Comput Methods Programs Biomed - Indirect measurement of the vascular endothelial glycocalyx layer thickness in human submucosal capillaries with a plug-in for ImageJ.


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CKGROUND: The thickness of vascular endothelial glycocalyx layer can be measured indirectly during a spontaneous leukocyte passage from oral submucosal capillaries in humans. The subsequent differences in red blood cell (RBC) column widths, before a spontaneous white blood cell passage (pre-WBC) and after a spontaneous WBC passage (post-WBC) can be used in off-line analysis to measure glycocalyx thickness: [pre-WBC width-post-WBC width]/2. We created and validated a semi-automatic plug-in for ImageJ to measure the endothelial glycocalyx layer thickness.METHODS: Video clips presenting human sublingual microvasculature were created with a side-stream dark field imaging device. Spontaneous leukocyte passages in capillaries were analyzed from video clips with ImageJ. The capillary glycocalyx layer thickness was measured by the indirect approach with two manual and two semi-automatic methods.RESULTS: There were no statistically significant differences between glycocalyx layer thicknesses measured with different methods, even though small inter-method differences in RBC column thicknesses could be detected. Inter-rater differences were systematically smaller with both semi-automatic methods. Intra-rater coefficient of variation [CV] (95% CI) was largest when measurements were made completely manually [9.2% (8.4-10.0)], but improved significantly with automatic image enhancement prior to manual measurement [7.2% (6.4-8.0)]. CV could be improved further when using semi-automatic analysis with an in-frame median filter radius of 1 pixel [5.8% (5.0-6.6)], or a median filter radius of 2 pixels [4.3% (3.5-5.1)].CONCLUSIONS: Semi-automatic analysis of glycocalyx decreased the intra-rater CV and the inter-rater differences compared to the manual method. On average, each of the four methods yielded equal results for the glycocalyx thickness. Being the only feasible bed side method in most clinical scenarios, indirect measurement of glycocalyx thickness with orthogonal polarization spectral imaging or side-stream dark field imaging device and our plug-in can advance the study of glycocalyx layer pathology in man.

Resumo Limpo

ckground thick vascular endotheli glycocalyx layer can measur indirect spontan leukocyt passag oral submucos capillari human subsequ differ red blood cell rbc column width spontan white blood cell passag prewbc spontan wbc passag postwbc can use offlin analysi measur glycocalyx thick prewbc widthpostwbc width creat valid semiautomat plugin imagej measur endotheli glycocalyx layer thicknessmethod video clip present human sublingu microvasculatur creat sidestream dark field imag devic spontan leukocyt passag capillari analyz video clip imagej capillari glycocalyx layer thick measur indirect approach two manual two semiautomat methodsresult statist signific differ glycocalyx layer thick measur differ method even though small intermethod differ rbc column thick detect interrat differ systemat smaller semiautomat method intrarat coeffici variat cv ci largest measur made complet manual improv signific automat imag enhanc prior manual measur cv improv use semiautomat analysi infram median filter radius pixel median filter radius pixel conclus semiautomat analysi glycocalyx decreas intrarat cv interrat differ compar manual method averag four method yield equal result glycocalyx thick feasibl bed side method clinic scenario indirect measur glycocalyx thick orthogon polar spectral imag sidestream dark field imag devic plugin can advanc studi glycocalyx layer patholog man

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