Pneumatosis Intestinalis, Pneumoperitoneum, Pneumoretroperitoneum, Pneumomediastinum, and Pneumobilia After Pembrolizumab Therapy: A Case Report.

Immune checkpoint inhibitors (ICIs) are becoming increasingly popular in treating cancers resistant to traditional chemotherapy. While ICIs have shown promise in treating cancer, the class of drugs also comes with certain risks, such as the development of pneumatosis intestinalis (PI) in rare cases. Pembrolizumab, an ICI that inhibits programmed cell death protein 1 (PD-1), has, in some rare instances, caused PI. Patients with ICI-induced PI may also present with pneumoperitoneum, pneumoretroperitoneum, pneumomediastinum, and pneumobilia. In the current report, we describe the presentation and management of a 50-year-old female with initial complaints of diffuse abdominal pain, constipation, abdominal distension, nausea, and decreased urine output approximately six months after beginning pembrolizumab and two months after the most recent dose of pembrolizumab. Subsequent CT imaging revealed massive PI with pneumoperitoneum, pneumoretroperitoneum, pneumomediastinum, and pneumobilia suspected to be secondary to pembrolizumab. Here, we discuss the possible mechanisms of ICI-induced PI and evaluate the management of patients presenting with PI and pneumoperitoneum.

Immune and metabolic perturbations in COVID-19-associated pulmonary mucormycosis: A transcriptome analysis of innate immune cells.

BACKGROUND AND OBJECTIVES: The mechanisms underlying COVID-19-associated pulmonary mucormycosis (CAPM) remain unclear. We use a transcriptomic analysis of the innate immune cells to investigate the host immune and metabolic response pathways in patients with CAPM. PATIENTS AND METHODS: We enrolled subjects with CAPM (n = 5), pulmonary mucormycosis (PM) without COVID-19 (n = 5), COVID-19 (without mucormycosis, n = 5), healthy controls (n = 5) without comorbid illness and negative for SARS-CoV-2. Peripheral blood samples from cases were collected before initiating antifungal therapy, and neutrophils and monocytes were isolated. RNA sequencing was performed using Illumina HiSeqX from monocytes and neutrophils. Raw reads were aligned with HISAT-2 pipeline and DESeq2 was used for differential gene expression. Gene ontology (GO) and metabolic pathway analysis were performed using Shiny GO application and R packages (ggplot2, Pathview). RESULTS: The derangement of core immune and metabolic responses in CAPM patients was noted. Pattern recognition receptors, dectin-2, MCL, FcRγ receptors and CLEC-2, were upregulated, but signalling pathways such as JAK-STAT, IL-17 and CARD-9 were downregulated; mTOR and MAP-kinase signalling were elevated in monocytes from CAPM patients. The complement receptors, NETosis, and pro-inflammatory responses, such as S100A8/A9, lipocalin and MMP9, were elevated. The major metabolic pathways of glucose metabolism-glycolysis/gluconeogenesis, pentose phosphate pathway, HIF signalling and iron metabolism-ferroptosis were also upregulated in CAPM. CONCLUSIONS: We identified significant alterations in the metabolic pathways possibly leading to cellular iron overload and a hyperglycaemic state. Immune responses revealed altered recognition, signalling, effector functions and a pro-inflammatory state in monocytes and neutrophils from CAPM patients.

Spatial Coherence Approaches to Distinguish Suspicious Mass Contents in Fundamental and Harmonic Breast Ultrasound Images.

When compared to fundamental B-mode imaging, coherence-based beamforming, and harmonic imaging are independently known to reduce acoustic clutter, distinguish solid from fluid content in indeterminate breast masses, and thereby reduce unnecessary biopsies during a breast cancer diagnosis. However, a systematic investigation of independent and combined coherence beamforming and harmonic imaging approaches is necessary for the clinical deployment of the most optimal approach. Therefore, we compare the performance of fundamental and harmonic images created with short-lag spatial coherence (SLSC), M-weighted SLSC (M-SLSC), SLSC combined with robust principal component analysis with no M-weighting (r-SLSC), and r-SLSC with M-weighting (R-SLSC), relative to traditional fundamental and harmonic B-mode images, when distinguishing solid from fluid breast masses. Raw channel data acquired from 40 total breast masses (28 solid, 7 fluid, 5 mixed) were beamformed and analyzed. The contrast of fluid masses was better with fundamental rather than harmonic coherence imaging, due to the lower spatial coherence within the fluid masses in the fundamental coherence images. Relative to SLSC imaging, M-SLSC, r-SLSC, and R-SLSC imaging provided similar contrast across multiple masses (with the exception of clinically challenging complicated cysts) and minimized the range of generalized contrast-to-noise ratios (gCNRs) of fluid masses, yet required additional computational resources. Among the eight coherence imaging modes compared, fundamental SLSC imaging best identified fluid versus solid breast mass contents, outperforming fundamental and harmonic B-mode imaging. With fundamental SLSC images, the specificity and sensitivity to identify fluid masses using the reader-independent metrics of contrast difference, mean lag one coherence (LOC), and gCNR were 0.86 and 1, 1 and 0.89, and 1 and 1, respectively. Results demonstrate that fundamental SLSC imaging and gCNR (or LOC if no coherence image or background region of interest is introduced) have the greatest potential to impact clinical decisions and improve the diagnostic certainty of breast mass contents. These observations are additionally anticipated to extend to masses in other organs.

Optical absorption spectra and corresponding in vivo photoacoustic visualization of exposed peripheral nerves.

Significance: Multispectral photoacoustic imaging has the potential to identify lipid-rich, myelinated nerve tissue in an interventional or surgical setting (e.g., to guide intraoperative decisions when exposing a nerve during reconstructive surgery by limiting operations to nerves needing repair, with no impact to healthy or regenerating nerves). Lipids have two optical absorption peaks within the NIR-II and NIR-III windows (i.e., 1000 to 1350 nm and 1550 to 1870 nm wavelength ranges, respectively) which can be exploited to obtain photoacoustic images. However, nerve visualization within the NIR-III window is more desirable due to higher lipid absorption peaks and a corresponding valley in the optical absorption of water. Aim: We present the first known optical absorption characterizations, photoacoustic spectral demonstrations, and histological validations to support in vivo photoacoustic nerve imaging in the NIR-III window. Approach: Four in vivo swine peripheral nerves were excised, and the optical absorption spectra of these fresh ex vivo nerves were characterized at wavelengths spanning 800 to 1880 nm, to provide the first known nerve optical absorbance spectra and to enable photoacoustic amplitude spectra characterization with the most optimal wavelength range. Prior to excision, the latter two of the four nerves were surrounded by aqueous, lipid-free, agarose blocks (i.e., 3% w/v agarose) to enhance acoustic coupling during in vivo multispectral photoacoustic imaging using the optimal NIR-III wavelengths (i.e., 1630 to 1850 nm) identified in the ex vivo studies. Results: There was a verified characteristic lipid absorption peak at 1725 nm for each ex vivo nerve. Results additionally suggest that the 1630 to 1850 nm wavelength range can successfully visualize and differentiate lipid-rich nerves from surrounding water-containing and lipid-deficient tissues and materials. Conclusions: Photoacoustic imaging using the optimal wavelengths identified and demonstrated for nerves holds promise for detection of myelination in exposed and isolated nerve tissue during a nerve repair surgery, with possible future implications for other surgeries and other optics-based technologies.

Adaptive enhancement of acoustic resolution photoacoustic microscopy imaging via deep CNN prior.

Acoustic resolution photoacoustic microscopy (AR-PAM) is a promising medical imaging modality that can be employed for deep bio-tissue imaging. However, its relatively low imaging resolution has greatly hindered its wide applications. Previous model-based or learning-based PAM enhancement algorithms either require design of complex handcrafted prior to achieve good performance or lack the interpretability and flexibility that can adapt to different degradation models. However, the degradation model of AR-PAM imaging is subject to both imaging depth and center frequency of ultrasound transducer, which varies in different imaging conditions and cannot be handled by a single neural network model. To address this limitation, an algorithm integrating both learning-based and model-based method is proposed here so that a single framework can deal with various distortion functions adaptively. The vasculature image statistics is implicitly learned by a deep convolutional neural network, which served as plug and play (PnP) prior. The trained network can be directly plugged into the model-based optimization framework for iterative AR-PAM image enhancement, which fitted for different degradation mechanisms. Based on physical model, the point spread function (PSF) kernels for various AR-PAM imaging situations are derived and used for the enhancement of simulation and in vivo AR-PAM images, which collectively proved the effectiveness of proposed method. Quantitatively, the PSNR and SSIM values have all achieve best performance with the proposed algorithm in all three simulation scenarios; The SNR and CNR values have also significantly raised from 6.34 and 5.79 to 35.37 and 29.66 respectively in an in vivo testing result with the proposed algorithm.

Technique for circuit exchange in high-risk patients on extracorporeal life support.

Exchanging circuits in patients undergoing extracorporeal life support (ECLS) for component failure can be a life-threatening endeavor if the patient has no native organ function and is fully dependent on ECLS. Traditional circuit exchanges involve replacing the entire circuit at once, leading to a sufficient loss of support that risks hemodynamic deterioration in the absence of mechanical ventilation or mechanical circulatory support. A staged approach is described using a parallel circuit configuration to reduce the risk of physiologic decompensation in patients with total dependence on ECLS. The approach involves splicing in a second primed circuit in parallel, transitioning flow incrementally from old to new circuit, and removing the old circuit. This approach provides hemodynamic and physiologic stability even in patients with absent underlying cardiopulmonary function.

Sex difference in clinical and procedural outcomes in patients undergoing coronary atherectomy: a systematic review and meta-analysis.

BACKGROUND: Rotational and orbital coronary atherectomy (CA) are commonly utilized to treat complex calcified coronary lesions. We conducted a meta-analysis to evaluate sex differences in procedural complications and clinical outcomes after CA. METHODS: PubMed, Google Scholar, and Cochrane databases were searched for all studies comparing sex differences in procedural and clinical outcomes following CA. The outcomes of interest were procedural complications (coronary dissection, stroke, major bleeding, coronary perforation, cardiac tamponade, and slow or no flow in target vessel) and the clinical outcomes (including early mortality, mid-term all-cause mortality, stroke, myocardial infarction, and target vessel revascularization). Pooled risk ratios (RRs) with their corresponding 95% confidence intervals (CIs) were calculated using the Mantel-Haenszel random-effects model. RESULTS: Six observational studies with 3517 patients (2420 men and 1035 women) were included in this meta-analysis. While there was no significant difference in the early mortality (RR, 1.14; 95% CI, 0.37-3.53; P = 0.83) between men and women, at a mean follow-up of 2.9 years, all-cause mortality was significantly higher in women (RR, 1.29; 95% CI, 1.11-1.49; P = 0.0009). Women had an increased risk of procedure-related stroke (RR, 3.98; 95% CI, 1.06-14.90; P = 0.04), coronary dissection (RR, 2.10; 95% CI, 1.23-3.58; P = 0.006), and bleeding (RR, 2.26; 95% CI, 1.30-3.93; P = 0.004), whereas the rates of coronary perforation, cardiac tamponade, and the risk of slow or no flow in the revascularized artery were similar in both. CONCLUSION: In our analysis, women undergoing CA are at increased risk of mid-term mortality and procedure-related complications including stroke, coronary dissection, and major bleeding.

Deep and Domain Transfer Learning Aided Photoacoustic Microscopy: Acoustic Resolution to Optical Resolution.

Acoustic resolution photoacoustic micros- copy (AR-PAM) can achieve deeper imaging depth in biological tissue, with the sacrifice of imaging resolution compared with optical resolution photoacoustic microscopy (OR-PAM). Here we aim to enhance the AR-PAM image quality towards OR-PAM image, which specifically includes the enhancement of imaging resolution, restoration of micro-vasculatures, and reduction of artifacts. To address this issue, a network (MultiResU-Net) is first trained as generative model with simulated AR-OR image pairs, which are synthesized with physical transducer model. Moderate enhancement results can already be obtained when applying this model to in vivo AR imaging data. Nevertheless, the perceptual quality is unsatisfactory due to domain shift. Further, domain transfer learning technique under generative adversarial network (GAN) framework is proposed to drive the enhanced image's manifold towards that of real OR image. In this way, perceptually convincing AR to OR enhancement result is obtained, which can also be supported by quantitative analysis. Peak Signal to Noise Ratio (PSNR) and Structural Similarity Index (SSIM) values are significantly increased from 14.74 dB to 19.01 dB and from 0.1974 to 0.2937, respectively, validating the improvement of reconstruction correctness and overall perceptual quality. The proposed algorithm has also been validated across different imaging depths with experiments conducted in both shallow and deep tissue. The above AR to OR domain transfer learning with GAN (AODTL-GAN) framework has enabled the enhancement target with limited amount of matched in vivo AR-OR imaging data.

Serum glucose-regulated protein 78 (GRP78) levels in COVID-19-associated mucormycosis: results of a case-control study.

BACKGROUND: In experimental models, the expression of glucose-regulated protein 78 (GRP78) in endothelial cells played a role in the pathogenesis of mucormycosis. However, the role of GRP78 in COVID-19-associated mucormycosis (CAM) has not been studied. We hypothesized that serum GRP78 levels are elevated in subjects with CAM. OBJECTIVE: To compare the serum GRP78 levels in subjects with CAM and COVID-19 controls without mucormycosis. DESIGN AND SETTING: We performed a hospital-based, case-control study between 1 April 2021 and 31 May 2021. PARTICIPANTS: We enrolled 24 subjects each of CAM and COVID-19 subjects without mucormycosis. We also measured serum GRP78 levels in ten healthy controls. EXPOSURE: The primary exposure studied was serum GRP78 concentration, estimated using a commercially available ELISA kit in stored serum samples. RESULTS: We found the mean ± standard deviation (SD) serum GRP78 levels significantly higher (p = 0.0001) among the CAM (374.3 ± 127.3 pg/mL) than the COVID-19 (246.4 ± 67.0 pg/mL) controls. The proportion of subjects with an abnormal GRP78 level (> mean [184.8 pg/mL] plus two SD [23.2 pg/mL] of GRP78 from healthy participants) was 87.5% and 45.8% in the CAM group and COVID-19 controls, respectively. Serum GRP78 level was independently associated with CAM (odds ratio 1.011; 95% confidence interval [1.002-1.019]) after adjusting for diabetes mellitus and hypoxemia during acute COVID-19. CONCLUSION: Serum GRP78 levels were significantly higher in CAM than in COVID-19 controls. Further studies are required to the role of GRP78 in the pathogenesis of CAM.

Right atrial appendage thrombus in a patient with a left atrial appendage occlusion device.

Management of right atrial appendage (RAA) thrombus is a clinical dilemma. We describe a case of incidentally found RAA thrombus in a patient with a left atrial appendage closure device (WATCHMAN). Options for the management of RAA thrombus include observation, anticoagulation, thrombolytics, or surgical extraction. Size, mobility, site of attachment of the thrombi, patient factors (e.g., bleeding risk), and other indications for anticoagulation may be used to guide management.

Photoacoustic imaging aided with deep learning: a review.

Photoacoustic imaging (PAI) is an emerging hybrid imaging modality integrating the benefits of both optical and ultrasound imaging. Although PAI exhibits superior imaging capabilities, its translation into clinics is still hindered by various limitations. In recent years, deeplearning (DL), a new paradigm of machine learning, is gaining a lot of attention due to its ability to improve medical images. Likewise, DL is also widely being used in PAI to overcome some of the limitations of PAI. In this review, we provide a comprehensive overview on the various DL techniques employed in PAI along with its promising advantages.

Another decade of photoacoustic imaging.

Photoacoustic imaging-a hybrid biomedical imaging modality finding its way to clinical practices. Although the photoacoustic phenomenon was known more than a century back, only in the last two decades it has been widely researched and used for biomedical imaging applications. In this review we focus on the development and progress of the technology in the last decade (2011-2020). From becoming more and more user friendly, cheaper in cost, portable in size, photoacoustic imaging promises a wide range of applications, if translated to clinic. The growth of photoacoustic community is steady, and with several new directions researchers are exploring, it is inevitable that photoacoustic imaging will one day establish itself as a regular imaging system in the clinical practices.

Broken Scalpel Blade During Emergent Cricothyroidotomy: An Unexpected Complication in a Critical Situation.

Scalpel-bougie cricothyroidotomy is the most common surgical procedure to obtain emergency airway access when routine methods fail. We present a case of a broken scalpel blade during emergency cricothyroidotomy further complicating respiratory access.

Comparison of ultrasound-guided intermediate vs subcutaneous cervical plexus block for postoperative analgesia in patients undergoing total thyroidectomy: A randomised double-blind trial.

BACKGROUND AND AIMS: Intermediate cervical plexus block (CPB) is a new procedure whose analgesic efficacy compared to superficial cervical plexus block is yet to be established. We compared the analgesic efficacy of superficial vs intermediate CPB for post-operative analgesia after thyroid surgery. METHODS: Forty-five patients with American Society of Anaesthesiologists' physical status 1 or 2 undergoing total thyroidectomy were recruited. Forty-four patients in superficial/subcutaneous CPB group (n = 22) and intermediate CPB (n = 22) received 20 mL 0.25% bupivacaine with adrenaline 100 µg bilaterally in ultrasound-guided superficial and intermediate cervical plexus block before induction of general anaesthesia., respectively. The primary outcome measure was the postoperative visual analogue scale (VAS) scores at 0, 2, 4, 6, 12 and 24. Secondary outcome measures included the total dose of rescue analgesic required, duration of postoperative analgesia and patient's satisfaction score. Statistical analysis was with the Mann-Whitney U test and independent t-test. RESULTS: The post-operative VAS scores were lower in intermediate CPB group compared to superficial CPB group at 2, 4, 6, 12, 18 and 24 h [P < 0.05]. Time tofirst rescue analgesic demand was prolonged 10.06 ± 3.62 h in intermediate group compared to 7.94 ± 3.62 h in superficial group [P = 0.017] and total analgesic consumption were lower in intermediate group (71.25 ± 16.70 µg) than the superficial group (101.25 ± 50.31 µg) [P = 0.011]. CONCLUSION: Ultrasound-guided intermediate CPB reduces post-operative pain scores, prolongs duration of analgesia and decreases demands for rescue analgesia compared to superficial CPB.

High resolution, label-free photoacoustic imaging of live chicken embryo developing in bioengineered eggshell.

Chicken embryos have been proven to be an attractive vertebrate model for biomedical research. They have helped in making significant contributions for advancements in various fields like developmental biology, cancer research and cardiovascular studies. However, a non-invasive, label-free method of imaging live chicken embryo at high resolution still needs to be developed and optimized. In this work, we have shown the potential of photoacoustic tomography (PAT) for imaging live chicken embryos cultured in bioengineered eggshells. Laser pulses at wavelengths of 532 and 740 nm were used for attaining cross-sectional images of chicken embryos at different developmental stages. Cross-sections along different depths were imaged to gain knowledge of the relative depth of different vessels and organs. Due to high optical absorption of vasculature and embryonic eye, images with good optical contrast could be acquired using this method. We have thus reported a label-free method of performing cross-sectional imaging of chicken embryos at high resolution demonstrating the capacity of PAT as a promising tool for avian embryo imaging.

Rapid detection of terbinafine resistance in Trichophyton species by Amplified refractory mutation system-polymerase chain reaction.

Dermatophytosis has gained interest in India due to rise in terbinafine resistance and difficulty in management of recalcitrant disease. The terbinafine resistance in dermatophytes is attributed to single nucleotide polymorphisms (SNPs) in squalene epoxidase (SE) gene. We evaluated the utility of amplified refractory mutation system polymerase chain reaction (ARMS PCR) for detection of previously reported point mutations, including a mutation C1191A in the SE gene in Trichophyton species. ARMS PCR was standardized using nine non-wild type isolates and two wild type isolates of Trichophyton species. Study included 214 patients with dermatophyte infection from March through December 2017. Antifungal susceptibility testing of isolated dermatophytes was performed according to CLSI-M38A2 guidelines. Among dermatophytes isolated in 68.2% (146/214) patients, Trichophyton species were predominant (66.4%). High (>2 mg/L, cut off) minimum inhibitory concentrations to terbinafine were noted in 15 (15.4%) Trichophyton mentagrophytes complex isolates. A complete agreement was noted between ARMS PCR assay and DNA sequencing. C to A transversion was responsible for amino acid substitution in 397th position of SE gene in terbinafine resistant isolates. Thus, the ARMS PCR assay is a simple and reliable method to detect terbinafine-resistant Trichophyton isolates.

Convolutional neural network for resolution enhancement and noise reduction in acoustic resolution photoacoustic microscopy.

In acoustic resolution photoacoustic microscopy (AR-PAM), a high numerical aperture focused ultrasound transducer (UST) is used for deep tissue high resolution photoacoustic imaging. There is a significant degradation of lateral resolution in the out-of-focus region. Improvement in out-of-focus resolution without degrading the image quality remains a challenge. In this work, we propose a deep learning-based method to improve the resolution of AR-PAM images, especially at the out of focus plane. A modified fully dense U-Net based architecture was trained on simulated AR-PAM images. Applying the trained model on experimental images showed that the variation in resolution is ∼10% across the entire imaging depth (∼4 mm) in the deep learning-based method, compared to ∼180% variation in the original PAM images. Performance of the trained network on in vivo rat vasculature imaging further validated that noise-free, high resolution images can be obtained using this method.

Enhanced contrast acoustic-resolution photoacoustic microscopy using double-stage delay-multiply-and-sum beamformer for vasculature imaging.

In acoustic-resolution photoacoustic microscopy (AR-PAM) systems, the lateral resolution in the focal zone of the ultrasound (US) transducer is determined by the numerical aperture (NA) of the transducer. To have a high lateral resolution, a large NA is used. However, the larger the NA, the smaller the depth of focus [DOF]. As a result, the lateral resolution is deteriorated at depths out of the focal region. The synthetic aperture focusing technique (SAFT) along with a beamformer can be used to improve the resolution outside the focal region. In this work, for image formation in AR-PAM, we propose the double-stage delay-multiply-and-sum (DS_DMAS) algorithm to be combined with SAFT. The proposed method is evaluated experimentally using hair targets and in vivo vasculature imaging. It is shown that DS_DMAS provides a higher resolution and contrast compared to other methods. For the B-mode images obtained using the hair phantom, the proposed method reduces the average noise level for all the depths by about 134%, 57% and 23%, compared to the original low- resolution, SAFT+DAS and SAFT+DMAS methods, respectively. All the results indicate that the proposed method can be an appropriate algorithm for image formation in AR-PAM systems.

Metabolizable Semiconducting Polymer Nanoparticles for Second Near-Infrared Photoacoustic Imaging.

Photoacoustic (PA) imaging in the second near-infrared (NIR-II) window (1000-1700 nm) holds great promise for deep-tissue diagnosis due to the reduced light scattering and minimized tissue absorption; however, exploration of such a noninvasive imaging technique is greatly constrained by the lack of biodegradable NIR-II absorbing agents. Herein, the first series of metabolizable NIR-II PA agents are reported based on semiconducting polymer nanoparticles (SPNs). Such completely organic nanoagents consist of π-conjugated yet oxidizable optical polymer as PA generator and hydrolyzable amphiphilic polymer as particle matrix to provide water solubility. The obtained SPNs are readily degraded by myeloperoxidase and lipase abundant in phagocytes, transforming from nonfluorescent nanoparticles (30 nm) into NIR fluorescent ultrasmall metabolites (≈1 nm). As such, these nanoagents can be effectively cleared out via both hepatobiliary and renal excretions after systematic administration, leaving no toxicity to living mice. Particularly these nanoagents possess high photothermal conversion efficiencies and emit bright PA signals at 1064 nm, enabling sensitive NIR-II PA imaging of both subcutaneous tumor and deep brain vasculature through intact skull in living animals at a low systematic dosage. This study thus provides a generalized molecular design toward organic metabolizable semiconducting materials for biophotonic applications in NIR-II window.

1064 nm acoustic resolution photoacoustic microscopy.

Photoacoustic imaging is a noninvasive imaging technique having the advantages of high-optical contrast and good acoustic resolution at improved imaging depths. Light transport in biological tissues is mainly characterized by strong optical scattering and absorption. Photoacoustic microscopy is capable of achieving high-resolution images at greater depth compared to conventional optical microscopy methods. In this work, we have developed a high-resolution, acoustic resolution photoacoustic microscopy (AR-PAM) system in the near infra-red (NIR) window II (NIR-II, eg, 1064 nm) for deep tissue imaging. Higher imaging depth is achieved as the tissue scattering at 1064 nm is lesser compared to visible or near infrared window-I (NIR-I). Our developed system can provide a lateral resolution of 130 µm, axial resolution of 57 µm, and image up to 11 mm deep in biological tissues. This 1064-AR-PAM system was used for imaging sentinel lymph node and the lymph vessel in rat. Urinary bladder of rat filled with black ink was also imaged to validate the feasibility of the developed system to study deeply seated organs.