Single blood cell Raman spectroscopy reveals elevated haemoglobin content in poikilocytosis

Abstract

Abnormally shaped red blood cells (poikilocytes) can cause serious health problems such as anemia and increase the risk of death. At present, the biochemical abnormalities in poikilocytes is not well understood, especially at the single cell level. In this study, confocal Raman spectroscopy revealed biochemical differences between single normal red blood cells (RBCs) and poikilocytes. Intragastric administration of nanoparticulate titanium dioxide (TiO2) was used to produce poikilocytes. Adult rats were administered by gavage 200mg/kg body weight TiO2 every other day for 20 days (low-dose, N=5) or 250mg/kg every day for 60 days (high-dose, N=5). Low and high-dose controls (N=5 each) were administered distilled water for equal durations. Raman spectroscopy was performed on individual RBCs of low-dose subjects using 514nm excitation and a confocal setup. Whole blood from high-dose subjects underwent a Complete Blood Count (CBC) and inductively coupled plasma mass spectrometry (ICP-MS). Acanthocytes and echinocytes, two types of poikilocytes, were observed from TiO2 subjects. RBCs were grouped into four types: normal RBCs from controls and normal-looking RBCs, Acanthocytes, and Echinocytes from TiO2 subjects. The intensities of Raman bands at 1637, 1585, 1559, 1372 and 1228cm-1 are larger in acanthocytes than normal and normal-looking RBCs. The 1342cm-1 band is larger in normal RBCs, acanthocytes and echinocytes than in normal-looking RBCs. Also, the 975cm-1 band is larger in acanthocytes than normal-looking RBCs. These bands are associated with oxygenated RBCs. Overall, poikilocytes, especially acanthocytes, carry more oxygen and haemoglobin and this is corroborated by CBC and ICPMS.