Wound Area Measurement with Digital Planimetry: Improved Accuracy and Precision with Calibration Based on 2 Rulers

<div><p>Introduction</p><p>In the treatment of chronic wounds the wound surface area change over time is useful parameter in assessment of the applied therapy plan. The more precise the method of wound area measurement the earlier may be identified and changed inappropriate treatment plan. Digital planimetry may be used in wound area measurement and therapy assessment when it is properly used, but the common problem is the camera lens orientation during the taking of a picture. The camera lens axis should be perpendicular to the wound plane, and if it is not, the measured area differ from the true area.</p><p>Results</p><p>Current study shows that the use of 2 rulers placed in parallel below and above the wound for the calibration increases on average 3.8 times the precision of area measurement in comparison to the measurement with one ruler used for calibration. The proposed procedure of calibration increases also 4 times accuracy of area measurement. It was also showed that wound area range and camera type do not influence the precision of area measurement with digital planimetry based on two ruler calibration, however the measurements based on smartphone camera were significantly less accurate than these based on D-SLR or compact cameras. Area measurement on flat surface was more precise with the digital planimetry with 2 rulers than performed with the Visitrak device, the Silhouette Mobile device or the AreaMe software-based method.</p><p>Conclusion</p><p>The calibration in digital planimetry with using 2 rulers remarkably increases precision and accuracy of measurement and therefore should be recommended instead of calibration based on single ruler.</p></div

Box plots of relative differences between the measured area and reference area expressed in percentage for the Visitrak device, the Silhouette Mobile device, the AreaMe software and for the digital planimetry methods based on two ruler calibration in 4 ranges of wound area: (A) very small (< 1 cm²), (B) small (1–2 cm²), (C) medium (2–8 cm²), and (D) large (> 8 cm²).

<p>Box plots of relative differences between the measured area and reference area expressed in percentage for the Visitrak device, the Silhouette Mobile device, the AreaMe software and for the digital planimetry methods based on two ruler calibration in 4 ranges of wound area: (A) very small (< 1 cm²), (B) small (1–2 cm²), (C) medium (2–8 cm²), and (D) large (> 8 cm²).</p

Box plot of relative errors of area measurement expressed in percentage for the digital planimetry based on one ruler or two ruler calibration techniques.

<p>Box plot of relative errors of area measurement expressed in percentage for the digital planimetry based on one ruler or two ruler calibration techniques.</p

Effect of hAM CCM on HUVECs migration assayed by scratch test.

<p>There are results of 160 measurements, 8 independent assays with 10 measurements for test and control each. Median values and (P25, P75) are shown (n = 8, p < 0.05). Detailed description of the assay is in Material and methods.</p

Growth factors in hAM CCM.

<p>Forty-one growth factors were quantitated by antibody array and the obtained values were combined into following growth factor families: EGF family (EGF-2, HB-EGF, EGF-R); FGF family (bFGF, FGF-4, FGF-6, FGF-7); Hematopoietic factors HF (MCSF, MCSF-R, SCF, SCF-R); IGF family (IGF-1, IGF-2, IGF-1SR); IGFBP family (IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, IGFBP-6); Neurotrophic factors NF (bNGF, GDNF, NT-3, NT-4); PDGF family (PDGF-AA, PDGF-AB, PDGF-BB, PDGF-Ra, PDGF-Rb); TGF family (TGF-α, TGF-β, TGF-β2, TGF-β3); Vasculogenic factors VF (PLGF, VEGF, VEGF-R3, VEGF-D, VEGF-R2); some growth factors are presented separately: AR; G-CSF; GM-CSF; HGF. Each growth factor fluorescence value (FV) was measured and calculated as described in Materials and methods. (n = 4) (p < 0.05).</p

The example of real time migration assay of stimulated and controlled HUVECs directly from the X-Celligence system.

<p>The assay was repeated seven times with similar result during 25h observation at 37^°C. The difference between migration curves for cells in cultures with presence of hAM CCM and in control medium was significant. (p < 0.05).</p

Effect of hAM CCM on chemotaxy indeks of BM MNCs.

<p>The chemotaxy index (CI) after 2.5 h at 37^°C incubation time was calculated by dividing the number of cells in lower chamber by the number of cells added to the upper chamber counted at the start of the test. Median values and interquartile range (P25, P75) are shown (n = 12, p < 0.05).</p

Effect of hAM CCM on HUVECs expression of cell adhesion molecule-1 (CD31) marker.

<p>Significantly lower expression is observed in presence of hAM CCM. Median values and interquartile ranges (P25, P75) are shown (n = 3, p < 0.05).</p