Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and the second leading cause of cancer death worldwide. The development of HCC leads to abnormalities in the structure and function of blood vessels, which further lead to high pressure and hypoxia in the tumor microenvironment (TME). The most common clinical methods for identifying HCC nowadays are magnetic resonance imaging (MRI), computed tomography (CT), and ultrasound. MRI can measure multifunctional parameters of the liver; however, it has significant limitations in imaging resolution and is costly. CT can image the blood vessels in the liver; however, it uses X-ray, thus increases the risk of cancer. Ultrasound imaging is widely used to evaluate HCC; however, its sensitivity and specificity are low. Therefore, a more complete and reliable technique to analyze the micro-vascular morphology of HCC and TME is urgently needed. Photoacoustic imaging is a rapidly developing imaging technology in recent years. It offers a wide range of potential applications in the field of medical imaging and can visualize the structure and function information of biological tissues without labeling of contrast agents or invasion. Photoacoustic imaging has high specificity and sensitivity in the diagnosis of HCC and can visualize functional imaging of tumors and morphological examination of blood vessels.

A mouse model of in situ liver cancer was established, and the bioluminescence signal was activated by an in vivo fluorescence imaging system to locate the tumor. The microvascular structure characteristics and oxygen saturation of normal liver lobules, tumor centers, and adjacent tumors were accurately observed using photoacoustic microscopy. The concentrations of oxygenated and deoxygenated hemoglobins were quantified using the spectroscopic separation method to calculate blood oxygen saturation. The photoacoustic images were converted into binary images, and the vascular signals were extracted for density and diameter analysis.

The results obtained using photoacoustic microscopy via two wavelengths (532 nm/559 nm) show that the blood vessels in the normal liver are evenly spaced and well differentiated, whereas large irregular vessels appear at the edges of the tumors, and the vascular joints are curved and dilated. The blood vessels inside the tumors are unevenly distributed and the branch diameter increases. The oxygen concentration in the blood around the tumors decreases, resulting in a hypoxic and high-pressure TME.

In microvascular monitoring of hepatocellular carcinoma, the photoacoustic imaging can provide high-resolution images, which can more accurately detect the morphology of tiny and abnormal blood vessels, improving the accuracy of early cancer detection. Through image analysis, indicators such as the density and diameter of microvessels and other information such as oxygenation level and metabolic activity of tumor tissues can be evaluated to assess the growth state of tumors and predict the degree of malignancy. Photoacoustic imaging demonstrates a high application potential for studying the development of HCC. It can provide further insights into the antiangiogenic therapy of tumors and the diagnosis of numerous liver-related diseases.