Influence mechanism of the temporal duration of laser irradiation on photoacoustic technique: a review.

Significance: In the photoacoustic (PA) technique, the laser irradiation in the time domain (i.e., laser pulse duration) governs the characteristics of PA imaging-it plays a crucial role in the optical-acoustic interaction, the generation of PA signals, and the PA imaging performance. Aim: We aim to provide a comprehensive analysis of the impact of laser pulse duration on various aspects of PA imaging, encompassing the signal-to-noise ratio, the spatial resolution of PA imaging, the acoustic frequency spectrum of the acoustic wave, the initiation of specific physical phenomena, and the photothermal-PA (PT-PA) interaction/conversion. Approach: By surveying and reviewing the state-of-the-art investigations, we discuss the effects of laser pulse duration on the generation of PA signals in the context of biomedical PA imaging with respect to the aforementioned aspects. Results: First, we discuss the impact of laser pulse duration on the PA signal amplitude and its correlation with the lateral resolution of PA imaging. Subsequently, the relationship between the axial resolution of PA imaging and the laser pulse duration is analyzed with consideration of the acoustic frequency spectrum. Furthermore, we examine the manipulation of the pulse duration to trigger physical phenomena and its relevant applications. In addition, we elaborate on the tuning of the pulse duration to manipulate the conversion process and ratio from the PT to PA effect. Conclusions: We contribute to the understanding of the physical mechanisms governing pulse-width-dependent PA techniques. By gaining insight into the mechanism behind the influence of the laser pulse, we can trigger the pulse-with-dependent physical phenomena for specific PA applications, enhance PA imaging performance in biomedical imaging scenarios, and modulate PT-PA conversion by tuning the pulse duration precisely.

Second Near-Infrared Macrophage-Biomimetic Nanoprobes for Photoacoustic Imaging of Neuroinflammation.

Neuroinflammation is a significant pathological event involving the neurodegenerative process associated with many neurological disorders. Diagnosis and treatment of neuroinflammation in its early stage are essential for the prevention and management of neurological diseases. Herein, we designed macrophage membrane-coated photoacoustic (PA) probes (MSINPs), with targeting specificities based on naturally existing target-ligand interactions for the early diagnosis of neuroinflammation. The second near-infrared dye, IR1061, was doped into silica as the core and was encapsulated with a macrophage membrane. In vitro as well as in vivo, the MSINPs could target inflammatory cells via the inflammation chemotactic effect. PA imaging was used to trace the MSINPs in a neuroinflammation mouse model and showed a great targeted effect of MSINPs in the prefrontal cortex. Therefore, the biomimetic nanoprobe prepared in this study offers a new strategy for PA molecular imaging of neuroinflammation, which can enhance our understanding of the evolution of neuroinflammation in specific brain regions.

WSA-MP-Net: Weak-signal-attention and multi-scale perception network for microvascular extraction in optical-resolution photoacoustic microcopy.

The unique advantage of optical-resolution photoacoustic microscopy (OR-PAM) is its ability to achieve high-resolution microvascular imaging without exogenous agents. This ability has excellent potential in the study of tissue microcirculation. However, tracing and monitoring microvascular morphology and hemodynamics in tissues is challenging because the segmentation of microvascular in OR-PAM images is complex due to the high density, structure complexity, and low contrast of vascular structures. Various microvasculature extraction techniques have been developed over the years but have many limitations: they cannot consider both thick and thin blood vessel segmentation simultaneously, they cannot address incompleteness and discontinuity in microvasculature, there is a lack of open-access datasets for DL-based algorithms. We have developed a novel segmentation approach to extract vascularity in OR-PAM images using a deep learning network incorporating a weak signal attention mechanism and multi-scale perception (WSA-MP-Net) model. The proposed WSA network focuses on weak and tiny vessels, while the MP module extracts features from different vessel sizes. In addition, Hessian-matrix enhancement is incorporated into the pre-and post-processing of the input and output data of the network to enhance vessel continuity. We constructed normal vessel (NV-ORPAM, 660 data pairs) and tumor vessel (TV-ORPAM, 1168 data pairs) datasets to verify the performance of the proposed method. We developed a semi-automatic annotation algorithm to obtain the ground truth for our network optimization. We applied our optimized model successfully to monitor glioma angiogenesis in mouse brains, thus demonstrating the feasibility and excellent generalization ability of our model. Compared to previous works, our proposed WSA-MP-Net extracts a significant number of microvascular while maintaining vessel continuity and signal fidelity. In quantitative analysis, the indicator values of our method improved by about 1.3% to 25.9%. We believe our proposed approach provides a promising way to extract a complete and continuous microvascular network of OR-PAM and enables its use in many microvascular-related biological studies and medical diagnoses.

Itaconic acid regulation of TFEB-mediated autophagy flux alleviates hyperoxia-induced bronchopulmonary dysplasia.

BACKGROUND: Premature infants often require oxygen supplementation, which can elicit bronchopulmonary dysplasia (BPD) and lead to mitochondrial dysfunction. Mitochondria play important roles in lung development, in both normal metabolism and apoptosis. Enhancing our comprehension of the underlying mechanisms in BPD development can facilitate the effective treatments. METHODS: Plasma samples from BPD and non-BPD infants were collected at 36 weeks post-menstrual age and used for metabolomic analysis. Based on hyperoxia-induced animal and cell models, changes in mitophagy and apoptosis were evaluated following treatment with itaconic acid (ITA). Finally, the mechanism of action of ITA in lung development was comprehensively demonstrated through rescue strategies and administration of corresponding inhibitors. RESULTS: An imbalance in the tricarboxylic acid (TCA) cycle significantly affected lung development, with ITA serving as a significant metabolic marker for the outcomes of lung development. ITA improved the morphological changes in BPD rats, promoted SP-C expression, and inhibited the degree of alveolar type II epithelial cells (AEC II) apoptosis. Mechanistically, ITA mainly promotes the nuclear translocation of transcription factor EB (TFEB) to facilitate dysfunctional mitochondrial clearance and reduces apoptosis in AEC II cells by regulating autophagic flux. CONCLUSION: The metabolic imbalance in the TCA cycle is closely related to lung development. ITA can improve lung development by regulating autophagic flux and promote the nuclear translocation of TFEB, implying its potential therapeutic utility in the treatment of BPD.

Targeted Cyclo[8]pyrrole-Based NIR-II Photoacoustic Tomography Probe for Suppression of Orthotopic Pancreatic Tumor Growth and Intra-abdominal Metastases.

Pancreatic cancer is highly lethal. New diagnostic and treatment modalities are desperately needed. We report here that an expanded porphyrin, cyclo[8]pyrrole (CP), with a high extinction coefficient (89.16 L/g·cm) within the second near-infrared window (NIR-II), may be formulated with an αvß3-specific targeting peptide, cyclic-Arg-Gly-Asp (cRGD), to form cRGD-CP nanoparticles (cRGD-CPNPs) with promising NIR-II photothermal (PT) therapeutic and photoacoustic (PA) imaging properties. Studies with a ring-array PA tomography system, coupled with analysis of control nanoparticles lacking a targeting element (CPNPs), revealed that cRGD conjugation promoted the delivery of the NPs through abnormal vessels around the tumor to the solid tumor core. This proved true in both subcutaneous and orthotopic pancreatic tumor mice models, as confirmed by immunofluorescent studies. In combination with NIR-II laser photoirradiation, the cRGD-CPNPs provided near-baseline tumor growth inhibition through PTT both in vitro and in vivo. Notably, the combination of the present cRGD-CPNPs and photoirradiation was found to inhibit intra-abdominal metastases in an orthotopic pancreatic tumor mouse model. The cRGD-CPNPs also displayed good biosafety profiles, as inferred from PA tomography, blood analyses, and H&E staining. They thus appear promising for use in combined PA imaging and PT therapeutic treatment of pancreatic cancer.

Early metabolic markers as predictors of respiratory complications in preterm infants with bronchopulmonary dysplasia.

BACKGROUND: Bronchopulmonary dysplasia (BPD), a common complication of premature birth, exerts considerable impact on the respiratory health of infants. This study aimed to identify the role of plasma metabolites in predicting respiratory outcomes in BPD-afflicted infants. METHODS: This was a case-control study including 15 BPD premature infants and 15 gestational age and birth weight matched no-BPD preterm infants. Plasma samples, obtained at 36 weeks postmenstrual age (PMA), were subjected to a comprehensive analysis of over 300 metabolites using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The respiratory outcomes of the infants were collected with the first 2 years of corrected postnatal age. RESULTS: The analysis revealed a significant upregulation of urea and downregulation of nine metabolites in BPD infants, including oxalacetic acid, cis-aconitic acid, itaconic acid, betaine, L-asparagine, L-alanine, picolinic acid, inositol, and purine (p < 0.05). These metabolites primarily pertained to the citrate cycle (TCA cycle), glyoxylate and dicarboxylate metabolism, and alanine, aspartate, and glutamate metabolism. Furthermore, seven metabolites demonstrated substantial predictive capacity for respiratory readmissions within the first two years of corrected postnatal age, achieving an area under curve (AUC) exceeding or equal to 0.8. These included chenodeoxycholic acid, dehydrolithocholic acid, glucaric acid, D-glucuronic acid, gamma-glutamylvaline, mevalonic acid, and 3-ureidopropionic acid. CONCLUSIONS: This study identified ten distinct plasma metabolites at 36 weeks PMA that differentiate BPD infants from their non-BPD counterparts, implicating three major metabolic pathways. Additionally, seven metabolites showed strong predictive value for heightened risk of respiratory readmission within two years, underscoring their potential utility in clinical prognostication and management strategies for BPD.

Relief of tumor hypoxia using a nanoenzyme amplifies NIR-II photoacoustic-guided photothermal therapy.

Hypoxia is a critical tumor microenvironment (TME) component. It significantly impacts tumor growth and metastasis and is known to be a major obstacle for cancer therapy. Integrating hypoxia modulation with imaging-based monitoring represents a promising strategy that holds the potential for enhancing tumor theranostics. Herein, a kind of nanoenzyme Prussian blue (PB) is synthesized as a metal-organic framework (MOF) to load the second near-infrared (NIR-II) small molecule dye IR1061, which could catalyze hydrogen peroxide to produce oxygen and provide a photothermal conversion element for photoacoustic imaging (PAI) and photothermal therapy (PTT). To enhance stability and biocompatibility, silica was used as a coating for an integrated nanoplatform (SPI). SPI was found to relieve the hypoxic nature of the TME effectively, thus suppressing tumor cell migration and downregulating the expression of heat shock protein 70 (HSP70), both of which led to an amplified NIR-II PTT effect in vitro and in vivo, guided by the NIR-II PAI. Furthermore, label-free multi-spectral PAI permitted the real-time evaluation of SPI as a putative tumor treatment. A clinical histological analysis confirmed the amplified treatment effect. Hence, SPI combined with PAI could offer a new approach for tumor diagnosing, treating, and monitoring.

Synthesis and antitumor activity evaluation of coumarin Mannich base derivatives.

Twenty-one new coumarin Mannich base derivatives (11a-u) were synthesized, which exhibited antiproliferation activities in HepG2 (liver cancer), A549 (lung cancer), MCF-7 (breast cancer), and HT-29 (colon cancer). Most of the target compounds showed the most potent activity against HepG2 cells compared with other cancer cells, compound 11g showed the strongest antiproliferative activity (2.10 µM) against HepG2, even superior to the positive control drug 5-FU(5.49 µM). The nitric oxide (NO) release of all compounds in HepG2 cells was determined, of which compound 11g showed high levels of NO release (10.8 µM). Notably, the solubility of compound 11g increased 13-fold compared with the lead 8. The preliminary cytotoxicity studies suggest that 11g had little effect on LO2 cells(normal liver cells, >50 µM). The effect of compound 11g on the apoptosis of HepG2 cells was also studied, and the results showed that the induction effect of compound 11g on apoptosis is a concentration-dependent manner. Our results indicate that compound 11g might be a promising lead for further studies.

NIR-II Photoacoustic Imaging-Guided Oxygen Delivery and Controlled Release Improves Photodynamic Therapy for Hepatocellular Carcinoma.

Hypoxia, a prominent hallmark of hepatocellular carcinoma (HCC), undermines curative outcomes, elevates recurrence rates, and fosters metastasis, particularly during photodynamic therapy (PDT) in clinical settings. Studies indicate that alleviating tumor hypoxia enhances PDT efficacy. However, persistent challenges, including suboptimal oxygen delivery efficiency and absence of real-time feedback on blood oxygen fluctuations during PDT, considerably impede therapeutic efficacy in tumor treatment. This study addresses these issues using near-infrared-II (NIR-II) photoacoustic (PA) imaging for tumor-targeted oxygen delivery and controlled release. For this purpose, a biomimetic oxygen delivery system designated BLICP@O2 is developed, which utilizes hybrid tumor cell membranes and thermosensitive liposomes as oxygen carriers, incorporating the NIR-II dye IR1048, photosensitizer chlorin e6 (Ce6), and perfluorohexane. Upon sequential irradiation at 1064 and 690 nm, BLICP@O2 exhibits significant photothermal and photodynamic effects. Photothermal heating triggers oxygen release, enhancing the photodynamic effect of Ce6. Blood oxygen changes during PDT are tracked by multispectral PA imaging. Enhanced PDT efficacy, mediated by hypoxia relief, is convincingly demonstrated both in vitro and in vivo. This work presents an imaging-guided, dual-wavelength programmed cascaded treatment strategy for tumor-targeted oxygen delivery and controlled release, with real-time efficacy monitoring using PA imaging, offering valuable insights for overcoming challenges in PDT-based cancer therapy.

Restoring the imaging quality of circular transducer array-based PACT using synthetic aperture focusing technique integrated with 2nd-derivative-based back projection scheme.

Circular-array-based photoacoustic computed tomography (CA-PACT) is a promising imaging tool owing to its broad acoustic detection coverage and fidelity. However, CA-PACT suffers from poor image quality outside the focal zone along both elevational and lateral dimensions. To address this challenge, we proposed a novel reconstruction strategy by integrating the synthetic aperture focusing technique (SAFT) with the 2nd derivative-based back projection (2nd D-BP) algorithm to restore the image quality outside the focal zone along both the elevational and lateral axes. The proposed solution is a two-phase reconstruction scheme. In the first phase, with the assistance of an acoustic lens, we designed a circular array-based SAFT algorithm to restore the resolution and SNR along the elevational axis. The acoustic lens pushes the boundary of the upper limit of the SAFT scheme to achieve enhanced elevational resolution. In the second phase, we proposed a 2nd D-BP scheme to improve the lateral resolution and suppress noises in 3D imaging results. The 2nd D-BP strategy enhances the image quality along the lateral dimension by up-converting the high spatial frequencies of the object's absorption pattern. We validated the effectiveness of the proposed strategy using both phantoms and in vivo human experiments. The experimental results demonstrated superior image quality (7-fold enhancement in elevational resolution, 3-fold enhancement in lateral resolution, and an 11-dB increase in SNR). This strategy provides a new paradigm in the PACT system as it significantly enhances the spatial resolution and imaging contrast in both the elevational and lateral dimensions while maintaining a large focal zone.

Handheld photoacoustic imaging of indocyanine green clearance for real-time quantitative evaluation of liver reserve function.

Preoperative assessment of liver function reserve (LFR) is essential for determining the extent of liver resection and predicting the prognosis of patients with liver disease. In this paper, we present a real-time, handheld photoacoustic imaging (PAI) system-based noninvasive approach for rapid LFR assessment. A linear-array ultrasound transducer was sealed in a housing filled with water; its front end was covered with a plastic wrap. This PAI system was first implemented on phantoms to confirm that the photoacoustic (PA) intensity of indocyanine green (ICG) in blood reflects the concentration of ICG in blood. In vivo studies on normal rabbits and rabbits with liver fibrosis were carried out by recording the dynamic PA signal of ICG in their jugular veins. By analyzing the PA intensity-time curve, a clear difference was identified in the pharmacokinetic behavior of ICG between the two groups. In normal rabbits, the mean ICG clearance rate obtained by PAI at 15 min after administration (PAI-R15) was below 21.6%, whereas in rabbits with liver fibrosis, PAI-R15 exceeded 62.0% because of poor liver metabolism. The effectiveness of the proposed method was further validated by the conventional ICG clearance test and pathological examination. Our findings suggest that PAI is a rapid, noninvasive, and convenient method for LFR assessment and has immense potential for assisting clinicians in diagnosing and managing patients with liver disease.

Circular array transducer based-photoacoustic/ultrasonic endoscopic imaging with tunable ring-beam excitation.

Photoacoustic/ultrasound endoscopic imaging is regarded as an effective method to achieve accurate detection of intestinal disease by offering both the functional and structural information, simultaneously. Compared to the conventional endoscopy with single transducer and laser spot for signal detection and optical excitation, photoacoustic/ultrasound endoscopic probe using circular array transducer and ring-shaped laser beam avoids the instability brought by the mechanical scanning point-to-point, offering the dual-modality imaging with high accuracy and efficiency. Meanwhile, considering the complex morphological environments of intestinal tracts in clinics, developing the probe having sufficient wide imaging distance range is especially important. In this work, we develop a compact circular photoacoustic/ultrasonic endoscopic probe, using the group of fiber, lens and home-made axicon, to generate relatively concentrated ring-shaped laser beam for 360° excitation with high efficiency. Furthermore, the laser ring size can be tuned conveniently by changing the fiber-lens distance to ensure the potential applicability of the probe in various and complex morphological environments of intestines. Phantom experimental results demonstrate imaging distance range wide enough to cover from 12 mm to 30 mm. In addition, the accessibility of the photoacoustic signals of molecular probes in ex vivo experiments at the tissue depth of 7 mm using excitation energy of 5 mJ has also been demonstrated, showing a high optical excitation efficiency of the probe.

Multifunctional Electrospinning Polyhydroxyalkanoate Fibrous Scaffolds with Antibacterial and Angiogenesis Effects for Accelerating Wound Healing.

To treat large-scale wounds or chronic ulcers, it is highly desirable to develop multifunctional wound dressings that integrate antibacterial and angiogenic properties. While many biomaterials have been fabricated as wound dressings for skin regeneration, few reports have addressed the issue of complete skin regeneration due to the lack of vasculature and hair follicles. Herein, an instructive poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P34HB) fibrous wound dressing that integrates an antibacterial ciprofloxacin (CIP) and pro-angiogenic dimethyloxalylglycine (DMOG) is successfully prepared via electrospinning. The resultant dressings exhibit suitable flexibility with tensile strength and elongation at break up to 4.08 ± 0.18 MPa and 354.8 ± 18.4%, respectively. The in vitro results revealed that the groups of P34HB/CIP/DMOG dressings presented excellent biocompatibility on cell proliferation and significantly promote the spread and migration of L929 cells in both transwell and scratch assays. Capillary-like tube formation is also significantly enhanced in the P34HB/CIP/DMOG group dressings. Additionally, dressings from the P34HB/CIP and P34HB/CIP/DMOG groups show a broad spectrum of antimicrobial action against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli. In vivo studies further demonstrated that the prepared dressings in the P34HB/CIP/DMOG group not only improved wound closure, increased re-epithelialization and collagen formation, as well as reduced inflammatory response but also increased angiogenesis and remodeling, resulting in complete skin regeneration and hair follicles. Collectively, this work provides a simple but efficient approach for the design of a versatile wound dressing with the potential to have a synergistic effect on the rapid stimulation of angiogenesis as well as antibacterial activity in full-thickness skin repair.

Self-supervised learning enables 3D digital subtraction angiography reconstruction from ultra-sparse 2D projection views: A multicenter study.

3D digital subtraction angiography (DSA) reconstruction from rotational 2D projection X-ray angiography is an important basis for diagnosis and treatment of intracranial aneurysms (IAs). The gold standard requires approximately 133 different projection views for 3D reconstruction. A method to significantly reduce the radiation dosage while ensuring the reconstruction quality is yet to be developed. We propose a self-supervised learning method to realize 3D-DSA reconstruction using ultra-sparse 2D projections. 202 cases (100 from one hospital for training and testing, 102 from two other hospitals for external validation) suspected to be suffering from IAs were conducted to analyze the reconstructed images. Two radiologists scored the reconstructed images from internal and external datasets using eight projections and identified all 82 lesions with high diagnostic confidence. The radiation dosages are approximately 1/16.7 compared with the gold standard method. Our proposed method can help develop a revolutionary 3D-DSA reconstruction method for use in clinic.

Photoacoustic image-guided biomimetic nanoparticles targeting rheumatoid arthritis.

The high level of reactive oxygen species (ROS) in the rheumatoid arthritis (RA) microenvironment (RAM) and its persistent inflammatory nature can promote damage to joints, bones, and the synovium. Targeting strategies that integrate effective RAM regulation with imaging-based monitoring could lead to improvements in the diagnosis and treatment of RA. Here, we report the combined use of small interfering RNAs (siRNAsT/I) and Prussian blue nanoparticles (PBNPs) to silence the expression of proinflammatory cytokines TNF-α/IL-6 and scavenge the ROS associated with RAM. To enhance the in vitro and in vivo biological stability, biocompatibility, and targeting capability of the siRNAsT/I and PBNPs, macrophage membrane vesicles were used to prepare biomimetic nanoparticles, M@P-siRNAsT/I. The resulting constructs were found to suppress tumor necrosis factor-α/interleukin-6 expression and overcome the hypoxic nature of RAM, thus alleviating RA-induced joint damage in a mouse model. The M@P-siRNAsT/I of this study could be monitored via near-infrared photoacoustic (PA) imaging. Moreover, multispectral PA imaging without the need for labeling permitted the real-time evaluation of M@P-siRNAsT/I as a putative RA treatment. Clinical microcomputed tomography and histological analysis confirmed the effectiveness of the treatment. We thus suggest that macrophage-biomimetic M@P-siRNAsT/I and their analogs assisted by PA imaging could provide a new strategy for RA diagnosis, treatment, and monitoring.

Near-Infrared-II Activatable Symbiotic 2D Carbon-Clay Nanohybrids for Dual Imaging-Guided Combinational Cancer Therapy.

Two-dimensional (2D) nanomaterials hold great potential for cancer theranostic applications, yet their clinical translation faces great challenges of high toxicity and limited therapeutic/diagnostic modality. Here, we have created a kind of symbiotic 2D carbon-2D clay nanohybrids, which are composed of a novel 2D carbon nanomaterial (carbon nanochips, or CNC), prepared by carbonizing a conjugated polymer polydiiodobutadiyne, and a 2D layered aluminosilicate clay mineral montmorillonite (MMT). Intriguingly, with the formation of the nanohybrids, MMT can help the dispersion of CNC, while CNC can significantly reduce the hemolysis and toxicity of MMT. The symbiotic combination of CNC and MMT also leads to a synergistic anti-cancer theranostic effect. CNC has a strong absorption and high photothermal conversion efficiency in the second near-infrared region (NIR-II, 1000-1700 nm), while MMT contains Fe3+ that can facilitate the generation of reactive oxygen species from highly expressed H2O2 in tumor microenvironment. The nanohybrids not only enable a synergy of photothermal therapy and chemodynamic therapy to suppress the extremely rapid growth of RM1 tumors in mice but also allow for dual photoacoustic and magnetic imaging to guide the drug delivery and NIR-II irradiation execution, hence establishing a highly efficient and biosafe "all-in-one" theranostic platform for precision nanomedicine.

Video-rate high-resolution single-pixel nonscanning photoacoustic microscopy.

Optical-resolution photoacoustic microscopy (OR-PAM) is widely utilized in biomedical applications because of its ability to noninvasively image biological tissues in vivo while providing high-resolution morphological and functional information. However, one drawback of conventional OR-PAM is its imaging speed, which is restricted by the scanning technique employed. To achieve a higher imaging frame rate, we present video-rate high-resolution single-pixel nonscanning photoacoustic microscopy (SPN-PAM), which utilizes Fourier orthogonal basis structured planar illumination to overcome the above-mentioned limitations. A 473 × 473 µm2 imaging field of view (FOV) with 3.73 µm lateral resolution and video-rate imaging of 30 Hz were achieved. In addition, in both in vitro cell and in vivo mouse vascular hemodynamic imaging experiments, high-quality images were obtained at ultralow sampling rates. Thus, the proposed high-resolution SPN-PAM with video-rate imaging speed provides new insights into high-speed PA imaging and could be a powerful tool for rapid biological imaging.

Depth-extended acoustic-resolution photoacoustic microscopy based on a two-stage deep learning network.

Acoustic resolution photoacoustic microscopy (AR-PAM) is a major modality of photoacoustic imaging. It can non-invasively provide high-resolution morphological and functional information about biological tissues. However, the image quality of AR-PAM degrades rapidly when the targets move far away from the focus. Although some works have been conducted to extend the high-resolution imaging depth of AR-PAM, most of them have a small focal point requirement, which is generally not satisfied in a regular AR-PAM system. Therefore, we propose a two-stage deep learning (DL) reconstruction strategy for AR-PAM to recover high-resolution photoacoustic images at different out-of-focus depths adaptively. The residual U-Net with attention gate was developed to implement the image reconstruction. We carried out phantom and in vivo experiments to optimize the proposed DL network and verify the performance of the proposed reconstruction method. Experimental results demonstrated that our approach extends the depth-of-focus of AR-PAM from 1mm to 3mm under the 4 mJ/cm2 light energy used in the imaging system. In addition, the imaging resolution of the region 2 mm far away from the focus can be improved, similar to the in-focus area. The proposed method effectively improves the imaging ability of AR-PAM and thus could be used in various biomedical studies needing deeper depth.