Organ preservation
Ischemia–reperfusion decreases protein tyrosine phosphorylation and p38 mitogen-activated protein kinase phosphorylation in rat lung transplants

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Abstract

Background:

Dramatic alterations of protein tyrosine phosphorylation have been found during the ischemia–reperfusion (IR) period of human lung transplantation. IR also induces activation of p38 mitogen-activated protein kinase (p38) in the heart and kidney. The objective of the present study was to determine whether these changes exist in a rat single-lung transplant model for further mechanistic investigations.

Methods:

Isogeneic lung transplantation was performed from Lewis (LEW) to LEW rats, whereas allogeneic transplantation was from LEW to Brown Norway (BN) rats. Blood gases and peak airway pressure were monitored. Lung tissues were collected after 6 hours of cold ischemic preservation, after 30 minutes of warm ischemia for lung implantation, and after 2 hours of reperfusion. Protein tyrosine kinase (PTK) and phosphatase (PTP) activities were measured. Protein tyrosine phosphorylation, Src PTK and p38 expression and p38 phosphorylation were examined by western blotting.

Results:

In both iso- and allografts, the lung function of transplants was very well preserved. Protein tyrosine phosphorylation, PTK and PTP activities were decreased significantly after 2 hours of reperfusion. Src protein level and phosphorylation of p38 were reduced after 2 hours of reperfusion.

Conclusions:

During the early IR period of lung transplantation, decreased protein tyrosine phosphorylation may be involved in apoptosis and other biologic changes. The lack of p38 activation suggests that activity of mitogen-activated protein kinase pathways in the lung transplantation setting may be different from other IR processes.

Section snippets

Animals

Inbred male Lewis (LEW; RT-1l; 250 to 350 g) rats and inbred male Brown Norway (BN; RT-1n; 250 to 350 g) rats were obtained from Charles River, Inc. (Montreal, Quebec, Canada). All animals received humane care in compliance with The Principles of Laboratory Animal Care, formulated by the Institute of Laboratory Animal Resources (Toronto, ON, Canada); the Guide for the Care and Use of Laboratory Animals (NIH Publication No. 86-23, revised 1985); and the Guide to the Care and Use of Experimental

Airway pressures, arterial blood gases and wet/dry lung ratio

To determine the impact of allogeneic vs isogeneic responses on lung function during reperfusion, arterial blood gases and wet/dry lung weight ratio were used to compare the lung oxygenation and degree of pulmonary edema after reperfusion between isogeneic and allogeneic transplants. Before harvest, the Pao2 of the donor lungs was >580 mmHg and, after 2 hours of reperfusion, the Pao2 of blood returning from of the donor lung alone was >430 mmHg. The oxygenation function of lung grafts was

Discussion

In this study, the status of protein tyrosine phosphorylation was examined in a rat single-lung transplant model that simulates the clinical lung transplant setting of cold ischemic preservation for 6 hours, followed by lung implantation (warm ischemia) and reperfusion. Both iso- and allografts showed satisfactory pulmonary function after reperfusion. A dramatic decrease in protein tyrosine phosphorylation was observed after a 2-hour period of reperfusion, similar to Src PTK activity and Src

Acknowledgements

The authors thank John Mates, Xiao-Hui Bai, Monika Lodyga and Julia Hamilton for technical assistance. M. Liu is a recipient of the Premier’s Research Excellence Award from the Government of Ontario, Canada.

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    Supported by the Ontario Thoracic Society, the Canadian Cystic Fibrosis Foundation and the Canada Institutes of Health Research (MT-13270, MOP-42465, MOP-77559).

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