Identification of oral squamous cell carcinoma markers MUC2 and SPRR1B downstream of TANGO

Abstract

Purpose: Transport and Golgi organization protein 1 (TANGO) promotes angiogenesis and lymphangiogenesis in oral squamous cell carcinoma (OSCC). To elucidate the underlying mechanisms, this study aims to identify and characterize elements downstream of TANGO that mediate its involvement in OSCC.

Methods: In this study, microarray analysis compared gene expression between control and TANGO-repressed HSC3 cells. Protein expression in 213 OSCC tissue samples was analyzed immunohistochemically.

Results: TANGO repression decreased or increased expression of Mucin 20 (MUC20) and small proline-rich protein 1B (SPRR1B), respectively. MUC20 increased the growth and invasiveness of OSCC cells via altered matrix metalloproteinase (MMP)-2 and E-cadherin expression and c-met phosphorylation. MUC20 induced angiogenesis and lymphangiogenesis by activating vascular endothelial growth factors A and C. In well-differentiated OSCC, SPRR1B expression was high (P = 0.0091) and correlated with keratinization markers and promoted proliferation by inducing mitogen-activated protein kinase p38 phosphorylation. MUC20 expression correlated significantly with clinical stage (P = 0.0024), lymph node metastasis (P = 0.0036), and number of blood and lymph vessels (P < 0.0001). MUC20-expressing cases had a significantly worse prognosis than non-expressing cases (P < 0.0001).

Conclusion: MUC20 and SPRR1B located downstream of TANGO may be useful molecular markers for OSCC.

Keywords: MUC20; Oral cancer; SPRR1B; TANGO.

Fig. 1

Expression and regulation of MUC20 and SPRR1B in OSCC cells. a Expression of TANGO, MUC20, and SPRR1B in control HSC3 cells and HSC3 cells with the downregulation of TANGO. b Differential expression of MUC20 and SPRR1B in HSC3 and HSC4 cells. c Expression levels of MUC20 and SPRR1B by anti-TANGO antibody or rhTANGO treatment in OSCC cells. *P < 0.01; **P < 0.001

Fig. 2

The effect of MUC20 on growth, invasion and migration of OSCC cells. a Expression change by knockdown or increase in MUC20. b, c Changes in OSCC cell proliferation, invasion and migration by MUC20 in HSC3 cells (b) and HSC4 cells (c). Altered MMP-2 and E-cadherin secretion and MET phosphorylation levels by MUC20 in HSC3 (d) and HSC4 cells (e). *P < 0.05; **P < 0.01; ***P < 0.001

Fig. 3

Regulation of angiogenic and lymphangiogenic potential of OSCC cells by MUC20. a, b Altered endothelial cell proliferation induced by treatment with conditioned medium from MUC20 siRNA-treated HSC3 cells (a) and MUC20 transfected HSC4 cells (b). c, d Transmigration of endothelial cells to HSC3 cells that suppressed MUC20 expression (c) or HSC4 cells that overexpressed MUC20 (D). e, f Effect of MUC20 derived from HSC3 cells (e) and HSC4 cells (f) on tube formation ability of endothelial cells. g, h Alterations in VEGF-A and VEGF-C secretion levels by MUC20 in HSC3 (g) and HSC4 cells (h). *P < 0.05; *P < 0.01; ***P < 0.001

Fig. 4

Functions of SPRR1B in OSCC cells. Altered expression of keratinocyte differentiation markers induced by SPRR1B in HSC3 and HSC4 cells

Fig. 5

Expression of MUC20 and SPRR1B in OSCC samples. a, b Relationship between the expression levels of TANGO and MUC20 (a) or SPRR1B (b) in OSCC. c, d Differential expression of MUC20 (c) and SPRR1B (d) in OSCC, dysplasia and normal mucosa. e, f Comparison of MUC20 (e) and SPRR1B (f) in OSCC, cervical SCC and CRC

Fig. 6

Immunostaining of MUC20 and SPRR1B in OSCC specimens. a Expression of MUC20 and SPRR1B in normal mucosa and OSCC. b Disease-free survival curve of MUC20-positive and SPRR1B-positive cases. c Schema of TANGO-related signals in OSCC