of hormone responses to insulin‐induced hypoglycemia. Insulin‐hypoglycemia (Actrapid HM 0.1 IU/kg, i.v. as a bolus) was induced in 17 male patients and in 11 age‐, gender‐, and weight‐matched healthy subjects. Concentrations of growth hormone (GH), prolactin (PRL) and cortisol were analyzed in plasma. PRL release after thyreoliberin stimulation (TRH, 200 g, i.v.) was determined in 21 patients with active forms of RA and in 12 control subjects to evaluate pituitary lactotropic response. In RA patients, basal concentrations of glucose, GH, PRL, and cortisol were in the normal range and they were comparable to those in the control group. Stress of hypoglycemia induced significant elevation of GH, PRL, and cortisol concentrations in all groups. Cortisol responses to hypoglycemia were comparable in patients and in control subjects. GH release during hypoglycemia was increased (p < 0.05) and PRL response was attenuated (p < 0.05) in RA patients versus control subjects. After TRH administration, PRL response was the same in patients as in healthy subjects. In conclusion, the present study revealed an altered hypothalamic‐pituitary function in patients with RA, namely, an enhanced somatotropic and reduced lactotropic activation in response to insulin‐induced hypoglycemia. Basal hormone levels and cortisol release during hypoglycemia were similar to those in healthy subjects.

Changes in gene expression of phenylethanolamine N-methyltransferase in the transplanted human heart.

Abstract: Heart transplantation (HTx) is an accepted treatment for precisely defined patients with chronic congestive heart failure; however, as a result of the procedure, the graft is completely denervated. Our study focused on the catecholamine biosynthetic pathway, that is, the production of epinephrine, which is known to have positive chronotropic and inotropic effects on the heart. mRNA levels of the phenylethanolamine N‐methyltransferase (PNMT), the enzyme catalyzing epinephrine synthesis in myocardial tissue, were determined in 18 patients (0 to 10 yr after HTx). Samples of myocardium were obtained from the right ventricle at the time of a routine endomyocardial biopsy performed for the diagnosis of graft rejection. Results were correlated with the following clinical parameters: heart rate, heart rate variability, blood pressure, graft systolic function, and the presence of the rejection. We observed that heart PNMT mRNA levels were significantly higher during the first 3 yr as compared to longer periods after HTx. Also, a decrease in the average heart rate and an increase in the heart rate variability were documented. Levels of the PNMT mRNA do not correlate with blood pressure, left ventricular systolic function at rest, and rejection. Thus, a gradual decrease of the heart rate and an increase in the heart rate variability after HTx is considered to be a sign of cardiac graft reinervation. We speculate that the increased PNMT transcription in human myocardium in early intervals after HTx reflects “autonomous sympathicotrophy.” A decrease in the PNMT gene expression with the number of years after HTx could be a consequence of the reinnervation process.

Repeated Stress‐Induced Stimulation of Catecholamine Response Is Not Followed by Altered Immune Cell Redistribution

Abstract: Stress response is considered an important factor in the modulation of immune function. Neuroendocrine hormones, including catecholamines, affect the process of immune cell redistribution, important for cell‐mediated immunity. This longitudinal investigation was aimed at evaluating the effect of repeated stress‐induced elevation of catecholamines on immune cell redistribution and expression of adhesive molecules. We assessed the responses of epinephrine (EPI), norepinephrine (NE), cortisol, changes in lymphocytes subpopulations, and percentages of CD11a+, CD11b+, and CD62L+ lymphocytes to a 20‐min treadmill exercise of an intensity equal to 80% of the individuals Vo2max. The exercise was performed before and after 6 weeks of endurance training consisting of a 1‐h run 4 times a week (ET) and after 5 days of bed rest (HDBR) in 10 healthy males. We did not observe any significant changes in the basal levels of EPI, NE, and cortisol in the plasma, nor in the immune parameters after ET and HDBR. The exercise test led to a significant (P < .001) elevation of EPI and NE levels after both ET and HDBR, a significant elevation (P < .01) of cortisol after HDBR, an increase in the absolute numbers of leukocytes, granulocytes, monocytes, CD3+, CD4+, CD8+, CD16+, CD19+ lymphocytes, percentage of CD11a+ and CD11b+ lymphocytes, and to a decrease of CD62L1 before, after ET, and after HDBR. We found comparable changes in all measured immune parameters after ET and HDBR. In conclusion, repeated stress‐induced elevation of EPI and NE was not associated with an alteration in immune cell redistribution found in response to the single bout of exercise.

Effects of Real and Simulated Microgravity on Response of Sympathoadrenal System to Various Stress Stimuli

Abstract: Changes in plasma levels of epinephrine (EPI) and norepinephrine (NE) were investigated in humans exposed to physical exercise (WL), to psychic stressor (mental arithmetic test, MAT), and to oral glucose administration (oGTT) before and during a stay in microgravity (real space flight, SF) or in simulated microgravity (head‐down bed rest, HDBR). A permanent cannula inserted into the cubital vein and a special appliance, Plasma‐03, were used for blood collection, plasma separation, and freezing of samples during SF. Plasma EPI, NE, dihydroxyphenylglycol (DHPG), and dihydroxyphenylalanine (DOPA) levels were measured by the high‐pressure liquid chromatography (HPLC) method. Basal plasma EPI, NE, DHPG, and DOPA levels were found within the range of control values during SF. Preflight WL produced high increase in plasma NE and moderate elevation of plasma EPI, DHPG, and DOPA levels. Exaggerated exercise induced increases in plasma NE, DHPG, EPI, and DOPA levels were demonstrated in real microgravity. A return to preflight responses of sympathoadrenal system was seen after the landing. Plasma EPI, NE, and DHPG responses to MAT were relatively small, but increased during SF. During the oGTT the plasma EPI levels were slightly reduced in microgravity. Similarly as in SF, WL in HDBR was followed by significantly exaggerated responses of plasma catecholamines. These results show that both somatic and psychological stressors are able to induce an increased activation of sympathoadrenal system during SF or simulated microgravity in HDBR.

Effects of Space Flight and −6° Bed Rest on the Neuroendocrine Response to Metabolic Stress in Physically Fit Subjects

Abstract: The aim of this study was to evaluate the association of plasma epinephrine (EPI) and norepinephrine (NE) responses to insulin‐induced hypoglycemia (ITT) 3 weeks before the space flight (SF), on the fifth day of SF, on days 2 and 16 after landing in the first Slovak astronaut, and before and on the fifth day of prolonged bed rest (BR) in 15 military aircraft pilots, aged 33.5 ± 1.4 years, body mass index (BMI) 26.5 ± 0.7 kg/m2, maximal oxygen uptake (VO2max) 55.2 ± 2.4 mL/kg/min, who volunteered for the study. ITT was induced by i.v. administrations of 0.1 IU/kg body weight insulin (Actrapid HM) in a bolus. Insulin administration led to a comparable hypoglycemia in preflight, actual flight conditions, and before and after bed rest. ITT led to a pronounced increase in EPI levels and moderate increase in NE in preflight studies. However, an evidently reduced plasma elevation of EPI was found after insulin administration during SF and during BR. Thus, during the real microgravity in SF and simulated microgravity in BR, ITT activates the adrenomedullary system to less extent that at conditions of the Earths gravitation. Post‐flight changes in EPI and NE did not differ from those of preflight values, since SF was relatively short (8 days) and the readaptation to Earths gravitation was fast. It seems that an increased blood flow in brain might be responsible for the reduced EPI response to insulin. Responses to ITT in physically fit subjects indicate the stimulus specificity of the deconditioning effect of 5 days of bed rest on the stress response.

Effects of Endurance Training on Endocrine Response to Physical Exercise after 5 Days of Bed Rest in Healthy Male Subjects

Abstract: The study was designed to evaluate how a bout of endurance training (ET) influences the endocrine response after head‐down bed rest (HDBR). Eleven healthy males completed the study, which consisted of a 6‐wk ET followed by 5 days of −6° head‐down HDBR. Treadmill exercise at 80% of pretraining maximal aerobic capacity (VO2max) was performed before and after ET as well as after HDBR. ET increased VO2max by 13%. The response of norepinephrine was attenuated after ET and exaggerated after HDBR (P < 0.001). The differences in epinephrine responses were not statistically significant. The responses of cortisol and plasma renin activity (PRA) were unchanged after ET and were enhanced after HDBR (P < 0.001). The response of growth hormone after HDBR was reduced (P < 0.05). Only the change in cortisol response was associated with the increment of VO2max after ET (r= 0.68, P < 0.01). Endurance training failed to completely prevent changes in endocrine responses seen after HDBR. Improvement of physical fitness was associated with an enhancement of the cortisol response to exercise following the period of bed rest.