In vivo assessment of changes to canine airway smooth muscle following bronchial thermoplasty with OR-OCT.

The inability to assess and measure changes to the airway smooth muscle (ASM) in vivo is a major challenge to evaluating asthma and its clinical outcomes. Bronchial thermoplasty (BT) is a therapy for asthma that aims to reduce the severity of excessive bronchoconstriction by ablating ASM. Although multiple long-term clinical studies of BT have produced encouraging results, the outcomes of BT treatment in practice have been variable, and questions remain regarding the selection of patients. Previously, we have demonstrated an imaging platform called orientation-resolved optical coherence tomography that can assess ASM endoscopically using an imaging catheter compatible with bronchoscopy. In this work, we present results obtained from a longitudinal BT study performed using a canine model (n = 8) and with the goal of investigating the use of orientation-resolved optical coherence tomography (OR-OCT) for measuring the effects of BT on ASM. We demonstrate that we are capable of accurately assessing ASM both before and in the weeks following the BT procedure using blinded matching to histological samples stained with Masson's trichrome (P < 0.0001, r2 = 0.79). Analysis of volumetric ASM distributions revealed significant decreases in ASM in treated airways (average cross-sectional ASM area: 0.245 ± 0.145 mm2 pre-BT and 0.166 ± 0.112 mm2 6 wk following BT). These results demonstrate that OR-OCT can provide clinicians with the feedback necessary to better evaluate ASM and its response to BT, and may potentially play an important role in phenotyping asthma and predicting which patients are most likely to respond to BT treatment.NEW & NOTEWORTHY The inability to assess ASM in vivo is a significant hurdle in advancing our understanding of airway diseases such as asthma, as well as evaluating potential treatments and therapies. In this study, we demonstrate that endoscopic OR-OCT can be used to accurately measure changes to ASM structure following BT. Our results demonstrate how this technology could occupy an important role in asthma treatments targeting ASM.

In vivo transplantation of autogenous marrow-derived cells following rapid intraoperative magnetic separation based on hyaluronan to augment bone regeneration.

INTRODUCTION: This project was designed to test the hypothesis that rapid intraoperative processing of bone marrow based on hyaluronan (HA) could be used to improve the outcome of local bone regeneration if the concentration and prevalence of marrow-derived connective tissue progenitors (CTPs) could be increased and nonprogenitors depleted before implantation. METHODS: HA was used as a marker for positive selection of marrow-derived CTPs using magnetic separation (MS) to obtain a population of HA-positive cells with an increased CTP prevalence. Mineralized cancellous allograft (MCA) was used as an osteoconductive carrier scaffold for loading of HA-positive cells. The canine femoral multidefect model was used and four cylindrical defects measuring 10 mm in diameter and 15 mm in length were grafted with MCA combined with unprocessed marrow or with MS processed marrow that was enriched in HA(+) CTPs and depleted in red blood cells and nonprogenitors. Outcome was assessed at 4 weeks using quantitative 3D microcomputed tomography (micro-CT) analysis of bone formation and histomorphological assessment. RESULTS: Histomorphological assessment showed a significant increase in new bone formation and in the vascular sinus area in the MS-processed defects. Robust bone formation was found throughout the defect area in both groups (defects grafted with unprocessed marrow or with MS processed marrow.) Percent bone volume in the defects, as assessed by micro-CT, was greater in defects engrafted with MS processed cells, but the difference was not statistically significant. CONCLUSION: Rapid intraoperative MS processing to enrich CTPs based on HA as a surface marker can be used to increase the concentration and prevalence of CTPs. MCA grafts supplemented with heparinized bone marrow or MS processed cells resulted in a robust and advanced stage of bone regeneration at 4 weeks. A greater new bone formation and vascular sinus area was found in defects grafted with MS processed cells. These data suggest that MS processing may be used to enhance the performance of marrow-derived CTPs in clinical bone regeneration procedures. Further assessment in a more stringent bone defect model is proposed.

Spatial and temporal dynamics of in vitro photodynamic cell killing: extracellular hydrogen peroxide mediates neighbouring cell death.

Photodynamic killing of a cell population is generally considered to result from direct effects that occur in each cell. In some scenarios this may be an over-simplification and the potential for cell-cell signaling processes to contribute to the response of a population to photodynamic stress is addressed in this paper. Photodynamic killing of EMT6 cells in culture was studied in time and space using computerized time-lapse microscopy. The rate of cell killing was dependent on the fluence with both rapid and slower processes evident, the proportion of the former increasing with fluence. The spatial distribution of cell death was non-random and for the slow cell killing process was found to occur preferentially in the vicinity of dead or dying cells, suggesting a local signaling process. An inhibitory effect of extracellular catalase indicated the involvement of hydrogen peroxide in the spread of cell death and NADPH oxidase was determined as the principal source of hydrogen peroxide. This cell signaling pathway was observed for membrane-bound and mitochondrial photosensitizers but not for a nuclear photosensitizer. These secondary cell signalling pathways extend the oxidative damage to cells in space and time.