Influence of buffy coat–derived putative endothelial progenitor cells on tumor growth and neovascularization in oral squamous cell carcinoma xenografts
- 17 Downloads
The aim of this murine in vivo study was to investigate whether buffy coat–derived putative endothelial progenitor cells (BCEPC) alter tumor growth and neovascularization in oral squamous cell carcinomas (OSCC).
Materials and methods
A murine xenograft model using the PCI-13 oral cancer cell line was deployed of which n = 24 animals received 2 × 106 BCEPC by transfusion whereas the control group (n = 24) received NaCl (0.9%) instead. Tumor size, volume, and capillary density were determined by sonography and measurement with a caliper. Immunohistochemical analysis was carried out with antibodies specific for Cytokeratins, Flt-4, Podoplanin, and Vimentin.
In the experimental group, systemic application of BCEPC significantly increased tumor volume to 362.49% (p = 0.0012) and weight to 352.38% (p = 0.0018) as well as vascular densities to 162.15% (p = 0.0021) compared with control tumors. In addition, BCEPC-treated xenografts exhibited higher Cytokeratin expression levels by a factor of 1.47 (p = 0.0417), Podoplanin by a factor of 3.3 (p = 0.0020) and Vimentin by a factor of 2.5 (p = 0.0001), respectively.
Immunohistochemical investigations support the notion that BCEPC transfusion influences neovascularization and lymphatic vessel density, thereby possibly promoting tumor progression. Future studies, which will include gene expression analysis, should help to define the possible role of BCEPC during OSCC progression in more detail.
Endothelial progenitor cells (EPCs) could serve as a target structure for the treatment of OSCC and possibly other solid tumors.
KeywordsOral cancer Endothelial progenitor cell Neovascularization Lymphangiogenesis
The authors thank Dr. J. Goldschmitt (Laboratory of the Clinic for Oral and Maxillofacial Surgery, University Medical Center Mainz, Germany) as well as R. Peldszus and G. Sadowski (Interdisciplinary Head & Neck Oncology Laboratory, Department of Otolaryngology, University Hospital Marburg, Germany) for their excellent technical support. We also thank PD Dr. M. Bette (Institute of Anatomy and Cell Biology, Philipps-Universität Marburg) for the excellent support during microscopic evaluation. Finally, we also thank Dr. A. Schmidt and members of the Institute of Pathology and Molecular Pathology (University Hospital Marburg) for their collegial support and the provision of reagents. This study is part of the thesis work of M.O.
The study was funded by the foundation for tumor research head and neck (Stiftung für Tumorforschung Kopf-Hals), Wiesbaden, Germany.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. The local ethics committee (Landesärztekammer Rheinland-Pfalz) approved all of the experiments with human material in this article (Ethikvotum 837.387.11 (7929)). All applicable international, national, and institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted. A corresponding animal test application (animal experiment G 12-1-037) for the experiments was approved by the Landesuntersuchungsamt Rheinland-Pfalz in Koblenz and thus meets all §8 Paragraph 3 (2) of the German Animal Welfare Act required preconditions.
Informed consent was obtained from all individual participants included in the study.
- 7.Ziebart T, Yoon C-H, Trepels T, Wietelmann A, Braun T, Kiessling F, Stein S, Grez M, Ihling C, Muhly-Reinholz M, Carmona G, Urbich C, Zeiher AM, Dimmeler S (2008) Sustained persistence of transplanted proangiogenic cells contributes to neovascularization and cardiac function after ischemia. Circ Res 103:1327–1334CrossRefGoogle Scholar
- 8.Ziebart T, Pabst A, Klein MO, Kämmerer P, Gauss L, Brüllmann D, Al-Nawas B, Walter C (2011) Bisphosphonates: restrictions for vasculogenesis and angiogenesis: inhibition of cell function of endothelial progenitor cells and mature endothelial cells in vitro. Clin Oral Investig 15:105–111CrossRefGoogle Scholar
- 9.Ratajska A, Jankowska-Steifer E, Czarnowska E, Olkowski R, Gula G, Niderla-Bielińska J, Flaht-Zabost A, Jasińska A (2016) Vasculogenesis and its cellular therapeutic applications. Cells Tissues OrgansGoogle Scholar
- 15.Ziebart T, Blatt S, Günther C, Völxen N, Pabst A, Sagheb K, Kühl S, Lambrecht T (2016) Significance of endothelial progenitor cells (EPC) for tumorigenesis of head and neck squamous cell carcinoma (HNSCC): possible marker of tumor progression and neovascularization? Clin Oral Investig 20:2293–2300CrossRefGoogle Scholar
- 16.Moschetta M, Mishima Y, Sahin I, Manier S, Glavey S, Vacca A, Roccaro AM, Ghobrial IM (2014) Role of endothelial progenitor cells in cancer progression. Biochim Biophys Acta 1846(1):26–39Google Scholar
- 17.Wildt S (2015) Hairless NOD scid mouse. 9018441Google Scholar
- 21.Cardiff RD, Miller CH, Munn RJ (2014) Manual immunohistochemistry staining of mouse tissues using the avidin-biotin complex (ABC) technique. Cold Spring Harb Protoc 2014:659–662Google Scholar
- 22.Schindelin J, Arganda-Carreras I, Frise E, Kaynig V, Longair M, Pietzsch T, Preibisch S, Rueden C, Saalfeld S, Schmid B, Tinevez J-Y, White DJ, Hartenstein V, Eliceiri K, Tomancak P, Cardona A (2012) Fiji: an open-source platform for biological-image analysis. Nat Methods 9:676–682CrossRefGoogle Scholar
- 30.Torimura T, Ueno T, Taniguchi E, Masuda H, Iwamoto H, Nakamura T, Inoue K, Hashimoto O, Abe M, Koga H, Barresi V, Nakashima E, Yano H, Sata M (2012) Interaction of endothelial progenitor cells expressing cytosine deaminase in tumor tissues and 5-fluorocytosine administration suppresses growth of 5-fluorouracil-sensitive liver cancer in mice. Cancer Sci 103(3):542–548CrossRefGoogle Scholar
- 34.Neuchrist C, Erovic BM, Handisurya A, Fischer MB, Steiner GE, Hollemann D, Gedlicka C, Saaristo A, Burian M (2003) Vascular endothelial growth factor C and vascular endothelial growth factor receptor 3 expression in squamous cell carcinomas of the head and neck. Head Neck 25:464–474CrossRefGoogle Scholar
- 35.Ge Q, Zhang H, Hou J, Wan L, Cheng W, Wang X, Dong D, Chen C, Xia J, Guo J, Chen X, Wu X (2018) VEGF secreted by mesenchymal stem cells mediates the differentiation of endothelial progenitor cells into endothelial cells via paracrine mechanisms. Mol Med Rep 17:1667–1675Google Scholar
- 38.Mermod M, Bongiovanni M, Petrova TV, Dubikovskaya EA, Simon C, Tolstonog G, Monnier Y (2017) Correlation between podoplanin expression and extracapsular spread in squamous cell carcinoma of the oral cavity using subjective immunoreactivity scores and semiquantitative image analysis. Head Neck 39:98–108CrossRefGoogle Scholar
- 41.Aruga N, Kijima H, Masuda R, Onozawa H, Yoshizawa T, Tanaka M, Inokuchi S, Iwazaki M (2018) Epithelial-mesenchymal transition (EMT) is correlated with patient’s prognosis of lung squamous cell carcinoma. Tokai J Exp Clin Med 43:5–13Google Scholar