Effect of Dimethyl Sulfoxide and Melatonin on the Isolation of Human Primary Hepatocytes.

The availability of fully functional human hepatocytes is critical for progress in human hepatocyte transplantation and the development of bioartificial livers and in vitro liver systems. However, the cell isolation process impairs the hepatocyte status and determines the number of viable cells that can be obtained. This study aimed to evaluate the effects of using dimethyl sulfoxide (DMSO) and melatonin in the human hepatocyte isolation protocol. Human hepatocytes were isolated from liver pieces resected from 10 patients undergoing partial hepatectomy. Each piece was dissected into 2 equally sized pieces and randomized, in 5 of 10 isolations, to perfusion with 1% DMSO-containing perfusion buffer or buffer also containing 5 mM melatonin using the 2-step collagenase perfusion technique (experiment 1), and in the other 5 isolations to standard perfusion or perfusion including 1% DMSO (experiment 2). Tissues perfused with DMSO yielded 70.6% more viable hepatocytes per gram of tissue (p = 0.076), with a 26.1% greater albumin production (p < 0.05) than those perfused with control buffer. Melatonin did not significantly affect (p > 0.05) any of the studied parameters, but cell viability, dehydrogenase activity, albumin production, urea secretion, and 7-ethoxycoumarin O-deethylase activity were slightly higher in cells isolated with melatonin-containing perfusion buffer compared to those isolated with DMSO. In conclusion, addition of 1% DMSO to the hepatocyte isolation protocol could improve the availability and functionality of hepatocytes for transplantation, but further studies are needed to clarify the mechanisms involved.

Incubation with dimethyl sulfoxide prior to cryopreservation improves functionality of thawed human primary hepatocytes.

BACKGROUND: Efficient cryopreservation of human hepatocytes is essential for their use in cell therapy. This study investigated the effects of adding melatonin and/or dimethyl sulfoxide (DMSO) to pre-incubation and/or freezing solutions on the viability and function of thawed human hepatocytes. METHODS: Isolated human hepatocytes were pre-incubated for 90 min at 4°C in Williams' Medium E (WEM), WEM containing 5 mM melatonin dissolved in DMSO, or WEM containing the equivalent amount of DMSO (1%). The hepatocytes were frozen in University of Wisconsin solution (UW) and 10% DMSO, with or without 5 mM melatonin. After thawing, viability, plating efficiency, mitochondrial dehydrogenase activity (MTT), and albumin and urea production were analyzed. RESULTS: Viability and plating efficiency were not affected by melatonin or DMSO in pre-incubation media. Unexpectedly, hepatocytes pre-incubated with DMSO had significantly higher MTT (29.7% vs. control, p<0.01), albumin (82.8% vs. control, p<0.05), and urea amounts (26.2% vs. control, p=0.06) than those incubated only with WEM. Hepatocytes pre-incubated in media containing melatonin had amounts between those of cells incubated with DMSO or only with WEM (p<0.05 for MTT and p>0.05 for albumin and urea values). Also, the addition of melatonin to the freezing media did not significantly improve any of the studied parameters (p>0.05). DISCUSSION: Adding 1% DMSO to pre-incubation media prior to the cryopreservation of human hepatocytes preserves hepatocyte function after thawing. These findings could be considered in current hepatocyte cryopreservation protocols.

Nasofibroscopia en el diagnóstico de las afeccciones de vías aéreas superiores del niño / Nasofibroscopy in the diagnóstic of the superiors airs pathways of the child

Las últimas décadas han traído ayudas diagnósticas muy importantes para el conocimiento y detección de algunas enfermedades obstructivas de las vías aéreas. La Nasofaringolaringoscopia flexible es actualmente uno de los mejores métodos diagnósticos para las lesiones desde nariz hasta región subglótica. Nosotros presentamos nuestra experiencia con 125 niños en los cuales en los cuales fue realizada Nasofibroscopia usando un Olympus DENI tipo P3 (Melville NY) en el período comprendido entre junio de 1995 y mayo 1996. Las indicaciones para este procedimiento fueron Cuadros respiratorios a repetición (22,51 por ciento), Adenoiditis y Disfonía (9,27 por ciento) respectivamente, Ronquera (8,69 por ciento), Respirador bucal y Estridor inspiratorio (7,28 por ciento), Sospecha de aspiración de cuerpo extraño (5,96 por ciento). La Adenoiditis fue el diagnóstico más frecuente (24,26 por ciento) del total de hallazgos. En los casos de lesiones papilomatosas o congestion de mucosa indujo realizar un total de 20 biopsias de las cuales un (40 por ciento correspondío a lesiones por virus de VPH