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Recurrent or metastatic peri-urethral pelvic malignancies are a difficult-to-treat entity. Re-resection is recommended when possible but is frequently unfavorable due to scar tissue, fibrosis, and obliteration of tissue planes following previous interventions such as surgical resection and/or radiation therapy. Curative options for patients that have unresectable cancer are limited. Cryo-ablation has been extensively studied in the treatment of unresectable renal, liver and lung malignancies and has the potential to provide definitive treatment for recurrent pelvic malignancy. There is a paucity of reports of salvage cryo-ablation in patients with recurrent pelvic malignancies, as most of these tumors are located close to critical structures that could be irreversibly injured by thermal ablation and are hence treated with some form of radiation therapy. But, for patients who fail surgical and radiation treatments, options are limited. Here, we describe two cases of regional tumor recurrence in the pelvis treated with percutaneous cryoablation using protective techniques to avoid thermal injury to adjacent structures. In each case, cryo-ablation was performed successfully despite extensive previous surgical and radiation interventions. Salvage cryo-ablation resulted in a positive clinical and imaging response with an improvement in quality of life and absence of recurrence on follow-up imaging which continues to persist at the writing of this manuscript about 8 and 12-months following treatment.

Objectives: The objective of the study was to retrospectively investigate the safety and efficacy of computerized tomography-guided microwave ablation (MWA) in the treatment of Stage I non-small cell lung cancers (NSCLCs). Material and Methods: This retrospective, single-center study evaluated 21 patients (10 males and 11 females; mean age 73.8 ± 8.2 years) with Stage I peripheral NSCLCs treated with MWA between 2010 and 2020. All patients were surveyed for metastatic disease. Clinical success was defined as absence of FDG avidity on follow-up imaging. Tumor growth within 5 mm of the original ablated territory was defined as local recurrence. Welch t-test and Fisher’s exact test were used for univariate analysis. Hazard ratio (HR) and odds ratio (OR) were determined using Cox regression and Firth logistic regression. Significance was P < 0.05. Data are expressed as mean ± standard deviation. Results: Ablated tumors had longest dimension 17.4 ± 5.4 mm and depth 19.7 ± 15.1 mm from the pleural surface. Median follow-up was 20 months (range, 0.6–56 months). Mean overall survival (OS) following lung cancer diagnosis or MWA was 26.2 ± 15.4 months (range, 5–56 months) and 23.7 ± 15.1 months (range, 3–55 months). OS at 1, 2, and 5 years was 67.6%, 61.8%, and 45.7%, respectively. Progression-free survival (PFS) was 19.1 ± 16.2 months (range, 1–55 months). PFS at 1, 2, and 5 years was 44.5%, 32.9%, and 32.9%, respectively. Technical success was 100%, while clinical success was observed in 95.2% (20/21) of patients. One patient had local residual disease following MWA and was treated with chemotherapy. Local control was 90% with recurrence in two patients following ablation. Six patients (28.6%) experienced post-ablation complications, with pneumothorax being the most common event (23.8% of patients). Female gender was associated with 90% reduction in risk of death (HR 0.1, P = 0.014). Tumor longest dimension was associated with a 10% increase in risk of death (P = 0.197). Several comorbidities were associated with increased hazard. Univariate analysis revealed pre-ablation forced vital capacity trended higher among survivors (84.7 ± 15.2% vs. 73 ± 21.6%, P = 0.093). Adjusted for age and sex, adenocarcinoma, and neuroendocrine histology trended toward improved OS (OR: 0.13, 0.13) and PFS (OR: 0.88, 0.37) compared to squamous cell carcinoma. Conclusion: MWA provides a safe and effective alternative to stereotactic brachytherapy resulting in promising OS and PFS in patients with Stage I peripheral NSCLC. Larger sample sizes are needed to further define the effects of underlying comorbidities and tumor biology.

Objectives: Percutaneous renal biopsy (PRB) plays a critical role in the work-up of renal parenchymal disease. Although it is considered a low-risk procedure, additional interventions may be required in about 7% of the cases following biopsy. The purpose of this study was to identify risk factors for major hemorrhage by microscopic analysis of the cores obtained following PRB, with an intent to enhance the sensitivity and specificity of the risk stratification process, especially in patients undergoing this procedure in an outpatient setting. Material and Methods: A retrospective review identified 17 of 179 patients (9.50%) with major hemorrhage following PRB between July 2014 and June 2019. Using propensity score matching, 26 controls (without major hemorrhage) were matched to 17 cases (with major hemorrhage). The biopsy cores obtained from the cases and controls were analyzed by a single pathologist for medullary, cortical, total (medullary + cortical) lengths, and the number of arcuate arteries (AAs). Medullary:cortical (M:C), cortical:total (C:T), and medullary:total (M:T) length ratios were then calculated. Results: A stratified version of logistic regression was used to test for an association between each of the variables identified on the cores and the probability of a major hemorrhage. The analysis revealed that there was a statistically significant association between the number of AAs per specimen with the risk of major hemorrhage (P = 0.0006). When 0, 1, or >2 AAs were identified, the frequency of major hemorrhage was 13.04%, 66.67%, and 75.00%, respectively. The odds of major hemorrhage were 6 times higher with one AA and (95% CI, 1.28–32.30) and 15 times higher with >2 AAs (95% CI, 1.41–169.57). No significant association was found between medullary length (P = 0.228), medulla:cortex (M:C) (P = 0.089), medulla:total (M:T) (P = 0.108), or cortex:total (C:T) (P = 0.112) length ratios and major hemorrhage. Conclusion: There was a strong and incremental correlation between major renal hemorrhage following PRB and the number of AAs per core specimen. Identification of AAs by the pathologist, while assessing for sample adequacy, in the US suite can help predict major hemorrhage in patients undergoing PRBs.

Roux-en-Y gastric bypass (RYGB) is considered the gold standard for weight loss surgery and is an effective, safe treatment for morbid obesity and associated metabolic derangements. Complications such as small bowel obstruction are rare with a reported incidence of 5%. Obstruction caused by hiatal herniation of the gastric pouch and alimentary limb occurs even less frequently. Prompt recognition and treatment are imperative as delayed intervention may result in significant morbidity. At the time of this manuscript there have only been four reported cases in the literature highlighting a paucity of clinical guidance for the recognition and management of this complication. Here we present a case of acute small bowel obstruction secondary to hiatal herniation of the gastric pouch and proximal Roux limb. Furthermore, we review the literature and discuss the key aspects for the management of this complication.

Insulinoma is a rare neuroendocrine tumor. It may occur sporadically or as part of the genetic tumor syndrome multiple endocrine neoplasia type 1 (MEN1). Diagnosis is challenging because of the small size of insulin producing tumors that lead to hyperinsulinemia. Advances in imaging modalities may provide more accurate diagnosis of primary tumors, metastasis, and tumor functional status. Advances allow for improved medical and surgical management with new tools for research of neuroendocrine tumors. Surgical excision of the primary tumor is often curative; however, insulinomas in MEN1 syndrome are often multifocal with a high rate of recurrence presenting unique challenges in management. Here, we present the case of a 34-year-old male with recurrent hypoglycemic episodes and hyperparathyroidism diagnosed with multiple pancreatic insulinomas secondary to MEN1. Furthermore, we provide a brief review of the literature and discuss the approach to diagnosis and management in patients with MEN1 syndrome and future areas of investigation.

CRISPR editing of muscle stem cells (MuSCs) with adeno-associated virus serotype-9 (AAV9) holds promise for sustained gene repair therapy for muscular dystrophies. However, conflicting evidence exists on whether AAV9 transduces MuSCs. To rigorously address this question, we used a muscle graft model. The grafted muscle underwent complete necrosis before regenerating from its MuSCs. We injected AAV9.Cre into Ai14 mice. These mice express tdTomato upon Cre-mediated removal of a floxed stop codon. About 28%–47% and 24%–89% of Pax7(+) MuSCs expressed tdTomato in pre-grafts and regenerated grafts (p > 0.05), respectively, suggesting AAV9 efficiently transduced MuSCs, and AAV9-edited MuSCs renewed successfully. Robust MuSC transduction was further confirmed by delivering AAV9.Cre to Pax7-ZsGreen-Ai14 mice in which Pax7(+) MuSCs are genetically labeled by ZsGreen. Next, we co-injected AAV9.Cas9 and AAV9.gRNA to dystrophic mdx mice to repair the mutated dystrophin gene. CRISPR-treated and untreated muscles were grafted to immune-deficient, dystrophin-null NSG.mdx4cv mice. Grafts regenerated from CRISPR-treated muscle contained the edited genome and yielded 2.7-fold more dystrophin(+) cells (p = 0.015). Importantly, increased dystrophin expression was not due to enhanced formation of revertant fibers or de novo transduction by residual CRISPR vectors in the graft. We conclude that AAV9 effectively transduces MuSCs. AAV9 CRISPR editing of MuSCs may provide enduring therapy.

Recombinant adeno-associated virus (AAV)-based gene delivery is a promising approach to treat muscle diseases. However, body-wide muscle delivery and pre-existing immune responses pose significant challenges to AAV muscle gene therapy. While the determinants of tissue tropism and immunogenicity of AAV are amenable to traditional molecular engineering, the development of a muscle-specific, immunosilent AAV vector has remained elusive. Recent advances in understanding the relationship between capsid structural motifs and functional domains have created exciting developments in the search for a muscle-specific AAV. Novel approaches to generate unique AAV properties through forced evolution have resulted in capsids with improved immune properties and/or muscle-targeting efficiency. Optimization of the gene cassette to restrict expression to mature muscle fibers provides another level of control. These reengineered AAV vectors have the potential to greatly increase efficacy and reduce off-target effects for body-wide muscle gene therapy. In this chapter, we discuss recent advances in the development of a next-generation, muscle-specific AAV vector.

We studied time-dependent changes in muscle optical properties during degeneration and regeneration using polarization-sensitive optical coherence tomography (PSOCT). Excised canine muscle transplants in a xenograft mouse model were imaged ex vivo from 3- to 112-day post-transplantation. PSOCT images were quantified to evaluate post-transplantation changes of three optical/structural properties: attenuation, birefringence and fiber alignment. The birefringence and fiber alignment decreased after transplantation until 20∼30-day and recovered thereafter. The attenuation coefficient showed a reversed trend over the same period of time. These results suggest that optical properties could be used for monitoring skeletal muscle degeneration and regeneration.

Duchenne muscular dystrophy (DMD) is a X-linked, progressive childhood myopathy caused by mutations in the dystrophin gene, one of the largest genes in the genome. It is characterized by skeletal and cardiac muscle degeneration and dysfunction leading to cardiac and/or respiratory failure. Adeno-associated virus (AAV) is a highly promising gene therapy vector. AAV gene therapy has resulted in unprecedented clinical success for treating several inherited diseases. However, AAV gene therapy for DMD remains a significant challenge. Hurdles for AAV-mediated DMD gene therapy include the difficulty to package the full-length dystrophin coding sequence in an AAV vector, the necessity for whole-body gene delivery, the immune response to dystrophin and AAV capsid, and the species-specific barriers to translate from animal models to human patients. Capsid engineering aims at improving viral vector properties by rational design and/or forced evolution. In this review, we discuss how to use the state-of-the-art AAV capsid engineering technologies to overcome hurdles in AAV-based DMD gene therapy.

The Frank-Starling mechanism, whereby increased diastolic filling leads to increased cardiac output, depends on increasing the sarcomere length ( Ls) of cardiomyocytes. Ventricular stiffness increases with advancing age, yet it remains unclear how such changes in compliance impact sarcomere dynamics in the intact heart. We developed an isolated murine heart preparation to monitor Ls as a function of left ventricular pressure and tested the hypothesis that sarcomere lengthening in response to ventricular filling is impaired with advanced age. Mouse hearts isolated from young (3–6 mo) and aged (24–28 mo) C57BL/6 mice were perfused via the aorta under Ca2+-free conditions with the left ventricle cannulated to control filling pressure. Two-photon imaging of 4-{2-[6-(dioctylamino)-2-naphthalenyl]ethenyl}1-(3-sulfopropyl)-pyridinium fluorescence was used to monitor t-tubule striations and obtain passive Ls between pressures of 0 and 40 mmHg. Ls values (in μm, aged vs. young, respectively) were 2.02 ± 0.04 versus 2.01 ± 0.02 at 0 mmHg, 2.13 ± 0.04 versus 2.23 ± 0.02 at 5 mmHg, 2.21 ± 0.03 versus 2.27 ± 0.03 at 10 mmHg, and 2.28 ± 0.02 versus 2.36 ± 0.01 at 40 mmHg, indicative of impaired sarcomere lengthening in aged hearts. Atomic force microscopy nanoindentation revealed that intact cardiomyocytes enzymatically isolated from aged hearts had increased stiffness compared with those of young hearts (elastic modulus: aged, 41.9 ± 5.8 kPa vs. young, 18.6 ± 3.3 kPa; P = 0.006). Impaired sarcomere lengthening during left ventricular filling may contribute to cardiac dysfunction with advancing age by attenuating the Frank-Starling mechanism and reducing stroke volume.

Gene therapy is on the verge of producing viable treatments for numerous diseases. Millions of years of viral evolution has made virus an ideal system to deliver foreign genes to host cells. Here we discuss the most common viral vectors used in gene therapy such as retrovirus, lentivirus, adenovirus, adeno-associated virus, herpes virus, and poxvirus. For each viral vector, we review the virus biology, viral vector development and relevant clinical applications.

Duchenne muscular dystrophy (DMD) is a lethal X-linked childhood muscle wasting disease caused by mutations in the dystrophin gene. Nanobiotechnology-based therapies (such as synthetic nanoparticles and naturally existing viral and nonviral nanoparticles) hold great promise to replace and repair the mutated dystrophin gene and significantly change the disease course. While a majority of DMD nanotherapies are still in early preclinical development, several [such as adeno-associated virus (AAV)-mediated systemic micro-dystrophin gene therapy] are advancing for phase I clinical trials. Recent regulatory approval of Ataluren (a nonsense mutation read-through chemical) in Europe and Exondys51 (an exon-skipping antisense oligonucleotide drug) in the United States shall offer critical insight in how to move DMD nanotherapy to human patients. Progress in novel, optimized nano-delivery systems may further improve emerging molecular therapeutic modalities for DMD. Despite these progresses, DMD nanotherapy faces a number of unique challenges. Specifically, the dystrophin gene is one of the largest genes in the genome while nanoparticles have an inherent size limitation per definition. Furthermore, muscle is the largest tissue in the body and accounts for 40% of the body mass. How to achieve efficient bodywide muscle targeting in human patients with nanomedication remains a significant translational hurdle. New creative approaches in the design of the miniature micro-dystrophin gene, engineering of muscle-specific synthetic AAV capsids, and novel nanoparticle-mediated exon-skipping are likely to result in major breakthroughs in DMD therapy. WIREs Nanomed Nanobiotechnol 2018, 10:e1472. doi: 10.1002/wnan.1472 This article is categorized under: Biology-Inspired Nanomaterials > Protein and Virus-Based Structures Therapeutic Approaches and Drug Discovery > Emerging Technologies.

The Frank-Starling mechanism, whereby increased cardiac filling leads to increased cardiac output, is dependent on cardiomyocyte sarcomere length (Ls). The relationship between Ls and left-ventricular pressure (LVP) in living (non-fixed) myocardium is poorly defined. We examined the Ls versus LVP relationship in isolated perfused mouse hearts using 2-photon fluorescence microscopy. To determine a fluorescent indicator of Ls, isolated cardiomyocytes were labeled with Di-8-ANEPPS (membrane dye labeling t-tubules) and MitoTracker Deep Red (mitochondrial stain) and imaged using confocal microscopy. The resulting fluorescence signals were correlated to transmitted light Ls obtained under resting conditions and in the presence of 10 mM caffeine to induce cellular contracture. Fast-Fourier Transform and linear regression analyses illustrated that Di-8-ANEPPS (r2=0.97) and MitoTracker (r2=0.95) were both well-correlated to transmitted light Ls. Perfused hearts were loaded with Di-8-ANEPPS with the left-ventricle cannulated (via the left atrium) to alter LVP. Imaging was performed on an inverted microscope with the left-ventricular wall against the coverglass, allowing sub-epicardial cardiomyocytes to be imaged while their longitudinal axis was parallel to the focal plane. Measurements were performed under passive conditions (Ca-free, 25°C) with aortic pressure 1 mmHg higher than LVP at each pressure examined. Sarcomere length (μm) increased with LVP, with values of 2.02±0.01 at 0 mmHg, 2.24±0.02 at 10 mmHg, 2.29±0.02 at 20 mmHg, and 2.33±0.02 at 40 mmHg. These values are consistent with data in mammalian hearts fixed at varying LVP, and illustrate that Ls is more dynamic over lower pressure ranges (0-20 mm Hg) than higher pressure ranges (20-40 mm Hg). We conclude that the Ls-LVP relationship can be determined accurately in the living heart using 2-photon fluorescence microscopy, which should prove valuable for investigating disease-associated changes in passive properties of the heart.