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Jana Ježková,1 Viktória Ďurovcová,1 Laszlo Wenchich,2,3 Hana Hansíková,3 Jiří Zeman,3 Václav Hána,1 Josef Marek,1 Zdeňka Lacinová,4,5 Martin Haluzík,4,5 Michal Kršek11Third Department of Medicine, First Faculty of Medicine, Charles University and General University Hospital, Prague, Czech Republic; 2Institute of Rheumatology, Prague, Czech Republic; 3Department of Pediatrics and Adolescent Medicine, First Faculty of Medicine, Charles University and General University Hospital, Prague, Czech Republic; 4Institute of Medical Biochemistry and Laboratory Diagnostic, First Faculty of Medicine, Charles University and General University Hospital, Prague, Czech Republic; 5Centre for Experimental Medicine, Institute for Clinical and Experimental Medicine, Prague, Czech RepublicCorrespondence: Jana JežkováThird Department of Medicine, First Faculty of Medicine, Charles University and General University Hospital, U Nemocnice 1128 02 Praha 2, Prague, Czech RepublicTel +420 60 641 2613Fax +420 22 491 9780Email fjjezek@cmail.czPurpose: Cushing’s syndrome is characterized by metabolic disturbances including insulin resistance. Mitochondrial dysfunction is one pathogenic factor in the development of insulin resistance in patients with obesity. We explored whether mitochondrial dysfunction correlates with insulin resistance and other metabolic complications.Patients and methods: We investigated the changes of mRNA expression of genes encoding selected subunits of oxidative phosphorylation system (OXPHOS), pyruvate dehydrogenase (PDH) and citrate synthase (CS) in subcutaneous adipose tissue (SCAT) and peripheral monocytes (PM) and mitochondrial enzyme activity in platelets of 24 patients with active Cushing’s syndrome and in 9 of them after successful treatment and 22 healthy control subjects.Results: Patients with active Cushing’s syndrome had significantly increased body mass index (BMI), homeostasis model assessment of insulin resistance (HOMA-IR) and serum lipids relative to the control group. The expression of all investigated genes for selected mitochondrial proteins was decreased in SCAT in patients with active Cushing’s syndrome and remained decreased after successful treatment. The expression of most tested genes in SCAT correlated inversely with BMI and HOMA-IR. The expression of genes encoding selected OXPHOS subunits and CS was increased in PM in patients with active Cushing’s syndrome with a tendency to decrease toward normal levels after cure. Patients with active Cushing’s syndrome showed increased enzyme activity of complex I (NQR) in platelets.Conclusion: Mitochondrial function in SCAT in patients with Cushing’s syndrome is impaired and only slightly affected by its treatment which may reflect ongoing metabolic disturbances even after successful treatment of Cushing’s syndrome.Keywords: Cushing’s syndrome, insulin resistance, mitochondrial enzyme activity, gene expression

In the interpretation of the effect of a drug or intervention in a single patient, it is desirable to know the spontaneous fluctuations of the variable under study. A few studies have determined pulmonary haemodynamic variability in acute and short-term intervals. It seems reasonable to adopt a confidence limit of 6 mmHg or 22% for PPA changes to be significant. The long-term evolution is difficult to determine because of the unavoidable patient selection for repeat examinations several years apart, and because of the necessary treatment. In slight or moderate pulmonary hypertension the changes in PPA with time seem to be of small magnitude: less than 1 mmHg per year. [ABSTRACT FROM PUBLISHER]

Objective: Radiation therapy is one of the treatment options for pituitary adenomas. The most common side effect associated with Leksell gamma knife (LGK) irradiation is the development of hypopituitarism. The aim of this study was to verify that hypopituitarism does not develop if the maximum mean dose to pituitary is kept under 15 Gy and to evaluate the influence of maximum distal infundibulum dose on the development of hypopituitarism., Design and Methods: We followed the incidence of hypopituitarism in 85 patients irradiated with LGK in 1993-2003. The patients were divided in two subgroups: the first subgroup followed prospectively (45 patients), irradiated with a mean dose to pituitary <15 Gy; the second subgroup followed retrospectively 1993-2001 and prospectively 2001-2009 (40 patients), irradiated with a mean dose to pituitary >15 Gy. Serum TSH, free thyroxine, testosterone or 17β-oestradiol, IGF1, prolactin and cortisol levels were evaluated before and every 6 months after LGK irradiation., Results: Hypopituitarism after LGK irradiation developed only in 1 out of 45 (2.2%) patients irradiated with a mean dose to pituitary <15 Gy, in contrast to 72.5% patients irradiated with a mean dose to pituitary >15 Gy. The radiation dose to the distal infundibulum was found as an independent factor of hypopituitarism with calculated maximum safe dose of 17 Gy., Conclusion: Keeping the mean radiation dose to pituitary under 15 Gy and the dose to the distal infundibulum under 17 Gy prevents the development of hypopituitarism following LGK irradiation.

The effects of dietary supplementation with fat of different fatty acid profile and chronic intermittent hypoxia (CIH) on the fatty acid composition of serum and heart lipids were analysed. Adult male Wistar rats were fed a standard non-fat diet enriched with 10 % of lard, fish oil (n-3 PUFA) or maize oil (n-6 PUFA) for 10 weeks. After 4 weeks on the diets, each group was divided in two subgroups, either exposed to CIH in a barochamber (7000 m, twenty-five exposures) or kept at normoxia. In normoxic rats, the fish oil diet increased the level of conjugated dienes. The n-6:n-3 PUFA ratio in serum TAG, phospholipids (PL), cholesteryl esters (CE) and heart TAG, PL and diacylglycerols (DAG) followed the ratio in the fed diet (in the sequence maize oil>lard>fish oil). In heart TAG, PL and DAG, 20 : 4n-6 and 18 : 2n-6 were replaced by 22 : 6n-3 in the fish oil group. The main fatty acid in CE was 20 : 4n-6 in the lard and maize oil groups whereas in the fish oil group, half of 20 : 4n-6 was replaced by 20 : 5n-3. CIH further increased 20 : 5n-3 in CE in the fish oil group. CIH decreased the n-6:n-3 PUFA ratio in serum CE, heart TAG, PL and DAG in all dietary groups and stimulated the activity of catalase in the maize and fish oil groups. In conclusion, PUFA diets and CIH, both interventions considered to be cardioprotective, distinctly modified the fatty acid profile in serum and heart lipids with specific effects on conjugated diene production and catalase activity.

Objective: Pharmacological treatment with dopaminergic agonists (DA) is the treatment of choice for prolactinomas. Surgical and radiation treatment is also indicated in certain situations. We describe our 12-year experience in treating prolactinomas with the Leksell gamma knife (LGK)., Design: We followed 35 prolactinoma patients (25.7% microprolactinomas, 74.3% macroprolactinomas) treated with LGK irradiation. The mean follow-up period was 75.5 months. Prior to LGK irradiation, patients were treated with DA and 10 of them (28.6%) underwent neurosurgery. Indications for LGK irradiation were: DA intolerance (31.4%), DA resistance (45.7%) and efforts to reduce the DA dose or shorten the period of administration (22.9%). Pituitary function was monitored regularly at 6-month intervals. The central radiation dose range was 40-80 Gy (median 70 Gy), and the minimal peripheral dose was 20-49 Gy (median 34 Gy)., Results: Normoprolactinaemia was achieved in 37.1% of the patients who discontinued DA and in 42.9% of patients who continued DA treatment after LGK irradiation. The median time to prolactin normalization after discontinuation of DA was 96 months. No relapse was seen in any patient. After LGK irradiation, the prolactinoma stopped growing or decreased in size in all but one patient (97.1%)., Conclusion: LGK treatment resulted in normoprolactinaemia in 80.0% of the patients, all of whom had failed pharmacological treatment due to DA resistance or intolerance. After achieving normoprolactinaemia, no relapse of hyperprolactinaemia was observed in any patient. The size of the adenoma decreased even in those patients in whom it was not changed by previous DA treatment.

We examined the influence of dietary fatty acid (FA) classes on the expression of protein kinase C (PKC) delta and epsilon in relation to the cardioprotective effects of chronic intermittent hypoxia (CIH). Adult male Wistar rats were fed a nonfat diet enriched with 10% lard (saturated FA [SFA]), fish oil (n-3 polyunsaturated FA [n-3 PUFA]), or corn oil (n-6 PUFA) for 10 weeks. After 4 weeks on the diet, each group was divided into two subgroups that were either exposed to CIH in a barochamber (7000 m, 8 hrs/ day) or kept at normoxia for an additional 5-6 weeks. A FA phospholipid profile and Western blot analysis of PKC were performed in left ventricles. Infarct size was assessed in anesthetized animals subjected to 20-min coronary artery occlusion and 3-hr reperfusion. CIH decreased the n-6/n-3 PUFA ratio in all groups by 23% independently of the initial value set by various diets. The combination of n-3 diet and CIH had a stronger antiarrhythmic effect during reperfusion than the n-3 diet alone; this effect was less pronounced in rats fed the n-6 diet. The normoxic n-6 group exhibited smaller infarctions (by 22%) than the n-3 group. CIH decreased the infarct size in n-3 and SFA groups (by 20% and 23%, respectively) but not in the n-6 group. Unlike PKC epsilon, the abundance of PKC delta in the myocardial particulate fraction was increased by CIH except for the n-6 group. Myocardial infarct size was negatively correlated (r=- 0.79) with the abundance of PKC delta in the particulate fraction. We conclude that lipid diets modify the infarct size-limiting effect of CIH by a mechanism that involves the PKC delta-dependent pathway.

In a man of 56 years--a victim of traffic accident, contusions in both temporal lobes of the brain and a small subdural and subarachnoid haemorrhage were diagnosed by CT and MR examination. According to the findings of petechial haemorrhages in the corpus callosum a diffuse axonal injury was clinically diagnosed. After 2 months the patient died. Excisions of corpus callosum and the brain-stem were examined by histological staining including Palmgren@s method and by immunohistochemical detection of neuron-specific enolase (NSE) in damaged axons. Clinical findings were compared with morphological findings, i.e. autopsy, histological and immunohistochemical examination. The possibilities of clinical diagnostics of diffuse axonal injury are discussed.

The aim was to determine whether increased oxidative stress during the adaptation to chronic intermittent hypoxia (CIH) plays a role in the induction of improved cardiac ischemic tolerance. Adult male Wistar rats were exposed to CIH in a hypobaric chamber (7,000 m, 8 h/day, 5 days/wk, 24-30 exposures). Half of the animals received antioxidant N-acetylcysteine (NAC; 100 mg/kg) daily before the exposure; the remaining rats received saline. Control rats were kept under normoxia and treated in a corresponding manner. One day after the last exposure (and/or NAC injection), anesthetized animals were subject to 20 min of coronary artery occlusion and 3 h of reperfusion for determination of infarct size. In parallel subgroups, biochemical analyses of the left ventricular myocardium were performed. Adaptation to CIH reduced infarct size from 56.7 +/- 4.5% of the area at risk in the normoxic controls to 27.7 +/- 4.9%. NAC treatment decreased the infarct size in the controls to 42.0 +/- 3.4%, but it abolished the protection provided by CIH (to 41.1 +/- 4.9%). CIH decreased the reduced-to-oxidized glutathione ratio and increased the relative amount of PKC isoform-delta in the particulate fraction; NAC prevented these effects. The expression of PKC-epsilon was decreased by CIH and not affected by NAC. Activities of superoxide dismutase, catalase, and glutathione peroxidase were affected by neither CIH nor NAC treatment. It is concluded that oxidative stress associated with CIH plays a role in the development of increased cardiac ischemic tolerance. The infarct size-limiting mechanism of CIH seems to involve the PKC-delta-dependent pathway but apparently not the increased capacity of major antioxidant enzymes.

Objective: The Leksell gamma knife (LGK) is one of the treatment options for pituitary adenomas. We report on our long-term experience treating acromegaly using LGK., Design: Since 1993 we have followed 96 acromegaly patients through periods of from 12 to 120 months. The mean follow-up period was 53.7 +/- 26.8 months. Seventy-two patients were treated with neurosurgery prior to LGK; for 24 LGK was the primary treatment. Thirteen patients were irradiated twice, due to persistent activity of the adenoma or its residue. Pituitary functions were tested at 6-month intervals, post-irradiation. The target tumour volume for radiosurgery was between 93.3 and 12 700 mm3 (median 1350 mm3)., Results: Fifty per cent of the patients achieved mean GH < 2.5 microg/l within 42 months, normalized their IGF-I within 54 months, and achieved GH suppression in the oral glucose tolerance test (oGTT) < 1 microg/l with normal IGF-I within 66 months. LGK effectiveness was dependent on initial adenoma hormonal activity (GH and IGF-I serum levels), not on the size of the adenoma. Patients with primary neurosurgery followed by LGK irradiation had better outcomes than those with LGK alone. Irradiation arrested all adenoma growth, causing tumour shrinkage in 62.3% of patients. Twenty-six developed hypopituitarism when irradiated by 15 Gy (or more) on functional peritumoral pituitary tissue. No hypopituitarism appeared using lower doses., Conclusions: In acromegaly, LGK is a useful adjunct to primary neurosurgery when treating post-surgical residues because it can limit the duration of medical therapy. It can be used as a primary therapy when neurosurgery is not possible.

The histologica and immunohistochemical examination of cervical spinal cord were used in the group of 11 deceased of violent death. The positivity of neuron-specific enolase (NSE) in axons of cervical spinal cord was examined, and findings in the group of deceased on craniocervical injury (7 cases) and the group of deceased without cranio-cervical trauma (4 cases) were compared. While a routine histological examination including Palmgren's staining showed no significant differences between bouth groups, we noticed an increased frequency of appearance of NSE deceased with craniocervical injury.

Background: Cushing's syndrome (CS) is associated with central adiposity, insulin resistance and impaired glucose homeostasis. Adipose tissue is thought to regulates glucose homeostasis via circulating adipokines, such as resistin, leptin and adiponectin, although their role in the insulin resistance associated with CS has not been established., Design: We examined the relationship between insulin resistance and adipokine levels in CS patients. We compared plasma levels of resistin, leptin and adiponectin in 10 women and four men patients with CS, with 14 health subjects matched for age, gender and body mass index. A subgroup of three women and four men with pituitary-dependent CS were re-examined at least 9 months after curative surgery., Results: CS patients had significantly more truncal fat and less lean body mass as assessed by DEXA compared to control subjects. Total cholesterol, triglycerides and insulin resistance, as calculated using the homeostasis model assessment of insulin resistance (HOMA-R), was significantly increased in CS patients. Of the adipokines measured, only resistin was significantly different between female CS patients and female control subjects (5.05 +/- 0.56 vs. 2.91 +/- 0.39 micro g/l, P = 0.015). Curative surgery significantly reduced total body fat and truncal fat, leptin, total and low-density lipoprotein (LDL) cholesterol, glucose and HOMA-R. A reduction in both resistin and adiponectin was also observed but the differences between pre- and post-treatment levels did not achieve statistical significance., Conclusion: Here we report for the first time that resistin levels are significantly elevated in CS patients and may be important in the insulin resistance associated with glucocorticoid excess.

Unlabelled: Hypercortisolism is associated with a high risk of sickness rate and death rate particularly in view of facilitated arteriosclerotic processes. It is most frequently induced by drug therapy, but endogenous hypercortisolism (Cushing's syndrome) may serve as a suitable model of the effect of hypercortisolism on vascular wall. Our cohort included the following groups of patients and control individuals: 1. a group of patients with florid so far untreated Cushing's syndrome--14 patients, 2. a control group to these patients--16 individuals, 3. a group recently operated on and healed-up patients with Cushing's syndrome--8 patients, 4. a group of previous of previous cured-up patients with Cushing's syndrome--27 subjects, 5. a control group to those patients of group 4--17 persons. The following differences were found between the respective groups: 1. the ultrasonographic examination of carotid arteries demonstrated sclerotic plates or carotid stenosis in 21.3% of patients with florid Cushing's syndrome and 41.4% in patients with corticolism having been cured-up against 11.7% in the relevant control group; 2. the examination of skin microcirculation by the laser-doppler method revealed a lower velocity of perfusion increase during examination of postocclusion hyperemia in patients with florid Cushing's syndrome and hypercortisolism having been cured-up against a control group (CUSH., P < 0.04; previous cured-up, P < 0.02) as well as thermally-induced hyperemia (CUSH., P < 0.03; formerly cured-up, P < 0.04); 3. the laboratory examination of patients with florid Cushing's syndrome revealed higher values of LDL-cholesterol (P < 0.05) and total cholesterol (P < 0.001), malonyldialdehyde as an indicator of increased formation of oxygen radicals (P < 0.05) and oromucoid, the protein of acute phase, signaling a chronic inflammation (P < 0.05); 4. in patients who previously suffered from hypercortisolism increased levels of fibrinogen (P < 0.03) and the cytoadhesive molecule ICAM-1 (P < 0.05) were accompanied by decreased levels of the growth factor of vascular endothelia (VEGF) (P < 0.05) against patients with florid Cushing's syndrome., Conclusion: The findings of the examinations performed indicate that increased incidence of arteriosclerotic processes is present in patients with the florid Cushing's syndrome as well as in those who have suffered from Cushing's syndrome before.

Objective: This study is intended to contribute to a determination of the sensitivity of preserved hypophyseal function to focal radiation in pituitary adenomas., Methods: We compared two subgroups of patients followed up for a median of 5 years after gamma knife surgery (GKS). Subgroup 1 (n = 30) showed postirradiation hypopituitarism. Subgroup 2 (n = 33) was continually eupituitary. These subgroups were taken from a previously published study relating to a larger group of 163 patients with pituitary adenomas treated by GKS and evaluated after a median follow-up period of 2 years. A relatively high treatment dose was used in this larger group (median, 20 Gy to the tumor margin for growth control in nonfunctioning adenomas; median, 35 Gy for hypersecreting adenomas). Early results approached those of microsurgery, and there were only a few side effects. In the present study, we compared 16 different variables in the same two subgroups to discover the relationships that caused a delayed appearance of postirradiation hypopituitarism. The main pretreatment and treatment parameters were measured on reconstructed treatment plans. This database was used for statistical evaluation., Results: The relationship between the mean dose and the volume of functioning hypophysis was stronger in terms of worsening of pituitary function than that of the spot dose to different intrasellar structures. We found that for our group of patients, the safe mean dose of radiation to the hypophysis was 15 Gy for gonadotropic and thyrotropic functions and 18 Gy for adrenocorticotropic function. The worsening of pituitary function was also significantly dependent on the dose to different anatomic levels of the infundibulum, but we did not succeed in fully characterizing this relationship. In addition, we discovered significant levels of dependency of postirradiation pituitary damage to different pretreatment and treatment variables., Conclusion: Knowledge of the radiation tolerance of functioning pituitary structures subjected to GKS can ensure better preservation of pituitary function after irradiation. This is valid for the group of patients we studied. Our study's findings can be used as a guideline for GKS treatment of new patients with pituitary adenomas, and it can serve for comparison with the experience of other gamma knife centers.

Adult male Wistar rats were exposed to intermittent high altitude hypoxia of 7000 m simulated in a hypobaric chamber for 8 h/day, 5 days a week; the total number of exposures was 25. The concentration of individual phospholipids and their fatty acid (FA) profile was determined in right (RV) and left (LV) ventricles. Adaptation to hypoxia decreased the concentration of diphosphatidytglycerol (DPG) in hypertrophied RV by 19% and in non-hypertrophied LV by 12% in comparison with normoxic controls. Chronically hypoxic hearts exhibited lower phospholipid n-6 polyunsaturated FA(PUFA) content mainly due to decreased linoleic acid (18:2n-6), which was opposed by increased n-3 PUFA mainly due to docosahexaenoic acid (22:6n-3) in phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylinositol (PI). The content of arachidonic acid (20:4n-6) was unchanged in total phospholipids, but in PC it was increased in both ventricles (by 22%) and in PE decreased in LV only (by 20%). Chronic hypoxia increased the un-saturation index of PC and PE in both ventricles. The content of monounsaturated FA (MUFA) was increased and 18:2n-6 decreased in DPG. The proportion of saturated FA was increased in PC and PI of hypoxic RV but not LV. The FA composition of phosphatidylserine was not altered in hypoxic ventricles. It is concluded that chronic hypoxia led to only minor changes in individual phospholipid concentration in rat ventricular myocardium, but markedly altered their FA profile. These changes, in particular the greater incorporation of n-3 PUFA into phospholipids and increased un-saturation index, may lead to a better preservation of membrane integrity and thereby contribute to improved ischemic tolerance of chronically hypoxic hearts.

Bacillus firmus (a Gram-positive nonpathogenic and harmless bacterium), was shown to be a strong polyclonal activator of mouse B lymphocytes as estimated by ELISA testing of Ig concentrations in culture supernatants after incubation of BALB/c mouse splenocytes with inactivated bacillus. Synthesis of all main Ig classes and all IgG subclasses was stimulated in vitro, the considerable effect on IgA formation being the most interesting feature. B cell stimulation was T cell dependent, as was demonstrated by the effect of B. firmus on all Ig isotypes and by comparison of lymphocyte response of nu/nu mice and heterozygous nu/+ mice. The effect of B. firmus on splenocyte proliferation was stimulatory or suppressive depending on the dose of the bacterium. Increased synthesis of IFN-gamma and IL-10 (detected by ELISA in splenocyte culture supernatants) showed probable stimulation of Th1 and Th2 subpopulations. Considering the stimulatory effect on IgA formation and macrophage stimulation, B. firmus seems to be a prospective mucosal adjuvant and/or probiotic.

Caspases are cysteine proteases which participate in different stages of apoptosis. Apoptosis, cell death, programmed by its nucleus is associated also with a number of diseases and tissue damage. To this process increasing attention is paid also in the sphere of forensic medicine, in particular to make use of the diagnostic contribution in investigation of cause of death, vital reaction and time when the injury developed. In damaged tissues by immunohistochemical methods caspase activity was assessed. Caspase 8 activity (Flice) was detected in the heart muscle in congenital heart disease in a two-week old infant but also in ischaemia in a 58-year old woman with thrombosis of the coronary artery or in acute circulatory failure after an overdosage of pervitin in a 39-year-old drug addict. In the liver there was a positive finding in hepatocytes in chronic inflammatory changes caused by chronic alcohol abuse. An early caspase 8 reaction after an injury is suggested by positive findings in skeletal muscles of the neck and larynx in a 47-year-old man who was strangled. A positive reaction was proved only at the site of the line caused by the strangulation tool. However macroscopically nor by common histological staining the muscle tissue did not display any signs of injury at the mentioned sites. The results suggest that immunohistochemical evidence of caspase 8 can be used as a suitable complementary examination not only for detection of damaged tissue but also for evaluation of early stages of the vital reaction.

During early stages of chronic hypoxia under experimental conditions in the lungs alveolar macrophages are activated and in the latter an increased amount of matrix-metalloproteinase-collagenolytic enzymes is produced. Their presence was assessed in the present study in the lungs of infants who died suddenly (SIDS) and also in subjects who died from a violent death incl. acute and protracted external suffocation. The positive findings of these matrixins, in particular MMP-9 (gelatinase) were assessed in alveolar macrophages not only after protracted suffocation but also in the majority of sudden infant deaths. These findings supplement the mosaic of changes which indicate that sudden infant deaths are preceded by a period when the infant was exposed to hypoxia.

The objective of the study was to explain the effect of autolysis on immunohistochemical detection of neurone-specific enolase (NSE), beta-amyloid protein precursor (beta-APP) and ubiquitine in cerebral tissue. The examination was made in 6 deceased subjects without mechanical injury of the CNS and 6 subjects with a craniocerebral injury who survived from 6 hours to 3 days. In all deceased subjects the post-mortem examination was made within 24 hours after death. For immunohistochemical examination tissue excisions were taken from standard sites of the brain. The first tissue excisions were immersed into 10% formol after a post-mortem interval of 24 hours. The remaining tissue slices were subjected to autolysis at room temperature and gradually immersed into formol after 24-hour intervals, the longest post-mortem interval being 168 hours, i.e. 7 days. For visualization of the linked primary antibody the biotin-streptavidin system labelled with alkaline phosphatase was selected. In the group of 6 subjects who died after a craniocerebral injury in 4 instances axonal lesions were detected, i.e. axonal oedema or formation of retraction spheroids. The damaged axons were positive on examination with all investigated antibodies, whereby it was possible even after a 168-hour post-mortem interval to differentiate damaged and not damaged axons. In the group of 6 subjects without mechanical injury of the CNS in 5 instances axonal oedema was found, however, it was not positive with anti-NSE antibodies nor with anti-beta-APP. After the 24-hour post-mortem interval in this group in 3 instances ubiquitine positivity was found in axons but already after a post-mortem interval exceeding 2 days the axons were ubiquitine positive in all 6 subjects. Lumpy deposits of this substance could be detected in axons also beyond axonal structures.

Construction, performance characteristics and applications of a carbon paste thallium(III) ion-selective electrode are described. The electrode, which is based on ion-associate compounds formed between cetylpyridinium and chlorothallate(III) complexes dissolved in tricresyl phosphate as pasting liquid, showed near-Nernstian response over the concentration range of 5.8 x 10(-6) - 2.9 x 10(-3) mol/L. Potentiometric titrations of thallium(III) with cetylpyridinium chloride were affected by higher concentrations of excess halides, probably due to the formation of higher halogenothallates.

Immunohistological evidence of glycophorin from paraffin sections of tissues fixed with formol, makes its possible to visualize membranes of preserved erythrocytes and fragments after their disintegration. This fact permits even after development of post-mortem changes to differentiate actual extravasation, manifested by a positive finding of glycophorin from mere infiltration of tissue by blood pigment after death, where glycophorin is lacking. The method is very sensitive and can reveal even very discrete haemorrhage. From different grades of erythrocyte disintegration in haematomas in the same subject sometimes conclusions can be drawn also on the different time of development of traumatic tissue changes.

Blood group substances A, B, H were detect in placental tissue paraffin cuts (after 10% formaline fixation) by immunohistochemistry using indirect immunoperoxidase-two-layers technology or biotin-streptavidin complex signed by alkaline phosphates. Both fetal and maternal part of placenta are to be investigated which enables their group markers to be identified. Immunohistochemical detection of the A, B, H blood group substances in placenta can give conclusion about the blood group of fetus and of fetus and of mother as well.

The possibility of the use of heteropoly anions formed by addition of phosphate to the solutions of either molybdate or tungstate in the determination of phosphorus by the simple method of potentiometric titration was studied. The heteropoly anions are titrated on the ion-pairing principle using a titrant containing a lipophilic counter-ion, i.e. 1-(ethoxycarbonyl)pentadecyltrimethylammonium bromide (Septonex), the titration being monitored by carbon paste electrode, although other liquid membrane-based electrodes can also be used. Calibration plots of the titrant end-point consumption vs. concentration are not linear, but allow one to evaluate the content of phosphorus. Similar procedures, when optimized, should be elaborated for determination of numerous other elements forming heteropoly anions.

Skin injury alive produces quick and distinct increase of free histamine and serotonin. A fluorimetric method for detection of these compounds in tissues has been the only one described in the literature up to now but it needed a time consuming extraction and big amount of primary material (about 3 g of skin devoid of subcutaneous fat tissue) as well. The authors succeeded in substituting the method and developed a voltametric detection of histamine and serotonin using the computerized ECO-TRIBO-Polarograph. It enabled to omit the extraction of skin compounds and to decreased the amount of primary tissue to a quarter. The detection of free histamine and serotonin levels is presumed to enable a more accurate determination of the time of injury, particulary in short interval before death; it presents, with a simultaneous of glycophorin, a convenient complementation of immunohistochemical investigation of fibronectin.

Evidence of an early vital reaction after a skin injury is sometimes of decisive importance in forensic practice. Attention is devoted increasingly to immunohistochemical evidence of substances, the presence of which in the borders of the wounds signalizes an intravital origin of the injury. In this context evidence was focused on detection of glycophorine at sites of haematomas of the skin and subcutaneous layer and to fibronection in the borders of injuries and in haematomas. To visualize the two substances in paraffin tissue sections, the immunohistochemical method of indirect immunoperoxidase reaction was selected where in the first layer specific sera were used wither against glycophorine A, B or against fibronectin. Glycophorine is a sialoglycoprotein which is part of the red cell membrane. It can be detected at the site of haematomas and after the disintegration of erythrocytes during post-mortem autolysis. Conversely it is lacking where membrane structures were not present. This makes it possible even during advanced stages of post-mortem changes to differentiate actual haematomas from mere post-mortem inhibition of tissue by blood pigment (e.g. in hypostasis) where glycophorine cannot be detected. Fibronectin, another glycoprotein substance can, already in the early stage of injury, form the basis for a netlike structure in the shape of fine "strings". Detection of fibronectin suggests the intravital development of injury and in the authors cases it could be detected already within several minutes after the development of the injury.

Commonly used serological methods i. e. absorption-elution test and mixed agglutination do not always allow discriminate the nature of blood group substances A, B, H in hair investigation. The finding need not sometimes be unequivocal. A completion of forensic traditional serological methodology by indirect immunoperoxidase technique enables to eliminate doubts about interpretation of results. There is possible to identify blood group substances A, B, H in preserved medullary core of hair in this way.

Using two-dimensional echocardiography (ECHO), the authors examined within one week preceding right-sided catheterization of the right ventricle (RV) 44 patients. Of three tested ECHO techniques for calculation of right ventricular volumes and/or function the best correlations were obtained by the method of evaluation of area-length of the right ventricle from two orthogonal projections (apical 4-cavity and subcostal with visualization of the outflow portion of the RV). The following correlations were assessed: telediastolic volume--r = 0.831, telesystolic volume--r = 0.815 stroke volume--r = 0.810 and ejection fraction (EF)--r = 0.752 (p less than 0.001 for all correlations). The tested method enables us at the same time to screen at least existing right ventricular dysfunction (sensitivity 0.68; specificity 0.82). In the presence of tricuspidal insufficiency (TI) increase in right ventricular function occurs which may mask its existing failure (dependence of EF on the mean pressure level in pulmonary artery was proved only in patients without TI--r = 0.594). We assume that information on right ventricular function is important not only in diseases affecting primarily the right heart but also in advanced stages of left-sided failure. Follow-up of the natural course of the disease and the effect of provided treatment by the non-invasive ECHO method is thus useful not only in research but also in clinical practice.

We employed two-dimensional echocardiography for the assessment of right ventricular (RV) volumes and/or function in a series of 44 patients. The results of three different echocardiographic approaches were compared with the data obtained from single-plane RV angiography following ultrasound within a 7-day interval. Only the echocardiographic area length method with two orthogonal imaging planes employed (apical 4-chamber and subcostal projections) yielded the beneficial results. The correlations between echocardiographic and angiographic RV volume estimates were rather high (end-diastolic volume: r = 0.83, end-systolic volume: r = 0.82, stroke volume: r = 0.81) and satisfactory in ejection fraction (r = 0.75). Using the method mentioned, the differentiation of patients with an angiographic evidence of RV failure (echocardiographic ejection fraction less than 0.55) from those without it was possible with a sensitivity of 0.68 and a specificity of 0.82. Concerning the clinical impact of the presented study, we can recommend the technique in question as a screening procedure for the detection of changes in RV function exceeding 12% (95% confidence limits).

The effect of nitrates was monitored in 16 patients with precapillary, and in 12 patients with postcapillary pulmonary hypertension (PH). The patients had haemodynamic examination on acute administration of 10 mg of isosorbide nitrate (ID) in infusion and after 2-month therapy with 120 mg of isosorbide dinitrate retard daily. Acute ID administration decreased filling pressure of both ventricles, pulmonary and aortic pressure, pulmonary and systemic resistance and increased blood flow. No significant difference was found between the two types of PH. Long-term administration resulted in a statistically significant decrease in filling pressure (in both ventricles) and pulmonary pressure, even though the decrease was less pronounced than those seen following acute administration. Again, no statistical differences were noted between the two types of PH. While acute administration of nitrates decreases both preload and afterload, chronic administration decreases only preload. The implications of this finding for therapeutic practice are discussed.

Using the pulsed Doppler system with a high reparative frequency (HPRF), the authors made within 1 hour and 3 days after diagnostic right-sided catheterization 38 measurements of the trans-tricuspid pressure gradient in 31 consecutive patients. To the gradient calculated from the absolute velocity of flow through the jet of tricuspid regurgitation the authors added two different arbitrarily determined pressure values in the right atrium to obtain the systolic pressure in the right ventricle--RVSP (which assuming the absence of an obstruction between the right ventricle and pulmonary artery equals the systolic pressure in this artery). The calculated RVSP from the Doppler assessment correlated highly significantly with the directly manometrically obtained values of the same indicator (r = 0.973; p less than 0.001). An even closer correlation was obtained when using a differentiated estimate of the pressure in the right atrium/according to the filling of the jugular veins) (r = 0.977; p less than 0.001). From the RVSP values the authors tried to predict the presence of pulmonary hypertension (PH) diagnosed from the median pressure in the pulmonary artery (PAP greater than or equal to 20 mm Hg which was considered conclusive). Using the arbitrarily assessed range of systolic PH greater than 35 mm Hg, the sensitivity of the ultrasonographic estimate was 79% with a 60% specificity, while when the criterium of greater than 40 mm Hg was used, the corresponding values were 64% and 90%.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of 2-month treatment with isosorbide dinitrate (120 mg day-1), nifedipine (2 x 20 mg day-1) and their combination has been assessed in 16 patients with mild to moderate chronic cardiac failure. Isosorbide dinitrate decreased right atrial (-23%), pulmonary wedge (-20%) and pulmonary arterial (-17%) pressures but did not significantly change either cardiac output or systemic and pulmonary vascular resistance. Nifedipine increased cardiac output (+13%) and decreased systemic and pulmonary vascular resistance (both -17%) with no change of pressures. Combined therapy with both drugs decreased ventricular filling pressures (-8% and -15%), systemic (-20%) and pulmonary (-13%) arterial pressures, increased cardiac output (+26%) and decreased both systemic (-29%) and pulmonary (-29%) vascular resistances. Changes during exercise were almost the same as at rest. The effect of both drugs was more pronounced in patients with more severely pathological haemodynamic measurements before treatment. We conclude that combined treatment with both preload- and afterload-reducing agents can preserve or even potentiate a favourable haemodynamic effect of individual drugs.

The effect of isosorbide dinitrate (ID) on the development of pulmonary hypertension (PH) has been examined in 18 patients with idiopathic diffuse interstitial lung fibrosis (IDILF) during two years of treatment. All patients responded favourably to acutely administered ID by a decrease in pulmonary vascular resistance (PVR) by 38 +/- 13% and of the pulmonary arterial pressure (PAP) by 35 +/- 11%. Two years ID therapy led to sustained haemodynamic improvement in patients in whom blood gases were stable PAP was reduced from 38 +/- 5 to 30 +/- 6 mmHg and PVR from 483 +/- 197 to 364 +/- 175 dyne s cm-5. No haemodynamic changes were noted in the subjects with progressive hypoxaemia whereas steady deterioration of central haemodynamics was recorded in those patients who died during the follow-up. It is concluded that some patients with IDILF and PH may benefit from long-term ID treatment, but that the outcome of this treatment cannot be reliably predicted from the results of the initial first ID administration.

The authors examined a group of 50 patients incl. 28 (56%) with pulmonary hypertension (PH) at rest, mostly postcapillary. Using a Doppler device, they measured the acceleration time (ACT) in the outflow tract of the right ventricle and trunk of the pulmonary artery. Under conditions of concurrent Doppler and direct, catheterization measurements the authors found a close correlation between ACT and the mean pressure in the pulmonary artery (r = -0.88), which was, however, not confirmed after a 1-10 day interval (r = = -0.64). The authors conclude that Doppler ACT reflects only the actual pressure in the pulmonary circulation. The authors found a close correlation between ACT values in the right ventricular outflow tract and the pulmonary artery (r = 0.95), and no significant difference. Comparison of Doppler diagnosis of PH with former echocardiographic methods revealed unequivocally the priority of the Doppler method. Failure of echocardiography is most probably associated with the postcapillary type of PH in the group of patients and does not rule out the validity of original findings in precapillary PH. Rapid and easy assessment along with a simpler interpretation is in favour of the Doppler technique.

The authors compared the sensitivity of the simple dipyridamole test (DP test) and the dipyridamole test combined with an isometric load (DP + HG test). Evaluation by means of two-dimensional echocardiography was used. The sensitivity and specificity of the DP test, in case of a 50% or greater stenosis of one or several coronary arteries, as a criterion of classification of the group was 45% and 100%, as compared with 72% and 100% in the combined DP + HG test. The correct diagnosis was confirmed by a selective reference coronarographic findings in all patients. The electrocardiographic changes alone in both tests were positive for IHD only in 9 and 17% resp. Ergometry was more sensitive (44%) than electrocardiographically evaluated changes, using the mentioned intervention tests, but it did not attain the sensitivity of the echocardiographic evaluation. The work indicates that temporary changes of the regional kinetics evaluated by two-dimensional echocardiography are a highly specific manifestation of myocardial ischaemia which developed during a load, and these changes are detected sooner and more frequently than associated electrocardiographic changes.

Natural history of 99 patients with angiographically defined left ventricular aneurysm (LVA) has been studied. Congestive heart failure (CHF) was predominant in 52.5% patients. Mean follow-up was 94 months. The cumulative survival at 1, 5, 7, 10 years were 92, 65, 60 and 28% respectively. Left ventricular end-diastolic pressure averaged 2.8 kPa (LVEDP), ejection fraction (EF) 31%, contractile segment ejection fraction (CSEF) 45%, stroke work 6.7 mj. g-1, diastolic compliance 0.36 kPa-1 and passive elastic modulus 64.1 kPa. High risk groups of patients were those with LVEDP more than 3.3 kPa, CSEF less then 40%, EF less then 20% and with extent of LVA more then 40% of the diastolic area of the left ventricle. The Cox analysis of survival indicated following variables predicted outcome: functional impairment due to CHF, LVEDP, LVEDVI and number of vessels diseased.

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The authors used the pulse and continuous-wave Doppler technique to perform 51 measurements of stroke volume and cardiac output in 37 persons. Results were compared with those obtained using Fick's principle of thermodilution (immediately subsequent or simultaneous). Regression analysis showed a close correlation between Doppler and haemodynamic values - r = 0.86 for stroke volume and 0.94 for cardiac output (p less than 0.001). Using the paired t-test, the methods did not differ significantly. Standard deviation of Doppler stroke volume values from reference haemodynamic values was 10%, and only two measurements differed by more than 25%. Doppler determination of cardiac output can be therefore recommended as an alternative method in patients with high-quality echo Doppler recordings.

Duplex Doppler echocardiography with high pulse repetition frequency was used to study: group A - 7 patients with mitral stenosis (MS) during cardiac catheterization, and group B - 26 patients with MS in whom the Doppler examination was performed 24 h-3 months prior to catheterization. Using the peak flow velocity within the stenotic flow jet, the pressure drop and mitral valve area (MVA) were calculated and compared with data obtained invasively. The inter-observer variability of the two parameters was 12% and 5%, respectively. The correlation between Doppler pressure drop and invasive measurement was excellent in group A (r = 0.83), while less close in group B (r = 0.71). However, using only MVA estimates (group B), it was possible to recognize most of MS with catheterization MVA less than or equal to 1.0 cm2/m2 (sensitivity and predictive value both of 90%). Therefore, even nonsimultaneous quantitation of MS seems valuable in pre-operative patient evaluation, adding a new information to hitherto existing echocardiographic criteria.

Unlabelled: Survival rates were determined for a group of 136 patients in whom left ventricular aneurysm was determined by angiography. They were treated medically (99 patients) or surgically (37 patients). Congestive heart failure was predominant in all patients. The two groups did not differ in regard to clinical and haemodynamic data except for a more extensive coronary artery disease in the surgical group. Survival rates at the 5, 7 and 9 years were 62.5%, 52.5% and 37.5% in the medicaly treated group and 50%, 40% and 32.5% in the surgical group. Functional improvement at least one Functional Class NYHA was present in 75% patients postoperatively, and in 27% of patients treated medically., Conclusion: In patients with left ventricular aneurysm with predominant congestive heart failure surgical treatment improved quality of life but did not increase long-term survival.