Lymphangiogenesis assessed using three methods is related to tumour grade, breast cancer subtype and expression of basal marker.

Lymphangiogenesis is a potential indicator of cancer patients' survival. However, there is no standardisation of methodologies applied to the assessment of lymphatic vessel density. In 156 invasive ductal breast cancers (T  1/N+/M0), lymphatic and blood vessels were visualised using podoplanin and CD34, respectively. Based on these markers expression, four parameters were assessed: (i) distribution of podoplanin-stained vessels (DPV) - the percentage of fields with at least one lymphatic vessel (a simple method proposed by us), (ii) lymphatic vessel density (LVD), (iii) LVD to microvessel density ratio (LVD/MVD) and (iv) the expression of podoplanin in cancer-associated fibroblasts. Next, we estimated relations between the above-mentioned parameters and: (i) breast cancer subtype, (ii) tumour grade, and (iii) basal markers expression. We found that intensive lymphangiogenesis, assessed using all studied methods, is positively related to high tumour grade, triple negative or HER2 subtype and expression of basal markers. Whereas, the absence of podoplanin expression in fibroblasts of cancer stroma is related to luminal A subtype, low tumour grade or lack of basal markers expression. Distribution of podoplanin-stained vessels, assessed by a simple method proposed by us (indicating the percentage of fields with at least one lymphatic vessel), might be used instead of the "hot-spot" method.