Characteristic gene expression induced by polyurethane foam/spheroid culture of hepatoma cell line, Hep G2 as a promising cell source for bioartificial liver

BACKGROUND/AIMS: Polyurethane foam (PUF)/ spheroid-culture can improve liver-specific functions of hepatoma cell line, Hep G2. Therefore, gene expression profile in the PUF/spheroid culture is hypothesized to be different from that in the monolayer culture. The aim of this study is to clarify the characteristic gene expression in PUF/spheroid-cultured Hep G2 cells, as a cell source for bioartificial liver (BAL), using microarray analysis. METHODOLOGY: Morphological change and liver specific functions of ammonia removal rate and albumin synthesis rate of Hep G2 were compared between in a monolayer or PUF/spheroid culture. Microarray analysis was performed using cDNA microarrays made in Hitachi Software Engineering Co., Ltd., (Yokohama, Japan), which contains a total of 1,281 cDNA clones. RESULTS: The ammonia removal rate of Hep G2 spheroids increased to 369%, and the albumin synthesis rate of Hep G2 spheroids also increased 311% when compared with monolayer culture. In addition, the ammonia removal capacity of primary human hepatocytes in the PUF/spheroid culture was superior to that in the monolayer culture. The microarray analysis demonstrated that the PUF/spheroid-cultured Hep G2 cells expressed 39 up-regulated (more than 3.0-fold) and 31 down-regulated (less than 0.333-fold) genes. Among the 70 genes differentially expressed in PUF/spheroid cultured Hep G2 cells, subsets of glutathione S-transferase- and angio-tensin-related genes were drastically up-regulated, on the other hand, subsets of assigned for growth factor, glucocorticoid, and stress response, were down-regulated. CONCLUSIONS: Hepatoma cell line, Hep G2 cells in the PUF/spheroid culture is a promising hepatocyte source for BAL. Microarray analysis revealed a number of characteristic genes altered by the PUF/spheroid.     

Catheter ablation of fatal ventricular tachyarrhythmias storm in acute coronary syndrome—role of Purkinje fiber network

Ventricular fibrillation (VF) or ventricular tachycardia (VT) storm is a life-threatening arrhythmia. Antiarrhythmic drugs (AADs) are not necessarily effective to rescue life from such conditions. Catheter ablation (CA) targeting triggering premature ventricular contractions (PVCs) of VF or VT that originates from Purkinje fiber network (PFN) is reported to be effective, especially in idiopathic patients. However, in condition of acute coronary syndrome (ACS), the efficacy of CA is not well understood. To clarify the usefulness of CA as an alternative way to AADs, we performed CA in four patients with VF or VT storm. The Purkinje potential was seen just before the myocardial ventricular wave during sinus rhythm that became more prominent and double components during the initiating PVC at the targeted area. Following CA, spontaneous episodes of VF or VT were no longer observed. CA is an efficacious way to bail out PFN-related VF or VT storm even in ACS.     

Evaluation of a hybrid artificial liver using a polyurethane foam packed-Bed culture system in dogs

BACKGROUND AND AIMS. We developed a polyurethane foam packed-bed culture system of hepatocyte spheroids as a hybrid artificial liver (PUF-HAL), which was effective for recovery from liver failure in rat experiments. In this report, the design of a scaled-up PUF-HAL for dogs is described and evaluated using a dog acute liver failure model. METHODS. Warm ischemic liver failure was induced with a portocaval shunt in each dog. The dogs were divided into two groups: (1) a control group (N = 4), in which each dog was attached to a PUF-HAL without hepatocytes for 9 h, and (2) a HAL group (N = 5), in which each dog was attached to a PUF-HAL with hepatocytes. Blood pressure, blood ammonia, blood glucose, serum creatinine, and other parameters related to liver function were compared between the two groups. RESULTS. In the HAL group, blood ammonia and serum creatinine levels were significantly lower, and blood pressure and blood glucose levels significantly higher, than those in the control group. CONCLUSIONS. The scaled-up PUF-HAL developed for large animals is useful as a liver support system in the dog acute liver failure model.     

Inter- and intratumoral variability in magnetic resonance imaging of pleomorphic adenoma: an attempt to interpret the variable magnetic resonance findings

OBJECTIVES: The purpose of our study was to describe the various magnetic resonance (MR) findings of pleomorphic adenoma and to interpret these findings. METHODS: MR studies of 33 pleomorphic adenomas and 13 malignant tumors in the major salivary glands were reviewed. RESULTS: High signal intensity on short-inversion-time inversion recovery (STIR) and T2-weighted (T2W) images, progressive enhancement on dynamic MR images, and high apparent diffusion coefficient (ADC) values on diffusion-weighted (DW) images reflected myxoid-dominant components in pleomorphic adenomas. Hypercellularity with less-myxoid stroma showed reduced signal intensity on STIR and T2W images and also reduced ADC values on DW images, and the peak of time versus signal intensity curves (TICs) was reached earlier on dynamic MR images. CONCLUSIONS: The MR images of hypercellularity components in pleomorphic adenoma overlap with those of malignant parotid tumors. Detecting myxoid components by STIR, T2W, DW, and dynamic MR images is useful for predicting whether salivary gland tumors are pleomorphic adenoma or not.     

Long-term outcome of iron-endocytosing cultured corneal endothelial cell transplantation with magnetic attraction

The long-term efficacy and safety of transplanting iron-endocytosing cultured corneal endothelial cells (CECs) with magnetic attraction were evaluated. Rabbit corneas were subjected to cryo-injury to detach CECs. Cultured rabbit CECs (RCEC) were exposed to spherical iron powder and then injected into the anterior chamber, after which a neogium magnet was fixed on the eyelid for 24 hr to attract the cells to Descemet's membrane (RCEC-iron group, n=4). An RCEC group (cryo-injury and injection of normal cultured RCEC, n=4) and a Cryo group (cryo-injury without injection of RCEC, n=4) served as controls. Intraocular pressure was measured for 12 months after surgery. Corneal findings on slit lamp biomicroscopy, RCEC density, the electro-retinogram (ERG), and residual iron in the ocular tissues were evaluated at final assessment. Intraocular pressure did not increase in any group throughout 12 months of observation. At the final assessment, the average corneal edema score of the RCEC-iron group was significantly lower than that of the RCEC or Cryo groups (p=0.021). The average CEC density of the RCEC-iron group was 2581+/-230 cells mm(-2) (mean+/-SD), whereas no CECs were observed on the inner surface of the central cornea in the RCEC and Cryo groups. No significant differences of the ERG (a- and b-wave amplitudes, and b-wave/a-wave ratio) were detected among the groups. Iron powder was not detected by Berlin blue staining in the ocular tissues of the RCEC-iron group. Apoptotic cells were not observed in the endothelium by terminal transferase-mediated nick-end labeling. Transplanted iron-endocytosing RCEC remained viable for 12 months after surgery. There were no detectable ocular complications after the transplantation of iron-endocytosing cultured RCEC. Magnetic attachment of iron-endocytosing CECs can be an effective and safe method for corneal endothelial repair.     

Ammonia-mediated LTP inhibition: effects of NMDA receptor antagonists and L-carnitine

Because hyperammonemia is thought to contribute to the pathogenesis of hepatic encephalopathy, we examined the effects of ammonia on ATP levels, neuronal morphology, and synaptic function in rat hippocampal slices. Although ammonia did not alter ATP levels supported by 10 mM glucose, ammonia significantly depressed ATP levels in the presence of 3.3 mM glucose or 10 mM pyruvate, suggesting effects on respiratory energy metabolism. Ammonia also impaired synaptic function and neuronal integrity sustained by pyruvate. In 10 mM glucose, ammonia inhibited the induction and maintenance of long-term potentiation (LTP) in a concentration-dependent fashion. These inhibitory effects of ammonia were overcome by L-carnitine. DL-APV, an antagonist of NMDA receptors, also diminished the effects of ammonia on ATP levels and LTP induction, indicating that ammonia impairs neuronal function via altered metabolism and untimely NMDA receptor activation. These results suggest that L-carnitine and NMDA receptor antagonists have the potential to preserve neuronal function during hyperammonemia.     

Therapeutic angiogenesis in the ischemic canine heart induced by myocardial injection of naked complementary DNA plasmid encoding hepatocyte growth factor

OBJECTIVE: We investigated the efficacy of directly injecting a plasmid with complementary DNA encoding human hepatocyte growth factor into ischemic canine myocardium to induce angiogenesis. METHODS: Four weeks after ligation of the left anterior descending coronary artery, 125 microg of a complementary DNA plasmid encoding the gene for either hepatocyte growth factor (n = 8) or LacZ (transfection control group, n = 8) was injected directly into the myocardium at the border between the normal tissue and the infarction. Eight other dogs were used as a sham control group. Regional thickening fraction, which indicated contractile function, and blood flow in the normal (circumflex branch territory) and ischemic areas were evaluated under dobutamine administration just before and 4 weeks after transfection. The animals were killed, and capillary numbers in both areas were assessed. These data in the ischemic area were evaluated as the percentage of those in the normal. RESULTS: The number of myocardial capillaries in the ischemic area was successfully increased to approximately 140% of usual in the hepatocyte growth factor group, whereas no change was observed in the other groups (P =.0017 by analysis of variance). Furthermore, regional thickening fraction and blood flow in the ischemic area, which had deteriorated after coronary ligation, showed significant improvement in the hepatocyte growth factor group relative to the other groups (thickening fraction P <.0001 by analysis of variance, blood flow P =.0005 by analysis of variance). CONCLUSIONS: These results support the efficacy of the direct injection of plasmid complementary DNA encoding human hepatocyte growth factor to induce therapeutic angiogenesis in the ischemic myocardium.     

Dissolution of human cholesterol gallstones in vitro with bile salts. A scanning and stereoscanning electron microscopic study.

This report deals with the in vitro dissolution of cholesterol gallstones by bile salts, as observed with the scanning electron microscope. Scanning electron microscopic techniques permit excellent visualization of the substructure of these stones and hence allow insight into the structural basis of gallstone dissolution. The small faceted, mixed cholesterol gallstone was used since this is the most common type clinically. Scanning electron microscopy revealed that the gallstone has four structurally different zones; an outer surface, a cortex of compactly layered cholesterol plates, an inner layer of cholesterol mixed with other components, and a center or nidus. The cortex is of particular importance in terms of gallstone dissolution with bile salts, since the interior of the stone remains unaffected until this zone is removed. The speed and completeness of dissolution of the stone depends mainly upon the thickness of the cortex and in part upon the content of noncholesterol components in the interior of the stone. Chenodeoxycholate and cholate behave similarly in terms of the dissolution process as observed by scanning electron microscopy. The solubilization process is very similar to that observed in vitro with solvents such as ether and ethanol. Stones treated with 400 mM cholate were completely solubilized in 3 weeks. The process described in this paper is probably similar to that which occurs in vivo in spontaneous gallstone dissolution in man and in patients treated with bile salts for gallstone dissolution.