2.6. Analysis of microcirculatory parameters

We analysed the parameters (a) microvascular flow index; (b) tissue angioarchitecture scores; and (c) capillary density. All parameters were validated in previous studies.18, 19, 20, 21, 23 Scoring of the microcirculation was performed by one investigator who was blinded for the clinical data (LdW). Another investigator with extensive experience in scoring microcirculation (AWK) provided close supervision and was available as an independent arbiter when there was doubt during scoring. AWK also performed an additional analysis of 10% of the data, which demonstrated high agreement with the initial analysis by LdW. We qualitatively described the angioarchitecture that was observed at the surgical sites. All videos that could not be scored according to the normal tissue angioarchitecture scores, were discussed and described by at least two investigators.

The microvascular flow was assessed to determine perfusion. Flow characteristics were assessed using the MFI. The MFI was developed and validated by Boerma and coworkers and tested for reproducibility.23 This score is based on the determination of the predominant type of flow in four quadrants. This scoring system quantifies the microcirculatory perfusion as absent (0), intermittent (1), sluggish (2) or normal (3), providing an index for microcirculatory blood flow velocity. The MFI score per region was the score that occurred the most. When the MFI was 3 in all quadrants, this was considered a normally perfused microvasculature. Perfusion was reduced when the MFI was 1 or 2, and perfusion was absent when we observed an MFI of 0.

Tissue angioarchitecture is the morphology or lay‐out of the vascular network. This scoring method was devised and validated to provide rapid recognition of subepithelial vascular patterns and showed a high agreement between observers in a previous study with complete agreement in 93% of frames and ICC between observers of 0.78.18 Three types of predefined sub‐epithelial vascular patters may be recognized and classified with a score 1, 2, or 3.18 In score 1 capillary loops are predominantly observed. This is considered to be the most favorable angioarchitecture score, which is most frequently seen in premenopausal women with a thicker vaginal epithelium.20 The capillary loops are necessary to supply the avascular epithelium with oxygen and nutrients. In score 2, capillary loops and vascular networks are both seen. In score 3, the vascular network without capillary loops is seen. Score 3 angioarchitercure is associated with a thin epithelium of the vaginal wall. Figure ​Figure11 shows these three types of angioarchitecture. When the angioarchitecture did not meet one of these predefined patterns, we qualitatively described the angioarchitecture.

Screenshots of CytoCam videos with different types of angioarchitecture. Each image represents an area of 1.55 × 1.16 mm. Left: angioarchitecture score 1. Middle: angioarchitecture score 2. Right: angioarchitecture score 3

When the vascularization was classified as a microvascular architecture score of 1 or 2 (ie, capillary loops were identifiable), the video was judged suitable for the assessment of capillary density. The capillary density was determined by counting the number of capillary loops per visual field and expressed as the mean number of capillary loops per square millimeter (cpll/mm²). The frame of each image was 1.55 mm × 1.16 mm, resulting in a total area of 1.8 mm². The capillary density score per region was calculated by averaging the frames and divided by 1.8 to obtain the unit cpll/mm². Assessment of capillary density was performed in the same way in previous studies regarding the vaginal microcirculation.18, 19, 20