Search

The significance of plasminogen activation during the tympanic membrane (TM) healing is known mainly from studies performed on knock-out mice. In the previous study, we reported activation of genes coding proteins of plasminogen activation and inhibition system in rat’s TM perforation healing. The aim of the present study was the evaluation of protein products expressed by these genes and their tissue distribution using Western blotting and immunofluorescent method, respectively, during 10-day observation period after injury. Otomicroscopical and histological evaluation were employed to assess the healing process. The expression of urokinase plasminogen activator (uPA) and its receptor (uPAR) were significantly upregulated in the proliferation phase, with subsequent gradual attenuation during remodeling phase of healing process, when keratinocyte migration was weakening. The expression of plasminogen activator inhibitor type 1 (PAI-1) also showed the highest levels during the proliferation phase. The increase of tissue plasminogen activator (tPA) expression was observed during the whole observation period, with the highest activity during the remodeling phase. Immunofluorescence of these proteins was present mainly in migrating epithelium. Our study found that plasminogen activation (uPA, uPAR, tPA) and inhibitory (PAI-1) molecules form a well-structured regulatory system of the epithelial migration that is critical to the healing of TM after its perforation.

Purpose Exaggerated release of proinflammatory mediators during sepsis contributes to inadequate vasodilatation and depressed myocardial contractility, which lead to development of shock and circulatory collapse. The aim of the study was to evaluate the effect of IL-6 and aging on activation of intracellular signaling pathways in the myocardium induced by bacterial lipopolysaccharide (LPS) administration. Material/methods LPS was injected intraperitoneally to male 3- and 24-month old mice with systemic IL-6 gene knock-out (IL-6KO) and the reference strain (WT). LPS was given intraperitoneally in single low (0.1 mg/kg) or high (10 mg/kg) dose, or in two doses (0.1 + 10 mg/kg) with 24-h delay. The expression and phosphorylation of STAT3, ERK1/2, Akt1/2/3 proteins in the left ventricular myocardium was evaluated after 24 h using Western blotting. Results Low LPS dose induced higher STAT3 phosphorylation only in old IL-6KO mice, not affecting ERK1/2 and Akt1/2/3 phosphorylation in any group. High LPS dose upregulated STAT3 phosphorylation similarly in all groups, reduced ERK1/2 expression in young WT mice and upregulated Akt1/2/3 expression and phosphorylation in young IL-6KO mice. Pretreatment with low LPS dose attenuated phosphorylation of STAT3 in both old groups and phosphorylation of Akt1/2/3 in young IL-6KO group. Two-dose approach also significantly potentiated ERK1/2 phosphorylation in both old groups. Conclusions Obtained results show that IL-6 deficiency alters the activity of intracellular signaling pathways: JAK/STAT in old and Akt in young LPS-treated mice. This may indicate that lack of IL-6 attenuates Akt-related cytoprotective effect of pretreatment with low LPS dose in young but not in aged animals.

Chronic peripheral elevation of interleukin 6 (IL-6) in humans is associated with cognitive deficits. 4- and 24-month-old IL-6-deficient C57BL/6J (IL-6KO) and reference wild-type (WT) mice were tested in an object recognition test. Discrimination ratios and recognition indexes were significantly lower in 4-month-old IL-6KO and in 24-month-old WT mice vs 4-month-old WT animals. Their discrimination ratios had negative values and recognition indexes were below 50% indicating inability to differentiate the novel from the familiar object after 1-hour delay. In 24-month-old IL-6KO mice recognition index reached 53.17% indicating that their recognition memory was not worsened with age in comparison with younger IL-6-deficient animals. Results of holeboard and elevated plus maze indicated that this effect was memory specific. Inborn IL-6 deficiency attenuated recognition memory in 4-month-old mice and did not altered recognition memory in aged animals. IL-6 signalling may constitute a target for development of the protection against memory disturbances connected with IL-6 overexpression. Electronic supplementary material The online version of this article (10.1007/s10519-019-09959-6) contains supplementary material, which is available to authorized users.

Introduction Inflammatory mediators play an important role in development and progression of cardiovascular disease. Both adrenergic stimulation and high levels of interleukin-6 (IL-6) indicate an unfavorable outcome in patients with myocardial infarction or heart failure. Understanding the interaction between β-adrenergic stimulation and IL-6 in the myocardium may contribute to developing more effective treatments. The aim of this study was to verify the role of IL-6 in the effects of β-adrenergic stimulation in activating selected intracellular signaling pathways in mouse myocardium. Material and methods Experiments were performed on 12-week-old male mice: 16 C57BL/6JIL6‑/‑TMKopf (IL-6 KO) and 17 C57BL/6J (WT). Animals received intraperitoneal injections of isoproterenol (ISO, 50 mg/kg) or placebo (0.9% NaCl) once a day for 16 days. The phosphorylation of STAT3 (signal transducer and activator of transcription 3), ERK1/2 (extracellular-regulated kinases 1/2), Akt1/2/3, p-38, c-Raf and expression of SOCS3 (suppressor of cytokine signaling 3), PIAS1/3 (protein inhibitors of activated STAT) was assessed by western blotting in the myocardium 24 h after the last injection. Evaluation of gene expression downstream of these pathways was performed by real-time PCR. Results Chronic ISO treatment leads to increased fibrosis of the myocardium in mice lacking IL-6, which is accompanied by increased activity of ERK1/2, p38 and reduced expression of SOCS3. Administration of ISO in IL-6 KO animals intensified gene expression of proteins activated by MAPK/ERK (myc; CEBPB; BMP4; Fasn; Tank), while it reduced expression of genes repressed by ERK 1/2 (Wisp1, Wnt1). Conclusions IL-6 plays an important role in regulating the activation of MAPK pathways in the mouse myocardium in response to chronic β-adrenergic stimulation.

Interleukin 6 (IL6) and p53 are linked by mutual regulatory mechanisms and are both upregulated in aging. The aim of this study was to evaluate the effects of aging and IL6 on expression of p53 in the mouse heart. Male C57BL6/J wild-type and IL6 knockout mice at the age of 4-5 months (young adult) and 24-30 months (old) were used. Myocardial expression of proteins such as p53, p21, Mdm2, and phospho-Akt/Akt was estimated using Western blotting and expression of p53 and p21 mRNA using real-time polymerase chain reaction. Expression of p53 protein was lower in IL6 knockout hearts than in wild-type hearts. Aging caused significant upregulation of p53 protein level; however, it was significantly higher in old wild-type hearts than in old IL6 knockout hearts (p < .05). Similar p53 mRNA levels in all groups implied IL6 influence on age-related proteasomal degradation of p53. Localization of p53 mainly in the extranuclear compartment and lack of p21 upregulation in aged hearts may suggest quenched transcriptional activity of p53 despite increased abundance of p53. We conclude that lack of IL6 attenuates expression of p53 protein in the hearts of young mice and diminishes its accumulation with aging by post-transcriptional mechanisms; however, this is not related to altered phenotype of aging heart.

The significance of interleukin 6 (IL-6) in long-term reference memory was tested in the Morris water maze (MWM) in 4-month-old C57BL/6J IL-6-deficient (IL-6 KO) and control mice. Three-day learning measured by escape latency time to find the hidden platform was comparable in both genotypes. In a single probe trial performed 7 days later, without the platform, latency to the platform site and path length to the target place were significantly shorter (p

Activation of PPARs may be involved in the development of heart failure (HF). We evaluated the relationship between expression of PPARγin the myocardium during coronary artery bypass grafting (CABG) and exercise tolerance initially and during follow-up. 6-minute walking test was performed before CABG, after 1, 12, 24 months. Patients were divided into two groups (HF and non-HF) based on left ventricular ejection fraction and plasma proBNP level. After CABG, 67% of patients developed HF. The mean distance 1 month after CABG in HF was397±85 m versus420±93 m in non-HF. PPARγmRNA expression was similar in both HF and non-HF groups. 6MWT distance 1 month after CABG was inversely correlated with PPARγlevel only in HF group. Higher PPARγexpression was related to smaller LVEF change between 1 month and 1 year (R=0.18,p<0.05), especially in patients with HF. Higher initial levels of IL-6 in HF patients were correlated with longer distance in 6MWT one month after surgery and lower PPARγexpression. PPARγexpression is not related to LVEF before CABG and higher PPARγexpression in the myocardium of patients who are developing HF following CABG may have some protecting effect.

TNFα and PPARγ are important modulators of metabolism, inflammation, and atherosclerosis. Coronary artery disease is the leading cause of heart failure (HF). The aim of the study was to assess whether polymorphisms of the TNFα (-308G>A) and PPARG2 (Pro12Ala) genes are associated with the risk of developing HF by patients with ischemic heart disease. Methods. 122 patients without HF (aged 63 ± 8.8 years, 85% males) with confirmed coronary artery disease qualified for coronary bypass grafting were enrolled in the study. After the procedure, they were screened for cardiac parameters. Those with elevated NT-proBNP or diminished left ventricular ejection fraction during follow-up were assigned to the HF group (n=78), and the remaining ones to the non-HF group (n=44). The TNFα -308G>A and PPARG2 Pro12Ala polymorphisms were detected using the TaqMan method. Results. The distributions of TNFα -308G>A and PPARG2 Pro12Ala did not differ between the HF and non-HF groups (-308G>A: 16% vs. 11.4% of alleles; Pro12Ala: 23.9% vs. 20.5% of alleles, respectively). IL-6 concentration in the plasma of TNFα A-allele carriers at months 1 and 12 after CABG was higher in the HF group compared to the non-HF group (1 month after CABG: 5.3 ± 3.4 vs. 3.1 ± 2.9, pα concentration or other parameters related to HF. Conclusions. Our study did not reveal any correlation between the PPARG2 Pro12Ala and TNFα -308G>A polymorphisms and development of HF in patients with ischemic heart disease after coronary bypass grafting.

TNFα and PPARγ are important modulators of metabolism, inflammation, and atherosclerosis. Coronary artery disease is the leading cause of heart failure (HF). The aim of the study was to assess whether polymorphisms of the TNFα (-308G>A) and PPARG2 (Pro12Ala) genes are associated with the risk of developing HF by patients with ischemic heart disease. Methods. 122 patients without HF (aged 63 ± 8.8 years, 85% males) with confirmed coronary artery disease qualified for coronary bypass grafting were enrolled in the study. After the procedure, they were screened for cardiac parameters. Those with elevated NT-proBNP or diminished left ventricular ejection fraction during follow-up were assigned to the HF group (n=78), and the remaining ones to the non-HF group (n=44). The TNFα -308G>A and PPARG2 Pro12Ala polymorphisms were detected using the TaqMan method. Results. The distributions of TNFα -308G>A and PPARG2 Pro12Ala did not differ between the HF and non-HF groups (-308G>A: 16% vs. 11.4% of alleles; Pro12Ala: 23.9% vs. 20.5% of alleles, respectively). IL-6 concentration in the plasma of TNFα A-allele carriers at months 1 and 12 after CABG was higher in the HF group compared to the non-HF group (1 month after CABG: 5.3 ± 3.4 vs. 3.1 ± 2.9, pA polymorphisms and development of HF in patients with ischemic heart disease after coronary bypass grafting.

Background Our approach was to determine the influence of a single systemic administration of AM281, synthetic cannabinoid structurally similar to SR141716A, on recognition memory in rats. Methods To assess the influence of AM281 on acquisition of information the compound was given intraperitoneally once, at the doses of 0.1, 0.5, 1.0 or 2.0 mg/kg, 15 min before learning trial (T1) and in order to evaluate its influence on consolidation process AM281 was given at indicated doses, immediately after T1 trial in an object recognition test. Since cannabinoids may alter motor function and affect anxiety, the influence of AM281 on psychomotor activity and anxiety was evaluated in an open-field and elevated plus maze test, respectively. Results Administration of AM281 at the doses: 0.1, 0.5 and 1.0 mg/kg significantly improved acquisition of information, while 0.1 and 0.5 mg/kg of AM281 significantly facilitated consolidation process. Not only did AM281 not affect locomotor and exploratory activity, but also anxiety. Conclusion This is the first evidence that AM281 exerts facilitatory effect on recognition memory in rats. This effect seems to be memory specific because no alterations in animals’ psychomotor activity and anxiety were observed.

Objectives Type III collagen plays significant role in skin wound healing, forming provisional matrix guiding the inflammatory cells and fibroblasts into the wound site. Our preliminary study performed on rat's tympanic membrane (TM) using Rat Wound Healing RT2 Profiler PCR Array revealed up-regulated expression of collagen type III α1 chain mRNA also during healing of TM. This study was undertaken to confirm and evaluate collagen type III protein expression and distribution during TM healing process. Methods Sixty rats were used, of which 10 served as controls and the others had their TM perforated. The experimental animals were divided into five subgroups on the basis of time points (03, 06, 09, 14, 20 day after injury). Videootoscopy and histology were employed to assess morphology of the healing process. The expression of collagen type III was evaluated using Western blot analysis and its tissue localization was determined by the immunohistochemical method. Results The expression of collagen type III remained on the same level as in control TM up to day 06. On day 09 abrupt (p = 0.01) increase of the collagen type III expression was observed and it maintained on the same level to the end of observation period. In perforated TM collagen type III was detected in the healing area along the perforation border and around dilated blood vessels. On day 14 and 20 collagen type III was found in the connective tissue filling up the TM previous defect. Conclusions Taking into consideration our recent and previous data, as well as results obtained by other authors, is seems possible that the increase of collagen type III expression in the late stage of TM healing contributes to proper scar formation.

Objectives Our preliminary study performed on perforated rat's tympanic membrane (TM) using Rat Wound Healing RT2 Profiler PCR Array showed significantly increased levels of mRNA for collagens type I and V. Enhanced expression of those genes does not assure that their protein products are indeed present, and in what quantity. Therefore, this study was undertaken to analyze the collagen type I and V content in the healing TM. Methods Sixty rats were used, of which 10 served as controls and the others had their TM perforated. The experimental animals were divided into five subgroups on the basis of time points (03, 06, 09, 14, 20 day after injury). Videootoscopy and histology were employed to assess the morphology of the healing process. The expression of collagen type I and V was evaluated using Western blot analysis. Tissue localization of collagens was determined by the immunofluorescence method. Results The collagen type I expression was three times higher on the third day after injury and remained on that level for whole period of observation, up to day 20. The increase of the collagen type V expression was gradual, reaching the highest level on day 14 following injury. In comparison to the control TM statistically significant increase in the level of expression was observed starting from day 09 to the end of observation period. In healing TM immunofluorescent labeling of collagen type I and V was seen on the surface of remnants of previous lamina propria and in the loose proliferating fibrous tissue. On day 20 immunofluorescence was present mainly on the surface of thin connective tissue layers forming the scar in the place of previous perforation. Conclusion Although the collagens type I and V are present only in subepithelial layer in the normal rat's TM they play significant role in TM healing process.

Significance of interleukin 6 (IL-6) deficiency in cognitive processes was evaluated in 4- and 24-month-old C57BL/6J IL-6-deficient (IL-6 KO) and control (WT) mice in Morris water maze (MWM), holeboard test (HB) and elevated plus maze (EPM). During 3-day learning escape latency time (ELT) was longer in IL-6 KO than in WT mice, however their swimming was slower, floating longer, and path length did not differ. The comparison of ELT and the distance traveled between the first and the third learning day within each group revealed significant decrease of ELT in all groups with the highest difference in 4-month-old WT mice, and significant decrease of distance traveled only in both groups of WT mice. In a single probe trial, performed 24 h after the last learning session, there were no major differences in the absolute values of ELT, but ELT turned out to be significantly shorter in both IL-6 KO groups, when it was compared to the ELT on the last learning day, indicating on better memory retrieval. In HB test only significant increase in number of rearings in aged WT mice, and in EPM significant prolongation of open arm time and higher number of open arm entries in 4-month-old IL-6 KO mice were observed. Results of HB and EPM tests showed that alterations of learning and reference memory observed in MWM were specific to cognition. Attenuation of learning ability in young adult IL-6-deficient mice assessed in MWM suggests that physiological level of IL-6 is involved in mechanisms engaged in proper memory formation, and it may also indicate on the importance of IL-6 signaling in brain development. Maintained on similar level in both 4- and 24-month-old IL-6 KO mice learning ability and its attenuation in 24-month-old vs 4-month-old WT mice indicates on slower age-related memory decline in mice not expressing IL-6. Better performance of IL-6 KO mice in the probe trial points to their reference memory improvement and may also indicate that IL-6 plays a role in mechanism responsible for cognitive flexibility.

Aging is related to gradual increase of interleukin 6 (IL-6) plasma level that affects peroxisome proliferator-activated receptor (PPAR) expression. We evaluated age-related changes in cardiac expression of PPARα, its coactivator PGC-1α, and selected downstream proteins in mice with systemic IL-6 knockout (IL6KO). Male C57BL6/J wild-type (WT) and IL6KO mice were used at the age of 16-20 weeks (young) and 24-30 months (senescent). Echocardiography and electron microscopy were applied to assess the function and ultrastructure of the heart. Western blotting and quantitative real-time PCR were used to estimate protein and mRNA levels of selected genes. PPARα expression in the myocardium of young IL6KO animals is lower and remains unchanged with aging, whereas in WT mice it declines with age and in both senescent groups it is equal. We observed aging-related upregulation of PGC-1α and less pronounced decline of Sirt3 in IL6KO animals; the level of cytochrome C was significantly decreased in IL6KO group only, suggesting disturbed mitochondrial function, which was not sufficient to evoke obvious changes in cardiac performance and function assessed by echocardiography. IL-6 and aging are involved in regulation of PPARα and PGC-1α expression and may influence the mitochondrial function.

Cannabinoids constitute a varied group of lipophilic substances able to infiltrate the blood-brain barrier and influence neuronal processes. Clinical observations supported by experimental data have revealed that these compounds exert a deleterious effect on cognitive processes. The present study was carried out to determine the influence of a single systemic administration of CP55,940, a potent synthetic agonist of cannabinoid receptors, on spatial memory retrieval assessed in a Morris water maze.C57BL/6J male mice were submitted to three consecutive days of training to find a hidden platform in the water maze. CP55,940 was given intraperitoneally once, at doses of 0.025, 0.125 or 0.25mg/kg on the fourth day, 30min before testing memory retrieval, and in separate groups before testing psychomotor activity and anxiety level in a hole-board test.CP55,940 only at the highest dose of 0.25mg/kg significantly altered all parameters used to assess spatial memory. It increased the latency in the first crossing of the former platform location (target area), decreased the number of target area crossings and shortened the time spent in the target quadrant. Moreover, CP55,940 at doses of 0.25 and 0.125mg/kg attenuated motor and exploratory activity in hole-board test.Since the attenuated psychomotor activity after a dose of 0.125mg/kg did not interfere with memory retrieval, we assume that the impairment of spatial memory observed after the highest dose of CP55,940 (0.25mg/kg) was exerted by its influence on cognitive processes, however, the impact on locomotion could not be excluded.

Introduction Tympanic membrane (TM) perforations are commonly seen in clinical practice as a result of trauma or in the course of otitis media. The TM is a unique structure suspended in air which makes its healing processes different than in the skin wounds. The aim of the study was otoscopical and histological evaluation of the rat's TM healing process. Material and methods 56 male Wistar rats were used for the study. Fifty of them had TMs perforated bilaterally using CO 2 laser, additional 6 served as a controls. The animals were sacrificed on either day 1, 2, 3, 6 and 10 post injury. Process of healing was assessed otoscopicaly, subsequently TM were dissected and processed for histological evaluation. Results At day 6 after perforation half and on day 10 all of TM were healed. On the first day, in histological evaluation, focal thickening of the epithelial layer was observed at some distance from the edge of perforation, on the side of annulus. On the following day proliferation of epithelium covering outer surface of TM on the side of the malleus handle and annulus was clearly visible. An eosinophilic mass containing macrophages and granulocytes was seen in front of the migrating epithelium. On day 3–6 migrating epithelium reached the edge of perforation. Proliferation of the connective tissue layer followed the epithelium. Conclusions The present results indicate that the squamous epithelium covering the outer surface of TM constitutes the first layer which restores continuity of TM. The proliferation of the connective tissue occurs in the direct vicinity of the proliferating and migrating epithelium.

Mature cell release takes place through the hypothe tical barrier- the bone marrow-blood barrier. It co nsists of blood vessels and cells that surround them. Such ce lls are, among others, megakaryocytes. They are involved in the release of platelets or other cellular componen ts of blood. The amount and the development of megakaryocytes and thus the maturity of barrier dep ends on many factors. One of them is IL-6, which gene is located on the short arm of chromosome 7 (7p15-p21). The study was performed on wild-type mice (C57B4/6J) and on mice lacking the gene encoding IL-6 (IL-6 KO-C57B4/6J IL-6tm1 Kopf-/-). Bone marrow of all animals in both groups has been collected, fixed and staine d. Megakaryocytes were identified in each microscopic specimen using immunohistochemical reaction with the CD61 antibody. Then all specimens were subjected to histomorphometric analysis. Statistically significa nt differences in the total number and size of maga kariocytes was noted, which affects bone marrow-blood barrier functions. This could be used in the conventional t reatment of cancer, respiratory diseases and coagulation dis orders and in the treatment using nanoparticles.

Background Interleukin 6 (IL-6) may be involved in regulation of cardiac lipid metabolism and mitochondrial function through its influence on peroxisome proliferator-activated receptors (PPARs). In this study we evaluated the impact of the physiological level of IL-6 on the expression of PPARα and PGC-1α in the heart and the effect of lack of this cytokine on high-fat diet (HFD) induced lipotoxicity. Methods Male C57BL6/J wild type (WT) and IL-6 knock-out (IL-6KO) mice were used. 20 animals of each genotype were fed with HFD for 15–18weeks. Cardiac function was assessed using echocardiography and cardiomyocyte ultrastructure was examined using electron microscopy. QT-PCR and Western blotting were applied to estimate the expression of PPARα and PGC-1α at the transcriptional and protein levels. Results At baseline WT and IL-6KO mice had similar size and function of the left ventricle. HFD induced similar left ventricular hypertrophic response in both groups without causing heart failure, but only WT animals had increased resting ejection fraction of the LV. Ultrastructure of HFD groups showed markers of lipotoxicity, that were more pronounced in IL-6KO group. In basal conditions IL-6KO animals had lower PPARα and similar PGC-1α expression as compared to WT. HFD induced downregulation of both PPARα and PGC-1α in WT animals, while in IL-6KO mice this effect was constrained. Conclusion IL-6 is involved in basal regulation of PPARα and PGC-1α expression in cardiomyocytes. The lack of this cytokine promotes high-fat diet induced lipotoxicity but without overt manifestations of cardiac failure.

Purpose In recent years several reports have suggested involvement of interleukin 6 (IL-6) in beta-adrenergic effects on myocardium, particularly in enhancement of STAT3 phosphorylation (downstream signal transducer of IL-6). Here we present a study of isoproterenol effects on hearts of IL-6 deficient mice. Methods Male 12 week old C57Bl6/J mice and age and sex matched mice from IL-6 knockout strain (C57B16/J IL6-/− ) received a single intraperitoneal bolus of either isoproterenol (15 mg/kg) or placebo (0.9% NaCl) and were sacrificed after 1 or 24 hours (n=8 in each group). Another group of mice from both genotypes received a three-day isoproterenol treatment (20 mg/kg every 8h). Activation of STAT3 and MEK/ERK pathways were assessed after a single dose of isoproterenol by means of western blotting. Results After injection of placebo a significantly lower level of STAT3 phosphorylation was observed in IL-6 KO animals. This difference was abolished after isoproterenol both at 1 and 24-hour time points. Isoproterenol produced potent and rapid activation of both STAT3 and MEK/ERK pathways that returned to the levels of placebo treated controls after 24 hours. Lack of IL-6 did not affect phosphorylation of ERKs. Three-day treatment with isoproterenol caused significant increase of indices of RV and LV hypertrophy in both WT and IL-6 KO animals with no significant differences between genotypes. Conclusion IL-6 is not necessary for isoproterenol induced STAT3 phosphorylation, but may affect activation of this pathway by mild non-specific stimuli. Lack of IL-6 does not affect activation of MEK/ERK pathway nor cardiac hypertrophy by beta-adrenergic agonists.

IL-6 is a biologically active cytokine released during exercise by contracting skeletal muscles. It appears to be highly involved in the regulation of muscles energy substrate utilization. Whether an ablation of IL-6 (IL-6 KO) in mice subjected to a single bout of exercise affects lipid and/or glucose metabolism is currently unknown. In the present study we examined fatty acid (FAT/CD36, FABPpm, FATP-1, FATP-4) as well as glucose (GLUT-1, GLUT-4) transporters expression in IL-6 KO mice. In addition, intramuscular glycogen and lipid content was also evaluated. The expression of all fatty acid transporters (FAT/CD36: +25 %; FATP-1: +31 %; FABPpm: +12.7 %; FATP-4: +7.2 %) was increased in muscles from IL-6 KO mice compared to wild type (WT) mice. Accordingly intramuscular lipid content was also increased in these muscles (FFA: +38 %; DAG: +36 % and TAG: +160 %). Interestingly, IL-6 deficiency had only minor effect on glucose transporters expression (GLUT-1: -4 %, and GLUT-4: -5.1 %), with no apparent difference in muscular glycogen content. A single bout of exercise increased the glucose transporters (GLUT-1: +8 %; GLUT-4: +15 %) as well as FA transporters (FAT/CD36: +13 %; FABPpm: +4.5 %; FATP: +2.5 %, FATP-4: +10 %) expression but only in WT skeletal muscles. In muscles from IL-6 KO mice exercise induced changes only in glucose (GLUT-1: +20 %; GLUT-4: +35 %) but not in the content of FA transporters. Concomitantly, IL-6 KO mice displayed shorter time toward exhaustion with more pronounced reductions in intramuscular lipid and glycogen content. We may speculate, that IL-6 deficiency provokes more pronounced glucose utilization over lipid oxidation during a single bout of exhausting exercise.

Previous studies have reported the upregulation of CCN proteins early after acute heart injury. The aim of the present work was to evaluate the expression of the CCN1 and CCN2 proteins and their regulation by angiotensin II in the atrial myocardium of a chronically failing heart. Male adult mice were subjected to ligation of the left coronary artery to produce myocardial infarction (the MI group), and 16 of them were treated for 12 weeks with the AT1 receptor antagonist telmisartan (the MI-Tel group). Sham-operated mice served as controls. The expression of proteins was evaluated by immunohistochemistry 12 weeks after the operation. In shamoperated mice, stainings for CCN1 and CCN2 proteins were positive within atrial cardiomyocytes. CCN1-positive reaction revealed diffused cytoplasmic localization, while CCN2 was present mainly within the perinuclear cytoplasm. CCN1 was upregulated in the MI group, while CCN2 remained at basal level. Telmisartan prevented the upregulation of CCN1 and decreased CCN2 level. We compared the experimental data with the expression of CCN1 and CCN2 proteins in human right atrial appendages. We found an inverse, but not significant, relation between the level of either protein and the left ventricular ejection fraction. This suggests a similar atrial regulation of CCN1 and CCN2 expression also in humans. We conclude that in the murine atria, CCN1 and CCN2 proteins are expressed constitutively. In chronic heart failure, CCN proteins tend to be upregulated, which may be related to the action of angiotensin II.

Ghrelin is a peptide of 28 amino acids that transmits appetite related signals from peripheral organs to the brain. The main source of ghrelin is stomach. The regulation of ghrelin secretion is still unknown. The finding that fasting and food intake, respectively increase and decrease the secretion of ghrelin suggests that this hormone may be a bridge connecting somatic growth with energy metabolism and appears to play an important role in the alteration of energy homeostasis and body weight in pathophisiological conditions. The purpose of this study was the evaluation of gastric ghrelin immunoreactivity and ghrelin plasma concentration in male Wistar rats with hyperthyroidism. Experimental model of hyperthyroidism was induced by intraperitoneal injection of levothyroxine at the dose of 80 microg/kg daily over 21 days. At the end of experiment the animals were anaesthetized, blood was taken from abdominal aorta to determinate plasma ghrelin concentration by RIA and then the animals underwent resection of distal part of stomach. Immunohistochemical study were performed using monoclonal specific antybodies against ghrelin. Hyperthyroidism was a reason of increase of gastric mucosal ghrelin - immunoreactivity, accompanied by a significant decreased of ghrelin plasma concentration. Those observations may indicate, that chronic administration of L-thyroxine cause the change of ghrelin plasma concentration in rats, probably via direct influence on gastric X/A-like cells, but this effect is not responsible for hyperphagia associated with hyperthyroidism.

It has been reported that cannabinoids may cause overeating in humans and in laboratory animals. Although, endogenous cannabinoids and their receptors (CB1) have been found in the hypothalamus, and recently also in gastrointestinal tract, the precise mechanism of appetite control by cannabinoids remains unknown. Recently, ghrelin--a hormone secreted mainly from the stomach X/A-like cells was proposed to be an appetite stimulating agent. The aim of this study was the evaluation of the influence of a single ip injection of a stable analogue of endogenous cannabinoid--anandamide, R-(+)-methanandamide (2.5 mg/kg) and CP 55,940 (0.25 mg/kg), an exogenous agonist of CB1 receptors, on ghrelin plasma concentration and on ghrelin immunoreactivity in the gastric mucosa of male Wistar rats. Four hours after a single injection of both cannabinoids or vehicle, the animals were anaesthetized and blood was taken from the abdominal aorta to determinate plasma ghrelin concentration by RIA. Subsequently, the animals underwent resection of distal part of stomach. Immunohistochemical study of gastric mucosa, using the EnVision method and specific monoclonal antibodies against ghrelin was performed. The intensity of ghrelin immunoreactivity in X/A-like cells was analyzed using Olympus Cell D image analysis system. The attenuation of ghrelin-immunoreactivity of gastric mucosa, after a single injection of R-(+)-methanandamide and CP 55,940 was accompanied by a significant increase of ghrelin plasma concentration. These results indicate that stimulation of appetite exerted by cannabinoids may be connected with an increase of ghrelin secretion from gastric X/A-like cells.

Interleukin-6 (IL-6) has been shown to be involved in the pathogenesis of several bone diseases characterized by a negative balance between bone resorption and formation. The aim of the study was to estimate serum markers of bone turnover: osteoclast-derived tartrate-resistant acid phosphatase form 5a (TRACP 5b) reflecting resorption, and osteocalcin as a marker of bone formation in IL-6 knock-out mice to assess the role of IL-6 in the pathogenesis of thyrotoxicosis-related disturbances of bone metabolism. The study was performed on forty, 14-15 weeks old, female mice: C57BL/6J (wild-type; WT) and C57BL/6J (IL6-/-Kopf) (IL-6 knock-out; IL6KO). The mice were randomly divided into 4 groups with 10 mice in each one: 1. WT mice in thyrotoxicosis (WT-thx), 2. WT controls (WT-ctrl), 3. IL6KO mice with thyrotoxicosis (IL6KO-thx), and 4. IL6KO controls. Experimental model of hyperthyroidism was induced by intraperitoneal injection of levothyroxine at a dose of 1 microg/g, daily over 21 days. The serum levels of TRACP 5b and osteocalcin were determined by ELISA. Serum concentration of TRACP 5b (median and interquartile ranges) were significantly increased in both groups of mice with thyrotoxicosis: WT [28.2(18.8-41.6) U/l] and IL6KO [26.4(23.0-31.2) U/l] as compared to the respective controls. Osteocalcin serum levels in IL6KO-thx mice [111.9(103.1-175.6) ng/ml] were significantly elevated in comparison to WT-thx animals [46.1(32.5-58.9) ng/ml]. In summary, the results of the present study suggest that IL-6 plays a crucial role in thyrotoxicosis-related disturbances of bone turnover in mice, determining the imbalance between bone resorption and bone formation caused by excess of thyroid hormones predominantly by inhibition of bone formation.

Background Aging is related to declined cardiac hemodynamic function. As pumping performance may be significantly related to slowed ventricular depolarization and non-synchronous contraction, we hypothesized that aging may cause dysfunction of intercalated disc (ID), which is the structure responsible for intercellular electrical communication between cardiomyocytes. Methods Male C57BL/6J mice were used for the study at two ages: 4 and 24 months. Electrocardiographic recording was made to analyze the time of ventricular depolarization. Then mice were killed, and the hearts were harvested for examination in transmission electron microscopy (TEM) and immunofluorescence imaging. The expression of connexin 43 (Cx43), N-cadherin, and β-catenin in the myocardium of the left ventricle was evaluated using Western blotting. Results In senescent mice, analysis of averaged QRS complex showed its significant prolongation. At the ultrastructural level, we found frequent disruptions of the ID (affecting 29 ± 5% of them), mainly at the site of adherens junction, with relatively preserved desmosomal intercellular connections and diminished number of gap junctions. Western blotting revealed significantly decreased abundance of Cx43 protein in aged animals, which may cause slowed impulse propagation through the gap junctions and contribute to the observed electrocardiographic alterations. The level of RNA for Cx43 is similar between young and old animals, which suggests a post-transcriptional mechanism of Cx43 protein downregulation. Conclusions Our study shows age-related disorganization of ID, which may be responsible for slowed conduction of the depolarization wave within the heart, and supports the hypothesis of cardiac dysfunction in senescence.

We analyzed the role of interleukin 6 (IL-6) in modulation of the pattern of mice spontaneous activity. Wild type (WT) and IL-6 deficient mice of both sexes, young and aging, were housed individually and various types of their activity were recorded and analyzed with the Phenorack system in their home cages during 72 hours-long sessions. All investigated groups of mice were active mainly during the dark phase of the 24-hours cycle. Generally, the IL-6 deficient animals were more active than their WT controls and females of both genotypes more active than males. Aging mice were less active than the sex and genotype-matched young animals. The independent variables (age, sex and genotype) strongly interacted, which suggests that the modulatory influence of IL-6 on mice behavior may be different in males and females and that it changes during aging. We conclude that under normal physiological conditions signaling of IL-6 via its receptor participates in modulation of the basic pattern of activity. This modulation differs in males and females and changes with aging.

Background CCN family of proteins has been implicated in various processes in cardiovascular physiology and pathology, including angiogenesis, regeneration and fibrosis. In this study we assessed long term changes of CCN1 and CCN2 gene products abundance in the failing ventricular myocardium. Methods Male, 12–14-weeks-old C57BL6/J and C57BL6/J IL−6−/− mice were used. To assess short term changes, a transient reversible ischemia model was utilized. Heart failure was caused by ligation of anterior descending coronary artery. The presence of systolic dysfunction was confirmed by echocardiography and left ventricular ANP RNA expression. Molecular analysis was performed on left ventricular samples from animals sacrificed 12–14 weeks after infarction. Western blotting and QT-PCR were used to investigate abundance of CCN proteins and RNAs, respectively. Results Short ischemia resulted in marked increase of CCN1 transcript. However, three months after myocardial infarction (MI), remote myocardium showed a markedly increased expression of CCN1 protein, but not RNA. In the case of CCN2, the RNA was distinctly up-regulated, whereas the protein presented only modest, non-significant increase in failing myocardium. Expression of CCN2 RNA closely correlated with expression of ANP. Long-term telmisartan administration after infarction decreased the expression of CCN1 protein. Interleukin 6 (IL-6) deficiency caused increased CCN2 protein abundance in control animals, but the difference was absent after MI. Infarction did not increase CCN1 protein in the hearts of IL-6 deficient mice. Conclusion CCN genes are activated in heart failure. Their regulation is multidimensional both transcriptional and posttranscriptional. The involved pathways include angiotensin II and IL-6.

The involvement of dopaminergic projection to the hippocampus in the facilitatory effect of cholecystokinin-unsulphated octapeptide (CCK-8 us ) and caerulein (CER) on memory motivated affectively was investigated in male rats. CCK-8 us and CER were given subcutaneously at the doses of 10 μg kg−1and 0.5 μg kg−1, respectively, immediately after a single learning trial in a passive avoidance situation, after bilateral 6-OHDA lesions to the dentate gyrus of the hippocampus. In order to protect noradrenergic neurones against destruction by neurotoxin, 30 min before surgery rats were pre-treated intraperitoneally with 25 mg kg−1of desmethylimipramine, an inhibitor of noradrenaline uptake. Bilateral 6-OHDA lesions to the hippocampus significantly attenuate the facilitatory effect of CCK-8 us and CER on retention of passive avoidance behaviour evaluated 24 h after the learning trial. Neither, destruction of dopaminergic endings in the hippocampus, nor application of CCK-8 us and CER changed the spontaneous psychomotor activity of rats estimated in an `open field' test. These results may indicate that the facilitatory effect of CCK-8 us and CER on memory motivated affectively is, in part, mediated by dopaminergic projection from the ventral tegmental area to the dentate gyrus of the hippocampus. 2000 Academic Press@p$hr

It has been found in our laboratory that the positive influence of angiotensin II (AII) and its 3-7 fragment [AII(3-7)] on learning and memory processes is mediated by the excitatory amino acids, since it was abolished by NMDA receptor antagonists. The purpose of the present study was to investigate whether bilateral disruption of glutamatergic temporo- entorhinal connections may have an influence on the facilitatory effect of both angiotensin peptides on memory motivated affectively. The bilateral transections of temporo-entorhinal connections were made in 27 male rats 10 days before testing the influence of intracerebroventricular AII and AII(3-7) injection on retrieval of a passive avoidance response. Twenty-seven additional rats served as sham-operated controls. Twenty-five lesioned and 25 sham-operated animals were accepted to the final analysis. AII and its 3-7 fragment significantly improved the retrieval process in sham-operated groups of rats. Bilateral disruption of temporo-entorhinal connections totally abolished the facilitatory effect of both angiotensins on recall of information in a passive avoidance situation. Moreover, bilateral disruption of temporo-entorhinal connections markedly but not significantly attenuated crossings of squares, evaluated in an open field test, without an influence on rearings and bar approaches. These results may suggest that in the facilitatory effect of AII and AII(3-7) on memory motivated affectively involves reciprocal glutamatergic connection between lateral entorhinal cortex and temporal cortex.

The involvement of dopaminergic projection to the central amygdala in the facilitatory effect of cholecystokinin unsulphated octapeptide (CCK-8US) and caerulein (CER) on memory motivated affectively was investigated in male rats. CCK-8US and CER were administered subcutaneously at the doses of 10 μg kg−1and 0.5 μg kg−1, respectively, immediately after a single learning trial in a passive avoidance situation, after bilateral 6-OHDA lesions to the central amygdala. Bilateral 6-OHDA lesions to the central amygdala totally abolished the facilitatory effect of CCK-8US and CER on retention of passive avoidance behaviour evaluated 24 h after the learning trial. These results may indicate that the facilitatory effect of CCK-8US and CER on memory motivated affectively is mediated by dopaminergic projection from ventral tegmental area to the central amygdala.1999 Academic Press@p$hr

Objectives The goal of this work was to identify genes, known to be involved in the skin wound healing, that express differentially in the healthy and injured tympanic membrane (TM), and designate the molecules potentially beneficial for treatment of TM perforation. The molecular mechanisms controlling the course of TM regeneration are far from being elucidated. Methods Twenty rats had their tympanic membranes perforated, while four served as a control. Animals were sacrificed on either days 1, 2, 3, 5 and 10 post injury, and TMs were immediately dissected and frozen in liquid nitrogen. Total TM RNA was isolated and reversely transcribed. qPCR was performed using Rat Wound Healing RT2 Profiler PCR Array (QIAGEN) containing primers for 84 genes. Results Statistically significant changes in the expression of 42 genes were found in various stages of TM healing. The increased expression of genes taking part in the inflammatory reaction (interleukin 6, granulocyte and macrophage chemotactic proteins) was observed from day 2. The expression of several genes of extracellular matrix components and their remodeling enzymes was also changed. Among growth factor genes: Vegfa, Igf1 and Hbegf showed increased expression at the beginning of the healing process, while Hgf expression was highest on day 3. Conclusions Several changes in the expression of genes involved in remodeling of extracellular matrix point to important role of connective tissue in TM healing. The molecules accelerating this process, like HbEGF and HGF, seem to be good candidates for further evaluation of their possible use in clinical treatment.

Chronic heart failure often leads to worsening of the renal function. Mediators of this process include inflammatory and neuroendocrine factors. CCN1 (Cyr 61), a member of growth factor-inducible immediate early genes, which modulates inflammation and fibrogenesis, is excreted with urine in the early phase of acute renal injury and may be involved in the pathogenesis of the cardiorenal syndrome. The aim of the study was to evaluate CCN1 protein abundance and localization in the kidney of IL-6-deficient C57BL/6J (IL-6 KO) mice and respective wild-type (WT) animals in basal conditions and in animals with chronic heart failure twelve weeks after myocardial infarction. Age- and sex-matched mice from both strains subjected to sham operation served as controls. One group of WT animals subjected to myocardial infarction was treated with antagonist of AT1 receptor telmisartan over 12 weeks. Abundance and localization of CCN1 protein in kidney were assessed with Western blotting and immunohistochemistry, respectively. In all groups the strongest immunohistochemical reaction for CCN1 was observed in distal convoluted tubules and in smaller arteries, however, the total expression of CCN1 protein was lower in IL-6 KO mice in comparison to WT animals. The main difference in CCN1 distribution between the examined genotypes was lack of reaction in internal renal medulla and very weak reaction in proximal convoluted tubules in IL-6 KO mice. Experimental heart failure only slightly attenuated the expression of CCN1 protein in the kidney of WT mice and had no effect in IL-6 KO mice. Although, blockade of AT1 receptor did not alter CCN1 protein expression in kidneys of WT mice after myocardial infarction, it significantly changed its CCN1 distribution in the renal tubular system.

Interleukin-6 has been shown to cause imbalance between bone resorption and formation in thyrotoxicosis. The aim of the present study was an attempt to estimate the influence of estrogens on thyrotoxicosis-related disturbances in bone turnover in relation to RANKL-RANK/osteoprotegerin system in IL-6 deficient mice. The study was performed on 56, 12-13 weeks old, female mice: C57BL/6J (wild-type; WT) and C57BL/6J (IL6-/-Kopf) (IL-6 knock-out; IL6KO). The mice were randomly divided into 8 groups with 7 mice in each one: 1. WT controls, 2. IL6KO controls, 3. WT mice with thyrotoxicosis, 4. IL6KO mice with thyrotoxicosis, 5. WT ovariectomized, 6. IL6KO ovariectomized, 7. WT ovariectomized mice with thyrotoxicosis, and 8. IL6KO ovariectomized mice with thyrotoxicosis. Experimental model of menopause was evoked by bilateral ovariectomy carried out in 8-9 weeks old mice. Thyrotoxicosis was induced by intraperitoneal injection of levothyroxine at a dose of 1 μg/g daily over 21 days. The serum levels of TRACP5b, osteocalcin, OPG, and RANKL were determined by ELISA. RANKL serum concentrations were elevated significantly in all groups of ovariectomized mice as compared to respective controls, however, in a minor degree in IL6KO thyrotoxic mice as compared to wild-type animals. Osteoprotegerin serum levels were significantly increased in all thyrotoxic groups of mice except ovariectomized IL6KO animals. To sum up, the results of the present study suggest that IL-6 plays a key role in stimulation of RANKL-RANK/OPG system and this effect is strongly enhanced in conditions of accelerated bone turnover such as thyrotoxicosis and/or estrogen depletion.

High density of cannabinoid receptors type 1 (CB1) in the brain suggests that endocannabinoid system plays an important role in the functioning of the central nervous system. Natural and synthetic cannabinoids are known to attenuate learning and memory processes. The adverse effects of cannabinoids are reversed by SR141716A, at first reported to be a selective CB1 receptor antagonist, later shown to possess also inverse agonist properties. The present study was performed in an attempt to determine the influence of different doses of AM251, a member of the same cannabinoid group as SR141716A, on recognition memory evaluated in an object recognition test. Because cannabinoids may alter motor function and affect anxiety, the influence of AM251 on psychomotor activity and anxiety was assessed in an "open-field" test and elevated plus maze, respectively. While the lowest dose of AM251 (1.0 mg/kg) significantly improved recognition memory, higher doses (2.5 mg/kg and 5.0 mg/kg) did not have an influence on it. Moreover, AM251 did not affect anxiety but in the highest dose significantly attenuated psychomotor activity in rats. The main finding of the present study indicates that AM251, at the dose of 1.0 mg/kg, improves recognition memory in rats without alteration of their psychomotor activity and anxiety. The pro-cognitive effect exerted by compounds belonging like AM251 to diarylpyrazole group may be beneficial in therapeutic use of these compounds, especially in patients with cognitive dysfunctions.

Ghrelin is a peptide of 28 amino acids that transmits appetite related signals from peripheral organs to the brain. The main source of ghrelin is stomach. The regulation of ghrelin secretion is still unknown. The finding that fasting and food intake, respectively increase and decrease the secretion of ghrelin suggests that this hormone may be a bridge connecting somatic growth with energy metabolism and appears to play an important role in the alteration of energy homeostasis and body weight in pathophisiological conditions. The purpose of this study was the evaluation of gastric ghrelin immunoreactivity and ghrelin plasma concentration in male Wistar rats with hyperthyroidism. Experimental model of hyperthyroidism was induced by intraperitoneal injection of levothyroxine at the dose of 80 microg/kg daily over 21 days. At the end of experiment the animals were anaesthetized, blood was taken from abdominal aorta to determinate plasma ghrelin concentration by RIA and then the animals underwent resection of distal part of stomach. Immunohistochemical study were performed using monoclonal specific antybodies against ghrelin. Hyperthyroidism was a reason of increase of gastric mucosal ghrelin - immunoreactivity, accompanied by a significant decreased of ghrelin plasma concentration. Those observations may indicate, that chronic administration of L-thyroxine cause the change of ghrelin plasma concentration in rats, probably via direct influence on gastric X/A-like cells, but this effect is not responsible for hyperphagia associated with hyperthyroidism.

Interleukin 6 (IL-6) is a pleiotropic cytokine involved in both inflammatory reaction and myocardial response to stress. Its effects largely depend on the concentration of the soluble receptors (sIL-6R and sgp130). We investigated the production of IL-6, sIL-6R and sgp130 by the heart during ischemia and reperfusion. Methods: The levels of IL-6 were determined in blood of 34 patients with first myocardial infarction (STEMI), left anterior descending (LAD) artery occlusion, otherwise normal coronaries, without significant co-morbidities and 16 comparable subjects with stable ischemic heart disease and lesion in LAD. Blood samples from coronary sinus (CS) and aorta (Ao) were drawn before percutaneous intervention (PCI), immediately after and at the end of the procedure. Venous blood from 30 healthy volunteers served as control. Results: STEMI patients presented high IL-6 concentrations that increased further after reperfusion when its levels in CS became significantly higher than in Ao. In both groups prior to the PCI there were significantly higher concentrations of sIL-6R in Ao than in CS. This difference disappeared immediately after reperfusion. STEMI patients who experienced cardiovascular complications had higher IL-6 concentration and higher transcardiac sIL-6R gradient than patients with event-free hospitalisation. This association was confirmed in multivariate logistic regression analysis. Myocardial infarction increases concentration of IL-6 that is further elevated by reperfusion. A transcardiac gradient of sIL-6R during ischemia may indicate that large amounts of soluble IL-6 receptors are bound to the infarcted heart and thus affect signal transduction. IL-6 and initial sIL-6R gradient may portend complications in STEMI patients.

Interleukin 6 (IL-6) is a pleiotropic cytokine that is highly expressed in response to ischemia and reperfusion. It has dichotomous roles in the heart, functioning both as an inflammatory mediator as well as a protective agent. The aim of this study was to evaluate the effect of IL-6 deficiency on the expression of apoptotic regulatory proteins under both baseline conditions and following induction of ischemia and reperfusion in the mouse heart. C57BL/6J IL-6−/− TMKopf (IL6KO) and C57BL/6J mice (WT) were subjected to 30 minutes of local reversible myocardial ischemia in vivo or a sham operation. The expression of Bcl-2, Bax and STAT3 in the heart was assessed by western blotting. Under both baseline conditions and following the sham operation, IL-6 deficiency was associated with reduced expression of Bcl-2 and Bax. The TUNEL-FITC, Evans blue and tetrazolium chloride staining of the hearts following ischemia and reperfusion revealed similar injury in operated IL6KO and WT animals. There was increased STAT3 phosphorylation in operated mice regardless of the genotype. Bcl-2 and Bax expression was also comparable between the mouse strains following ischemia and reperfusion. In summary, these results indicated that IL-6 deficiency affected the basal expression of apoptotic regulators, but this did not profoundly alter the extent of reperfusion injury or apoptosis in the mouse heart following ischemia and reperfusion.

Diabetes causes changes in the myocardium, which are often called diabetic cardiomyopathy. This condition has been extensively investigated in animal models with high glucose levels. Nevertheless, it has not been investigated whether moderate hyperglycemia, in the absence of other features of metabolic syndrome, may also cause similar changes in the heart. The aim of the study was to assess changes in the myocardium in an animal model of mild type 1 diabetes. Moderate hyperglycemia was induced in 8- to 10-week-old male C57BL6J mice by 5 intraperitoneal injections of streptozotocin (40 mg/kg). After 16 weeks, they were sacrificed, and left ventricle (LV) dimensions and extent of cardiac fibrosis were assessed by morphometry. The abundance of CCN proteins in LVsamples was assessed using western blotting, while activity of metalloproteinase 2 was established in zymography. Real time PCR was used to investigate the expression of transforming growth factor beta1 (TGFbeta1) and atrial natriuretic peptide. Mice with moderate hyperglycemia presented comparable cardiac dimensions with fibrosis and hypertrophy parameters as the non-diabetic controls. However, the abundance of profibrotic CCN2 protein was significantly increased in hyperglycemic animals (1.67 +/- 0.28 vs. 1 +/- 0.47, p < 0.05). Interestingly, this change was independent from the TGFbeta1 expression, as its RNA abundance was similar in both groups. Moderate hyperglycemia also caused an increase in the activity of the metalloproteinase 2 (1.21 +/- 0.17 vs. 1 +/- 0.07, p < 0.05). Despite diabetes, no profound changes in cardiac morphology were found. In our animal model, moderate hyperglycemia caused activation of a profibrotic gene expression program, which was counterbalanced by the increase of metalloproteinase activity.

Coronary heart disease remains one of the main problems in healthcare systems in western countries. Despite a vast improvement in revascularisation techniques in recent years, we still encounter many patients, who are not suitable for conventional revasularisation methods. A delivery of proangiogenic substances like VEGF or FGF to cardiac muscle was thought to help these patients. Nevertheless, in order to induce angiogenesis in heart in safe and efficient manner further studies of mechanism regulating vessel growth are necessary. This interest resulted in still increasing number of papers dealing with angiogenesis in heart. Still, the results of clinical studies were in large part discouraging. Therefore novel angiogenic substances are currently under investigation. The great expectations are associated with proteins affecting multiple levels and processes involved in vessel growth and maturation. These include among others PIGF and CYR61. Other promising approach is the induction of angiogenesis by stem cells and endothelial progenitor cells. Hopefully these efforts will soon reveal therapeutic methods, which will be applicable in patients with severe ischemic heart disease disqualified from conventional revascularisation procedures.

We have used transgenic male mice not expressing interleukin-6 (IL-6) [C57BL/6J(IL-6/-tm Kopf)] in object recognition test to assess the role of endogenous IL-6 in recognition memory. Wild-type controls showed better memory than IL-6 knock-out mice. Results of our experiment suggest that endogenous IL-6 may play an important role in the process of recognition memory in mice.

The purpose of the present study was to evaluate the effect of a single intraperitoneal injection of a stable analogue of endogenous cannabinoid anandamide - R-(+)-methanandamide (2.5 mg/kg) and CP 55,940 (0.25 mg/kg), an egzogenous CB1 receptor-agonist, on the calcitonin (CT) immunoreactivity of the thyroid parafollicular (C) cells. Four hours after injection with both cannabinoids CT immunoreactivity, evaluated with an avidin-biotin peroxidase complex method by means of rabbit antibodies against CT, was seen to be enhanced in the parafollicular cells in comparison to those of the control group. In thyroids taken from cannabinoid-treated rats the majority of follicles, particularly those located peripherally were large in size, and had low epithelium. Moreover, dilatation of the blood vessels was observed. These changes were accompanied by a significant decrease in CT plasma level, without changes in calcium concentrations. This is the first evidence that a single injection of the cannabinoids R-(+)-methanandamide and CP 55,940 significantly decreases the activity of thyroid C cells.

The aim of this study was to evaluate the significance of measuring calcitonin (CT) plasma concentrations in patients with simple and hyperthyroid goitre treated surgically. Eighty four patients who underwent operations during the years 2000-2002 were analysed. Plasma concentrations of CT were determined by commercially available radioimmunoassay on the day of hospitalisation. Elevated concentrations of CT were found in 8 patients: in 5 out of 26 (19.2%) and in 3 out of 33 (9.0%) patients with Graves' disease and with multinodular goitre, respectively. No major differences in concentrations of CT were observed in patients with simple goitre. Postoperative morphological analysis of pathologically changed hyperactive thyroids showed the presence of enlarged C cells distributed either in small groups or even singly with weakening immunohistochemical reaction for CT. These observations may point to the possibility of a relationship between the functional state of the thyroid gland and the activity of C cells.

The role of the parafollicular (C) cells, the second most important cells in the thyroid gland, has not hitherto been clarified. They are considered to be disperse neuroendocrine cells of the APUD system and synthesise and release many of the regulatory peptides. Few publications are concerned with the evaluation of the structure and function of C cells in the thyroid gland or the probable relationship between these cells and the follicular cells in physiological and pathological conditions. For this reason immunohistochemical investigations were carried out into the activity of the C cells in rats in an experimental model of hyperthyroidism caused by chronic thyroxine influence. This C-cell activity was then evaluated. Differences in the quantity, distribution and calcitonin immunoreactivity of C cells were observed in hyperthyroid rats in comparison to the control group, together with a significant diminution of plasma TSH and calcitonin levels. Our preliminary study may indicate a functional interaction between follicular and parafollicular cells in the thyroid gland.

Cannabinoids are known to attenuate learning and memory in both humans and animals. In rodents, disruptive effect of cannabinoids on memory, reversed by SR 141716, a specific CB(1) receptor antagonist, was shown in behavioral tests based on conditioning. There are no data concerning the influence of cannabinoids on recognition memory. Recently, the improvement of recognition memory in cannabinoid CB(1) receptor knock-out mice was reported. Therefore, the purpose of the present study was to determine whether a stable analogue of endogenous cannabinoid anandamide, R-(+)-methanandamide (0.25 and 2.5 mg/kg, i.p.) and a potent CB(1) receptor agonist, CP 55,940 (0.025 and 0.25 mg/kg i.p.) affect recognition memory in rats evaluated in an object recognition test, based on discrimination between the familiar and a new object presented at 1h interval. Because cannabinoids at the higher doses can produce motor inhibition, the influence of both compounds on psychomotor activity was evaluated in an open field test. CP 55,940 and R-(+)-methanandamide, at both doses given once, 15 min before the learning trial, significantly attenuated recognition memory, measured by the difference in exploration of a new object and a duplicate of the familiar object. Moreover, CP 55,940 at the higher dose significantly attenuated ambulation, and bar approaches, and at both doses also rearings, evaluated in an open field, performed immediately after an object recognition test, while R-(+)-methanandamide at both doses did not alter locomotor and exploratory activity of rats. This is the first evidence that cannabinoids impair recognition memory in rats.

In our laboratory, the positive influence of angiotensin II and its 3-7 fragment on learning and memory processes was found to be mediated by excitatory amino acids, because it was abolished by N-methyl-D-aspartate (NMDA) receptor antagonists. The purpose of the present study was to investigate whether bilateral disruption of glutamatergic temporo-entorhinal connections may have an influence on the facilitatory effect of both angiotensin peptides on recognition memory. The bilateral transections of temporo-entorhinal connections were made in 32 male rats 10 days before testing the effect of intracerebroventricular AII or AII(3-7) injection on the recognition of objects evaluated in an object-recognition test. Thirty additional rats served as sham-operated controls. The final analysis was based on 29 lesioned and 26 sham-operated animals. AII and its 3-7 fragment significantly improved object recognition in the sham-operated groups of rats. Bilateral disruption of temporo-entorhinal connections totally abolished the facilitatory effect of both angiotensins on object recognition. Moreover, bilateral disruption of temporo-entorhinal connections significantly attenuated crossings of squares and rearings, without affecting bar approaches and defecation evaluated in an open-field test. These results may suggest that the facilitatory recognition memory effect of AII and AII(3-7) requires a reciprocal glutamatergic connection between the lateral entorhinal cortex and the temporal cortex.

It has been found in our laboratory that the positive influence of vasopressin (AVP) on memory processes is mediated by excitatory amino acids, since it was abolished by NMDA receptor antagonists. The purpose of the present study was to investigate whether bilateral transections of glutamatergic temporo–entorhinal connections may have an influence on the facilitatory effect of AVP on retrieval process of a passive avoidance behaviour. The bilateral transections of temporo–entorhinal connections were made in male Wistar rats 10 days before testing of the influence of intracerebroventricular AVP (1 μg per rat) injection on memory, evaluated in a passive avoidance task. Although AVP significantly facilitated the retrieval process both in sham-operated and in lesioned groups of rats, bilateral disruption of temporo–entorhinal connections significantly attenuated the facilitatory effect of AVP on the retrieval process. Moreover, bilateral transections of temporo–entorhinal connections failed to affect motor activity, such as crossings of squares, without an influence on rearings and bar approaches evaluated in an open field test. These results may suggest that in the facilitatory effect of AVP on the retrieval process is involved a reciprocal glutamatergic connection between the lateral entorhinal cortex and the temporal cortex.