Cochlear repair by transplantation of human cord blood CD133+ cells to nod-scid mice made deaf with kanamycin and noise.

We investigated the fate of human cord blood CD133+ hematopoietic stem cells (HSC) transplanted intravenously (IV) into irradiated nod-scid mice previously made deaf by ototoxic treatment with kanamycin and/ or intense noise, to verify whether HSC engraft the cochlea and contribute to inner ear restoration, in vivo. We tested the presence of HLA.DQalpha1 by PCR, used for traceability of engrafted cells, finding evidence that HSC migrated to various host tissues, including the organ of Corti (OC). By histology, antibody and lectin-staining analysis, we confirmed that HSC IV transplantation in mice previously damaged by ototoxic agents correlated with the repair process and stimulation ex novo of morphological recovery in the inner ear, while the cochlea of control oto-injured, nontransplanted mice remained seriously damaged. Dual color FISH analysis also provided evidence of positive engraftment in the inner ear and in various mouse tissues, also revealing small numbers of heterokaryons, probably derived from fusion of donor with endogenous cells, for up to 2 months following transplantation. These observations offer the first evidence that transplanted human HSC migrating to the inner ear of oto-injured mice may provide conditions for the resumption of deafened cochlea, emerging as a potential strategy for inner ear rehabilitation.

Gentamicin-induced cytotoxicity involves protein kinase C activation, glutathione extrusion and malondialdehyde production in an immortalized cell line from the organ of corti.

The objective of the present study was to investigate the biochemical mechanisms underlying gentamicin cytotoxicity in OC-k3 cells derived from an immortalized cell line developed from the organ of Corti of transgenic mice. Administration of 50 microM gentamicin significantly reduced cell proliferation and viability, as well as initiating morphological changes associated with apoptosis. Protein kinase C (PKC) alpha activity was increased in gentamicin-treated cells, peaking 15 min after dosing (+138.2%). This PKCalpha increase was followed by a rise of glutathione (GSH) efflux and a concomitant 29% decrease in intracellular GSH levels at 30 min. PKCalpha-specific inhibitors blocked these cytotoxic effects. Gentamicin also increased malondialdehyde levels, while N-acetylcysteine, GSH and ascorbic acid prevented gentamicin-induced cell death. These findings suggest that gentamicin cytotoxicity involves a production of intracellular reactive oxygen species and a concomitant PKC-dependent fall of intracellular scavenging abilities (GSH), events that together drive cells to undergo apoptosis.

Determination of histamine in the whole blood of colon cancer patients.

The aim of the present work is to investigate whether histamine assay could be useful in detecting the presence of primary cancer. The high-performance liquid chromatographic (HPLC)-based o-phthalaldialdehyde (OPA) histamine derivatization assay was investigated with respect to several variables, dramatization reagent concentration, organic solvent requirement, derivatization time and counter-ion effect on chromatographic separation. The OPA histamine assay, in the absence of added -SH groups, was found to detect histamine in whole blood samples with relative standard deviations <14% and recoveries not less than 90%. The assay showed high selectivity towards other aminic-containing compounds and a detection limit of 18 nM of histamine was evaluated. Calibration curves in the range 50-500 nM were obtained by using histamine standards in 0.1 M HCl with a regression coefficient value (r(2)) of 0.9969. In order to assess the usefulness of this assay in primary tumor monitoring, two groups of individuals, 29 controls and 29 colon cancer patients were selected, and serum levels of histamine, carcinogen embrionary antigen (CEA), carcinogen antigen 19.9 (CA19.9), and tumor staging, were determined. A significant histamine reduction (P=0.028) between controls (180.12+/-70.4 nM) and patients (134.5+/-90.3 nM) was found, and a cut-off value of 157.5 nM was extrapolated as intercept point of sensitivity and specificity curves. Fifty percent of patients showed a histamine value below the cut-off, while 45.8 and 8.3% of patients were positive for CEA and CA19.9, respectively. No correlation was found between Tumor Node Metastasis staging and histamine amount, indicating that this marker is not related to the tumor mass. Our data suggest that histamine level, together with other classical tumor markers, could be a potentially interesting tumor marker in colon cancer monitoring.