Deep Learning for Automated Contouring of Gross Tumor Volumes in Esophageal Cancer.

Purpose: The aim of this study was to propose and evaluate a novel three-dimensional (3D) V-Net and two-dimensional (2D) U-Net mixed (VUMix-Net) architecture for a fully automatic and accurate gross tumor volume (GTV) in esophageal cancer (EC)-delineated contours. Methods: We collected the computed tomography (CT) scans of 215 EC patients. 3D V-Net, 2D U-Net, and VUMix-Net were developed and further applied simultaneously to delineate GTVs. The Dice similarity coefficient (DSC) and 95th-percentile Hausdorff distance (95HD) were used as quantitative metrics to evaluate the performance of the three models in ECs from different segments. The CT data of 20 patients were randomly selected as the ground truth (GT) masks, and the corresponding delineation results were generated by artificial intelligence (AI). Score differences between the two groups (GT versus AI) and the evaluation consistency were compared. Results: In all patients, there was a significant difference in the 2D DSCs from U-Net, V-Net, and VUMix-Net (p=0.01). In addition, VUMix-Net showed achieved better 3D-DSC and 95HD values. There was a significant difference among the 3D-DSC (mean ± STD) and 95HD values for upper-, middle-, and lower-segment EC (p<0.001), and the middle EC values were the best. In middle-segment EC, VUMix-Net achieved the highest 2D-DSC values (p<0.001) and lowest 95HD values (p=0.044). Conclusion: The new model (VUMix-Net) showed certain advantages in delineating the GTVs of EC. Additionally, it can generate the GTVs of EC that meet clinical requirements and have the same quality as human-generated contours. The system demonstrated the best performance for the ECs of the middle segment.

Integrated Histotripsy and Bubble Coalescence Transducer for Thrombolysis.

After the collapse of a cavitation bubble cloud, residual microbubbles can persist for up to seconds and function as weak cavitation nuclei for subsequent pulses in a phenomenon known as cavitation memory effect. In histotripsy, the cavitation memory effect can cause bubble clouds to repeatedly form at the same discrete set of sites. This effect limits the efficacy of histotripsy-based tissue fractionation. Our previous studies have indicated that low-amplitude bubble-coalescing (BC) ultrasound sequences interleaved with high-amplitude histotripsy pulses can coalesce the residual bubbles into one large bubble quickly. This reduces the cavitation memory effect and may increase treatment efficacy. Histotripsy has been investigated for thrombolysis by breaking up clots into debris smaller than red blood cells. However, this treatment has low efficacy for aged or retracted clots. In this study, we investigate the use of histotripsy with BC to improve the efficacy of treatment of retracted clots. An integrated histotripsy and bubble-coalescing (HBC) transducer system with specialized electronic driving system was built in-house. One high-amplitude (32 MPa), one-cycle histotripsy pulse followed by 36 low-amplitude (2.4 MPa), one-cycle BC pulses formed one HBC sequence. Results indicate that HBC sequences successfully generated a flow channel through the retracted clots at scan speeds of 0.2-0.5 mm/s. The channel size created using the HBC sequence was 128% to 480% larger than that created using histotripsy alone. The clot debris particles generated during HBC treatments were within the tolerable range. These results illustrate the concept that BC improves the treatment efficacy of histotripsy thrombolysis for retracted clots.

Integrated Histotripsy and Bubble Coalescence Transducer for Rapid Tissue Ablation.

Residual bubbles produced after collapse of a cavitation cloud provide cavitation nuclei for subsequent cavitation events, causing cavitation to occur repeatedly at the same discrete set of sites. This effect, referred to as cavitation memory, limits the efficiency of histotripsy soft tissue fractionation. Besides passively mitigating cavitation memory by using a low pulse repetition frequency (~1 Hz), an active strategy was developed by our group. In this strategy, low-amplitude ultrasound sequences were used to stimulate coalescence of residual bubbles. The goal of this work is to remove cavitation memory and achieve rapid, homogeneous lesion formation using a single phased array transducer. A 1-MHz integrated histotripsy and bubble coalescing (BC) transducer system with a specialized electronic driving system was built in house. High-amplitude ( MPa) histotripsy pulses and subsequent low-amplitude (~1-2 MPa) BC sequences were applied to a red blood cell tissue-mimicking phantom at a single focal site. Significant reduction of the cavitation memory effect and increase in the fractionation rate were observed by introducing BC sequence. Effects of BC pulsing parameters were further studied. The optimal BC parameters were then utilized to homogenize a mm2 region at high rate.

Real-time monitoring of controllable cavitation erosion in a vessel phantom with passive acoustic mapping.

Cavitation erosion in blood vessel plays an important role in ultrasound thrombolysis, drug delivery, and other clinical applications. The controllable superficial vessel erosion based on ultrasonic standing wave (USW) has been used to effectively prevent vessel ruptures and haemorrhages, and optical method is used to observe the experiments. But optical method can only work in transparent media. Compared with standard B-mode imaging, passive acoustic mapping (PAM) can monitor erosion in real time and has better sensitivity of cavitation detection. However, the conventionally used PAM has limitations in imaging resolution and artifacts. In this study, a unique PAM method that combined the robust Capon beamformer (RCB) with the sign coherence factor (SCF) was proposed to monitor the superficial vessel erosion in real time. The performance of the proposed method was validated by simulations. In vitro experiments showed that the lateral (axial) resolution of the proposed PAM was 2.31±0.51 (3.19±0.38) times higher than time exposure acoustics (TEA)-based PAM and 1.73±0.38 (1.76±0.48) times higher than RCB-based PAM, and the cavitation-to-artifact ratio (CAR) of the proposed PAM could be improved by 22.5±3.2dB and 7.1±1.2dB compared with TEA and RCB-based PAM. These results showed that the proposed PAM can precisely monitor the superficial vessel erosion and the erosion shift after USW modulation. This work may have the potential of developing a useful tool for precise spatial control and real-time monitoring of the superficial vessel erosion.

Neuroprotective Effect of Low-Intensity Pulsed Ultrasound Against MPP+-Induced Neurotoxicity in PC12 Cells: Involvement of K2P Channels and Stretch-Activated Ion Channels.

Parkinson's disease is the second most common neurodegenerative disease. It is characterized by the loss of dopaminergic neurons in the substantia nigra pars compacta. 1-Methyl-4-phenylpyridinium (MPP+) is a dopaminergic neuronal toxin that is widely used in constructing Parkinson's disease models in vitro. Low-intensity pulsed ultrasound (LIPUS) is a non-invasive therapeutic approach that has neuromodulation and neuroprotective effects in the central neural system; however, whether LIPUS can provide protection for dopaminergic neurons against MPP+-induced neurocytotoxicity remains unknown. In this study, we found that pre-treatment with LIPUS (1 MHz, 50 mW/cm2, 20% duty cycle and 100-Hz pulse repetition frequency, 10 min) inhibited MPP+-induced neurotoxicity and mitochondrial dysfunction in PC12 cells. LIPUS decreased MPP+-induced oxidative stress by modulating antioxidant proteins, including thioredoxin-1 and heme oxygenase-1, and prevented neurocytotoxicity via the phosphoinositide 3-kinase (PI3K)-Akt and ERK1/2 pathways. Furthermore, these beneficial effects were attributed to the activation of K2P channels and stretch-activated ion channels by LIPUS. These data indicate that LIPUS protects neuronal cells from MPP+-induced cell death through the K2P channel- and stretch-activated ion channel-mediated downstream pathways. The data also suggest that LIPUS could be a promising therapeutic method in halting or retarding the degeneration of dopaminergic neurons in Parkinson's disease in a non-invasive manner.

Low-Intensity Pulsed Ultrasound Enhances Nerve Growth Factor-Induced Neurite Outgrowth through Mechanotransduction-Mediated ERK1/2-CREB-Trx-1 Signaling.

Enhancing the action of nerve growth factor (NGF) is a potential therapeutic approach to neural regeneration. To facilitate neural regeneration, we investigated whether combining low-intensity pulsed ultrasound (LIPUS) and NGF could promote neurite outgrowth, an essential process in neural regeneration. In the present study, PC12 cells were subjected to a combination of LIPUS (1 MHz, 30 or 50 mW/cm2, 20% duty cycle and 100-Hz pulse repetition frequency, 10 min every other day) and NGF (50 ng/mL) treatment, and then neurite outgrowth was compared. Our findings indicated that the combined treatment with LIPUS (50 mW/cm2) and NGF (50 ng/mL) promotes neurite outgrowth that is comparable to that achieved by NGF (100 ng/mL) treatment alone. LIPUS significantly increased NGF-induced neurite length, but not neurite branching. These effects were attributed to the enhancing effects of LIPUS on NGF-induced phosphorylation of ERK1/2 and CREB and the expression of thioredoxin (Trx-1). Furthermore, blockage of stretch-activated ion channels with Gd3+ suppressed the stimulating effects of LIPUS on NGF-induced neurite outgrowth and the downstream signaling activation. Taken together, our findings suggest that LIPUS enhances NGF-induced neurite outgrowth through mechanotransduction-mediated signaling of the ERK1/2-CREB-Trx-1 pathway. The combination of LIPUS and NGF could potentially be used for the treatment of nerve injury and neurodegenerative diseases.

Precise spatial control of cavitation erosion in a vessel phantom by using an ultrasonic standing wave.

In atherosclerotic inducement in animal models, the conventionally used balloon injury is invasive, produces excessive vessel injuries at unpredictable locations and is inconvenient in arterioles. Fortunately, cavitation erosion, which plays an important role in therapeutic ultrasound in blood vessels, has the potential to induce atherosclerosis noninvasively at predictable sites. In this study, precise spatial control of cavitation erosion for superficial lesions in a vessel phantom was realised by using an ultrasonic standing wave (USW) with the participation of cavitation nuclei and medium-intensity ultrasound pulses. The superficial vessel erosions were restricted between adjacent pressure nodes, which were 0.87 mm apart in the USW field of 1 MHz. The erosion positions could be shifted along the vessel by nodal modulation under a submillimetre-scale accuracy without moving the ultrasound transducers. Moreover, the cavitation erosion of the proximal or distal wall could be determined by the types of cavitation nuclei and their corresponding cavitation pulses, i.e., phase-change microbubbles with cavitation pulses of 5 MHz and SonoVue microbubbles with cavitation pulses of 1 MHz. Effects of acoustic parameters of the cavitation pulses on the cavitation erosions were investigated. The flow conditions in the experiments were considered and discussed. Compared to only using travelling waves, the proposed method in this paper improves the controllability of the cavitation erosion and reduces the erosion depth, providing a more suitable approach for vessel endothelial injury while avoiding haemorrhage.

Apoptosis Induced by Microbubble-Assisted Acoustic Cavitation in K562 Cells: The Predominant Role of the Cyclosporin A-Dependent Mitochondrial Permeability Transition Pore.

Acoustic cavitation of microbubbles has been described as inducing tumor cell apoptosis that is partly associated with mitochondrial dysfunction; however, the exact mechanisms have not been fully characterized. Here, low-intensity pulsed ultrasound (1 MHz, 0.3-MPa peak negative pressure, 10% duty cycle and 1-kHz pulse repetition frequency) was applied to K562 chronic myelogenous leukemia cells for 1 min with 10% (v/v) SonoVue microbubbles. After ultrasound exposure, the apoptotic index was determined by flow cytometry with annexin V-fluorescein isothiocyanate/propidium iodide. In addition, mitochondrial membrane potential (ΔΨm) was determined with the JC-1 assay. Translocation of apoptosis-associated protein cytochrome c was evaluated by Western blotting. We found that microbubble-assisted acoustic cavitation can increase the cellular apoptotic index, mitochondrial depolarization and cytochrome c release in K562 cells, compared with ultrasound treatment alone. Furthermore, mitochondrial dysfunction and apoptosis were significantly inhibited by cyclosporin A, a classic inhibitor of the mitochondrial permeability transition pore; however, the inhibitor of Bax protein, Bax-inhibiting peptide, could not suppress these effects. Our results suggest that mitochondrial permeability transition pore opening is involved in mitochondrial dysfunction after exposure to microbubble-assisted acoustic cavitation. Moreover, the release of cytochrome c from the mitochondria is dependent on cyclosporin A-sensitive mitochondrial permeability transition pore opening, but not formation of the Bax-voltage dependent anion channel complex or Bax oligomeric pores. These data provide more insight into the mechanisms underlying mitochondrial dysfunction induced by acoustic cavitation and can be used as a basis for therapy.