This study was aimed to investigate the expression and clinicopathologic significance of Gli1 and Gli2, 2 factors of Hedgehog(Hh) signaling pathway, in non-Hodgkin's lymphoma (NHL). Gli1 and Gli2 mRNA and protein in 18 cases of NHL and 10 cases of reactive lymphadenitis were amplified and identified by real-time PCR, and were assayed by immunohistochemical staining respectively. The results showed that (1) Gli1 and Gli2 mRNA in NHL group (RQ 2.05, 2.31) were expressed higher than that in reactive lymphadenitis group (RQ 0.82, 0.89). Gli1 mRNA activated level was positively related with Gli2 (r = 0.63, p < 0.01). In addition, Gli2 also positively correlated to clinical stages of NHL (p = 0.03), but the expressions of Gli1 and Gli2 mRNA had no significant correlation to B symptoms, blood β(2)-microglobulin, age and sex. (2) The positive expression rate of Gli1 and Gli2 protein in NHL group were 80% and 68% respectively, which were extremely higher than that in reactive lymphadenitis group. Gli1 protein level was positively related with Gli2 (r = 0.62, p < 0.05). Both Gli1 and Gli2 protein expression positively correlated to clinical staging of NHL (p = 0.05, p = 0.01). It is concluded that the Gli1 and Gli2 of Hh signaling pathway have been found to higher express in patients with NHL, and have significance for clinical staging and predicting prognosis of NHL. To further investigate the role of Hh signaling pathway in NHL will contribute to elucidate the occurrence and development of NHL, and provide a favorable method for therapy of NHL.
Adult ; Aged ; Female ; Hedgehog Proteins ; metabolism ; Humans ; Kruppel-Like Transcription Factors ; metabolism ; Lymphoma, Non-Hodgkin ; metabolism ; pathology ; Male ; Middle Aged ; Neoplasm Staging ; Nuclear Proteins ; metabolism ; Signal Transduction ; Transcription Factors ; metabolism ; Zinc Finger Protein GLI1 ; Zinc Finger Protein Gli2

OBJECTIVETo investigate the role of the hedgehog (HH) signaling pathway transcription factor glioma-associated oncogene hoinolog 1 (GLI-1) in EGF-regulated enhancement of the invasiveness of the prostate cancer ARCaP(E) cell line in vitro.

METHODSThe expressions of EGFR and GLI-1 in prostate cancer ARCaP(E) cells were analyzed by immunofluorescence staining. ARCaP(E) cells were treated with EGF at 100 ng/ml, followed by detection of the changes in cell morphology and invasiveness, as well as in the expressions of p-ERK, ERK and GLI-1. Migration transwell assay was used to determine the effects of 100 ng/ml EGF and GLI-1 antagonist GANT61 on the invasiveness of the ARCaP(E) cells.

RESULTSBoth EGFR and GLI-1 were expressed in the ARCaP(E) cells. EGF induced morphological transition of epithelial-like ARCaP(E) cells to mesenchymal-like cells, increased their in vitro invasiveness, and significantly upregulated the expressions of p-ERK and GLI-1 in the ARCaP(E) cells (P<0.05). GANT61 significantly inhibited the in vitro invasiveness of the ARCaP(E) cells and reduced the enhancing effect of EGF on their invasiveness (P<0.05).

CONCLUSIONThe results from ARCaP(E) cells shed light on the cross-talk of the HH pathway with the EGF/ERK signaling pathway. GLI-1 might be responsible for EGF-regulated enhancement of the invasiveness of ARCaP(E) cells in vitro.

Cell Line, Tumor ; Epidermal Growth Factor ; metabolism ; Humans ; Male ; Prostatic Neoplasms ; metabolism ; pathology ; Signal Transduction ; Transcription Factors ; genetics ; metabolism ; Zinc Finger Protein GLI1

OBJECTIVETo study the expression of Smo protein and the downstream transcription factor Gli1 protein in Sonic hedgehog signal transduction pathway in gastric carcinoma.

METHODSA tissue microarray was constructed using 85 gastric carcinoma and 25 normal gastric mucosa specimens. The expression of Smo and Gli1 proteins were detected immunohistochemically and the correlation between their expression in gastric carcinoma was analyzed.

RESULTSOnly weak expression, if any, of Smo and Gli1 proteins was detected in normal gastric mucosa, but in papillary adenocarcinoma, tubular adenocarcinoma and poorly differentiated adenocarcinoma, their expressions were significant increased as the differentiation degree was lowered. Smo protein expression in gastric carcinoma was significantly correlated with that of Gli1 protein with correlation coefficient of 0.989 (P<0.001).

CONCLUSIONThe abnormal activity of Sonic hedgehog signal transduction pathway may play an important role in the occurrence of papillary adenocarcinoma, tubular adenocarcinoma and poorly differentiated adenocarcinoma, and this abnormality is associated with Smo protein overexpression, which upregulates the expression of the downstream transcription factor Gli1 protein.

Adenocarcinoma ; metabolism ; pathology ; physiopathology ; Adult ; Aged ; Female ; Hedgehog Proteins ; physiology ; Humans ; Immunohistochemistry ; Male ; Middle Aged ; Receptors, G-Protein-Coupled ; biosynthesis ; Signal Transduction ; Smoothened Receptor ; Stomach Neoplasms ; metabolism ; pathology ; physiopathology ; Transcription Factors ; biosynthesis ; Zinc Finger Protein GLI1

OBJECTIVETo investigate the effect of Hedgehog (HH) pathway on proliferation and in vitro tumorigenicity of gastric cancer cell lines.

METHODSThe expression of SHH, PTCH, SMO, SUFU and GLI1 in seven cell lines were tested by RT-PCR. siRNA targeting GLI1 mRNA was transfected into MKN28 cells. Cell proliferation and in vitro tumorigenicity were examined by CCK8 and soft agar colony formation test.

RESULTSSHH in six gastric cancer cell lines was up-regulated. Expression of PTCH in KATOIII cell lines and expression of SUFU in MKN28 and KATOIII were reduced. GLI1 siRNA significantly inhibited the expression of GLI1 in MKN28 cell line. Growth rate and colony formation rate of MKN28 cells treated with GLI1 siRNA were significantly lower than those of control cells (all P <0.001).

CONCLUSIONSHH signaling pathway is widely activated in gastric cancer cell lines. The activation of HH signaling pathway promotes the growth of MKN28 cells.

Cell Line, Tumor ; Cell Proliferation ; Gastric Mucosa ; cytology ; Hedgehog Proteins ; metabolism ; Humans ; Oncogene Proteins ; metabolism ; RNA, Small Interfering ; Signal Transduction ; Stomach Neoplasms ; metabolism ; pathology ; Trans-Activators ; metabolism ; Zinc Finger Protein GLI1

OBJECTIVETo investigate the effect of cyclopamine on metastatic ability of human esophageal cancer EC109 cells and explore the possible mechanism.

METHODSTranswell chamber assay and angiogenesis assay were used to examine the metastatic ability, invasiveness and angiogenesis of EC109 cells treated with cyclopamine for 48 h. The expression of Gli-1 mRNA was detected using RT-PCR, and Western blotting was used to examine the protein expressions of Gli-1, matrix metalloproteinase-9 (MMP-9) and vascular endothelial growth factor (VEGF).

RESULTSInhibition of the hedgehog signaling pathway by cyclopamine suppressed the migration, invasion, and angiogenesis of EC109 cells. Cyclopamine treatment significantly lowered the expression of Gli-1 mRNA (P<0.05) and the protein expressions of Gli-1, MMP-9 and VEGF (P<0.05).

CONCLUSIONCyclopamine can significantly inhibit the metastatic capacity of EC109 cells possibly by down-regulating MMP-9 and VEGF expression as a result of Gli-1 inhibition.

Cell Line, Tumor ; Esophageal Neoplasms ; metabolism ; pathology ; Gene Expression Regulation, Neoplastic ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Neoplasm Metastasis ; RNA, Messenger ; genetics ; Signal Transduction ; drug effects ; Transcription Factors ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism ; Veratrum Alkaloids ; pharmacology ; Zinc Finger Protein GLI1

OBJECTIVETo investigate the effect of Helicobacter Pylori lipopolysaccharide (Hp-LPS) on expression of Gli and Ptch-1 proteins in sonic hedgehog (Shh) signaling pathway of gastric mucosa GES-1 cells.

METHODSThe LPS was extracted from Hp by hot phenol water method, and then the concentration of LPS was detected by the kinetic turbidimetric assay. GES-1 cells were stimulated by different concentrations of Hp-LPS (0, 1, 10, 20, 30 and 40 μg/ml). The inhibition rates of cell growth were measured by MTT assay after treated with Hp-LPS for 24 h. The expression of Gli and Ptch-1 proteins were determined by Western Blot.

RESULTSMTT assay showed that the inhibition rates of GES-1 cell growth after treatment by different concentrations of Hp-LPS (1, 10, 20, 30 and 40μg/ml) were 25.8% ± 2.7%, 34.2% ± 3.1 %, 46.3% 3.4%, 60.8% ± 2.1% and 82.9% ± 2.8% respectively (r=0.985, P<0.001). Western blot showed that the expressions of Gli and Ptch-1 proteins were decreased after Hp-LPS treatment (0, 1, 10, 20, 30 and 40 μg/ml): the relative expression values of Gli were 1.286 ± 0.180, 0.963 ± 0.067, 0.850 ± 0.085, 0.566 ± 0.058, 0.549 ± 0.056 and 0.377 ± 0.047, respectively (r=-0.945, P<0.001); those of Ptch-1 were 1.688 ± 0.088, 1.466 ± 0.061, 1.170 ± 0.065, 1.042 ± 0.064, 0.648 ± 0.057 and 0.482 ± 0.074, respectively (r=-0.985, P<0.001).

CONCLUSIONHp-LPS can decrease the related protein expression of Shh signaling pathway, which indicates that Hp may interfere with the function of Shh signaling pathway in gastric mucosa via the effect of its LPS.

Cells, Cultured ; Epithelial Cells ; drug effects ; Gastric Mucosa ; cytology ; Hedgehog Proteins ; metabolism ; Humans ; Lipopolysaccharides ; administration & dosage ; pharmacology ; Patched Receptors ; Patched-1 Receptor ; Receptors, Cell Surface ; metabolism ; Signal Transduction ; Transcription Factors ; metabolism ; Zinc Finger Protein GLI1

OBJECTIVE: To explore the regulatory role of SOX2-OT in migration of lung squamous cell carcinoma H520 cells and the underlying mechanisms. METHODS: Wound- healing and Transwell migration assays were performed to examine the changes in migration and invasion capacity of lung squamous cell line H520, which expressed higher levels of SOX2-OT than other lung cancer cell lines, following RNA interference-mediated SOX2-OT knockdown. The transcription levels of epithelial-mesenchymal transition (EMT)-related components was detected by qRT-PCR and immunoblotting. Gli1 gain-of-function analysis was performed in H520 cells with SOX2-OT knockdown and the changes in EMT phenotype of the cells were examined. miR-200c mimic and inhibitor were used to analyze the mechanism by which SOX2-OT positively regulates Gli1 and the mediating role of SOX2. RESULTS: SOX2-OT knockdown significantly lowered the invasiveness and migration capacity of H520 cells and caused changes in EMT phenotype of the cells. Overexpression of Gli1, which was positively regulated by SOX2-OT, reversed the inhibitory effect of SOX2-OT knockdown on migration of H520 cells. Transfection of the cells with miR-200c inhibitor effectively reversed SOX2-OT knockdown-induced down-regulation of SOX2. CONCLUSION The SOX2-OT/SOX2 axis positively regulates migration of lung squamous H520 cells via Gli1-mediated EMT.
Humans ; Epithelial-Mesenchymal Transition/genetics* ; Zinc Finger Protein GLI1/metabolism* ; Cell Line, Tumor ; Cell Movement/genetics* ; Lung Neoplasms/genetics* ; MicroRNAs/metabolism* ; Gene Expression Regulation, Neoplastic ; Cell Proliferation/genetics* ; Neoplasm Invasiveness/genetics* ; SOXB1 Transcription Factors/metabolism*

OBJECTIVETo investigate whether the sonic hedgehog signaling pathway is involved in the development of human fetal prostate, and to evaluate the changing staining patterns of its molecules, sonic hedgehog (SHH), patchedl (PTC1), smoothened (SMO), and GLI1, in the human fetal prostate at various gestation stages.

METHODSFifteen human fetal prostate specimens at various developmental stages (10 - 39 weeks) were included in this study. SHH, PTC1, SMO and GLI1 were detected in all the specimens by immunohistochemical technique. All the slides were observed and assessed under the light microscope.

RESULTSSHH, PTC1, SMO and GLI1 could be detected in human fetal prostate tissues, and their expression formed two surges, the former at week 16, and the latter at week 28. The staining of SHH and SMO was distributed only in the ductal epithelium but not in the stroma. The expression of PTC1 and GLI1 could be found mainly in the epithelium, with minimal staining in the stroma.

CONCLUSIONThe sonic hedgehog signaling pathway is involved in the development of the human fetal prostate. The high expression of its molecules at early gestation stages might be associated with the induction of prostatic buds, while their abundant expression at later gestation stages might be related to the prostate ductal branching, growth, differentiation and morphogenesis.

Gene Expression Regulation, Developmental ; physiology ; Hedgehog Proteins ; biosynthesis ; Humans ; Male ; Oncogene Proteins ; biosynthesis ; Patched Receptors ; Prostate ; embryology ; metabolism ; Receptors, Cell Surface ; biosynthesis ; Receptors, G-Protein-Coupled ; biosynthesis ; Signal Transduction ; physiology ; Smoothened Receptor ; Trans-Activators ; biosynthesis ; Zinc Finger Protein GLI1

OBJECTIVETo study the role and possible mechanism of glioma-associated oncogene-1 (Gli1) in regulating the cell invasion and migration of pancreatic cancer cells.

METHODSQuantitative real-time (qRT) -PCR was used to detect the effect of siRNA interference on Gli1, murine double minute 2 (MDM2) and p53 genes. Cell invasion and migration assays were used to observe the effect of Gli1, MDM2 and p53 silence on cell invasion and migration in p53 wild-type Capan-2 pancreatic cancer cells, respectively. Meanwhile, immunoblotting (IB) was used to detect the protein level of matrix metalloproteinase (MMP) -9, phospho-excelluar signal-regulated kinase (pERK) and phosphorylation protein kinase B (pAKT) in Gli1-silencing Capan-2 cells. The data were analyzed by paired t-test.

RESULTSqRT-PCR showed that the expression of Gli1, MDM2 and p53 is down-regulated 70.5% and 74.5%, 61.8% and 65.3%, and 73.8% and 78.2% after siRNA interference, compared with the mock and siRNA control groups, respectively. Cell invasion (94 ± 8) and migration (143 ± 8) in p53 wild-type Capan-2 cells transfected with Gli1siRNA were significantly decreased, compared with the siRNA control group (150 ± 7, 190 ± 10) (t = 6.584, P = 0.022; t = 8.266, P = 0.014) , while MDM2 silence inhibited cell invasion (experiment group:85 ± 12, control group: 138 ± 6) and migration (experiment group: 127 ± 9, control group:180 ± 10) in the same cells, respectively (t = 5.097, P = 0.036;t = 4.860, P = 0.040). However, cell invasion (experiment group: 153 ± 11, control group: 106 ± 7) and migration (experiment group: 209 ± 13, control group: 164 ± 8) in p53-silencing Capan-2 cells were significantly enhanced (t = 4.669, P = 0.043; t = 4.990, P = 0.038). IB showed that Gli1 silence down-regulated MMP-9 but not pERK and pAKT protein expression.

CONCLUSIONGli1 might contribute to the cell invasion and migration in pancreatic cancer via the regulation of MDM2, p53 and MMP-9 expression.

Animals ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation ; Matrix Metalloproteinase 9 ; metabolism ; Mice ; Neoplasm Invasiveness ; Oncogene Proteins ; genetics ; metabolism ; Pancreas ; metabolism ; Pancreatic Neoplasms ; metabolism ; pathology ; Proto-Oncogene Proteins c-akt ; metabolism ; Proto-Oncogene Proteins c-mdm2 ; metabolism ; RNA, Small Interfering ; genetics ; Trans-Activators ; genetics ; metabolism ; Transfection ; Tumor Suppressor Protein p53 ; metabolism ; Zinc Finger Protein GLI1

OBJECTIVETo study the clinicopathological significance and relationship of Gli1, MDM2 and p53 expression in human pancreatic cancer.

METHODSThe expression of Gli1, MDM2 and p53 proteins in 57 paired paraffin embedded pancreatic ductal adenocarcinoma (PDAC) specimens and adjacent non-cancerous pancreatic tissues was detected by immunohistochemistry. The relationship between their expression and clinicopathological characters was analyzed. Quantitative real-time PCR (qRT-PCR) was used to examine the expression of Gli1 mRNA level in 14 paired fresh PDAC specimens and adjacent non-cancerous pancreatic tissues. siRNA interference were used to further detect the close relationship among them.

RESULTSIHC showed the expression of Gli1 (50.9%), MDM2 (57.9%) and p53 (56.1%) was increased in 57 cases of pancreatic cancer compared to that in paired normal pancreatic tissues (33.3%, 26.3% and 17.5% respectively, t = 2.413, 2.848 and 2.960, all P < 0.05). Gli1 expression was positively associated with tumor TNM stage (χ(2) = 8.211, P = 0.004), invasion depth (χ(2) = 4.247, P = 0.039) and MDM2 expression (r = 0.299, χ(2) = 5.105, P = 0.024), while expression of MDM2 and p53 was associated with tumor invasion depth (χ(2) = 5.182, P = 0.023) and TNM stage (χ(2) = 5.696, P = 0.017), respectively. Univariate and multivariate analysis revealed that Gli1 was an independent adverse prognostic indicator for patients with PDAC (RR = 2.290, 95%CI: 1.051-4.992, P = 0.037), and patients with Gli1 and MDM2 co-expression had a significantly poorer overall survival than patients with their negative expression (P = 0.034). Gli1 mRNA expression was much higher in 14 cases of PDAC than that in adjacent normal pancreatic tissues (t = 2.926, P = 0.012). In p53 mutant AsPC-1 cells, Gli1 knockdown down regulated MDM2, but had no effect on p53 expression, whereas Gli1 knockdown down regulated MDM2 and up regulated p53 protein levels in p53 wild-type Capan-2 cells.

CONCLUSIONSGli1, MDM2 and p53 are overexpressed in PDAC, and are benefit for predicting patients' prognosis. Gli1can regulate MDM2 and wild-type p53 expression. Their co-expression might coordinately contribute to the development and progression of PDAC.

Adult ; Aged ; Carcinoma, Pancreatic Ductal ; metabolism ; Female ; Gene Expression Regulation, Neoplastic ; Humans ; Male ; Middle Aged ; Oncogene Proteins ; metabolism ; Pancreatic Neoplasms ; metabolism ; Prognosis ; Proto-Oncogene Proteins c-mdm2 ; metabolism ; RNA, Messenger ; metabolism ; Trans-Activators ; metabolism ; Tumor Suppressor Protein p53 ; metabolism ; Zinc Finger Protein GLI1