Development of a reproducible porcine model of infected burn wounds.

Severe burns are traumatic and physically debilitating injuries with a high rate of mortality. Bacterial infections often complicate burn injuries, which presents unique challenges for wound management and improved patient outcomes. Currently, pigs are used as the gold standard of pre-clinical models to study infected skin wounds due to the similarity between porcine and human skin in terms of structure and immunological response. However, utilizing this large animal model for wound infection studies can be technically challenging and create issues with data reproducibility. We present a detailed protocol for a porcine model of infected burn wounds based on our experience in creating and evaluating full thickness burn wounds infected with Staphylococcus aureus on six pigs. Wound healing kinetics and bacterial clearance were measured over a period of 27 d in this model. Enumerated are steps to achieve standardized wound creation, bacterial inoculation, and dressing techniques. Systematic evaluation of wound healing and bacterial colonization of the wound bed is also described. Finally, advice on animal housing considerations, efficient bacterial plating procedures, and overcoming common technical challenges is provided. This protocol aims to provide investigators with a step-by-step guide to execute a technically challenging porcine wound infection model in a reproducible manner. Accordingly, this would allow for the design and evaluation of more effective burn infection therapies leading to better strategies for patient care.

Lumbar muscle atrophy caused by harness replacement in a chronic calf model of total artificial heart implantation.

The postoperative care of animals implanted with mechanical circulatory support devices is complex. The standard of care requires continuous monitoring of hemodynamic parameters post implant, wound care, and maintenance of the animal's well-being, but also includes controlling the animal's biomechanics under conditions of continuous restraint and harnessing. In such studies, a harness provides secure fixation of the exteriorized device driveline and pressure lines and aids animal handling (lifting, position adjustment, and assistance with standing up). Harnessing is a key element in large-animal surgery. It affects the animal's conditions, safety, and post-procedure troubleshooting and thus may drastically worsen postoperative outcomes if improperly handled. Here we report a case associated with an unplanned harness replacement in a chronic animal model implanted with the Cleveland Clinic continuous-flow total artificial heart. Inadvertent changes to the harness resulted in posture change caused by muscular atrophy of the calf's spine that had been under long-term harness support.

Median Sternotomy or Right Thoracotomy Techniques for Total Artificial Heart Implantation in Calves.

The choice of optimal operative access technique for mechanical circulatory support device implantation ensures successful postoperative outcomes. In this study, we retrospectively evaluated the median sternotomy and lateral thoracotomy incisions for placement of the Cleveland Clinic continuous-flow total artificial heart (CFTAH) in a bovine model. The CFTAH was implanted in 17 calves (Jersey calves; weight range, 77.0-93.9 kg) through a median sternotomy (n = 9) or right thoracotomy (n = 8) for elective chronic implantation periods of 14, 30, or 90 days. Similar preoperative preparation, surgical techniques, and postoperative care were employed. Implantation of the CFTAH was successfully performed in all cases. Both methods provided excellent surgical field visualization. After device connection, however, the median sternotomy approach provided better visualization of the anastomoses and surgical lines for hemostasis confirmation and repair due to easier device displacement, which is severely limited following right thoracotomy. All four animals sacrificed after completion of the planned durations (up to 90 days) were operated through full median sternotomy. Our data demonstrate that both approaches provide excellent initial field visualization. Full median sternotomy provides larger viewing angles at the anastomotic suture line after device connection to inflow and outflow ports.

Anatomy of the bovine ascending aorta and brachiocephalic artery found unfavorable for total artificial heart implant.

The biocompatibility assessment of the Cleveland Clinic continuous-flow total artificial heart is an important part of the device developmental program. Surgical and postoperative management are key factors in achieving optimal outcomes. However, the presence of vascular anatomical abnormalities in experimental animal models is often unpredictable and may worsen the expected outcomes. We report a technical impediment encountered during total artificial heart implantation complicated by unfavorable bovine anatomy of the ascending aorta and brachiocephalic arterial trunk.

Environmental monitoring for Aspergillus fumigatus in association with an immunosuppressed rabbit model of pulmonary aspergillosis.

Aspergillus fumigatus causes life-threatening pneumonia in immunocompromised patients. Conidia, the infectious form of the organism, are handled in a biologic safety cabinet under BSL2 conditions. However because germinated conidia form noninfectious hyphae in tissue, we hypothesized that rabbits inoculated intratracheally would grow A. fumigatus in their lungs but that the environment would remain free of this fungus, potentially permitting maintenance of infected animals under ABSL1 conditions. We performed a surveillance study for the presence of A. fumigatus in the environment before proceeding with antifungal therapy studies of experimental pulmonary aspergillosis. The expected outcome included absence of A. fumigatus in the environment, stool, and blood and presence in rabbit lungs. Female SPF New Zealand white rabbits were immunosuppressed and inoculated intratracheally (n = 4) or intraesophageally (n = 2) with 1.25 × 10(8) conidia of A. fumigatus. Feces, pan liners, and walls were sampled daily during the 11-d experiment, and blood was sampled on days 2, 6, and 8 after inoculation. Samples were cultured on 5% Sabouraud glucose agar plates. Lungs were weighed and scored for hemorrhagic infarcts and homogenized for culture on 5% Sabouraud glucose agar and trypticase soy agar plates. Blood cultures, rabbit stool, and environmental swabs were all negative for A. fumigatus whereas the lungs inoculated intratracheally demonstrated 4.5 × 10(2) ± 0.8 × 10(2) CFU/g of A. fumigatus. Therefore, neutropenic rabbits with experimental invasive pulmonary aspergillosis do not shed conidia of A. fumigatus and can be safely housed under ABSL1 conditions after inoculation.

Inhibitors of gamma-secretase block in vivo and in vitro T helper type 1 polarization by preventing Notch upregulation of Tbx21.

Notch receptors are processed by gamma-secretase acting in synergy with T cell receptor signaling to sustain peripheral T cell activation. Activated CD4+ T cells differentiate into T helper type 1 (TH1) or TH2 subsets. Molecular cues directing TH1 differentiation include expression of the TH1-specific transcription factor T-bet, encoded by Tbx21. However, the regulation of Tbx21 remains incompletely defined. Here we report that Notch1 can directly regulate Tbx21 through complexes formed on the Tbx21 promoter. In vitro, gamma-secretase inhibitors extinguished expression of Notch, interferon-gamma and Tbx21 in TH1-polarized CD4+ cells, whereas ectopic expression of activated Notch1 restored Tbx21 transcription. In vivo, administration of gamma-secretase inhibitors substantially impeded TH1-mediated disease progression in the mouse experimental autoimmune encephalomyelitis model of multiple sclerosis. Thus, using gamma-secretase inhibitors to modulate Notch signaling may prove beneficial in treating TH1-mediated autoimmunity.