Limits...
Downregulated NM23-H1 expression is associated with intracranial invasion of nasopharyngeal carcinoma.

Liu SJ, Sun YM, Tian DF, He YC, Zeng L, He Y, Ling CQ, Sun SH - Br. J. Cancer (2008)

Bottom Line: NM23-H1 expression was significantly lower in 5-8F cells compared with that in 6-10B cells.Moreover, patch-clamp and transwell chamber were adopted to investigate the invading potential-associated biological dynamic mechanisms in the two cell lines, and Ca(2+) current and motility were significantly elevated in 5-8F cells compared with that in 6-10B cells.Berberine, an inhibitor of Ca(2+) current, could substantially increase the expression of NM23-H1 and decrease 5-8F cell motility.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Integrative Medicine, Traditional Chinese Medicine University of Hunan, Changsha, People's Republic of China.

ABSTRACT
Because the focus of nasopharyngeal carcinoma (NPC) is very close to intracranial organs, it often makes incursions into cranial cavity. Identification of intracranial invasion-associated indicators will provide potential therapeutic targets for NPC patients with intracranial invasion. In this regard, Human Xpro HC-plus cancer-related gene chip was utilised to screen intracranial invasion-associated genes for NPC from the biopsied primary focus tissue samples. In all, 8 upregulated and 23 downregulated genes were obtained. VEGF165 and MMP-9, the two upregulated genes, and NM23-H1, the downregulated one, were further confirmed by immunohistochemistry, quantitative real-time PCR and western blot. Invasion-associated cellular and nude mouse models were subsequently employed to study the biological properties of NM23-H1. NM23-H1 expression was significantly lower in 5-8F cells compared with that in 6-10B cells. Moreover, patch-clamp and transwell chamber were adopted to investigate the invading potential-associated biological dynamic mechanisms in the two cell lines, and Ca(2+) current and motility were significantly elevated in 5-8F cells compared with that in 6-10B cells. Berberine, an inhibitor of Ca(2+) current, could substantially increase the expression of NM23-H1 and decrease 5-8F cell motility. The specificity of berberine on NM23-H1 and cell motility was confirmed by RNAi assay.

Show MeSH

Related in: MedlinePlus

Validation of NM23-H1, VEGF165 and MMP-9 expression in the tissue samples of primary focus of NPC. (A) Validation with IHC (H&E, × 10). A1, A3, A5: NPC tissues with no invasion; A2, A4, A6: NPC tissues with intracranial invading signs. (B) Validation by the quantitative real-time PCR. Glyceraldehyde 3-phosphate dehydrogenase served as control. Relative mRNA levels of NM23-H1, VEGF165 or MMP-9 mRNA/GAPDH are expressed as the relative abundance. C1: NPC tissues with no invasion, C5: NPC tissues with intracranial invading signs. (C) Validation with western blot. Tissues lysates from two groups were subjected to western immunoblotting with anti-NM23-H1, VEGF165 or MMP-9 antibody, and blot was reprobed with anti-GAPDH to verify equal loading.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2361469&req=5

fig2: Validation of NM23-H1, VEGF165 and MMP-9 expression in the tissue samples of primary focus of NPC. (A) Validation with IHC (H&E, × 10). A1, A3, A5: NPC tissues with no invasion; A2, A4, A6: NPC tissues with intracranial invading signs. (B) Validation by the quantitative real-time PCR. Glyceraldehyde 3-phosphate dehydrogenase served as control. Relative mRNA levels of NM23-H1, VEGF165 or MMP-9 mRNA/GAPDH are expressed as the relative abundance. C1: NPC tissues with no invasion, C5: NPC tissues with intracranial invading signs. (C) Validation with western blot. Tissues lysates from two groups were subjected to western immunoblotting with anti-NM23-H1, VEGF165 or MMP-9 antibody, and blot was reprobed with anti-GAPDH to verify equal loading.

Mentions: From the above 31 differentially expressed genes, we chose three genes that is likely to be associated with intracranial invasion of NPC, including NM23-H1, MMP-9 and VEGF165 for validation. NM23-H1 has been shown as a tumour metastasis suppressor in gastric carcinoma, ovarian cancer and other cancers (Kantor et al, 1993; Freije et al, 1997; Fan et al, 2003). MMP-9 has been demonstrated to play a key role in the invading process of cancerous cells (Hofmann et al, 2005). Elevated VEGF165 expression might be closely associated with the metastatic potentiality of malignant cells (Cianchi et al, 2001; Su et al, 2006). To confirm our microarray findings, we used immunohistochemistry (IHC), quantitative real-time PCR and western blot to determine the expression of NM23-H1, MMP-9 and VEGF165 in NPC tissues with and without intracranial invasion. As shown in Figure 2, both mRNA and protein levels of NM23-H1 are significantly lower in NPC tissues with intracranial invasion compared with that without invasion. Meanwhile, we found that both mRNA and protein levels of MMP-9 and VEGF165 are apparent higher in NPC tissues with intracranial invasion (Figure 2).


Downregulated NM23-H1 expression is associated with intracranial invasion of nasopharyngeal carcinoma.

Liu SJ, Sun YM, Tian DF, He YC, Zeng L, He Y, Ling CQ, Sun SH - Br. J. Cancer (2008)

Validation of NM23-H1, VEGF165 and MMP-9 expression in the tissue samples of primary focus of NPC. (A) Validation with IHC (H&E, × 10). A1, A3, A5: NPC tissues with no invasion; A2, A4, A6: NPC tissues with intracranial invading signs. (B) Validation by the quantitative real-time PCR. Glyceraldehyde 3-phosphate dehydrogenase served as control. Relative mRNA levels of NM23-H1, VEGF165 or MMP-9 mRNA/GAPDH are expressed as the relative abundance. C1: NPC tissues with no invasion, C5: NPC tissues with intracranial invading signs. (C) Validation with western blot. Tissues lysates from two groups were subjected to western immunoblotting with anti-NM23-H1, VEGF165 or MMP-9 antibody, and blot was reprobed with anti-GAPDH to verify equal loading.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2361469&req=5

fig2: Validation of NM23-H1, VEGF165 and MMP-9 expression in the tissue samples of primary focus of NPC. (A) Validation with IHC (H&E, × 10). A1, A3, A5: NPC tissues with no invasion; A2, A4, A6: NPC tissues with intracranial invading signs. (B) Validation by the quantitative real-time PCR. Glyceraldehyde 3-phosphate dehydrogenase served as control. Relative mRNA levels of NM23-H1, VEGF165 or MMP-9 mRNA/GAPDH are expressed as the relative abundance. C1: NPC tissues with no invasion, C5: NPC tissues with intracranial invading signs. (C) Validation with western blot. Tissues lysates from two groups were subjected to western immunoblotting with anti-NM23-H1, VEGF165 or MMP-9 antibody, and blot was reprobed with anti-GAPDH to verify equal loading.
Mentions: From the above 31 differentially expressed genes, we chose three genes that is likely to be associated with intracranial invasion of NPC, including NM23-H1, MMP-9 and VEGF165 for validation. NM23-H1 has been shown as a tumour metastasis suppressor in gastric carcinoma, ovarian cancer and other cancers (Kantor et al, 1993; Freije et al, 1997; Fan et al, 2003). MMP-9 has been demonstrated to play a key role in the invading process of cancerous cells (Hofmann et al, 2005). Elevated VEGF165 expression might be closely associated with the metastatic potentiality of malignant cells (Cianchi et al, 2001; Su et al, 2006). To confirm our microarray findings, we used immunohistochemistry (IHC), quantitative real-time PCR and western blot to determine the expression of NM23-H1, MMP-9 and VEGF165 in NPC tissues with and without intracranial invasion. As shown in Figure 2, both mRNA and protein levels of NM23-H1 are significantly lower in NPC tissues with intracranial invasion compared with that without invasion. Meanwhile, we found that both mRNA and protein levels of MMP-9 and VEGF165 are apparent higher in NPC tissues with intracranial invasion (Figure 2).

Bottom Line: NM23-H1 expression was significantly lower in 5-8F cells compared with that in 6-10B cells.Moreover, patch-clamp and transwell chamber were adopted to investigate the invading potential-associated biological dynamic mechanisms in the two cell lines, and Ca(2+) current and motility were significantly elevated in 5-8F cells compared with that in 6-10B cells.Berberine, an inhibitor of Ca(2+) current, could substantially increase the expression of NM23-H1 and decrease 5-8F cell motility.

View Article: PubMed Central - PubMed

Affiliation: Faculty of Integrative Medicine, Traditional Chinese Medicine University of Hunan, Changsha, People's Republic of China.

ABSTRACT
Because the focus of nasopharyngeal carcinoma (NPC) is very close to intracranial organs, it often makes incursions into cranial cavity. Identification of intracranial invasion-associated indicators will provide potential therapeutic targets for NPC patients with intracranial invasion. In this regard, Human Xpro HC-plus cancer-related gene chip was utilised to screen intracranial invasion-associated genes for NPC from the biopsied primary focus tissue samples. In all, 8 upregulated and 23 downregulated genes were obtained. VEGF165 and MMP-9, the two upregulated genes, and NM23-H1, the downregulated one, were further confirmed by immunohistochemistry, quantitative real-time PCR and western blot. Invasion-associated cellular and nude mouse models were subsequently employed to study the biological properties of NM23-H1. NM23-H1 expression was significantly lower in 5-8F cells compared with that in 6-10B cells. Moreover, patch-clamp and transwell chamber were adopted to investigate the invading potential-associated biological dynamic mechanisms in the two cell lines, and Ca(2+) current and motility were significantly elevated in 5-8F cells compared with that in 6-10B cells. Berberine, an inhibitor of Ca(2+) current, could substantially increase the expression of NM23-H1 and decrease 5-8F cell motility. The specificity of berberine on NM23-H1 and cell motility was confirmed by RNAi assay.

Show MeSH
Related in: MedlinePlus