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Gli1 maintains cell survival by up-regulating IGFBP6 and Bcl-2 through promoter regions in parallel manner in pancreatic cancer cells.

Xu XF, Guo CY, Liu J, Yang WJ, Xia YJ, Xu L, Yu YC, Wang XP - J Carcinog (2009)

Bottom Line: Cyclopamine inhibited cell proliferation at low concentration and induced apoptosis at high concentration.Effect of RNA interference (RNAi) for Gli1 to cell survival is mainly due to proliferation inhibition though involved in apoptosis.The transcription factor Gli1 bound to promoter regions of Bcl-2 and IGFBP6 genes and the levels of IGFBP6, proliferating cell nuclear antigen (PCNA) and Bcl-2 messenger RNA (mRNA) were decreased as well as Gli1 mRNA significantly by cyclopamine or RNAi in cultured pancreatic cancer cells (p < 0.01).

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Gastroenterology, The Tenth Hospital, Tongji University, Shanghai, China. wangxp1965@yahoo.com.cn.

ABSTRACT

Background: Aberrant activation of Hedgehog (Hh) signaling pathway has been reported to be related to malignant biological behavior of pancreatic cancer but its mechanism is unclear yet. Since IGF pathway and Bcl-2 family are involved in proliferation and apoptosis of pancreatic cancer cells, we hypothesize that they are possibly associated with Hh pathway.

Materials and methods: We studied the relationship of Shh-Gli1 signaling pathway with proliferation and apoptosis of pancreatic cancer cells and the regulation of transcription factor Gli1 to insulin-like growth factor binding protein 6 (IGFBP6) and Bcl-2 genes at the level of transcription.

Results: Sonic hedgehog (Shh), Smoothened (Smo), patched and Gli1 were expressed in pancreatic cancer cells. Cyclopamine inhibited cell proliferation at low concentration and induced apoptosis at high concentration. Effect of RNA interference (RNAi) for Gli1 to cell survival is mainly due to proliferation inhibition though involved in apoptosis. The transcription factor Gli1 bound to promoter regions of Bcl-2 and IGFBP6 genes and the levels of IGFBP6, proliferating cell nuclear antigen (PCNA) and Bcl-2 messenger RNA (mRNA) were decreased as well as Gli1 mRNA significantly by cyclopamine or RNAi in cultured pancreatic cancer cells (p < 0.01). Finally PCNA, IGFBP6 and Bcl-2 mRNA were upregulated as well as Shh or Gli1 in pancreatic cancer tissues (p < 0.01).

Conclusions: Our study reveals that Gli1 maintained cell survival by binding the promoter regions and facilitating transcription of IGFBP6 and Bcl-2 genes in a parallel manner in pancreatic cancer cells and suggests it may be one of the mechanisms of Shh-Gli1 signaling pathway in pancreatic cancer.

No MeSH data available.


Related in: MedlinePlus

Binding of Gli1 to Bcl-2 and IGFBP6 genes by XChIP-PCR assay. A: total DNA sheared for 6 rounds of 10 s pulses (350 W, 60 s intervals). (M: BL200 DNA Marker; 1: BxPC3; 2: AsPC-1; 3: Panc-1; 4: SW1990.); B: a, b, c and d show expression of Gli1 protein compared with negative controls (a', b', c' and d') by IP-western assay; C: results of XChIP-PCR. the lanes of a1, a2, a3 and a4 were PCR products of positive control DNA templates; that of b1, b2, b3 and b4 were PCR products of DNA templates from Gli1 XChIP; that of c1, c2, c3 and c4 were negative controls
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Figure 0005: Binding of Gli1 to Bcl-2 and IGFBP6 genes by XChIP-PCR assay. A: total DNA sheared for 6 rounds of 10 s pulses (350 W, 60 s intervals). (M: BL200 DNA Marker; 1: BxPC3; 2: AsPC-1; 3: Panc-1; 4: SW1990.); B: a, b, c and d show expression of Gli1 protein compared with negative controls (a', b', c' and d') by IP-western assay; C: results of XChIP-PCR. the lanes of a1, a2, a3 and a4 were PCR products of positive control DNA templates; that of b1, b2, b3 and b4 were PCR products of DNA templates from Gli1 XChIP; that of c1, c2, c3 and c4 were negative controls

Mentions: In order to determine whether Hh signaling pathway is activated in pancreatic cancer cell lines, RT-PCR assay was used to examine the expression of Shh, Patched, Smo and Gli1 mRNA in BxPC-3, AsPC-1, Panc-1 and SW1990 cells. The results showed that mRNA of Shh, Patched, Smo and Gli1 genes were expressed significantly [Figure 1]. Moreover, the IP-western assay was carried out as described in the Material and Methods section. As expected, the presences of Gli-1 proteins were detected in BxPC-3, AsPC-1, Panc-1 and SW1990 cells [Figure 5B].


Gli1 maintains cell survival by up-regulating IGFBP6 and Bcl-2 through promoter regions in parallel manner in pancreatic cancer cells.

Xu XF, Guo CY, Liu J, Yang WJ, Xia YJ, Xu L, Yu YC, Wang XP - J Carcinog (2009)

Binding of Gli1 to Bcl-2 and IGFBP6 genes by XChIP-PCR assay. A: total DNA sheared for 6 rounds of 10 s pulses (350 W, 60 s intervals). (M: BL200 DNA Marker; 1: BxPC3; 2: AsPC-1; 3: Panc-1; 4: SW1990.); B: a, b, c and d show expression of Gli1 protein compared with negative controls (a', b', c' and d') by IP-western assay; C: results of XChIP-PCR. the lanes of a1, a2, a3 and a4 were PCR products of positive control DNA templates; that of b1, b2, b3 and b4 were PCR products of DNA templates from Gli1 XChIP; that of c1, c2, c3 and c4 were negative controls
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2746911&req=5

Figure 0005: Binding of Gli1 to Bcl-2 and IGFBP6 genes by XChIP-PCR assay. A: total DNA sheared for 6 rounds of 10 s pulses (350 W, 60 s intervals). (M: BL200 DNA Marker; 1: BxPC3; 2: AsPC-1; 3: Panc-1; 4: SW1990.); B: a, b, c and d show expression of Gli1 protein compared with negative controls (a', b', c' and d') by IP-western assay; C: results of XChIP-PCR. the lanes of a1, a2, a3 and a4 were PCR products of positive control DNA templates; that of b1, b2, b3 and b4 were PCR products of DNA templates from Gli1 XChIP; that of c1, c2, c3 and c4 were negative controls
Mentions: In order to determine whether Hh signaling pathway is activated in pancreatic cancer cell lines, RT-PCR assay was used to examine the expression of Shh, Patched, Smo and Gli1 mRNA in BxPC-3, AsPC-1, Panc-1 and SW1990 cells. The results showed that mRNA of Shh, Patched, Smo and Gli1 genes were expressed significantly [Figure 1]. Moreover, the IP-western assay was carried out as described in the Material and Methods section. As expected, the presences of Gli-1 proteins were detected in BxPC-3, AsPC-1, Panc-1 and SW1990 cells [Figure 5B].

Bottom Line: Cyclopamine inhibited cell proliferation at low concentration and induced apoptosis at high concentration.Effect of RNA interference (RNAi) for Gli1 to cell survival is mainly due to proliferation inhibition though involved in apoptosis.The transcription factor Gli1 bound to promoter regions of Bcl-2 and IGFBP6 genes and the levels of IGFBP6, proliferating cell nuclear antigen (PCNA) and Bcl-2 messenger RNA (mRNA) were decreased as well as Gli1 mRNA significantly by cyclopamine or RNAi in cultured pancreatic cancer cells (p < 0.01).

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Gastroenterology, The Tenth Hospital, Tongji University, Shanghai, China. wangxp1965@yahoo.com.cn.

ABSTRACT

Background: Aberrant activation of Hedgehog (Hh) signaling pathway has been reported to be related to malignant biological behavior of pancreatic cancer but its mechanism is unclear yet. Since IGF pathway and Bcl-2 family are involved in proliferation and apoptosis of pancreatic cancer cells, we hypothesize that they are possibly associated with Hh pathway.

Materials and methods: We studied the relationship of Shh-Gli1 signaling pathway with proliferation and apoptosis of pancreatic cancer cells and the regulation of transcription factor Gli1 to insulin-like growth factor binding protein 6 (IGFBP6) and Bcl-2 genes at the level of transcription.

Results: Sonic hedgehog (Shh), Smoothened (Smo), patched and Gli1 were expressed in pancreatic cancer cells. Cyclopamine inhibited cell proliferation at low concentration and induced apoptosis at high concentration. Effect of RNA interference (RNAi) for Gli1 to cell survival is mainly due to proliferation inhibition though involved in apoptosis. The transcription factor Gli1 bound to promoter regions of Bcl-2 and IGFBP6 genes and the levels of IGFBP6, proliferating cell nuclear antigen (PCNA) and Bcl-2 messenger RNA (mRNA) were decreased as well as Gli1 mRNA significantly by cyclopamine or RNAi in cultured pancreatic cancer cells (p < 0.01). Finally PCNA, IGFBP6 and Bcl-2 mRNA were upregulated as well as Shh or Gli1 in pancreatic cancer tissues (p < 0.01).

Conclusions: Our study reveals that Gli1 maintained cell survival by binding the promoter regions and facilitating transcription of IGFBP6 and Bcl-2 genes in a parallel manner in pancreatic cancer cells and suggests it may be one of the mechanisms of Shh-Gli1 signaling pathway in pancreatic cancer.

No MeSH data available.


Related in: MedlinePlus