Laser spectroscopic imaging techniques have received tremendous attention in the field of cancer diagnosis due to their high sensitivity, high temporal resolution, and short acquisition time. However, the limited tissue penetration of the laser is still a challenge for the in vivo diagnosis of deep-seated lesions. Nanomaterials have been universally integrated with spectroscopic imaging techniques for deeper cancer diagnosis in vivo. The components, morphology, and sizes of nanomaterials are delicately designed, which could realize cancer diagnosis in vivo or in situ. Considering the enhanced signal emitting from the nanomaterials, we emphasized their combination with spectroscopic imaging techniques for cancer diagnosis, like the surface-enhanced Raman scattering (SERS), photoacoustic, fluorescence, and laser-induced breakdown spectroscopy (LIBS). Applications of the above spectroscopic techniques offer new prospects for cancer diagnosis.

In liver tumor surgery, the recognition of tumor margin and radical resection of microcancer focis have always been the crucial points to reduce postoperative recurrence of tumor. However, naked-eye inspection and palpation have limited effectiveness in identifying tumor boundaries, and traditional imaging techniques cannot consistently locate tumors in real time. As an intraoperative real-time navigation imaging method, NIR fluorescence imaging has been extensively studied for its simplicity, reliable safety, and superior sensitivity, and is expected to improve the accuracy of liver tumor surgery. In recent years, the research focus of NIR fluorescence has gradually shifted from the first near-infrared window (NIR-I, 700–900 nm) to the second near-infrared window (NIR-II, 1000–1700nm). Fluorescence imaging in NIR-II reduces the scattering effect of deep tissue, providing a preferable detection depth and spatial resolution while significantly eliminating liver autofluorescence background to clarify tumor margin. Developing fluorophores combined with tumor antibodies will further improve the precision of fluorescence-guided surgical navigation. With the development of a bunch of fluorophores with phototherapy ability, NIR-II can integrate tumor detection and treatment to explore a new therapeutic strategy for liver cancer. Here, we review the recent progress of NIR-II fluorescence technology in liver tumor surgery and discuss its challenges and potential development direction.

Cells are highly sensitive to their geometrical and mechanical microenvironment that directly regulate cell shape, cytoskeleton and organelle, as well as the nucleus morphology and genetic expression. The emerging two-dimensional micropatterning techniques offer powerful tools to construct controllable and well-organized microenvironment for single-cell level investigations with qualitative analysis, cellular standardization, and in vivo environment mimicking. Here, we provide an overview of the basic principle and characteristics of the two most widely-used micropatterning techniques, including photolithographic micropatterning and soft lithography micropatterning. Moreover, we summarize the application of micropatterning technique in controlling cytoskeleton, cell migration, nucleus and gene expression, as well as intercellular communication.

Measurement of blood flow velocity is key to understanding physiology and pathology in vivo. While most measurements are performed at the middle of the blood vessel, little research has been done on characterizing the instantaneous blood flow velocity distribution. This is mainly due to the lack of measurement technology with high spatial and temporal resolution. Here, we tackle this problem with our recently developed dual-wavelength line-scan third-harmonic generation (THG) imaging technology. Simultaneous acquisition of dual-wavelength THG line-scanning signals enables measurement of blood flow velocities at two radially symmetric positions in both venules and arterioles in mouse brain in vivo. Our results clearly show that the instantaneous blood flow velocity is not symmetric under general conditions.

Temporary spinal cord stimulation (tSCS) can effectively reduce the pain and severity of postherpetic neuralgia (PHN). However, there are no effective and objective methods for predicting the effects of tSCS on PHN. Laser speckle contrast imaging (LSCI) is frequently used in neurology to evaluate the effectiveness of treatment. To assess the accuracy of LSCI in predicting the impact of tSCS on PHN, 14 adult patients receiving tSCS treatments for spinal nerve-innervated (C6-T2) PHN participated in this observational study. Visual analog scale (VAS) assessments and LSCI blood flow images of the fingers were recorded after the tSCS procedure. The results showed that the VAS scores of all patients decreased significantly. Moreover, the blood flow index (BFI) values were significantly higher than they were before the procedure. Increased blood flow and pain alleviation were positively correlated. The findings indicated that spinal nerve PHN (C6-T2) was significantly reduced by tSCS. Pain alleviation by tSCS was positively correlated with increased blood flow in the hand. The effect of tSCS on PHN may thus be predicted using an independent and consistent indicator such as LSCI.

In this work, we present an intravascular dual-mode endoscopic system capable of both intravascular photoacoustic imaging (IVPAI) and intravascular optical coherence tomography (IVOCT) for recognizing spontaneous coronary artery dissection (SCAD) phantoms. IVPAI provides high-resolution and high-penetration images of intramural hematoma (IMH) at different depths, so it is especially useful for imaging deep blood clots associated with imaging phantoms. IVOCT can readily visualize the double-lumen morphology of blood vessel walls to identify intimal tears. We also demonstrate the capability of this dual-mode endoscopic system using mimicking phantoms and biological samples of blood clots in ex vivo porcine arteries. The results of the experiments indicate that the combined IVPAI and IVOCT technique has the potential to provide a more accurate SCAD assessment method for clinical applications.

Photodynamic therapy (PDT) has limited effects in treating metastatic breast cancer. Immune checkpoints can deplete the function of immune cells; however, the expression of immune checkpoints after PDT is unclear. This study investigates whether the limited efficacy of PDT is due to upregulated immune checkpoints and tries to combine the PDT and immune checkpoint inhibitor to observe the efficacy. A metastatic breast cancer model was treated by PDT mediated by hematoporphyrin derivatives (HpD-PDT). The anti-tumor effect of HpD-PDT was observed, as well as CD4+T, CD8+T and calreticulin (CRT) by immunohistochemistry and immunofluorescence. Immune checkpoints on T cells were analyzed by flow cytometry after HpD-PDT. When combining PDT with immune checkpoint inhibitors, the antitumor effect and immune effect were assessed. For HpD-PDT at 100mW/cm² and 40, 60 and 80J/cm², primary tumors were suppressed and CD4+T, CD8+T and CRT were elevated; however, distant tumors couldn’t be inhibited and survival could not be prolonged. Immune checkpoints on T cells, especially PD1 and LAG-3 after HpD-PDT, were upregulated, which may explain the reason for the limited HpD-PDT effect. After PDT combined with anti-PD1 antibody, but not with anti-LAG-3 antibody, both the primary and distant tumors were significantly inhibited and the survival time was prolonged, additionally, CD4+T, CD8+T, IFN-γ+CD4+T and TNF-α+CD4+T cells were significantly increased compared with HpD-PDT. HpD-PDT could not combat metastatic breast cancer. PD1 and LAG-3 were upregulated after HpD-PDT. Anti-PD1 antibody, but not anti-LAG-3 antibody, could augment the antitumor effect of HpD-PDT for treating metastatic breast cancer.

Fluorescence imaging in the second near-infrared window (NIR-II, 900–1880nm) with less scattering background in biological tissues has been combined with the confocal microscopic system for achieving deep in vivo imaging with high spatial resolution. However, the traditional NIR-II fluorescence confocal microscope with separate excitation focus and detection pinhole makes it possess low confocal efficiency, as well as difficultly to adjust. Two types of upgraded NIR-II fluorescence confocal microscopes, sharing the same pinhole by excitation and emission focus, leading to higher confocal efficiency, are built in this work. One type is fiber-pinhole-based confocal microscope applicable to CW laser excitation. It is constructed for fluorescence intensity imaging with large depth, high stabilization and low cost, which could replace multiphoton fluorescence microscopy in some applications (e.g., cerebrovascular and hepatocellular imaging). The other type is air-pinhole-based confocal microscope applicable to femtosecond (fs) laser excitation. It can be employed not only for NIR-II fluorescence intensity imaging, but also for multi-channel fluorescence lifetime imaging to recognize different structures with similar fluorescence spectrum. Moreover, it can be facilely combined with multiphoton fluorescence microscopy. A single fs pulsed laser is utilized to achieve up-conversion (visible multiphoton fluorescence) and down-conversion (NIR-II one-photon fluorescence) excitation simultaneously, extending imaging spectral channels, and thus facilitates multi-structure and multi-functional observation.

Nicotinamide adenine dinucleotide (NADH) is a cofactor that serves to shuttle electrons during metabolic processes such as glycolysis, the tricarboxylic acid cycle, and oxidative phosphorylation (OXPHOS). NADH is autofluorescent, and its fluorescence lifetime can be used to infer metabolic dynamics in living cells. Fiber-coupled time-correlated single photon counting (TCSPC) equipped with an implantable needle probe can be used to measure NADH lifetime in vivo, enabling investigation of changing metabolic demand during muscle contraction or tissue regeneration. This study illustrates a proof of concept for point-based, minimally-invasive NADH fluorescence lifetime measurement in vivo. Volumetric muscle loss (VML) injuries were created in the left tibialis anterior (TA) muscle of male Sprague Dawley rats. NADH lifetime measurements were collected before, during, and after a 30s tetanic contraction in the injured and uninjured TA muscles, which was subsequently fit to a biexponential decay model to yield a metric of NADH utilization (cytoplasmic vs protein-bound NADH, the A1τ1/A2τ2 ratio). On average, this ratio was higher during and after contraction in uninjured muscle compared to muscle at rest, suggesting higher levels of free NADH in contracting and recovering muscle, indicating increased rates of glycolysis. In injured muscle, this ratio was higher than uninjured muscle overall but decreased over time, which is consistent with current knowledge of inflammatory response to injury, suggesting tissue regeneration has occurred. These data suggest that fiber-coupled TCSPC has the potential to measure changes in NADH binding in vivo in a minimally invasive manner that requires further investigation.

Adenine nucleotide translocator (ANT) is a mitochondrial protein involved in the exchange of ADP and ATP across the mitochondrial inner membrane. It plays a crucial role in cellular energy metabolism by facilitating the transport of ATP synthesized within the mitochondria to the cytoplasm. The isoform ANT1 predominately expresses in cardiac and skeletal muscles. Mutations or dysregulation in ANT1 have been implicated in various mitochondrial disorders and neuromuscular diseases. We aimed to examine whether ANT1 deletion may affect mitochondrial redox state in our established ANT1-deficient mice. Hearts and quadriceps resected from age-matched wild type (WT) and ANT1-deficient mice were snap-frozen in liquid nitrogen. The Chance redox scanner was utilized to perform 3D optical redox imaging. Each sample underwent scanning across 3–5 sections. Global averaging analysis showed no significant differences in the redox indices (NADH, flavin adenine dinucleotide containing-flavoproteins Fp, and the redox ratio Fp/(NADH+Fp) between WT and ANT1-deficient groups. However, quadriceps had higher Fp than hearts in both groups (p=0.0004 and 0.01, respectively). Furthermore, the quadriceps were also more oxidized (a higher redox ratio) than hearts in WT group (p=0.004). NADH levels were similar in all cases. Our data suggest that under non-stressful physical condition, the ANT1-deficient muscle cells were in the same mitochondrial state as WT ones and that the significant difference in the mitochondrial redox state between quadriceps and hearts found in WT might be diminished in ANT1-deficient ones. Redox imaging of muscles under physical stress can be conducted in future.