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Stanniocalcin-1 Reduces Tumor Size in Human Hepatocellular Carcinoma.

Yeung BH, Shek FH, Lee NP, Wong CK - PLoS ONE (2015)

Bottom Line: Lentiviral-based STC1 overexpression in Hep3B and MHCC-97L cells however showed inhibitory action on the pro-migratory effects of IL-6 and IL-8 and reduced size of tumor spheroids.The inhibitory effect of STC1 on tumor growth was confirmed in vivo using the stable STC1-overexpressing 97L cells on a mouse xenograft model.Collectively, the anti-inflammatory and pro-apoptotic functions of STC1 were suggested to relate its inhibitory effect on the growth of HCC cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Hong Kong Baptist University, Kowloon Tong, Hong Kong.

ABSTRACT
Growing evidence has revealed high expression levels of stanniocalcin-1 (STC1) in different types of human cancers. Numerous experimental studies using cancer cell lines demonstrated the involvement of STC1 in inflammatory and apoptotic processes; however the role of STC1 in carcinogenesis remains elusive. Hepatocellular carcinoma (HCC) an exemplified model of inflammation-related cancer, represents a paradigm of studying the association between STC1 and tumor development. Therefore, we conducted a statistical analysis on the expression levels of STC1 using clinicopathological data from 216 HCC patients. We found that STC1 was upregulated in the tumor tissues and its expression levels was positively correlated with the levels of interleukin (IL)-6 and IL-8. Intriguingly tumors with greater expression levels of STC1 (tumor/normal ≥ 2) were significantly smaller than the lower level (tumor/normal<2) samples (p = 0.008). A pharmacological approach was implemented to reveal the functional correlation between STC1 and the ILs in the HCC cell-lines. IL-6 and IL-8 treatment of Hep3B cells induced STC1 expression. Lentiviral-based STC1 overexpression in Hep3B and MHCC-97L cells however showed inhibitory action on the pro-migratory effects of IL-6 and IL-8 and reduced size of tumor spheroids. The inhibitory effect of STC1 on tumor growth was confirmed in vivo using the stable STC1-overexpressing 97L cells on a mouse xenograft model. Genetic analysis of the xenografts derived from the STC1-overexpressing 97L cells, showed upregulation of the pro-apoptotic genes interleukin-12 and NOD-like receptor family, pyrin domain-containing 3. Collectively, the anti-inflammatory and pro-apoptotic functions of STC1 were suggested to relate its inhibitory effect on the growth of HCC cells. This study supports the notion that STC1 may be a potential therapeutic target for inflammatory tumors in HCC patients.

No MeSH data available.


Related in: MedlinePlus

STC1 was a downstream target of IL6 and IL8.(A) STC1 gene expression was determined in a panel of HCC cell lines using qPCR. (B) STC1 protein levels in cell lysates and condition media of H2P and Hep3B cells were measured using ELISA. (C) Western blotting analysis of STC1, IL6, IL8 and ERK1/2 in H2P and Hep3B cell lines. (D) After overnight serum starvation, addition of 25 and 50 ng/ml of IL6 or IL8 in Hep3B cells for 30 min induced STC1 protein expression. The secretion (left) and gene expression (right) of both IL6 and IL8 were measured in Hep3B monolayer attached cells (E) and 10 days’ spheroids (F).
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pone.0139977.g002: STC1 was a downstream target of IL6 and IL8.(A) STC1 gene expression was determined in a panel of HCC cell lines using qPCR. (B) STC1 protein levels in cell lysates and condition media of H2P and Hep3B cells were measured using ELISA. (C) Western blotting analysis of STC1, IL6, IL8 and ERK1/2 in H2P and Hep3B cell lines. (D) After overnight serum starvation, addition of 25 and 50 ng/ml of IL6 or IL8 in Hep3B cells for 30 min induced STC1 protein expression. The secretion (left) and gene expression (right) of both IL6 and IL8 were measured in Hep3B monolayer attached cells (E) and 10 days’ spheroids (F).

Mentions: To characterize the role of STC1 in hepatocarcinogenesis, expression levels of STC1 in different human HCC cell lines were determined and in vitro functional assays were then conducted. The data revealed that low STC1 expression levels was detected in most of the HCC cell lines, but the expression level was greater in H2P cells (Fig 2A). Consistently, the data of ELISA showed significant high levels of STC1 protein detected in cell lysates and condition media of H2P cells (Fig 2B). To compare the expression of profiles of STC1, IL6, IL8 and pERK, protein lysates were prepared from Hep3B and H2P cells. H2P cells showed greater expression levels of IL-6, IL-8 and p-ERK1/2 (137F5) (Fig 2C). The western blot data of IL6 and IL8 support the correlation analysis of the clinicopathological data from the patient samples (Table 2) and the findings reported by Westerberg et al. [28], showing STC1 expression was induced by IL-6 via MAPK signaling. To investigate the possible cause-and-effect relationship between STC1 and the cytokines IL6 and IL8, a pharmacological approach was adopted to elucidate the possible interaction between IL-6, IL-8 and STC1 using Hep3B cells. Upon stimulation by 50 ng/mL of IL-6 or IL-8 for 30 min, STC1 protein expression was found to be upregulated (Fig 2D). On the contrary, to address the effects of STC1 on IL6/IL8 expression, STC1-overexpressed Hep3B cells (Hep3B/STC1) and the empty vector control Hep3B/pLenti were generated using lentiviral system. The overexpression of STC1 was verified using qPCR (data not shown) and western blotting (Figure A in S2 File). The STC1-overexpressing cells, either cultured in monolayers (Fig 2E) or spheroids (Fig 2F) condition, did not show noticeable effects on the secretion (left) and mRNA expression (right) of IL-6 or IL-8. Collectively, the data suggest that STC1 is a downstream target of IL6/IL8 in Hep3B cells.


Stanniocalcin-1 Reduces Tumor Size in Human Hepatocellular Carcinoma.

Yeung BH, Shek FH, Lee NP, Wong CK - PLoS ONE (2015)

STC1 was a downstream target of IL6 and IL8.(A) STC1 gene expression was determined in a panel of HCC cell lines using qPCR. (B) STC1 protein levels in cell lysates and condition media of H2P and Hep3B cells were measured using ELISA. (C) Western blotting analysis of STC1, IL6, IL8 and ERK1/2 in H2P and Hep3B cell lines. (D) After overnight serum starvation, addition of 25 and 50 ng/ml of IL6 or IL8 in Hep3B cells for 30 min induced STC1 protein expression. The secretion (left) and gene expression (right) of both IL6 and IL8 were measured in Hep3B monolayer attached cells (E) and 10 days’ spheroids (F).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4607425&req=5

pone.0139977.g002: STC1 was a downstream target of IL6 and IL8.(A) STC1 gene expression was determined in a panel of HCC cell lines using qPCR. (B) STC1 protein levels in cell lysates and condition media of H2P and Hep3B cells were measured using ELISA. (C) Western blotting analysis of STC1, IL6, IL8 and ERK1/2 in H2P and Hep3B cell lines. (D) After overnight serum starvation, addition of 25 and 50 ng/ml of IL6 or IL8 in Hep3B cells for 30 min induced STC1 protein expression. The secretion (left) and gene expression (right) of both IL6 and IL8 were measured in Hep3B monolayer attached cells (E) and 10 days’ spheroids (F).
Mentions: To characterize the role of STC1 in hepatocarcinogenesis, expression levels of STC1 in different human HCC cell lines were determined and in vitro functional assays were then conducted. The data revealed that low STC1 expression levels was detected in most of the HCC cell lines, but the expression level was greater in H2P cells (Fig 2A). Consistently, the data of ELISA showed significant high levels of STC1 protein detected in cell lysates and condition media of H2P cells (Fig 2B). To compare the expression of profiles of STC1, IL6, IL8 and pERK, protein lysates were prepared from Hep3B and H2P cells. H2P cells showed greater expression levels of IL-6, IL-8 and p-ERK1/2 (137F5) (Fig 2C). The western blot data of IL6 and IL8 support the correlation analysis of the clinicopathological data from the patient samples (Table 2) and the findings reported by Westerberg et al. [28], showing STC1 expression was induced by IL-6 via MAPK signaling. To investigate the possible cause-and-effect relationship between STC1 and the cytokines IL6 and IL8, a pharmacological approach was adopted to elucidate the possible interaction between IL-6, IL-8 and STC1 using Hep3B cells. Upon stimulation by 50 ng/mL of IL-6 or IL-8 for 30 min, STC1 protein expression was found to be upregulated (Fig 2D). On the contrary, to address the effects of STC1 on IL6/IL8 expression, STC1-overexpressed Hep3B cells (Hep3B/STC1) and the empty vector control Hep3B/pLenti were generated using lentiviral system. The overexpression of STC1 was verified using qPCR (data not shown) and western blotting (Figure A in S2 File). The STC1-overexpressing cells, either cultured in monolayers (Fig 2E) or spheroids (Fig 2F) condition, did not show noticeable effects on the secretion (left) and mRNA expression (right) of IL-6 or IL-8. Collectively, the data suggest that STC1 is a downstream target of IL6/IL8 in Hep3B cells.

Bottom Line: Lentiviral-based STC1 overexpression in Hep3B and MHCC-97L cells however showed inhibitory action on the pro-migratory effects of IL-6 and IL-8 and reduced size of tumor spheroids.The inhibitory effect of STC1 on tumor growth was confirmed in vivo using the stable STC1-overexpressing 97L cells on a mouse xenograft model.Collectively, the anti-inflammatory and pro-apoptotic functions of STC1 were suggested to relate its inhibitory effect on the growth of HCC cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Hong Kong Baptist University, Kowloon Tong, Hong Kong.

ABSTRACT
Growing evidence has revealed high expression levels of stanniocalcin-1 (STC1) in different types of human cancers. Numerous experimental studies using cancer cell lines demonstrated the involvement of STC1 in inflammatory and apoptotic processes; however the role of STC1 in carcinogenesis remains elusive. Hepatocellular carcinoma (HCC) an exemplified model of inflammation-related cancer, represents a paradigm of studying the association between STC1 and tumor development. Therefore, we conducted a statistical analysis on the expression levels of STC1 using clinicopathological data from 216 HCC patients. We found that STC1 was upregulated in the tumor tissues and its expression levels was positively correlated with the levels of interleukin (IL)-6 and IL-8. Intriguingly tumors with greater expression levels of STC1 (tumor/normal ≥ 2) were significantly smaller than the lower level (tumor/normal<2) samples (p = 0.008). A pharmacological approach was implemented to reveal the functional correlation between STC1 and the ILs in the HCC cell-lines. IL-6 and IL-8 treatment of Hep3B cells induced STC1 expression. Lentiviral-based STC1 overexpression in Hep3B and MHCC-97L cells however showed inhibitory action on the pro-migratory effects of IL-6 and IL-8 and reduced size of tumor spheroids. The inhibitory effect of STC1 on tumor growth was confirmed in vivo using the stable STC1-overexpressing 97L cells on a mouse xenograft model. Genetic analysis of the xenografts derived from the STC1-overexpressing 97L cells, showed upregulation of the pro-apoptotic genes interleukin-12 and NOD-like receptor family, pyrin domain-containing 3. Collectively, the anti-inflammatory and pro-apoptotic functions of STC1 were suggested to relate its inhibitory effect on the growth of HCC cells. This study supports the notion that STC1 may be a potential therapeutic target for inflammatory tumors in HCC patients.

No MeSH data available.


Related in: MedlinePlus