Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, PC97
A comparative study of erythrocyte membrane proteins in type 2 diabetic patients and healthy controls
J. Singh1, S. Kappala1, S. Raj bhandari2, P. Abel1
1. School of Pharmacy and School of Forensic and Investigative Sciences, University of Central Lancashire, Preston, Lancashire, United Kingdom. 2. Diabetes and Endocrinology, Lancashire Teaching Hospitals, Chorley, Lancashire, United Kingdom.
Type 2 Diabetes mellitus (T2DM) is a major metabolic disorder affecting more than 200 million people worldwide (Zimmet et al., 2001). Altered structural and functional properties of erythrocyte membrane proteins have been widely implicated in T2DM patients (McMillan et al., 1978). This study analysed the specific proteins of the erythrocyte membrane in T2DM patients compared to healthy controls. T2DM patients and healthy control subjects were recruited from the local hospital diabetic clinic and the University. Blood samples (10 ml per subject) were obtained from each subject and erythrocyte ghost membranes were isolated separately from each sample. Total erythrocyte membrane protein concentration of each sample was determined using bovine serum albumin (BSA) as standard. The protein components of the erythrocyte ghost membranes were determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) which was performed on a Bio-Rad mini gel system using 12 % ready Tris-HCl gel. The gels were stained using silver staining method and identification of protein bands was done using molecular weight marker on a Bio-Rad gel documentation system. The project had the relevant ethical clearance from LREC and the University Ethics Committee to undertake the study. This investigation analysed five proteins namely spectrin, ankyrin, band 3, protein 4.1 and glycophorin in 9 T2DM patients and 9 healthy controls for comparison. The protein quantification of each electrophoretic band was accomplished through scanning densitometry and each protein was expressed in terms of intensity of each band. Initial experiments have indicated significant (Student’s t-test; p<0.05; n=9) increases in the volumes of all five proteins in erythrocyte membranes of T2DM patients compared to healthy controls. Typically, the values (mean ± SEM) were - (A) spectrin- control 1,056 ± 92 and T2DM 3,302 ± 70, (B) ankyrin- control 1,107 ± 69 and T2DM 3,376 ± 40, (C) band 3 protein- control 1,525 ± 56 and T2DM 3,221 ± 28, (D) protein 4.1- control 1,646 ± 128 and T2DM 3,033 ± 23 and (E) glycophorin - control 2,072 ± 124 and T2DM 2,719 ± 20. These results provide evidence of profound quantitative and qualitative alteration of the erythrocyte membrane proteins in T2DM patients compared to healthy controls indicating functional implications of the diabetic patients.
Where applicable, experiments conform with Society ethical requirements