Page 120 - Effects of radiotherapy and hyperbaric oxygen therapy on oral microcirculation Renee Helmers

Chapter 6
Data analysis
The CCTools software was used to export image recordings in .avi file format. The
following parameters were analyzed form the exported clips: Fd, classification
of angioarchitecture, total and functional capillary density (TCD and FCD
respectively), microvascular flow index (MFI), and buccal blood vessel diameter
(Ø ).Angioarchitecturewasdividedinto3classtypes38:anarrayofcapillaryloops bv
(score 1), both capillary loops and vascular network (score 2) and vascular network without any loops (score 3). Frame selection for capillary density assessment was based on clarity, resolution and the absence of pressure artifacts.2 To determine TCD and FCD the capillary loops in each frame (area of 1.80 mm2) were counted. At completion of the density dataset the results were divided by 1.80 to convert to mean number of capillaries per millimeter squared (cpll/mm2). The type of flow per quadrant was described using the MFI scoring system: absent (score 0), intermittent (score 1), sluggish (score 2), or normal (score 3).6 Blood vessel diameter (Øbv) was analyzed in Adobe Photoshop (Adobe Photoshop CC 2020, Adobe Systems Incorporated, San Jose, California, USA) using a digital overlay diameter measuring tool (containing magnitudes of 5, 10, 20, 30, 40 and 50 mm) that was previously described elsewhere.12 The diameter magnitude of the 5 largest vessels per quadrant (20 total) were measured and averaged as Øbv per frame. Vessel morphology was screened throughout all time points for IR induced alterations in the form of telangiectasias and/or disorganization of vessel loops. An intraclass correlation coefficient of 0.982 (p<0.001) was computed from a sample subset of data on buccal and gingiva FCD to ascertain interrater reproducibility agreement between examiners (RH, NFS, and DMJM). Remaining datasets were randomized and analyzed at random for each parameter (Fd, angioarchitecture, MFI, and buccal Øbv) by the same investigator (RH).
Statistical analysis
Previous capillary density mean comparisons between healthy and irradiated oral mucosa demonstrated a large effect size of 0.86.12 In the present study comparative analysis of capillary density between each time point was performed using generalized linear model approach. With a total sample size of 32, a single-group repeated measures analysis of variance (ANOVA) with a 5% significance level has an 80% power to detect an effect size of 0.22. Finally, 34 patients were included in this study [Fig. 1]. Normality distribution of all data parameters (FCD, TCD, Øbv, Fd) was assessed by the Shapiro-Wilk test. Based on the outcome of the Shapiro-Wilk test, repeated measures
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