Bronchoscopy-Guided High-Power Microwave Ablation in an in vivo Porcine Lung Model.

Introduction: Percutaneous microwave ablation (MWA) is clinically accepted for the treatment of lung tumors and oligometastatic disease. Bronchoscopic MWA is under development and evaluation in the clinical setting. We previously reported on the development of a bronchoscopy-guided MWA system integrated with clinical virtual bronchoscopy and navigation and demonstrated the feasibility of transbronchial MWA, using a maximum power of 60 W at the catheter input. Here, we assessed the performance of bronchoscopy-guided MWA with an improved catheter (maximum power handling of up to 120 W) in normal porcine lung in vivo (as in the previous study). Methods: A total of 8 bronchoscopy-guided MWA were performed (n = 2 pigs; 4 ablations per pig) with power levels of 90 W and 120 W applied for 5 and 10 min, respectively. Virtual bronchoscopy planning and navigation guided transbronchial or endobronchial positioning of the MWA applicator for ablation of lung parenchyma. Following completion of ablations and post-procedure CT imaging, the lungs were harvested and sectioned for gross and histopathologic ablation analysis. Results: Bronchoscopy-guided MWA with applied energy levels of 90 W/5 min and 120 W/10 min yielded ablation zones with short-axis diameters in the range of 20-28 mm (56-116% increase) as compared to ∼13 mm from our previous study (60 W/10 min). Histology of higher-power and previous lower-power ablations was consistent, including a central necrotic zone, a thermal fixation zone with intact tissue architecture, and a hemorrhagic periphery. Catheter positioning and its confirmation via intra-procedural 3D imaging (e.g., cone-beam CT) proved to be critical for ablation consistency. Conclusion: Bronchoscopy-guided MWA with an improved catheter designed for maximum power 120 W yields large ablations in normal porcine lung in vivo.

Characterization of a Biochemical Mouse Model of Primary Aldosteronism for Thermal Therapies.

Introduction: Aldosterone-producing adenoma (APA) is the most common cause of endocrine-related hypertension but surgery is not always feasible. Current medical interventions are associated with significant side effects and poor patient compliance. New APA animal models that replicate basic characteristics of APA and give physical and biochemical feedback are needed to test new non-surgical treatment methods, such as image-guided thermal ablation. Methods: A model of APA was developed in nude mice using HAC15 cells, a human adrenal carcinoma cell line. Tumor growth, aldosterone production, and sensitivity to angiotensin II were characterized in the model. The utility of the model was validated via treatment with microwave ablation and characterization of the resulting physical and biochemical changes in the tumor. Results: The APA model showed rapid and relatively homogeneous growth. The tumors produced aldosterone and steroid precursors in response to angiotensin II challenge, and plasma aldosterone levels were significantly higher in tumor bearing mice two hours after challenge verses non-tumor bearing mice. The model was useful for testing microwave ablation therapy, reducing aldosterone production by 80% in treated mice. Conclusion: The HAC15 model is a useful tumor model to study and develop localized treatment methods for APA.

Directional microwave ablation in spine: experimental assessment of computational modeling.

BACKGROUND: Despite the theoretical advantages of treating metastatic bone disease with microwave ablation (MWA), there are few reports characterizing microwave absorption and bioheat transfer in bone. This report describes a computational modeling-based approach to simulate directional microwave ablation (dMWA) in spine, supported by ex vivo and pilot in vivo experiments in porcine vertebral bodies. MATERIALS AND METHODS: A 3D computational model of microwave ablation within porcine vertebral bodies was developed. Ex vivo porcine vertebra experiments using a dMWA applicator measured temperatures approximately 10.1 mm radially from the applicator in the direction of MW radiation (T1) and approximately 2.4 mm in the contra-lateral direction (T2). Histologic assessment of ablated ex vivo tissue was conducted and experimental results compared to simulations. Pilot in vivo experiments in porcine vertebral bodies assessed ablation zones histologically and with CT and MRI. RESULTS: Experimental T1 and T2 temperatures were within 3-7% and 11-33% of simulated temperature values. Visible ablation zones, as indicated by grayed tissue, were smaller than those typical in other soft tissues. Posthumous MRI images of in vivo ablations showed hyperintensity. In vivo experiments illustrated the technical feasibility of creating directional microwave ablation zones in porcine vertebral body. CONCLUSION: Computational models and experimental studies illustrate the feasibility of controlled dMWA in bone tissue.

PULMONARY AND COELOMIC MYCOSES DUE TO METARHIZIUM AND BEAUVERIA SPECIES IN REPTILES.

This report documents cases of fatal pulmonary mycosis caused by entomopathogenic fungi in the genera Metarhizium and Beauveria (Order Hypocreales) in a loggerhead sea turtle (Caretta caretta), a Chinese alligator (Alligator sinensis), two gopher tortoises (Gopherus polyphemus), a Cuvier's dwarf caiman (Paleosuchus palpebrosus), a false gharial (Tomistoma schlegelii), a green sea turtle (Chelonia mydas), and a Kemp's ridley sea turtle (Lepidochelys kempii), and a case of granulomatous coelomitis in a hawksbill sea turtle (Eretmochelys imbricata). Fungi identified in these cases included Beauveria bassiana, Beauveria brongniartii, Metarhizium anisopliae, Metarhizium robertsii, and one case of infection by a novel Metarhizium species. The animals were either housed at zoos or brought into rehabilitation from the wild. Although the majority of animals had comorbidities, the fungal infections were believed to be the primary cause of death. Fungal susceptibility testing was performed on two Beauveria spp. isolates, and revealed lower minimum inhibitory concentrations for itraconazole and voriconazole when compared to terbinafine and fluconazole. This case series demonstrates that a variety of reptile species from different orders are vulnerable to infection with Metarhizium, and multiple species of sea turtle are susceptible to infection with Beauveria.

Injectable Peptide Hydrogel Encapsulation of Mesenchymal Stem Cells Improved Viability, Stemness, Anti-Inflammatory Effects, and Early Stage Wound Healing.

Human-adipose-derived mesenchymal stem cells (hADMSCs) are adult stem cells and are relatively easy to access compared to other sources of mesenchymal stem cells (MSCs). They have shown immunomodulation properties as well as effects in improving tissue regeneration. To better stimulate and preserve the therapeutic properties of hADMSCs, biomaterials for cell delivery have been studied extensively. To date, hyaluronic acid (HA)-based materials have been most widely adopted by researchers around the world. PGmatrix is a new peptide-based hydrogel that has shown superior functional properties in 3D cell cultures. Here, we reported the in vitro and in vivo functional effects of PGmatrix on hADMSCs in comparison with HA and HA-based Hystem hydrogels. Our results showed that PGmatrix was far superior in maintaining hADMSC viability during prolonged incubation and stimulated expression of SSEA4 (stage-specific embryonic antigen-4) in hADMSCs. hADMSCs encapsulated in PGmatrix secreted more immune-responsive proteins than those in HA or Hystem, though similar VEGF-A and TGFß1 release levels were observed in all three hydrogels. In vivo studies revealed that hADMSCs encapsulated with PGmatrix showed improved skin wound healing in diabetic-induced mice at an early stage, suggesting possible anti-inflammatory effects, though similar re-epithelialization and collagen density were observed among PGmatrix and HA or Hystem hydrogels by day 21.

Pax5 and CD3 immunophenotyping of lymphoma in 2 central bearded dragons.

Two central bearded dragons (Pogona vitticeps), a 3-y-old male and a 5-y-old female, were diagnosed with different manifestations of lymphoma at the Kansas State Veterinary Diagnostic Laboratory between 2019 and 2020. The 3-y-old male was presented for postmortem evaluation and was in poor body condition. Microscopically, nearly all examined organs contained variable numbers of neoplastic round cells. Neoplastic cells in the stomach and liver had moderate immunoreactivity to CD3 consistent with multicentric T-cell lymphoma, and non-neoplastic lymphocytes infiltrating the stomach mass had strong immunoreactivity to Pax5. The 5-y-old female had an ulcerated oral mass located in the right lingual gingiva submitted as an excisional biopsy. Microscopically, the mass was composed of large numbers of neoplastic round cells in the epithelium and connective tissue that were strongly and diffusely positive for CD3 and frequently positive for Pax5, consistent with a dual-positive, localized, epitheliotropic T-cell lymphoma. Neoplastic and non-neoplastic lymphocytes did not stain with CD20 or CD79a. Neoplasms are increasingly reported as a cause of morbidity and mortality in reptiles. Our 2 cases illustrate various presentations of T-cell lymphoma and the effectiveness of CD3 and Pax5 immunohistochemistry in bearded dragons.

Domestic sheep and bighorn sheep carry distinct gammaherpesviruses belonging to the genus Macavirus.

The genus Macavirus of the subfamily Gammaherpesvirinae comprises two genetically distinct lineages of lymphotropic viruses. One of these lineages includes viruses that can cause malignant catarrhal fever (MCF), which are known as MCF viruses (MCFV). All MCFVs are genetically and antigenically related but carried by different hosts. In this study, we report the recognition of new MCFV carried by bighorn sheep. The virus was first identified in a bighorn sheep from Banff National Park, Alberta, Canada. Analysis of a conserved region of the viral DNA polymerase gene of the virus carried by this bighorn sheep showed 85.88% nucleotide identity to the MCFV carried by domestic sheep, ovine herpesvirus 2 (OvHV-2). Further investigation of bighorn samples obtained from animals in the US and Canada showed 98.87-100% identity to the DNA polymerase sequence of the first bighorn in the study. Phylogenetic analysis indicated that the MCFV carried by bighorn sheep is closely related but distinct from OvHV-2. Epidemiological and virulence features of the newly recognized MCFV are still unknown and warrant further investigation. Considering the current nomenclature for MCFVs, we suggest a tentative designation of ovine herpesvirus-3 (OvHV-3) for this newly identified bighorn sheep MCFV.

High copy number of ovine gammaherpesvirus 2 DNA associated with malignant catarrhal fever-like syndrome in a lamb.

Domestic and wild sheep are the natural reservoirs for ovine gammaherpesvirus 2 (OvHV-2), the causative agent of sheep-associated malignant catarrhal fever (SA-MCF). Virtually all adult sheep are infected with OvHV-2 under natural flock conditions, and infection is normally subclinical. MCF-like clinical signs and typical histologic lesions in sheep have been linked during case investigations at veterinary diagnostic laboratories; however, the confirmation of naturally occurring MCF in sheep is problematic. To date, the assays for detection of OvHV-2-specific antibodies or DNA are usually positive in sheep, regardless of health status, so mere detection of antibodies or the agent is of minimal diagnostic significance in this species. We document herein a naturally occurring MCF case in a 4-mo-old domestic lamb and demonstrate that the affected animal had 100-1,000 times more OvHV-2 copy numbers in tissues than in healthy adult and age-matched sheep. These results indicate that high copy numbers of viral DNA in tissues associated with characteristic lesions can be used to confirm the diagnosis of MCF in sheep.

Experimental induction of malignant catarrhal fever in pigs with ovine herpesvirus 2 by intranasal nebulization.

Malignant catarrhal fever (MCF), a frequently fatal herpesviral disease primarily of ruminant species, has been sporadically reported in pigs. All cases of naturally occurring porcine MCF reported to date have been linked to ovine herpesvirus 2 (OvHV-2), a gammaherpesvirus in the genus Macavirus carried by sheep. Experimental induction of MCF by aerosolization of the virus in nasal secretions collected from infected sheep has been successful in bison, cattle and rabbits. The goals of this study were to determine the susceptibility of pigs to MCF following experimental intranasal inoculation of OvHV-2, and to characterize the disease. Twelve pigs in four groups were nebulized with 10(5), 10(6), 10(7), or 10(8) DNA copies of OvHV-2 from sheep nasal secretions. Three control pigs were nebulized with nasal secretions from uninfected sheep. Three additional pigs were inoculated intravenously with 10(7) DNA copies of OvHV-2 to evaluate this route of infection with cell-free virus. Seven of twelve intranasally challenged pigs became infected with OvHV-2. Five of these seven, all in higher dose groups, developed MCF. Lesions resembled those reported in natural cases of porcine MCF. The most striking and consistent histological lesions were in trachea, lung, kidney and brain. These comprised mucopurulent tracheitis, interstitial pneumonia, necrotizing arteritis-periarteritis, and nonpurulent meningoencephalitis. No infection was established in the intravenously challenged or control groups. The study showed that MCF can be experimentally induced in pigs by aerosol challenge using sheep nasal secretions containing OvHV-2. Domestic pigs are a natural clinically susceptible host for sheep-associated MCF. They represent a useful, cost-effective model for MCF research.

Histologic and molecular identification of disseminated Histoplasma capsulatum in a captive brown bear (Ursus arctos).

A 33-year-old brown bear (Ursus arctos) was evaluated for chronic cough, partial anorexia, and lethargy in early fall of 2009. Radiographs revealed a generalized increase in interstitial density with focal lung field consolidation and air bronchograms more prevalent in the cranial lung lobes. Tracheal sputum and wash fluid grew mixed bacteria and 2 species of Candida on bacterial and fungal cultures, respectively. Serum was negative for antibodies to Aspergillus, Blastomyces, Coccidioides, and Histoplasma by semiquantitative radial immunodiffusion. Antimicrobial and antifungal treatment was administered. The bear died 1 month after entering hibernation. Gross necropsy revealed coalescent nodules and sheets of firm tan tissue covering pleural surfaces of the thoracic cavity and within pulmonary parenchyma, enlarged mesenteric lymph nodes, and intestinal ulcerations. Histopathology revealed granulomatous inflammation with intrahistiocytic yeast, consistent with Histoplasma organisms, in lung, diaphragm, mesenteric lymph nodes, intestine, and adrenal glands. Molecular analysis performed on DNA isolated from lung tissue, including conventional polymerase chain reaction (PCR) targeting the internal transcribed spacer region for the ribosomal RNA gene complex and real-time PCR targeting the gene encoding a unique region of M specific protein, identified the organism to be 100% identical to Histoplasma capsulatum with an average of 4.9 × 10(7) gene copies per gram of tissue. The present report describes histologic and molecular techniques for diagnosing histoplasmosis.

Inhibition of mammary tumor growth by a novel nontoxic retinoid: chemotherapeutic evaluation of a C-linked analog of 4-HPR-glucuronide.

Previous studies from our laboratory suggest that 4-HPROG, the O-glucuronide derivative of 4-HPR, has improved mammary cancer chemopreventive/ antitumor activities as well as reduced toxicity, as compared to 4-HPR. This O-linked glucuronide derivative is a substrate to the P-glucuronidase enzyme and may also undergo hydrolysis in vivo to the vitamin A metabolite, retinoic acid, that is toxic at high concentrations. In an effort to improve analog potency relative to its toxicity, the 4-HPROG's phenolic oxygen was replaced with a methylene group, thus preventing biological cleavage of the glucuronide moiety. The resulting C-linked analog, 4-HPR-C-glucuronide (4-HPRCG), cannot be hydrolyzed to 4-HPR. The results of this study show that 4-HPRCG is an effective chemotherapeutic agent that caused 49% regression of DMBA-induced mammary tumors in rats, while showing almost no side-effects that are often observed with other natural or synthetic retinoids, such as a reduction in blood retinol level, elevation in blood triglyceride (TG) level, and decrease in bone mineral content (BMC). These results suggest that 4-HPRCG should be considered as a better candidate for breast cancer treatment.

Hydrolysis of 4-HPR to atRA occurs in vivo but is not required for retinamide-induced apoptosis.

The retinamide, N-(4-hydroxyphenyl)retinamide (4-HPR), has shown promising anti-tumor activity, but it is unclear whether this compound is hydrolyzed to all-trans retinoic acid (atRA) and if so, whether this plays any role in its chemotherapeutic activity. To address this issue, the ability of 4-hydroxybenzylretinone (4-HBR), a carbon-linked analog of 4-HPR, to support growth in vitamin A-deficient (VAD) animals and to activate an atRA-responsive gene in vivo was compared to 4-HPR and atRA. Further, the non-hydrolyzable 4-HBR analog was used to determine whether the presence of the labile amide linkage in 4-HPR is essential for the induction of apoptosis in cultured MCF-7 breast cancer cells. Studies in VAD rats showed that 4-HPR, like atRA, supports animal growth and induces CYP26B1 mRNA expression in lung whereas 4-HBR does not. Analysis of plasma from 4-HPR- and atRA-treated VAD animals revealed the presence of atRA whereas it was not detected in plasma from animals given 4-HBR. To determine whether hydrolysis to atRA is necessary for apoptosis induced by 4-HPR in MCF-7 breast cancer cells, morphological and biochemical assays for apoptosis were performed. 4-HBR, like 4-HPR, induced apoptosis in MCF-7 cells. Apoptosis was not induced even at high concentrations of atRA, showing that 4-HPR and 4-HBR act in cells via a distinct signaling pathway. These results show that although limited hydrolysis of 4-HPR occurs in vivo, the ability to liberate atRA is not required for these 4-hydroxyphenyl retinoids to induce apoptosis in MCF-7 breast cancer cells. Thus the non-hydrolyzable analog, 4-HBR, may have significant therapeutic advantage over 4-HPR because it does not liberate atRA that can contribute to the adverse side effects of drug administration in vivo.