In vivo imaging of hemoglobin and melanin variations using photoacoustic tomography

Objectives In the photoacoustic imaging (PAI), hemoglobin and melanin images could be acquired by the absorption coefficient differences between deoxyhemoglobin (HbR), oxyhemoglobin (HbO₂) and melanin. In this study, we made melanin producing bacteria and visualized the bacteria using PAI.

Methods We engineered attenuated Salmonella typhimurium expressing tyrosine kinase, key enzyme in the process of melanin synthesis. U87MG human glioma cancer cells were injected subcutaneously into BALB/c nude mice. When U87MG reach 120 mm³, melanin-producing Salmonellae was i.v. injected in tumor-bearing mice. PAI was acquired by Nexus 128, fully 3D PA computed tomography scanner (ENDRA, Inc). Wavelength tuning range of tunable laser is from 680 to 950 nm. In order to calculate the HbR, HbO₂ and melanin in relative values, we modified existing formula as follow: [HbR;HbO2;Melanin]_(x,y,z)=(M^TM)^-1M^TФ(x,y,z) The PAI was acquired at five optical wavelengths which are 680, 750, 850, 900 and 920 nm. HbR, HbO₂ and melanin were separated each other from obtained images. Correlation between max values and time was evaluated by Spearman correlation analysis. The distribution of vessels and melanin were calculated and compared using the one sample t-test.

Results In the phantom study, the max value was continuously increased during 12 hours and coefficient of determination (r²) was 0.96 (p<0.01). The distribution of melanin from tumor with or without bacterial injection was 31.49 mm³ and 7.58±1.76 mm³, respectively. The distribution of melanin was significantly different (p<0.001).

Conclusions The melanin was excellently detected by PAI which has shown great potential for imaging molecular signatures of bacterial colonization in tumor.