Various studies have demonstrated that the lymphatic system is the additional route for solid tumor metastasis. Lymph nodes metastasis in Head and neck squamous cell carcinoma (HNSCC) is a major prognostic indicator for disease progression and a guide for therapeutic strategies. We conducted a study to compare intratumoral (IT) and peritumoral (PT) lymphatic vessel density (LVD) in HNSCC using lymphatic marker D2-40 and its correlation with lymph node metastasis, histological grading and other clinicopathological parameters. Fifty specimen of HNSCC with modified radical neck dissection tissue were included in the study group. Tissue from tumor, peritumoral tissue, tumor margin and all the lymph nodes were processed for paraffin wax blocks and histopathological diagnosis. Immunohistochemical profile of lymphatic vessels in intratumoral and peritumoral tissue was assessed by subjecting one section each from the tumor and peritumoral tissue to D2-40 immunostain. To determine LVD, four fields with the highest LVD (hot spots) were identified. The mean values were calculated by taking an average of all the measurements. The comparison of LVD between peritumoral and intratumoral area revealed significantly higher PT-LVD (P = 0.001). No significant association was seen between LVD, IT-LVD and PT-LVD and different age groups, gender, site of tumor, risk factors, size of tumor, tumor inflammation, pushing/infiltrating margin and stage of tumors. Significantly higher LVD, IT-LVD and PT-LVD was seen in association with lymph node metastasis. Both high intratumoral and peritumoral LVD were found significantly associated with the presence of lymph node metastasis, however lymphatic vessels were found to be significantly more numerous and larger in peritumoral areas as compared to intratumoral lymphatics. The specificity of D2-40 as a lymphatic endothelial marker was also confirmed. The results of our study support the possibility of using the determination of tumor lymphangiogenesis to identify patients of HNSCC who are at risk of developing the lymph node metastasis.
Head and neck squamous cell carcinoma Lymphatic vessel density Peritumoral Intratumoral Lymphangiogenesis D2-40 immunostain
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Compliance with Ethical Standards
Conflict of interest
All authors state that there are no conflicts of interests.
All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards.
Informed consent was obtained from all individual participants included in the study.
Siddiqui MS, Chandra R, Aziz A, Suman S (2012) Epidemiology and histopathological spectrum of head and neck cancers in bihar, a state of eastern India. Asian Pac J Cancer Prev 13:3949–3953CrossRefPubMedGoogle Scholar
Trivedi NP, Kekatpure VD, Trivedi NN, Kuriakose MA (2012) Head and neck cancer in India: Need to formulate uniform national treatment guideline? Indian J Cancer 49:6–10CrossRefPubMedGoogle Scholar
Mehrotra R, Singh M, Gupta RK, Singh M, Kapoor AK (2005) Trends of prevalence and pathological spectrum of head and neck cancers in North India. Indian J Cancer 42:89–93CrossRefPubMedGoogle Scholar
Stacker SA, Baldwin ME, Achen MG (2002) The role of tumor lymphangiogenesis in metastatic spread. FASEB J 16:922–934CrossRefPubMedGoogle Scholar
Pepper MS (2001) Lymphangiogenesis and tumor metastasis: Myth or reality? Clin Cancer Res 7:462–468PubMedGoogle Scholar
Tobler NE, Detmar M (2006) Tumor and lymph node lymphangiogenesis-impact on cancer metastasis. J Leukoc Biol 80:691–696CrossRefPubMedGoogle Scholar
Van der Auwera I, Cao Y, Tille JC, Pepper MS, Jackson DG, Fox SB et al (2006) First international consensus on the methodology of lymphangiogenesis quantification in solid human tumors. Br J Cancer 95:1611–1625CrossRefPubMedPubMedCentralGoogle Scholar
Fukunaga M (2005) Expression of D2-40 in lymphatic endothelium of normal tissues and in vascular tumors. Histopathology 46:396–402CrossRefPubMedGoogle Scholar
Bancroft JD, Layton C (2012) The hematoxylins and eosin. In: Suvarna SK, Layton C, Bancroft JD (eds) Theory and practice of histological techniques, 7th edn. Churchill Livingstone, New York, pp 173–186Google Scholar
Akhter M, Hossain S, Rahman QB, Molla MR (2011) A study on histological grading of oral squamous cell carcinoma and its co-relationship with regional metastasis. J Oral Maxillofac Pathol 15:168–176CrossRefPubMedPubMedCentralGoogle Scholar
Jackson P, Blythe D (2012) Immunohistochemical techniques. In: Suvarna SK, Layton C, Bancroft JD (eds) Theory and practice of histological techniques, 7th edn. Churchill Livingstone, New York, pp 381–426Google Scholar
Jemal A, Siegel R, Ward E, Hao Y, Xu J, Murray T et al (2008) Cancer statistics, 2008. CA Cancer J Clin 58:71–96CrossRefPubMedGoogle Scholar
Miyahara M, Tanuma J, Sugihara K, Semba I (2007) Tumor lymphangiogenesis correlates with lymph node metastasis and clinicopathologic parameters in oral squamous cell carcinoma. Cancer 110:1287–1294CrossRefPubMedGoogle Scholar
Xuan M, Fang YR, Wato M, Hata S, Tanaka A (2005) Immunohistochemical co-localization of lymphatics and blood vessels in oral squamous cell carcinomas. J Oral Pathol Med 34:334–339CrossRefPubMedGoogle Scholar
Zhao D, Pan J, Li XQ, Wang XY, Tang C, Xuan M et al (2008) Intratumoral lymphangiogenesis in oral squamous cell carcinoma and its clinicopathological significance. J Oral Pathol Med 37:616–625CrossRefPubMedGoogle Scholar
Frech S, Hormann K, Riedal F, Gotte K (2009) Lymphatic vessel density in correlation to lymph node metastasis in head and neck squamous cell carcinoma. Anticancer Res 29:1675–1679PubMedGoogle Scholar
Chung MK, Min JY, So YK, Ko YH, Jeong HS, Son YI et al (2010) Correlation between lymphatic vessel density and regional metastasis in squamous cell carcinoma of the tongue. Head Neck 32:445–451PubMedGoogle Scholar
Baradaranfar BH, Zahir ST, Atighechi S, Dadgarnia MH, Bayati F, Mohajerani MR (2011) Lymphatic metastasis in head and neck squamous cell carcinoma. Pak J Med Sci 27:303–306Google Scholar
Franchi A, Gallo O, Massi D, Baroni G, Santucci M (2004) Tumor lymphangiogenesis in head and neck squamous cell carcinoma: a morphometric study with clinical correlations. Cancer 101:973–978CrossRefPubMedGoogle Scholar
Audet N, Beasley NJ, Macmillion C, Jackson DG, Gullane PJ, Kamel-Reid S (2005) Lymphatic vessel density, nodal metastases, and prognosis in patients with head and neck cancer. Arch Otolaryngol Head Neck Surg 131:1065–1070CrossRefPubMedGoogle Scholar
Beasley NJ, Prevo R, Banerji S, Leek RD, Moore J, Cox G et al (2002) Intratumoral lymphangiogenesis and lymph node metastasis in head and neck cancer. Cancer Res 62:1315–1320PubMedGoogle Scholar
Kyzas PA, Geleff S, Batistatou A, Agnantis NJ, Stefanou D (2005) Evidence for lymphangiogenesis and its prognostic implications in head and neck squamous cell carcinoma. J Pathol 206:170–177CrossRefPubMedGoogle Scholar
Longatto Filho A, Oliveira TG, Pinheiro C, De Carvalho MB, Curioni OA, Mercante AM et al (2007) How useful is the assessment of lymphatic vascular density in oral carcinoma prognosis? World J Surg Oncol 5:140CrossRefPubMedPubMedCentralGoogle Scholar
Stinga AS, Stinga AC, Margaritescu C, Stepan AE, Barca A, Cruce M (2009) Lymphatic vessels in tumor invasion front of squamous cell carcinoma of the oral tongue. Ann RSCB 14:180–186Google Scholar
Yuan P, Temam S, El-Naggar A, Zhou X, Liu DD, Lee JJ et al (2006) Overexpression of podoplanin in oral cancer and its association with poor clinical outcome. Cancer 107:563–569CrossRefPubMedGoogle Scholar