Secondary structure alterations of RBC assessed by FTIR-ATR in correlation to 2,3-DPG levels in ApoE/LDLR Mice.

In the present study, we characterized the secondary structure alterations of intact red blood cells (RBCs) cytosol with special attention to the sex-related alterations in 8- and 24-week-old female and male ApoE/LDLR mice, compared to age-matched female and male C57BL/6J control animals. Results were obtained with previously established methodology based on Fourier transform infrared spectroscopy-attenuated total reflectance (FTIR-ATR). Additionally, we evaluated 2,3-DPG levels in the RBCs and showed its potential link to the hemoglobin (Hb) secondary structure alterations. Considering Hb structure alterations probed by FTIR-ATR, the ratio of turns to α-helices in 8-week-old ApoE/LDLR mice suggested more pronounced secondary structure alterations within the RBCs than in the age-matched control. Sex-related differences were observed solely in 24-week-old male ApoE/LDLR mice, which showed statistically significant increase in the secondary structure alterations compared to 24-week-old female ApoE/LDLR mice. Similar to the secondary structure alterations, no sex-related differences were observed in the levels of 2,3-DPG in RBCs, except for 24-week-old male ApoE/LDLR mice, which showed significantly higher levels compared to the age-matched female ApoE/LDLR mice. Considering the age-related alterations, we observed significant increases in the intracellular 2,3-DPG of RBCs with animals' age in all studied groups, except for female ApoE/LDLR mice, where a significant difference was not reported. This suggests the clear correlation between secondary structure of Hb alterations and 2,3-DPG levels for male and female murine RBC and proves a higher resistance of older female RBCs to the secondary structure changes with progression of atherosclerosis. Moreover, it may be concluded that higher 2,3-DPG levels in RBCs occurred in response to the secondary structure alterations of Hb in ApoE/LDLR mice.