Met-enkephalins in patients with inflammatory bowel diseases.

PURPOSE: Opioid peptides provide a link between the neuroendocrine and immune systems. They modify the inflammatory process through their effect on the synthesis and secretion of cytokines and on the proliferation of leukocytes to the inflammatory lesion. The evaluation analyzed changes in free met-enkephalin concentration values in the serum and colon mucosal biopsy specimens of patients with inflammatory bowel disease (IBD). MATERIAL AND METHODS: In serum and colon mucosal biopsy specimens, free met-enkephalin levels were determined in 43 patients with ulcerative colitis (UC) and 38 individuals with Crohn's disease (CD). The evaluation analyzed the effect of disease activity, inflammatory lesions of the colon and laboratory parameters, on the level of the investigated marker. The control group consisted of 45 healthy volunteers. RESULTS: Serum free met-enkephalin levels were depressed in patients with CD (85.4pg/ml) and UC (101.5pg/ml) as compared to the controls (119.4pg/ml). Met-enkephalin levels in colonic biopsies collected from inflammatory lesions in IBD patients were significantly higher as compared to sections without inflammatory lesions (6.59pg/mg vs. 2.89pg/mg, p < 0.01 in the CD group and 6.12pg/mg vs. 3.47pg/mg, p < 0.05 in the UC group) and their level correlated with disease activity. CONCLUSIONS: The present investigation is the first study that demonstrates changes in free met-enkephalin levels in IBD that may play a role in the pathogenesis and course of the disease. Further studies are necessary to assess the anti-inflammatory effect of opioid peptides.

Met-enkephalin in the liver as a marker of hepatocellular damage in chronic viral hepatitis type B and C.

PURPOSE: The aim of the study was to assess the liver Met-enkephalin concentration in chronic viral hepatitis type B and C as well as in liver cirrhosis in order to estimate the role of opioid system in pathogenesis of liver disease. MATERIAL AND METHODS: The concentration of Met-enkephalin was examined in liver tissue of 103 consecutive patients with chronic hepatitis type B and C. Control group consisted of uninfected patients. Met-enkephalin concentration was analyzed in relation to the degree of hepatic necroinflammatory activity and the extent of fibrosis estimated by histopathological examination of liver bioptates and compared to such parameters as age, sex and concomitant diseases. RESULTS: Significant differences in Met-enkephalin concentration were found between cases with advanced fibrosis (stage 3 and 4 acc. to Batts and Ludwig classification) and cases with fibrosis classified as stage 2 (p < 0.05). Met-enkephalin concentration was higher in HCV infected patients in comparison to HBV infected patients (p < 0.05) and uninfected controls (0.05 < p < 0.1). There wasn't found any correlation between Met-enkephalin level and necroinflammatory activity in the liver, age, sex and concomitant diseases. CONCLUSIONS: Met-enkephalin concentration measurement in the liver tissue seams to be a useful method for differentiation of stage 2 from stages 3 and 4 of severe liver fibrosis. There is increased concentration of Met-enkephalin in liver tissue in HCV infected patients in comparison to HBV infected or uninfected individuals. The degree of necroinflammatory activity in the liver as well as sex and age of patients with chronic hepatitis do not correlate with changes in opioid system.

Morphine-induced changes in the activity of proopiomelanocortin and prodynorphin systems in zymosan-induced peritonitis in mice.

We have shown that supplementation of proinflammatory agent with a high dose of morphine not only abolishes inflammation-related pain symptoms but also inhibits influx of leukocytes to the inflamed peritoneal cavity. Present investigations focused on effects of morphine on proopiomelanocortin and prodynorphin systems during zymosan-induced peritonitis. Males of SWISS mice were ip injected with zymosan (Z, 40 mg/kg) or zymosan with morphine (ZM, 20 mg/kg). At time 0 (controls) and 4 and 24h after stimulation, peritoneal leukocytes (PTLs) were counted, PTL levels of opioid peptides (beta-endorphin and dynorphin) measured by radioimmunoassays, while mRNAs coding their respective precursors (POMC and PDYN) and receptors (MOR and KOR) determined by QRT-PCR. Influx of inflammatory PTLs, mainly PMNs, was significantly delayed by morphine co-injection. Total levels of beta-endorphin and dynorphin corresponded with PTL numbers, while levels per cell were similar in all groups except of beta-endorphin, decreased in ZM at 4h. Levels of both peptides in peritoneal fluid were increased in Z and ZM groups at 4h, while at 24h only in case of beta-endorphin in Z group. POMC was increased only in ZM group at 4h of peritonitis, while PDYN in both Z and ZM groups at the same time. MOR mRNA was increased 24h after injection in Z and ZM groups, while KOR mRNA was similar in all groups except of decrease in Z at 24h. In conclusion, endogenous opioids and their receptors are involved in zymosan-induced peritonitis and affected in various ways by morphine co-injection.

Effect of tianeptine and fluoxetine on the levels of Met-enkephalin and mRNA encoding proenkephalin in the rat.

The purpose of the present study was to evaluate the effects of acute and repeated treatment with two antidepressant drugs (ADs) of opposite pharmacological profile, i.e. tianpetine (TIA, serotonin reuptake enhancer) and fluoxetine (FLU, serotonin reuptake inhibitor) on the levels of Met-Enkephalin, (Met-Enk, a member ofopioid peptide family, which has been suggested to play a role in the mechanism of action ADs) as well as on mRNA coding for proenkephalin (mRNA PENK) in various regions of the rat brain, pituitary, adrenal glands and plasma. Male Wistar rats were treated acutely or repeatedly (10 mg/kg p.o., twice daily for 14 days) with TIA or FLU. Tissue for biochemical experiments was taken 2 h after last dose of appropriate drug. The levels of Met-Enk were estimated by radioimmunoassay, mRNA PENK was measured using in situ hybridization. From the results obtained in the present study it may be concluded that repeated administration of TIA or FLU induced similar changes in the levels of Met-Enk in the rat hippocampus, striatum, hypothalamus and neurointermediate lobe of pituitary. Such an effect is interesting, especially if one takes into account the differences in pharmacological profile between these two antidepressant drugs. It may be suggested that serotonin level might not be crucial for inducing the alterations in the content of Met-Enk. Since we did not observe any changes in the levels of PENK mRNA in the studied rat brain regions after repeated administration of TIA or FLU, it seems that the observed changes in the levels of Met-Enk do not result from effects of these antidepressants on biosynthesis of PENK, but rather from alterations in the peptide release. Another interesting finding of the present study was that in the anterior lobe of pituitary, adrenal glands and plasma, repeated administration of TIA induced alterations in the contents of Met-Enk, while repeated administration of FLU remained without any effect. It is tempting to speculate that such a differentiation between the effects of these two antidepressants might be linked to the well known feature of TIA (but not FLU) which has been shown to reduce both basal and stress-evoked activity of the hypothalamic-pituitary-adrenal (HPA) axis.

Acute reserpine treatment alters the catecholamine, glucocorticoid and opioid response to walking exercise in sheep.

The aim of the study was to examine the effects of reserpine on the plasma levels of adrenaline, noradrenaline, cortisol and alpha-neoendorphin in sheep under control conditions and during walking exercise. One hour of walking (5 km/h) caused a significant increase in both catecholamines and cortisol between 10 and 30 min of stress, and transiently decreased the level of alpha-neoendorphin at the same time. Reserpine at the dose of 0.3 mg/kg i.v. given before stress significantly lowered the basal levels of all tested parameters. A combination of the reserpine and walking exercise significantly attenuated the stress-induced changes in the plasma level of measured hormones.

The kappa-opioid receptor is present in porcine ovaries: localization in granulosa cells.

Opioid peptides derived from several genes are present in the reproductive system and are known to be involved in the regulation of reproduction. This work shows that the level of alpha-neoendorphin which derives from prodynorphin changes in folliculogenesis. The lowest level was observed in the small follicles, increased in the medium, and reached the highest level in the preovulatory follicles. The presence of the kappa-opioid receptor in granulosa cells, obtained from follicles of varying sizes was demonstrated. The monoclonal anti-kappa-opioid receptor antibody was used to reveal the expression of kappa-opioid receptor. The results obtained suggest changes in alpha-neoendorphin levels and kappa-opioid receptor density during maturation of the follicles.

Responses of heat stressed chickens to exogenous reverse triiodothyronine (rT3).

Heat stress is accompanied by a decrease in basal metabolic rate and plasma thyroid hormones. Unlike 3,5,3'-triiodothyronine (T3) and thyroxine (T4), 3,3',5'-triiodothyronine (rT3) displays hypometabolic properties and antagonizes the hypermetabolic effect of T3. This study analyses the role of rT3 in heat (38-39 degrees C) stressed immature chickens. Two experiments which differed in frequency of rT3 injections (one or two times a day), duration of heat stress (72 or 48 h) and blood sampling were performed. The dose was 14 micrograms rT3/100 gb.wt./injection (s.c.). It has been shown that rT3 treatment aggravates heat stress symptoms (enhances circulating corticosterone, catecholamines and free fatty acids) and increases mortality. The critical survival time of the rT3 treated and heated birds was at first 24 h of stress. No more chickens died during the next days of the experiment despite the continuation of rT3 injection, suggesting that rT3 might disturb the adaptation to heat. Reverse T3 in heat stressed chickens led to the highest reduction in food consumption (69.9%) and body weight gain (14.0% compared to initial weight). The opposite effect in water consumption (216.9%) was observed. In a neutral environment, rT3 significantly suppressed body temperature 6 h after injection (40.4; control; 41.1 degrees C), confirming its hypometabolic properties. However, at the same time rT3 significantly enhanced body temperature in heat stress (43.03 versus heated control 42.56 degrees C). In addition, in rT3 treated birds decreased plasma triglycerides (TG; 24.3%) and increased plasma free fatty acids (FFA; neutral temperature; 26.4% heat stress: 57%) were demonstrated. A correlation between corticosterone and FFA (r = 0.52) shows that some of the FFA may originate from lipolysis since hormones of the pituitary-adrenocortical axis accelerate lipolysis. The remaining part of the increased FFA appears to be due to suppressed utilization of FFA as a consequence of hypometabolic properties of rT3. Low and negative relation between TG and FFA (r = -0.26; P < 0.05) may support such an assumption. The two times higher peak of corticosterone in the rT3 and the overheated group, as compared to the heated control, occurred at 6 h of heat stress and indicates that rT3 increases the unfavourable effect of high temperature. This was also confirmed by elevated plasma adrenaline and noradrenaline in rT3-injected and heated chickens (55.5 and 120%, respectively). However, a single and two times higher peak of adrenaline at 24 h of heat stress was observed in saline treated birds, but not in rT3 supplemented animals, suggesting that this difference might explain one of the factors responsible for high mortality. In conclusion, the results obtained demonstrate that physiological doses of rT3, a hypometabolic hormone, enhance the unfavourable effect of heat stress in chickens.

Endogenous opioids may modulate the activity of the hypothalamus-pituitary-adrenocortical axis in domestic fowl.

The activity of the hypothalamus-pituitary-adrenocortical (HPA) axis is under complex neuronal, hormonal and peptidergic control. In order to determine the role of the endogenous opioids in the modulation of the HPA axis in hens we have examined the changes of the plasma levels of Met-enkephalin, alpha-neo-endorphin, catecholamines and corticosterone during rest, stress (30 min of overcrowding) and after naltrexone pretreatment. Short overcrowding induced an increase of all parameters, but the time of response and duration of elevation was different. Plasma Met-enkephalin showed a biphasic response: decrease followed by increase at the end of stress. Catecholamines and corticosterone peaked at 30 min of overcrowding and the highest level of alpha-neo-endorphin has been noticed at 20 min after the experiment was started. Naltrexone (2 mg/kg b.w., i.v) diminished the responses to stress of all parameters and did not change their basal levels, except for corticosterone. These results indicate that the endogenous opioids may take part in the mediation of HPA activity in hens during resting and stressful situations.

Effects of nicotine on the concentration of native and cryptic Met- and Leu-enkephalin in peripheral tissues.

The distribution of the native pentapeptides, Met- and Leu-enkephalin, and their cryptic forms (larger enkephalin-containing peptides) in adrenal medulla, spleen, lung, salivary gland, vas deferens, heart, duodenum and jejunum were determined by radioimmunoassay. The proportion of total Met- and Leu-enkephalin represented by native pentapeptide varied markedly among these tissues. Also, the distribution of native and cryptic Met-enkephalin was distinct from that of Leu-enkephalin. Repeated short-term administration of nicotine, 0.1 mg/kg i.p. six times at 30 min intervals, produced significant changes in native and cryptic Met-enkephalin in adrenal medulla, jejunum, vas deferens, spleen and heart. This regimen of nicotine also affected the concentration of Leu-enkephalin in adrenal medulla, jejunum and spleen.