Ultrastructure of liver progenitor/oval cells in children with nonalcoholic steatohepatitis.

PURPOSE: Very interesting reports have appeared lately on the role of liver progenitor/oval cells in the morphogenesis and development of nonalcoholic steatohepatits (NASH) in adult patients and experimental animals. However, no literature data concerning pediatric patients have been available. Therefore, the purpose of the study was to evaluate the ultrastructure of the population of liver progenitor/oval cells in the biopsy material from children with previously clinocopathologically diagnosed NASH. MATERIAL/METHODS: Electron-microscopic examinations were conducted on fresh tissue samples collected from 10 children with NASH (aged 2-14 years), which were fixed with a solution of 2% paraformaldehyde and 2.5% glutaraldehyde in 0.1 M cacodylate buffer. RESULTS: Ultrastructural examinations of the liver progenitor/oval cells in children with NASH show a quite prominent number of these cells, especially their two types, hepatic progenitor cells (HPCs) and intermediate hepatocyte-like cells (IHCs), with intermediate bile-like cells being the least frequent. They were found to occur single or in clusters of two, seldom of three, and frequently in the areas of advanced liver fibrosis or close to them. Many times, these cells were accompanied by hepatocytes showing a varying degree of death, to total cell disintegration. Interesting was the presence of activated nonparenchymal liver cells, i.e. Kupffer cells/macrophages and hepatic stellate cells, frequently found to adhere to the hepatic oval cells. CONCLUSIONS: The current study suggests a marked involvement of the population of liver progenitor/oval cells, mainly HPCs and IHCs, in the development of nonalcoholic steatohepatitis in pediatric patients, especially in fibrosis progression.

Topiramate as a neuroprotectant in the experimental model of febrile seizures.

PURPOSE: The aim of the study wad to estimate a potentially neuroprotective effect of topiramate (TPM) in the experimental model of FS. MATERIAL AND METHODS: 24 young male rats divided in 4 groups were involved in the study. Febrile seizures were induced by placing the animals in 45 degrees C warm water bath for four consecutive days. TPM at the dose 80 mg/kg b.m. was administered: before the FS and immediately after the FS. FS group and control rats received only normal saline. Thereafter hippocampal slices were prepared to performing histological and morphometric examination. RESULTS: Morphometric investigations revealed that FS caused death of 60% of the neurons in sector CA1 and a half of them in sector CA3. Histological examinations of hippocampal slices showed that TPM at a dose of 80 mg/kg b.m., administered before the seizures, considerably improved CA1 and CA3 pyramidal cell survival. Similar neuroprotective effect, but in a markedly lesser degree was observed when TPM was administrated after the FS. CONCLUSIONS: Our findings seem to confirm that FS exert a strong destructive effect on the sensitive hippocampal neurons and on the neuroprotective properties of TPM in this process, which may have practical implications. It can be assumed that in children with recurrent and prolonged FS, prophylactic drug administration could prevent hippocampal sclerosis and development of symptomatic epilepsy.

Matrix metalloproteinases and their tissue inhibitors in children with chronic hepatitis B treated with lamivudine.

PURPOSE: To evaluate if measurements of MMP-2, MMP-9, TIMP-1 and TIMP-2 have clinical applicability as markers of liver fibrosis and to assess the effect of long-term lamivudine treatment on liver fibrosis in children with chronic hepatitis B (chB). MATERIAL AND METHODS: The observation was carried out on 41 children with biopsy proven chB (HBe/+/, HBVDNA/+/) who were nonresponders to previous IFNalpha therapy. Lamivudine was administered in the group of 29 children (3 mg/kg/day, maximum 100 mg/daily). The serum concentration of examined markers was measured with ELISA before and after 24 months of therapy. ROC analysis was used to calculate the power of the assays to detect advanced liver fibrosis (score >2 according to Batts & Ludwig). RESULTS: Serum TIMP-1 and TIMP-2 levels were significantly higher and MMP-9 lower in children with chB compared to controls. There was a significant positive correlation between serum MMP-2 and negative correlation between MMP-9 level and the stage of liver fibrosis. The ability of serum MMP-9 to differentiate children with mild fibrosis from those with advanced fibrosis was significant (AUC = 0.75; p = 0.03). Other serum markers did not allow a useful prediction. 2-year lamivudine treatment did not improve histological fibrosis but it caused significant decrease of serum TIMP-2 (p = 0.01) and increase of MMP-9 level (p = 0.00005). CONCLUSIONS: MMP-9 is a better serum fibrosis marker than MMP-2, TIMP-1 and TIMP-2 to diagnose children with advanced liver fibrosis. The significant decrease of TIMP-2 and increase of MMP-9 level during therapy suggest antifibrotic effect of lamivudine in children with chB.

Serum level of YKL-40 does not predict advanced liver fibrosis in children with chronic hepatitis B.

PURPOSE: The aim of the study was to evaluate the serum concentration of YKL-40 (human cartilage glycoprotein-39) in the assessment of fibrosis stage in children compared to biopsy and prior to antiviral treatment for chronic hepatitis B. MATERIAL AND METHODS: We determined serum level of YKL-40 (METRA, EIA kit, Quidel Corporation, San Diego, USA) after an overnight fast in 63 children (age range 4-17 years, mean 10 years) with biopsy-verified chronic HBeAg-positive hepatitis B. Fibrosis stage and inflammation grade were assessed in a blinded fashion according to Ishak et al. We defined advanced liver fibrosis as a score >2. Receiver operating characteristics (ROC) analysis was used to calculate the power of the assay to detect advanced liver fibrosis (AccuROC, Canada). RESULTS: Serum concentration of YKL-40 was significantly higher in patients with chronic hepatitis B compared to controls (n=16) (38.5 +/- 19.2 vs 27.9 +/- 8.75 ng/mL; p = 0.032). The ability of serum YKL-40 to differentiate children with advanced liver fibrosis (n=31; 49.2%) from those with mild fibrosis was not significant (AUC = 0.387 +/- 0.072, p = 0.12). This marker was not a good predictor of histologic inflammation either. CONCLUSION: Serum level of YKL-40 does not predict advanced liver fibrosis in children with chronic hepatitis B.

C-erb-B2 and Bcl-xl protein expression in Barrett's oesophagus in correlation with morphological parameters.

The aim of the study was to evaluate the correlation of c-erb-b2 and Bcl-xl expression in biopsy specimens of Barrett's oesophagus from 44 patients with morphological features. The examined group was subdivided into: negative for dysplasia, indefinite for dysplasia, positive for dysplasia-low grade, and adenocarcinoma with high grade dysplasia. Positive c-erb-B2 staining was found in 34.1% and Bcl-xl protein expression was observed in 96.9% of BE. The results showed increased c-erb-B2 and Bcl-xl protein expressions with progressive grades of dysplasia to adenocarcinoma. In conclusion, an evaluation of c-erb-B2 and Bcl-xl expression can be useful for the histopatologic diagnosis of BE and correct interpretation of dysplasia.

S-100 protein as marker of the blood-brain barrier disruption in children with internal hydrocephalus and epilepsy--a preliminary study.

S-100 is a structural protein of the central nervous system. An elevated level of S-100 in CSF is generally considered to be a marker of nervous tissue damage. The presence of this protein in blood serum points to the functional and/or morphological disruption of the blood-brain barrier. We measured S-100 in the cerebrospinal fluid and blood of children with two of the most often observed pathological states in child neurology--internal hydrocephalus and epilepsy. High levels of S-100 in CSF were detectable in children with internal hydrocephalus. Increased blood levels of S-100 protein were detectable in both groups of paediatric patients. Our preliminary results indicate neuronal damage in internal hydrocephalus and morphological and/or functional disturbances of the blood-brain barrier (their increased permeability) in both above mentioned disabilities.

[Studies of damaged processes in the nervous system and possibilities of neuroprotection]. / Badania procesów uszkodzenia ukladu nerwowego i mozliwosci neuroprotekcji.

The paper presents the results of experimental studies and clinical trials associated with mechanism underlying nervous system damage and pharmacological interventions to be employed in such processes. Abnormal lipid peroxidation was demonstrated in experimental seizures, epilepsy and cerebral stroke. The authors presented the mechanism of calcium channel blockers activity in epilepsy and their experience in employing such agents in epileptic patients. Results of studies on S-100 protein determinations as a marker of the blood-brain barrier damage in epilepsy and hydrocephalus were discussed, along with the employment of evoked potentials in diagnostic management of headaches and the effects of complex treatment of epilepsy in children using Nootropil. Selected data on experimental valproate encephalopathy were also presented. The authors believe that these studies contribute to the understanding of mechanism that are responsible for nervous system dysfunction, as well as to the evaluation and treatment of their effects.

Morphometric analysis of the cerebellar cortex capillaries in the course of experimental valproate encephalopathy and after chronic exposure to sodium valproate using transmission electron microscopy.

Morphometric analysis of the cerebellar cortex capillary cross-section area performed in experimental valproate encephalopathy using transmission electron microscopy showed that prolongation of VPA application resulted in more enhanced lumen narrowing manifested in gradual reduction in the mean value of the coefficient examined. After 6, 9 and 12 months of experiment this value was statistically different from that obtained in control subgroups, being respectively lower by approximately 22%, 48% and 65%. One month after terminating of chronic administration this value was close to the one found after 12 months of the study. Three months after the drug withdrawal the coefficient was higher by approximately 44% compared to the one after 12 months, which seemed to indicate an increase in capillary lumen patency. The morphometric analysis of the cerebellar cortex capillary cross-section area performed in the present study objectifies the results of qualitative ultrastructural investigations concerning the microcirculation of this CNS structure.

CD44 expression in colorectal cancer. An immunohistochemical study including correlation with cathepsin D immunoreactivity and some tumour clinicopathological features.

CD44 is a cell adhesion molecule involved in tumour growth and progression. This study was undertaken to evaluate the expression of CD44 standard protein in a series of 54 colorectal adenocarcinomas in correlation with cathepsin D immunoreactivity and some other clinicopathological variables. Formalin-fixed and paraffin-embedded tissues were investigated with anti-CD44 standard protein and anti-cathepsin D antibody. Immunolocalisation of CD44 protein and cathepsin D was performed using LSAB method. 13 (41.9%) out of 31 carcinomas without lymph-node metastases had positive CD44 expression, whereas only 6 (26.1%) out of 23 carcinomas with lymph-node metastases were found positive for CD44 expression. CD44 expression in carcinomas was positively correlated with tumour cells cathepsin D (p<0.01) immunostaining. statistically significant correlation was found between the expression of CD44 standard protein and the tumour site, age and sex of the patients. These results suggest that the standard-type CD44 protein lymph-node metastases, probably with cooperation of cathepsin D.

Ultrastructure of Purkinje cell perikarya and their dendritic processes in the rat cerebellar cortex in experimental encephalopathy induced by chronic application of valproate.

Long-term intragastric administration of the antiepileptic drug sodium valproate (Vuprol Polfa) to rats for 1, 3, 6, 9 and 12 months, once daily at the effective dose of 200 mg/kg body weight showed morphological evidence of encephalopathy, manifested by numerous nonspecific changes within Purkinje cell perikarya and their dendritic processes. The first ultrastructural abnormalities appeared after 3 months. They became more severe in animals with longer survival and were most pronounced after 12 months. The changes were maintained both 1 and 3 months after drug withdrawal. Mitochondria of Purkinje cell perikarya were most severely affected. Damage to mitochondria was accompanied by disintegration and fragmentation of granular endoplasmic reticulum, dilation of channels and cisterns of Golgi apparatus, enlargement of smooth endoplasmic reticulum elements including submembranous cisterns, and accumulation of profuse lipofuscin deposits. Frequently, Purkinje cells appeared as dark ischemic neurones, with focally damaged cellular membrane and features of disintegration. Swollen Bergmann's astrocytes were seen among damaged Purkinje cells or at the site of their loss. The general pattern of submicroscopic alterations of Purkinje cell perikarya suggested severe disorders in several intercellular biochemical extents, including inhibition of oxidative phosphorylation and abnormal protein synthesis, both of which could lead to lethal damage. Ultrastructural abnormalities within dendrites were characterized by damage to elements of smooth endoplasmic reticulum, which was considerably enlarged, with formation of large vacuolar structures situated deep in the dendroplasm. Mitochondrial lesions and alterations in cytoskeletal elements--disintegration of microtubules or even their complete loss--were also observed. The general pattern of abnormalities within the organelles and cytoskeletal elements of dendritic processes in Purkinje cells in the VPA chronic experimental model imply that there are disturbances in detoxication processes. Furthermore these changes were irreversible, as they were maintained after drug withdrawal.

Morphological examinations of hard tissues of periodontium and evaluation of selected processes of lipid peroxidation in blood serum of rats in the course of experimental periodontitis.

The problem of teeth loss as a result of periodontitis is growing continuously. In the study we aimed to show the correlation between the disease and lipid metabolism disorders. We performed morphological examinations of hard tissues of rats' periodontium in the course of experimental ligature-induced periodontitis and we demonstrated the destruction of alveolodental ligament. The following changes were observed: degenerative changes including necrosis within periodontium, progressive destruction of bone mass of alveolar process of the mandible in the region of inflammatory infiltration. Simultaneously, biochemical examinations of blood serum were performed revealing decrease of basic antioxidant enzymes activities: SOD, GSH-Px, GSH-R with simultaneous increase of MDA--the final product of lipid peroxidation.

Blind liver biopsy in children--diagnostic significance and complications in authors' own material.

The purpose of the study is the analysis of liver biopsy performed in patients hospitalized at 3rd Department of Pediatric Diseases of Medical University in Bialystok in 1993-1998 with particular regard to the assessment of indications and the incidence of complications connected with the procedure. The procedure was conducted on the group of 250 patients aged 1-17 years with the use of Menghini needles. The indications for diagnostic liver puncture included chronic infection with HBV (228 children), chronic infection with HCV (6 children), chronic hypertransaminasemia (13 children), hepatomegaly (2 children) and hyperbilirubinemia (1 child). Complications were observed in 3 patients (1.2%). Blind liver biopsy is an important diagnostic tool in pediatric hepatology but possible occurrence of life-threatening complications suggests that it be used with caution. The paper was presented as a part of hepatological session of XXVI Congress of Polish Pediatricians in Mikolajki on 6-9th June 1999.

Histological diagnosis of gastritis based on Sydney System.

The review of literature concerning gastritis, especially the chronic form has been carried out. Based on published data and own authors experience an application of the Sydney System in differential diagnosis of gastritis was presented.

Effects of long-term administration of the antiepileptic drug--sodium valproate upon the ultrastructure of hepatocytes in rats.

Chronic intragastric application (1, 3, 6, 9 and 12 months) of the antiepileptic drug--sodium valproate (VPA; Vupral "Polfa") to rats in the effective dose of 200 mg/kg b.w./day exerts hepatotoxic effect after 9 and 12 months of the experiment. The first ultrastructural changes in hepatocytes were observed after 3 months of the drug administration. These became more intense in the subsequent stages of the experiment, to be most pronounced after 12 months. The most striking changes were in the mitochondria (significant swelling, an increase in their number, degeneration of matrix and cristae, disruption of the outer mitochondrial membrane) and in peroxisomes (proliferation, enlargement and the presence of distinct nucleoids). Further alterations in hepatocytes manifested themselves in: microvesicular fatty change with cholesterolosis (cholesterol clefts), damage to the cellular membrane of the sinusoidal pole with dilation of the perisinusoidal space of Disse, presence of cystern-like cytoplasmic vacuoles in the sinusoidal region, filled with plasma-like material and focal cytoplasmic necrosis. The changes in hepatocytes coexisted with the swelling and activation of sinusoidal cells, endothelial cells and Kupffer cells. The author suggests that mitochondria and peroxisomes considerably contribute to the morphogenesis of hepatocyte damage by VPA in the chronic experimental model.

Cyclophosphamide in diffuse lung damage.

Some cyclophosphamide toxic effects on lung tissue are presented. Cyclophosphamide metabolism, pathogenesis of lung damage and morphological lung tissue changes caused by that agent were characterized. Attention was focused on BAL evaluation as a useful method in the monitoring of lung tissue damage degree.

Cyclophosphamide-induced changes of serum angiotensin converting enzyme activity and pulmonary microvessels ultrastructure.

The effect of cyclophosphamide (CP) on the ultrastructure of the lung tissue and the activity of angiotensin converting enzyme (ACE) in serum was evaluated in rats. The animals were given cyclophosphamide (CP) in a single intraperitoneal dose of 150 mg/kg b.w. ACE activity was evaluated in the blood serum collected from the left ventricle of the heart using the spectrophometric method. In all time subgroups, the CP-receiving animals showed a decrease in ACE activity. Ultrastructural examinations of CP-treated animals revealed increased adhesion of neutrophiles and monocytes to the damage endothelium of the alveolar septa vessels and focally accumulation of the platelets.

Effects of pentoxifylline on some peripheral blood parameters and haemostasis in acute pulmonary tissue injury after cyclophosphamide.

Cyclophosphamide (CP) is one of the widely used cytostatic drugs, with a strong toxic influence on pulmonary tissue. Experimental works have shown that a single high dose of CP causes injury to all elements of the interalveolar septum, especially to the alveolar endothelial and epithelial cells. The aim of this work was to evaluate the influence of pentoxifylline (PTXF) (30 mg/kg b.w.) on the ultrastructure of lungs capillaries and blood cell count as well as plasma fibrinogen levels in Wistar rats after intraperitoneal injection of 150 mg/b.w. CP. We established that in the doses applied PTXF had no statistically significant influence on the number of leukocytes and erythrocytes determined in the left ventricular blood of rats receiving CP, while the number of white cells from animals given PTXF only was higher than in controls. There was a smaller decrease in the number of platelets (p < 0.05) and smaller reduction in fibrinogen level (p < 0.01) in the serum of PTXF-CP animals than in the CP group. The results obtained suggest a protective effect of PTXF on CP induced changes, which have been evidenced in some of the parameters examined. Ultrastructural examinations found the lungs to be the organ of extramedullary thrombocytopoiesis in CP-treated animals and revealed that platelet accumulation in the system of lung capillaries was a potential cause of the decrease observed in the number of blood platelets following CP administration.

A comparison of the peritoneal cell population of pregnant rabbits after LPS or TNF-alpha induced septic shock.

Septic shock is a catastrophic consequence of invasive infection. Unfortunately, recent advances in surgical and medical sciences have not significantly reduced the overall mortality from septic shock. Bacterial antigens stimulate a cascade of cytokine release; each cytokine helps the host to overcome infection, but their excessive production causes them to trigger events that lead to septic syndrome and shock. Tumour necrosis factor (TNF-alpha) has a pivotal role in orchestrating the events leading to septic shock. Intraperitoneal administration of certain substances can increase the number and phagocytic activity of cells, which reach naturally the site of infection. Activation of the immunity cells in the peritoneal cavity and their immunocompetence are found to be responsible for the organism protection against abdominal cavity infections. Macrophages, lymphocytes and granulocytes of low activity in the non-stimulated peritoneal cavity become significant due to the influence of numerous biologically active substances. This study was designed to determine the peritoneal response to local administration of LPS or TNF-alpha in the course of experimental septic shock.

[Morphologic analysis of congenital central nervous system malformations in children from the first of life dying in the years 1986-1990]. / Analiza morfologiczna wad wrodzonych osrodkowego ukladu nerwowego u dzieci od 1. roku zycia zmarlych w latach 1986-1990.

An incidence and morphology of the CNS congenital malformations in newborn babies and infants were analysed in the consecutive autopsies carried out in 1986-1990, i.e. following Tscharnobyl disaster. The obtained results were compared to those seen in the two earlier periods (1976-1980 and 1981-1986). In 1986-1990, a percentage of autopsies showing congenital CNS malformations increased approximately by two-fold (15%). The highest percentage of such malformations in specific years of the analysed period was noted in 1990 (20%). Central nervous system malformations were more frequent in female sex (57%) than in male sex (43%). In 64% of cases newborn babies were affected. A percentage of CNS malformations coexisting with other congenital malformations increased to 40% in the analysed period of time (from 29.4% in 1976-1985). Meningomyelocele (41%), congenital hydrocephalus (21.5%), multiple anomalies in brain (14%), and anencephaly (12.6%) constituted the most frequent group of CNS malformations. In 1986-1990, an incidence of meningomyelocele increased by more than two-fold (if this anomaly coexisted with hydrocephaly, an increase in the incidence exceeded three-fold), and hydrocephaly as well as an increase in the incidence of anencephaly by 1.5 times in comparison with 1976-1985.

[Coexistence of some diseases and analysis of death causes based on autopsy examinations carried out in liver cirrhosis patients based on autopsy observations in 1976-1990]. / Wspólistnienie niektórych schorzen oraz analiza przyczyn zgonów u osób z marskoscia watroby w swietle obserwacji autopsyjnych w latach 1976-1990.

The 19,094 autopsy examinations carried out between 1976-1990 revealed 698 (3.65%) case of cirrhosis, of which 64.6% were men. During the last 5 years the percentage of coexistance of hepatoma (hepatocellular carcinoma) with cirrhosis was higher 5-year periods (5.8%; 5.4%). Moreover, the same changing interrelation was observed for other malignancies and cirrhosis-higher (15%) in the last period than in the proceeding years (11.1%; 11.3%). The severity of atherosclerotic changes and coexistance of peptic ulcers, gall bladder disease and productive pulmonary tuberculosis in cirrhotic patients were also assessed. Finally the direct causes of these patients' death were discussed.