title: [Cell viability assessment of cold-preserved (4 degrees C) and cryopreserved (-196 degrees C) allograft valves by flowcytometric analysis].
abstract: We introduced cryopreservation technique to store the allograft valve and subsequently commenced the use of the cryopreserved aortic allograft valve for aortic valve replacement. For the clinical use of the allograft valve, it is essential to evaluate the cell viability. We assessed cell viability of the allograft valve by flowcytometry (FCM), using a fluorescein diacetate (FDA)-propidium iodide (PI) stain. Pulmonary allograft valves were harvested from the cadavers with a consent. Of the 9 valves collected and sterilized in 4 degrees C antibiotic solution for 48 hours, 4 valves were stored under refrigeration (cold-preserved), and the remaining 5 valves were cryopreserved. The warm ischemic time from cardiac death to valve preservation was 7.3 +/- 5.4 hours. The cold-preserved valves were stored at 4 degrees C in the nutrient medium (TC-199, calf serum and HEPES buffer). The cryopreserved valves were frozen by a programmable freezer to -80 degrees C in the same nutrient medium containing 10% dimethylsulfoxide (DMSO), and subsequently stored in liquid nitrogen (-196 degrees C). We assessed the cell viability of the allograft valve leaflet, before storage and at 2 weeks, 1 month and 2 months after the storage. Suspensions of the valve leaflet fibroblasts were double-stained with FDA (1 micrograms/ml) and PI (1 microgram/ml) for flowcytometric analysis of the viable cells (cells showing a chromatic response to FDA but no response to PI). In the cold-preserved group, percentage of the viable cell was 90.3 +/- 3.6% before storage, 62.0 +/- 3.0% after 1 month of storage, and 50.3 +/- 3.7% after 2 months of storage.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Surgery, III, Nara Medical College, Japan.
authors.forename: K
authors.initials: K
authors.lastname: Niwaya
authors.forename: S
authors.initials: S
authors.lastname: Kitamura
authors.forename: K
authors.initials: K
authors.lastname: Kawachi
authors.forename: R
authors.initials: R
authors.lastname: Morita
authors.forename: S
authors.initials: S
authors.lastname: Taniguchi
authors.forename: T
authors.initials: T
authors.lastname: Seki
authors.forename: M
authors.initials: M
authors.lastname: Fukutomi
authors.forename: T
authors.initials: T
authors.lastname: Kawata
authors.forename: H
authors.initials: H
authors.lastname: Sakaguchi
mesh_terms: D001021:Aortic Valve / Q000166:cytology; D002470:Cell Survival / Q000502:physiology; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D005434:Flow Cytometry; D006351:Heart Valves / Q000166:cytology / Q000637:transplantation; D006801:Humans; D008297:Male; D008875:Middle Aged; D011664:Pulmonary Valve / Q000166:cytology