In vivo photoacoustic and photothermal cytometry for monitoring multiple blood rheology parameters

Abstract

Alterations of blood rheology (hemorheology) are important for the early diagnosis, prognosis, and prevention of many diseases, including myocardial infarction, stroke, sickle cell anemia, thromboembolism, trauma, inflammation, and malignancy. However, real-time in vivo assessment of multiple hemorheological parameters over long periods of time has not been reported. Here, we review the capabilities of label-free photoacoustic (PA) and photothermal (PT) flow cytometry for dynamic monitoring of hemorhelogical parameters in vivo which we refer to as photoacoustic and photothermal blood rheology. Using phenomenological models, we analyze correlations between both PT and PA signal characteristics in the dynamic modes and following determinants of blood rheology: red blood cell (RBC) aggregation, deformability, shape (e.g., as in sickle cells), intracellular hemoglobin distribution, individual cell velocity, hematocrit, and likely shear rate. We present ex vivo and in vivo experimental verifications involving high-speed PT imaging of RBCs, identification of sickle cells in a mouse model of human sickle cell disease and in vivo monitoring of complex hemorheological changes (e.g., RBC deformability, hematocrit and RBC aggregation). The multi-parameter platform that integrates PT, PA, and conventional optical techniques has potential for translation to clinical applications using safe, portable, laser-based medical devices for point-of-care screening of disease progression and therapy efficiency.

Figure 1

The principles of in vivo PA and PT blood rheology tests. (A) Schematics. The insets on right shows PA signal (top) and PT signal (bottom) after one laser pulse. (B) Phenomenological model: PA signal trace dynamics in different vessels after averaging many signals generated by a high pulse rate laser.

Figure 2

Ex vivo rheology tests using optical and PT techniques. (A) High-resolution (×100) optical images (left column) and pulsed PT signals (right column) from single normal RBC, RBC aggregates consisting of three RBCs, and low-deformable swollen spherocyte. (B) Typical pathological types of low-deformable RBCs. (C) High-resolution (×100) optical images (left column) and PT signals (right column) from sickle (top) and round (bottom) RBCs with HbS.

Figure 3

High-resolution PT imaging of Hb in live RBCs. (A) Direction of spatial PT scanning of RBCs. (B) PT image of biconcave RBCs (A, top) obtained with the vertical scanning; (C) PT image of RBCs with a ring or round shape obtained with horizontal scanning. (D) Spectral identification of Hb based on correlation of PT spectra (blue points) from the indicated area on the RBC image with the absorption spectrum of Hb (solid black line). (E) Distribution of Hb in nonswollen spherocytes. (F) Comparison of the heterogeneity in PT images of intact (left) and sickle (middle and right) RBCs.

Figure 4

PT multiplex thermal lens imaging of blood flow. (A) Optical images of animal model: mouse ear. (B) Single RBC in small capillary; magnification 60×. (C) Several RBCs in large capillary, magnification 20×. (D) Multi file RBC flow in microvessels; magnification 10×. Dash lines show the boundary of blood vessels. Top row: PT images of ear blood vessels; bottom row: optical images of mesentery blood vessel.

Figure 5

Integration of PT flow cytometry and high speed (1,000–5,000 fpame per second) high resolution transmission imaging of circulating blood in vivo. (A) Dynamic PT signals from blood flow before (top) and after (bottom) formation of circulating RBC aggregates. (B) PT signals (top) from the individual RBCs (middle, bottom)) moving with velocity ~1 mm/sec in small mesenteric vessel with diameter of 10 μm (used for estimation of Ht). Optical monitoring of alterations of RBC deformability under chemical impacts of Diamide (C) and Chlorpromazine (D).

Figure 6

Noninvasive real-time PA monitoring of blood rheology in mouse ear vessels. (A) PA signals from blood after injection of distillate water and Dextran producing complex rheological alterations of RBC deformability, Ht and RBC aggregation.