BACKGROUND:Lysinuric protein intolerance (LPI) is a multi-organ metabolic disorder characterized by the imbalance in absorption and excretion of cationic amino acids like lysine, ornithine and arginine. Infants with LPI typically present with recurrent vomiting, poor growth, interstitial lung disease or renal impairment. The early onset of pulmonary alveolar proteinosis (PAP) has been reported to be associated with a severe form of LPI. Treatment of PAP most commonly consists of whole-lung lavage (WLL) and in autoimmune PAP, granulocyte-macrophage colony stimulating factor (GM-CSF) administration. Nevertheless, GM-CSF therapy in LPI-associated PAP has not been scientifically justified.CASE PRESENTATION:We describe the case of an 8-month-old infant presenting with respiratory failure due to LPI associated with PAP, who was twice treated with WLL; firstly, while on veno-venous ECMO assistance and then by the use of a selective bronchial blocker. After the two treatments with WLL, she was weaned from daytime respiratory support while on initially subcutaneous, then on inhaled GM-CSF therapy.CONCLUSIONS:This case supports the notion that GM-CSF therapy might be of benefit in patients with LPI-associated PAP. Further studies are needed to clarify the exact mechanism of GM-CSF in patients with LPI-associated PAP.

Background Paediatric patients are often exposed to subtherapeutic levels or treatment failure of beta-lactams, and prolonged infusion may be beneficial. We aimed to investigate the efficacy and safety of extended infusion (EI; defined as >= 3 h) or continuous infusion vs. short, intermittent infusion (SI; defined as <= 60 min) of beta-lactams in patients <21 years of age.Methods A systematic review and meta-analysis was conducted to compare EI and continuous infusion with SI of beta-lactams in children. A systematic search was performed in MEDLINE (via PubMed), Embase, CENTRAL, and Scopus databases for randomised controlled trials (RCTs) and observational studies published from database inception up to August 22, 2023. Any comparative study concerned with mortality, clinical efficacy, adverse events, or plasma concentrations of beta-lactams for any infection was eligible. Case reports, case series, and patients aged >21 years were excluded. Odds ratios (OR) and median differences with 95% confidence intervals (CI) were calculated using a random-effects model. Risk of bias (ROB) was assessed using ROB2 and ROBINS-I tools. The protocol was registered with PROSPERO, CRD42022375397.Findings In total, 19,980 articles were screened, out of which 19 studies (4195 patients) were included in the meta-analysis. EI administration was associated with a significantly lower all-cause mortality in both RCTs and non-RCTs [OR 0.74; CI 0.55-0.99; I-2 = 0%; CI 0-58%]. Early microbiological eradication was higher with EI [OR 3.18; CI 2.24-4.51; I-2 = 0%; CI 0-90%], but the clinical cure did not differ significantly between the two groups [OR 1.20; CI 0.17-8.71; I-2 = 79%; CI 32-93%]. Achieving the optimal plasma level (50-100% fT > MIC) appeared favourable in the EI group compared to the SI. No significant differences were observed in the adverse events. The overall ROB was high because of the small sample sizes and clinically heterogeneous populations.Interpretation Our findings suggest that extended infusion of beta-lactams was associated with lower mortality and increased microbiological eradication and was considered safe compared to short-term infusion.

A primer hyperoxaluria ritka anyagcsere-betegség. A glioxilát anyagcserezavara miatt nagy mennyiségű oxalát keletkezik, amely – mivel rosszul oldódik – a szövetekben oxalátkicsapódáshoz, oxalosishoz vezet. Az oxalosis elsőként a veséket érinti, enyhébb formában urolithiasishoz, súlyos esetekben nephrocalcinosishoz és progresszív vesefunkció-romláshoz, végül végstádiumú veseelégtelenséghez vezet. A betegség típusát genetikai vizsgálattal lehet pontosan azonosítani. Klinikai megjelenése nagyon változatos. Enyhébb esetekben az oxaláttermelés csökkentése és kicsapódásának gátlása jelenthet terápiás lehetőséget, súlyosabb esetben vesepótló kezelésre lehet szükség. A betegség kóroki terápiáját egyelőre a májátültetés jelenti. Az oxalát kialakulásában kulcsszerepet játszó enzim RNS-interferencia útján történő gátlásán alapuló biológiai terápia új perspektívát jelenthet a betegség komplex kezelésében.

Whole or partial trisomy of the short arm of chromosome 9 (9p) is considered to be one of the more frequent chromosome abnormalities compatible with life. The duplication may affect various organs, however the most common symptoms are certain specific facial dysmorphisms and abnormalities of the fingers, toes and nails. A one month old boy presented with failure to thrive, jaundice, ventricular septal defect (VSD) and dysmorphic face. He displayed symptoms of heart failure. The cardiologic examination revealed a significant VSD, hypoplasia of the aortic arch, pulmonary hypertension, decompensated circulatory failure and moderate left ventricle dysfunction. Routine cytogenetic analysis revealed a supernumerary marker chromosome. Fluorescence in situ hybridization (FISH) identified this as the short arm of chromosome 9. The child's karyotype was determined as 47,XY,+der(9)dup(9)(p10p24)dn. Due to his worsening condition and the high risk of the operation, it was decided to forego the procedure. After a short palliative care the child passed away. The child's clinical presentation and the uncharacteristic severity of his condition show that chromosome abnormalities involving duplicated genetic material are extremely heterogeneous. Thus treatment of each child should be individualized and may also involve difficult ethical considerations.

Establishment of a proper hemodynamic monitoring system in order to achieve optimal care among critically ill patients is fundamental. In contrast to invasive patient-checking systems, which were introduced decades ago and used in both adult and pediatric intensive care, the non-invasive methods have become more popular in recent years due to technical advancements in intensive care and patient monitoring. This increase in popularity can be attributed to the higher degree of safety and reduced complication rates as well as to its being more economical. Our summary focuses on the ICON® patient monitoring system. This newly engineered, non-invasive tool is based on electrical cardiometry, and uses hemodynamic parameters in both neonatal and pediatric care as well as in adults. The operating principle is simple: the conductivity of the blood in the aorta shows time-dependent changes. Prior to the opening of the aortic valve, the orientation of the red blood cells (RBCs) is random, and it is not until the contraction of the aorta that the RBCs and the opening of the aortic valve achieve a parallel position. The tool senses the conductivity between four placed electrodes, and measures the stroke volume (SV) and cardiac output (CO), before calculating other additional parameters (eg.: systemic vascular resistance) by tracing the variation of bioimpedance according to changes in the heart cycle. The most important advantages of ICON® are the measurements that are made available immediately as well as continuously, and the low complication rate that originates from its non-invasive operation. ICON® is a new, promising hemodynamic device in the tool belt of intensive care. Due to the nature of the device, it is possible to evaluate the status of the patient on a continuous basis, allowing for optimal care. To identify the more accurate clinical indications further measures will be necessary. Orv Hetil. 2018; 159(44): 1775-1781.

Abstract: Establishment of a proper hemodynamic monitoring system in order to achieve optimal care among critically ill patients is fundamental. In contrast to invasive patient-checking systems, which were introduced decades ago and used in both adult and pediatric intensive care, the non-invasive methods have become more popular in recent years due to technical advancements in intensive care and patient monitoring. This increase in popularity can be attributed to the higher degree of safety and reduced complication rates as well as to its being more economical. Our summary focuses on the ICON® patient monitoring system. This newly engineered, non-invasive tool is based on electrical cardiometry, and uses hemodynamic parameters in both neonatal and pediatric care as well as in adults. The operating principle is simple: the conductivity of the blood in the aorta shows time-dependent changes. Prior to the opening of the aortic valve, the orientation of the red blood cells (RBCs) is random, and it is not until the contraction of the aorta that the RBCs and the opening of the aortic valve achieve a parallel position. The tool senses the conductivity between four placed electrodes, and measures the stroke volume (SV) and cardiac output (CO), before calculating other additional parameters (eg.: systemic vascular resistance) by tracing the variation of bioimpedance according to changes in the heart cycle. The most important advantages of ICON® are the measurements that are made available immediately as well as continuously, and the low complication rate that originates from its non-invasive operation. ICON® is a new, promising hemodynamic device in the tool belt of intensive care. Due to the nature of the device, it is possible to evaluate the status of the patient on a continuous basis, allowing for optimal care. To identify the more accurate clinical indications further measures will be necessary. Orv Hetil. 2018; 159(44): 1775–1781.

Colonic perforation is an uncommon but severe complication of colonoscopy. We present a case of a 10 -year-old girl who was treated conservatively for inflammatory bowel disease for 6 years. She underwent a diagnostic colonoscopy after which she developed acute respiratory failure. Urgent bedside imaging series revealed pneumothorax, pneumomediastinum and pneumoperitoneum. A chest tube was immediately inserted, and then she was intubated and ventilated. Laparotomy revealed a 1 cm wide perforation of the transverse colon. The rupture was closed and a terminal ileostomy was performed. The postoperative course was uneventful. To best of our knowledge no other case of pneumothorax, pneumomediastinum and pneumoperitoneum after colonoscopy in children has been reported so far in literature.

BACKGROUND:Continuous Glucose Monitoring (CGM) has become an increasingly investigated tool, especially with regards to monitoring of diabetic and critical care patients. The continuous glucose data allows the calculation of several glucose variability parameters, however, without specific application the interpretation of the results is time-consuming, utilizing extreme efforts. Our aim was to create an open access software [Glycemic Variability Analyzer Program (GVAP)], readily available to calculate the most common parameters of the glucose variability and to test its usability.METHODS:The GVAP was developed in MATLAB® 2010b environment. The calculated parameters were the following: average area above/below the target range (Avg. AUC-H/L); Percentage Spent Above/Below the Target Range (PATR/PBTR); Continuous Overall Net Glycemic Action (CONGA); Mean of Daily Differences (MODD); Mean Amplitude of Glycemic Excursions (MAGE). For verification purposes we selected 14 CGM curves of pediatric critical care patients. Medtronic® Guardian® Real-Time with Enlite® sensor was used. The reference values were obtained from Medtronic®(')s own software for Avg. AUC-H/L and PATR/PBTR, from GlyCulator for MODD and CONGA, and using manual calculation for MAGE.RESULTS:The Pearson and Spearman correlation coefficients were above 0.99 for all parameters. The initial execution took 30 minutes, for further analysis with the Windows® Standalone Application approximately 1 minute was needed.CONCLUSIONS:The GVAP is a reliable open access program for analyzing different glycemic variability parameters, hence it could be a useful tool for the study of glycemic control among critically ill patients.

Background and aims: One of the principal physiological processes of the immediate postnatal period is the rapid disappearance of the transpulmonary fluid. Delayed removal of the alveolar fluid results in neonatal respiratory distress. This process is mainly governed by the postnatal activation of the amilorid sensitive Na-channel (ENaC) and the basolateral Na+-K+-ATPase. The role of aquaporin (AQP) water channels was also suggested as AQP4 mRNA exhibits a sharp and transient rise on the first day of life. Aims: To highlight the potential role of AQP4 water channels in the reabsorption of fetal alveolar fluid based on the case of a neonate. Methods: A male neonate born by Caesarian section due to maternal neuroyelitis optica (NMO) was admitted at 38 weeks of gestation. Apgar scores were 6 and 9, birth weight was 3550 g. He developed a severe respiratory failure necessitating 100% FiO2 on HFOV, inhaled NO and repeated use of surfactant, without significant clinical effect. Infectious or developmental causes of RDS were excluded. Results: Serum AQP4 autoantibodies were tested because of maternal NMO at 1 week and after complete remission of the respiratory symptoms, at 3 weeks. The first result was positive, while AQP4 autoantibodies disappeared on the second test. Conclusions: The presented case is the first newborn in which AQP4 autoantibodies were transiently detected in the presence of spontaneously regressing severe respiratory failure. The defective function of these water channels is supposed to play a significant role in this unusual case of neonatal respiratory failure.

Background and aims The last decade gave clear evidence that hyper/hypoglycemia and glucose variability are associated with increased mortality in critically ill patients. Continuous glucose monitor (CGM) is a new device in paediatric critical care units (PICU) with clear advantages in glucose monitoring. The aim of our study was to survey the incidence of glucose regulation disorders in our PICU and specify the association between the PRISM III score and the glycemic variability [mean amplitude of glycemic action (MAGE)]. Methods We evaluated 22 children: mean age: 1.3 years, mean length of PICU stay: 18 days; 20/22 patients were on invasive mechanical ventilation; 6/22 needed vasoactive agent therapy. CGM duration: 1–12 days. Interstitial glucose level was monitored by Guardian® REAL Time CGM (Medtronic®). Reference glucose values were obtained from blood gas analyzer or point-of-care glucose analyzer. We used Spearman correlation to evaluate the association between PRISM III and the MAGE. Results Hypo- and hyperglycemia (CGM glucose < 55 mg/dl / CGM glucose > 180 mg/dl) were detected in 4.6% and 2.5% of measurements, respectively. The mean MAGE (meaningful excursion >45 mg/dl) and PRISM III were 78 mg/dl and 19. We found a significant correlation between PRISM III and MAGE (r = 0,55; p < 0.05). Pearson’s correlation coefficient (0.82) and Clarke Error Grid analysis (96% clinical accuracy) proved a good reliability of the CGM. Conclusions Glucose homeostasis disorders are frequent in the PICU; hypoglycemia being more commonly detected. Increased PRISM III score contributes significantly to the elevation of glucose variability.

Acta PaediatricaVolume 102, Issue 4 p. e178-e180 Short Communication Autonomic dysfunction of glucose homoeostasis in congenital central hypoventilation syndrome Gábor Marics, Corresponding Author Gábor Marics [email protected] First Department of Paediatrics, Semmelweis University, Budapest, Hungary Correspondence Gábor Marics, M.D., First Department of Paediatrics, Semmelweis University, Budapest, Bókay u. 53., H-1083, Hungary. Tel.: + 0036 1 334 3186 | Fax: + 0036 1 303 6077 | Email: [email protected]Search for more papers by this authorJeanne Amiel, Jeanne Amiel Unité INSERM U781, Faculté Paris-Descartes, Institut IMAGINE, Paris, FranceSearch for more papers by this authorBarbara Vatai, Barbara Vatai First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this authorCsaba Lódi, Csaba Lódi First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this authorBorbála Mikos, Borbála Mikos Bethesda Children's Hospital, Budapest, HungarySearch for more papers by this authorPéter Tóth-Heyn, Péter Tóth-Heyn First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this author Gábor Marics, Corresponding Author Gábor Marics [email protected] First Department of Paediatrics, Semmelweis University, Budapest, Hungary Correspondence Gábor Marics, M.D., First Department of Paediatrics, Semmelweis University, Budapest, Bókay u. 53., H-1083, Hungary. Tel.: + 0036 1 334 3186 | Fax: + 0036 1 303 6077 | Email: [email protected]Search for more papers by this authorJeanne Amiel, Jeanne Amiel Unité INSERM U781, Faculté Paris-Descartes, Institut IMAGINE, Paris, FranceSearch for more papers by this authorBarbara Vatai, Barbara Vatai First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this authorCsaba Lódi, Csaba Lódi First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this authorBorbála Mikos, Borbála Mikos Bethesda Children's Hospital, Budapest, HungarySearch for more papers by this authorPéter Tóth-Heyn, Péter Tóth-Heyn First Department of Paediatrics, Semmelweis University, Budapest, HungarySearch for more papers by this author First published: 10 December 2012 https://doi.org/10.1111/apa.12125Citations: 13Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onEmailFacebookTwitterLinkedInRedditWechat References 1Amiel J, Laudier B, Attie-Bitach T, Trang H, de Pontual L, Gener B, et al. Polyalanine expansion and frameshift mutations of the paired-like homeobox gene PHOX2B in congenital central hypoventilation syndrome. Nat Genet 2003; 33: 459–61. 2Farina MI, Scarani R, Po C, Agosto C, Ottonello G, Benini F. Congenital central hypoventilation syndrome and hypoglycaemia. Acta Paediatr 2012; 101: e92–6. 3Meissner T, Rabl W, Mohnike K, Scholl S, Santer R, Mayatepek E. Hyperinsulinism in syndromal disorders. Acta Paediatr 2001; 90: 856–9. 4Hennewig U, Hadzik B, Vogel M, Meissner T, Goecke T, Peters H, et al. Congenital central hypoventilation syndrome with hyperinsulinism in a preterm infant. J Hum Genet 2008; 53: 573–7. 5Battelino T, Bode BW. Continuous glucose monitoring in 2010. Int J Clin Pract Suppl 2011; 170: 10–5. 6Bridges BC, Preissig CM, Maher KO, Rigby MR. Continuous glucose monitors prove highly accurate in critically ill children. Crit Care 2010; 14: R176. 7Crofton PM, Midgley PC. Cortisol and growth hormone responses to spontaneous hypoglycaemia in infants and children. Arch Dis Child 2004; 89: 472–8. 8Pardal R, Lopez-Barneo J. Low glucose-sensing cells in the carotid body. Nat Neurosci 2002; 5: 197–8. 9Kiba T. Relationships between the autonomic nervous system and the pancreas including regulation of regeneration and apoptosis: recent developments. Pancreas 2004; 29: e51–8. 10Koyama Y, Coker RH, Stone EE, Lacy DB, Jabbour K, Williams PE, et al. Evidence that carotid bodies play an important role in glucoregulation in vivo. Diabetes 2000; 49: 1434–42. 11Cutz E, Ma TK, Perrin DG, Moore AM, Becker LE. Peripheral chemoreceptors in congenital central hypoventilation syndrome. Am J Respir Crit Care Med 1997; 155: 358–63. 12Dubreuil V, Ramanantsoa N, Trochet D, Vaubourg V, Amiel J, Gallego J, et al. A human mutation in Phox2b causes lack of CO2 chemosensitivity, fatal central apnea, and specific loss of parafacial neurons. Proc Natl Acad Sci USA 2008; 105: 1067–72. 13Nekrep N, Wang J, Miyatsuka T, German MS. Signals from the neural crest regulate beta-cell mass in the pancreas. Development 2008; 135: 2151–60. Citing Literature Volume102, Issue4April 2013Pages e178-e180 ReferencesRelatedInformation

Background The differential diagnosis of respiratory alkalosis (RA) includes a state called central neurogenic hyperventilation (CNH). In the few reported cases of CNH the etiology was a stimulation of the respiratory center by an infiltrative tumor in the cerebral pons. In some cases, a shift in the cerebral pH to acidic range was also hypothesized. Case Report We report the case of a six year-old boy with a known Pearson syndrome, a mitochondrial disorder affecting bone marrow, pancreas and renal tubules. He was admitted to our PICU with deteriorating mental status and compensated metabolic acidosis (lactic, hyperchloremic and tubular). On admission, blood gas analysis showed a pH of 7.30 with a disproportionately low compensating pCO2 of 10 mmHg (HCO3 4.9 mmol/L). Serum HCO3 normalized by substitution (21.0 mmol/L), when he developed a RA (pH 7.51, pCO2 24 mmHg) persisting over 48 hours, even during sleeping periods. After reviewing his previous blood gas results, this phenomenon was present for years. After excluding known etiologies of RA, we suspected CNH caused by intra-cerebral acidosis. The pH and HCO3 were lower, while lactate was higher in cerebro-spinal fluid than in serum. An MR spectroscopy confirmed cerebral lactate accumulation, showing a peak in the posterior cerebrum. Encephalopathy is not among the classic manifestations of Pearson syndrome. Conclusion We were able to demonstrate elevated local lactate level leading to intra-cerebral acidosis, stimulation of the respiratory center and causing long-standing hyperventilation. This phenomenon adds a new aspect to the complex clinical picture of mitochondrial disorders.

A sejtkapcsolatok a daganatokban megvaltoznak, kulonosen a tight junction (TJ) gerincet alkoto claudin (CLDN) feherje csalad komponensei, melynek 24 tipusa ismert emberben. Tobb daganatban elsőkent vizsgalatuk a CLDN expresszio eltereseit protein es mRNS szinten. Megallapitottuk, hogy az egyes CLDN-ok expresszioja jelentősen nő a nyelőcső rakban, a Barrett oesophagusban es adenocarcinomaban (ACC). A CLDN mintazat elkuloniti a hepatoblastoma es endometrialis carcinoma formait, a pancreas endocrin tumorait es a ductalis ACC-t, valamint a tudőtumorokat, a primer es metasztatikus majdaganatokat. Egyes extracellularis matrix (ECM) komponensek, igy az agrin, valamint a matrilin2 expresszioja es lokalizacioja megvaltozik a cirrhosisban es majrakokban. Fenti vizsgalatok jelentősege, hogy szamos daganatban feltarta a sejtkapcsolatok es ECM valtozasait es igazolta, hogy az elteresek jellemzők az egyes tumorokra, kovetik a kiindulasi szovet sejtkapcsolo strukturainak feherje osszetetelet, bar expressziojuk merteke valtozik. A vizsgalt daganatok CLDN mintazata diagnosztikus ertekű. A vizsgalatok igazoltak, hogy a TJ dinamikusan valtozo protein osszetetele a carcinogenesis soran bonyolult funkcionalis valtozast takar es az egyes osszetevők aranya es egymassal valo kapcsolata a dontő az intakt sejtkapcsolat kialakulasaban. | Cell junctions change in tumors, especially the components of the claudin (CLDN) protein family (24 types of which are known in humans) forming the backbone of tight junctions (TJ). We were first to study CLDN expression alterations in several tumors at protein and mRNA levels. Our studies confirmed that expression of certain CLDNs shows significant increase in oesophageal squamous cell carcinoma, Barrett?s oesophagus and adenocarcinoma (ACC). The CLDN pattern differentiates components of hepatoblastomas and 2 types of endometrial carcinomas, endocrine tumors of the pancreas, ductal ACCs, lung tumors, primary and metastatic liver tumors. Certain extracellular matrix (ECM) components, as agrin, matrilin-2 exhibited altered expression and location in cirrhosis and liver cancers. Our observations revealed TJ and ECM changes in several tumor types, confirming that the changes are characteristic to the tumor. The CLDN pattern of the studied tumors are of diagnostic significance. Our studies confirmed that the dynamically changing protein composition of TJs covers complex functional changes during carcinogenesis, and the proportion and interrelationship of various components are decisive in the development of intact TJs.

Previously, increased diameter and enhanced myogenic tone were seen after 2-week 45o head-up (HUT2) in the rat. We studied the reversibility and the effect of extended tilt on this phenomenon using two experimental groups: HUT2 plus 2-week horizontal (HUT2HOR2), and 4-week tilting (HUT4). 4-weeks in normal cages (NC4) served as control. Diameter of saphenous vein (SV) in 2-20 mm Hg pressure range, wall and media thickness, endothelial and smooth muscle cell densities, and cell proliferation were measured. The diameter of SV from HUT4 was significantly larger compared with HUT2HOR2 or NC4 within the whole pressure range both in Krebs-Ringer (870.4+/-21.3 vs. 778.2+/-24.9 and 771.6+/-28.1 microm at 10 mm Hg, respectively) and in Ca(2+)-free solution. Myogenic and norepinephrine-induced vascular tone, wall and media thickness did not differ among the three groups. Endothelial cell density decreased in HUT4 (10.7+/-1.2) vs. HUT2HOR2 (15.1+/-1.0) and NC4 (15.3+/-0.6), while that of smooth muscle was unchanged. No cell proliferation marker was seen. In conclusion, both increased diameter and enhanced myogenic tone of SV seen in HUT2 proved to be reversible. HUT4 resulted in increased SV diameter, similarly to HUT2, however, vascular tone was not amplified. This suggests that a prolonged orthostatic load may readjust the function of smooth muscle.

Heparan sulfate proteoglycans mediate cell adhesion and control the activities of numerous growth and motility factors. They play a critical role in carcinogenesis and tumor progression. Agrin is a large multidomain heparan sulfate proteoglycan associated with basement membranes in several tissues. The expression of agrin in the liver has recently been described under physiologic and pathologic conditions. However, little is known about its role in malignancies. We aimed to study the mRNA and protein expression of agrin in cholangiocarcinoma (CC) and focused on the differences between CC and hepatocellular carcinoma (HCC). Eighty surgically removed liver specimens were studied by immunohistochemistry. Representative samples were used for immunoblotting. mRNA expression was measured in 32 samples by real-time polymerase chain reaction. By immunohistochemistry, agrin was seen around bile ducts and blood vessels within the portal areas in the normal liver. Although no expression was found within the hepatic lobules, agrin was deposited in the neovascular basement membrane in HCCs. Agrin was abundant in the tumor-specific basement membrane in well-differentiated areas of CCs, whereas with immunostaining, it was fragmented, decreased, or it even disappeared in less differentiated areas and sites of infiltration. By real-time polymerase chain reaction, up-regulation of agrin expression was measured in HCCs compared with that in the normal liver. CC samples showed an even higher expression of agrin. Immunoblotting confirmed these findings. Our results indicate that agrin might play an important role in neoangiogenesis in human HCC, being a part of the newly formed vasculature. In CC, however, agrin might be involved in tumor progression.

The matrilins represent a new family of oligomeric proteins that are assumed to act as adapter molecules connecting other proteins and proteoglycans in the extracellular matrix. Matrilin-2, the largest member of the family, displays a broad tissue distribution. It incorporates into loose and dense connective tissue and becomes associated with some basement membranes. The aim of our study was to analyse the expression of matrilin-2 in two liver regeneration models and to identify its cellular origin. Liver regeneration was induced in rats by partial hepatectomy (PH) and by the 2-acetylaminofluorene (AAF)/partial hepatectomy (PH) experimental models. Formalin fixed, paraffin embedded tissue sections were used for immunohistochemistry applying a rabbit matrilin-2 polyclonal antibody. Matrilin-2 was detected in normal rat liver and partially hepatectomized liver in the portal area, but could not be demonstrated in the acini. Matrilin-2 mRNA expression was analysed by RT-PCR and in situ hybridization. In the AAF/PH model the oval cells but not the hepatocytes produced matrilin-2 mRNA. Increase in protein level in the AAF/PH regenerating liver model was demonstrated by Western blotting. The protein was present in the basement membrane zone around the tubules formed by oval cells. Our data show that hepatic oval cells produce matrilin-2, a novel ECM protein, suggesting that matrilin-2 is an important component of ECM during stem cell-driven liver regeneration.

04B. Molecular and cellular biology (b) Biliary tract pathophysiology$121 liver, allowed to adhere in culture and submitted to a catalytic system, which reduces oxygen concentration to less than 1% within 30 minutes.Gene expressions were assessed by quantitative real-time RT-PCR.Results: In vivo, 24 hours after the onset of arterial liver ischemia in rats, cholestatic changes in serum were detected together with a decrease in bile flow and in bile acid and bicarbonate output in bile.No regurgitation of fluorescent ursodeoxycholic acid was detected, eliminating bile duct leakage as a major mechanism of cholestasis in this model.VEGF, a hypoxia-regulated gene, was induced in both hepatocytes and cholangiocytes, as ascertained by immunohistochemistry. Concomitantly, hepatic mRNA levels of ntcp, bsep and mrp2 were significantly reduced (by 70%, 50% and 70%, respectively), whereas Cftr mRNA levels were increased (by more than 4 fold), even though ductular reaction was not yet present at this time.In vitro, hypoxia caused a decrease in ntcp, bsep and mrp2 transcripts in hepatocytes (by 70%, 80% and 70%, respectively), and an increase in cftr transcripts (by 3 fold), associated with a rise in cAMP (by more than 10 fold) in cholangiocytes.Conclusion: These results demonstrate that hypoxia induces differential regulations in the expression of hepatobiliary transporters.The downregulation of transporters involved in bile salt-dependent and -independent secretion in hepatocytes may contribute to cholestasis.By contrast the upregulation of CFTR in cholangiocytes may, together with increased cAME a major regulator of CFTR expression and activity, and with subsequent ductular proliferation, provide a defense mechanism.