Abstract
BackgroundThe tumor draining lymph node concept was first described in penile cancer for staging. Immunohistochemistry and histopathology evaluations are routinely used in clinical practice to examine lymph nodes for metastasis. However, these methods are time-consuming with low diagnostic accuracy and micro-metastases might be missed. In this study, we aim to evaluate detection of metastatic cells in draining lymph nodes by flow cytometry.MethodsTo assess the sensitivity of micro-metastasis detection by FACS (Fluorescence-activated cell sorting), HeLa cells were titrated into Peripheral blood mononuclear cells (PBMCs) and expression of pan-cytokeratin AE1/AE3 was analyzed. Single cell suspensions were separately prepared from 10 regional lymph nodes obtained from 5 patients with invasive penile cancer undergoing radical surgery and lymph node dissection. Lymph node dereived cells were examined for cell surface expression of EpCAM, E-cadherin and intracellular expression of pan-cytokeratin AE1/AE3 by FACS.ResultsTen lymph nodes from 5 penile cancer patients were investigated in a head-to-head comparison between FACS and pathology examination of sections. All metastatic lymph nodes verified by pathology examination were also identified by FACS. Two additional lymph nodes with micro-metastases were diagnosed by FACS only.ConclusionsFACS analyses of pan-cytokeratin AE1/AE3 stained single cells from tumor draining lymph nodes can be used to detect micro-metastases in patients with penile cancer patients.
Highlights
The tumor draining lymph node concept was first described in penile cancer for staging
We evaluated the use of EpCAM, E-cadherin, and cytokeratins for detection of metastasis in tumor draining lymph nodes from penile cancer patients by flow cytometry
Since there is a resemblance between HPV positive penile cancer cells and cervical cancer cells [19] we decided to use HeLa cells for initial set up of the flow cytometry protocol
Summary
The tumor draining lymph node concept was first described in penile cancer for staging. Routine histopathology exam of penile cancer is performed by visual examination of hematoxylin and eosin (H&E)-stained slices under microscope by pathologists This method is generally considered to be time-consuming and most. Our previous studies have demonstrated that flow cytometry can be used to detect micro-metastasis in lymph nodes from patients with colon, renal and head and neck cancer using cytokeratin antibodies [8,9,10]. Ectopic presence of epithelial cells in lymph nodes is considered as a result of metastasis [16] These cells can be detected using EpCAM antibody which recognizes an adhesion molecule expressed on the cell surface of most epithelial cells [8]. The expression of E-cadherin, a type I cell adhesion molecule, has been associated with metastasis [17, 18]
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