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Establishment and characterization of a novel untransfected corneal endothelial cell line from New Zealand white rabbits.

ABSTRACT: PURPOSE: To establish and characterize a novel untransfected corneal endothelial cell line from New Zealand white rabbits (NRCE cell line) for studies on corneal endothelial cells. METHODS: Primary culture was initiated with a pure population of NRCE cells from corneal endothelia by successive detachment and reattachment procedure of different durations, and cultured in 20% fetal bovine serum-containing DMEM/F12 media with several supplements. The cell line was characterized by chromosome analysis, fluorescence immunoassay and reverse transcription PCR. The tumorigenic potency of the cell line was examined by subcutaneous inoculation to nude mice. The biocompatibility of the cell line to denuded amnions was examined with routine microscopic and electron microscopic techniques. RESULTS: NRCE cells in primary culture proliferated to confluency in 25 days and has been subcultured to passage 227 to date. The novel NRCE cell line, with a steady growing rate in 20% bovine calf serum (BCS)-containing DMEM/F12 medium and a population doubling time of 40.32 h at passage 191, has been established. NRCE cells exhibited chromosomal aneuploidy but their modal chromosome number was still 44. The results of gene expression patterns of marker proteins and membrane transport proteins, combined with immunofluorescent localization patterns of cell junction proteins, indicated that NRCE cells retained normal corneal endothelial characteristics and normal expression pattern of functional proteins. Furthermore, these cells, without any tumorigenic potency, had excellent biocompatibility to denuded amnions in 20% BCS-containing DMEM/F12 medium, and formed confluent cell sheets attached tightly to denuded amnions. CONCLUSIONS: These results suggest that a novel untransfected NRCE cell line established here maintains normal corneal endothelial characteristics and potencies to form normal cell junctions and perform normal functions of transmembrane transport.

SUBMITTER: Fan T

PROVIDER: S-EPMC2690960 | biostudies-literature | 2009

REPOSITORIES: biostudies-literature

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Establishment and characterization of a novel untransfected corneal endothelial cell line from New Zealand white rabbits.

Fan Tingjun T Wang Dansheng D Zhao Jun J Wang Jing J Fu Yongfeng Y Guo Ruichao R

Molecular vision 20090529

<h4>Purpose</h4>To establish and characterize a novel untransfected corneal endothelial cell line from New Zealand white rabbits (NRCE cell line) for studies on corneal endothelial cells.<h4>Methods</h4>Primary culture was initiated with a pure population of NRCE cells from corneal endothelia by successive detachment and reattachment procedure of different durations, and cultured in 20% fetal bovine serum-containing DMEM/F12 media with several supplements. The cell line was characterized by chro ...[more]

PMID: 19503740

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Project description:BackgroundBuprenorphine is one of the most used analgesics for postoperative pain in rabbits. The recommended dose in rabbits (0.01-0.05 mg/kg) is the same for intravenous (IV), intramuscular (IM), and subcutaneous (SC) administration, despite lack of pharmacokinetic data. Five male and five female New Zealand White rabbits (mean ± SD body weight 3.1 ± 0.3 kg) were administered 0.05 mg/kg buprenorphine by the IV, IM and SC routes and 0.1 mg/kg by the SC route, in a cross-over design with two-week wash-out periods between treatments. Blood was collected before, and up to 8 h post buprenorphine injection, for determination of serum levels by UPHLC-MS/MS.ResultsThe area under the time concentration curve (AUC0-t) was lower after SC (398 ± 155 ng/mL/min) than IM (696 ± 168 ng/mL/min, p < 0.001) and IV (789 ± 189 ng/mL/min, p < 0.001) administration. The maximum serum concentration was lower after SC (2.2 ± 1.4 ng/mL) than after IM (11 ± 3.2 ng/mL) administration (p < 0.001). The bioavailability was lower after SC (50 ± 19%) than after IM (95 ± 21%) administration (p = 0.006). The elimination half-life was longer after SC (260 ± 120 min) than after IM (148 ± 26 min, p = 0.002) as well as IV (139 ± 33 min) injection (p < 0.001). An increase in the SC dose from 0.05 to 0.1 mg/kg resulted in an increase in the area under the time concentration curve of 50% in female (p = 0.022) and 165% in male rabbits (p < 0.001). The bioavailability did not change in the females (36 ± 14%, p = 0.6), whereas it increased in the males (71 ± 23%, p = 0.008).ConclusionsThe lower bioavailability of 0.05 mg/kg buprenorphine after SC administration could explain the lack of efficacy seen in clinical pain studies in rabbits, using this route. For immediate pain relief, IV or IM administration is therefore be recommended, whereas SC administration may be useful to sustain analgesic serum levels, once efficient pain relief has been achieved. The current data do not support an increase in dose to compensate for the lower SC bioavailability.

Dataset Information

Establishment and characterization of a novel untransfected corneal endothelial cell line from New Zealand white rabbits.

Publications

Establishment and characterization of a novel untransfected corneal endothelial cell line from New Zealand white rabbits.

Similar Datasets

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