Differential Expression Of Osteopontin Research Articles

Differential osteopontin expression in human osteoblasts derived from iliac crest and alveolar bone and its role in early stages of angiogenesis.

In our previous study, we revealed significant differences of osteopontin (OPN) gene expression in primary human osteoblasts (HOBs) derived from iliac crest bone (iHOBs) and alveolar bone (aHOBs). The present study aims at assigning this discriminative expression to a possible biologic function. OPN is known to be involved in several pathologic and physiologic processes, among others angiogenesis. Therefore, we studied the reaction of human umbilical vein endothelial cells (HUVECs) to HOB-derived OPN regarding angiogenesis. To this end, human primary explant cultures of both bone entities from ten donors were established. Subsequent transcription analysis detected higher gene expression of OPN in iHOBs compared to aHOBs, thereby confirming the results of our previous study. This difference was particularly apparent when cultures were derived from female donors. Hence, OPN protein expression as well as the angiogenic potential of OPN was analyzed, originating from HOBs of one female donor. In accordance to the gene expression level, secreted OPN was more abundant in the supernatant of iHOBs than in aHOBs. Moreover, secreted OPN was found to stimulate migration of HUVECs, but not proliferation or tube formation. These results indicate an involvement in very early stages of angiogenesis and a functional distinction of OPN from HOBs derived from different bone entities.

Differential expression of osteopontin, and osteoprotegerin mRNA in epicardial adipose tissue between patients with severe coronary artery disease and aortic valvular stenosis: association with HDL subclasses

BackgroundPrevious studies suggest a relationship of the epicardial adipose tissue (EAT) with progression and calcification of the atherosclerotic plaque; however, it is unknown if this tissue expresses genes that may participate on these processes and if the expression of these genes is regulated by high-density lipoprotein (HDL) subclasses.MethodsTo explore this possibility, we determined the mRNA expression by qPCR of a pro-calcifying gene (osteopontin (OPN)), and two anti-calcifying genes (osteoprotegerin (OPG) and osteonectin (ON)), in biopsies of EAT obtained from 15 patients with coronary artery disease (CAD) determined by angiography, and 15 patients with diagnostic of aortic valve stenosis but without CAD as control group. We determined the distribution and composition of HDL subclasses by electrophoresis and their statistical relationship with the gene expression in EAT.ResultsEAT from CAD patients showed a higher expression level of OPN and OPG than control group, whereas ON expression was similar between groups. Large HDL subclasses were cholesterol-poor in CAD patients as estimated by the cholesterol-to-phospholipid ratio. A linear regression model showed an independent association of OPN expression with HDL3a-cholesterol, and OPG expression with the relative proportion of HDL3b protein. Logistic analysis determined that OPN expression was positively associated with the presence of atherosclerotic plaqueConclusionOPN, ON, and OPG genes are transcribed in EAT; to the exception of ON, the level of expression was different in CAD patients and control group, and correlated with some HDL subclasses, suggesting a new role of these lipoproteins.

Gene expression profiling demonstrates differential expression of osteopontin in follicular thyroid carcinomas compared to normal thyroid tissue in dogs

Follicular thyroid carcinoma (FTC) is an aggressive tumour in dogs with little known about its molecular pathogenesis. The overall goal of this study was to examine FTC and normal thyroid tissue gene expression. Microarray analysis was performed on a pilot group of five FTC-affected dogs and four healthy dogs, and then osteopontin validated with quantitative polymerase chain reaction (PCR) and immunohistochemistry (IHC) of thyroid tissue from non-invasive FTC, invasive FTC and healthy dogs. On microarray analysis, 489 transcripts were differentially expressed between FTC and normal thyroid: 242 transcripts were down-regulated and 247 were up-regulated. Osteopontin expression was markedly increased in tumour tissue compared to normal thyroid tissue. Quantitative PCR and IHC confirmed differential expression of osteopontin in both tumour types (invasive and non-invasive) compared to normal thyroid tissue. There is justification for further investigation of osteopontin as a potential molecular marker for screening and monitoring of canine FTC.

Up-regulation of osteopontin in aortic dissection

Objective To identify the differential expression of osteopontin (OPN) between the aorta specimens from the aortic dissection (AD) and the normal controls. Methods Twelve descending aorta specimens from patients with acute Stanford B dissection were taken during surgical operation, and 12 normal descending aorta specimens were taken from multi-organ donors. The GAPDH gene was used as control, and mRNA and protein expression of OPN was detected by using real-time polymerase chain reaction (PCR) and immunohistochemisty, respectively. Three low-powered fields of each specimen in beth groups were randomly chosen to count the cells positive for OPN protein. Results The expression levels of OPN in AD group was 5. 56 folds as those in the control group (P ＜0. 0l ). The protein expression levels of OPN was 3. 03 folds as those in the control group ( P ＜ 0. 01 ). Conclusion The gene and protein expression levels of OPN were down-regulated in AD group as compared with normal controls. OPN is one of the markers to identify smooth muscle cells as synthetic phenotype. The up-regulation of OPN may contribute to the development of AD. Key words: Aortic dissection; Osteopontin; Etiology

Nectin-like Molecule 1 Inhibits the Migration and Invasion of U251 Glioma Cells by Regulating the Expression of An Extracellular Matrix Protein Osteopontin

To investigate the molecular mechanism of nectin-like molecule 1 (NECL1) inhibiting the migration and invasion of U251 glioma cells. We infected U251 glioma cells with adeno-nectin-like molecule 1 (Ad-NECL1) or empty adenovirus (Ad). Transwell and wound healing assays were performed to observe the migration of U251 cells incubated with the cell supernatant from Ad-NECL1 or Ad infected U251 cells. DNA microarray was applied to screen the gene expression profile after the restoration of NECL1 in U251 glioma cell lines. The differential expression of osteopontin (OPN), a gene related to migration and invasion, was further analyzed with semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), Western blot, and immunohistochemistry. The restoration of NECL1 inhibited migration of U251 cells significantly (P<0.05). Altogether 195 genes were found differentially expressed by microarray, in which 175 were up-regulated and 20 down-regulated, including 9 extracellular matrix proteins involved in the migration of cells. Both mRNA and protein expressions of OPN, the most markedly reduced extracellular matrix protein, were found decreased in U251 cells after restoration of NECL1. Immunohistochemical assay also detected an increase of OPN in glioma tissues, related with the progressing of malignant grade. A link might exist between NECL1 and the extracellular matrix protein OPN in inhibiting the migration and invasion of U251 glioma cells.

The implication of osteopontin ( OPN) expression and genetic polymorphisms of OPN promoter in oral carcinogenesis

Osteopontin (OPN) is an extracellular matrix protein in hard tissues. The polymorphism in promoter region of OPN gene correlates to different gene expression and might implicate potential roles in tumor progression and metastasis. Immunohistochemistry (IHC) was utilized to detect the OPN expression in 58 oral squamous cell carcinoma (OSCC) tissues and adjacent normal oral mucosa. The differential OPN expression was further analyzed in relation to clinico-pathological features. Genomic DNA was obtained from isolated leukocytes of blood samples of OSCC patients (n=100), and healthy individuals (n=97) from Taiwan. The OPN gene polymorphism was analyzed by direct sequencing. Our result showed OPN expression was significantly higher in OSCC tissues than in the paired adjacent normal tissues (p<0.01). The expression of OPN was significantly associated with nodal metastasis and the more advanced clinical stage (p<0.05). More prevalent -156 insGG/insGG genotype and -443 T/T genotype was found in OSCC patients (p<0.05). A significant difference in -443T/-156GG/-66T and -443C/-156G/-66T haplotypes between OSCC and controls (p<0.05) was also noted. The OPN expression in tumor tissues significantly correlated with -156 insGG/insGG and -156 G/G+insGG/G genotypes (p<0.05). The conclusion is tissue OPN expression correlates to OSCC progression. -156 insGG/insGG genotype is associated with OSCC susceptibility and higher OPN expression.

EF1A1-actin interactions alter mRNA stability to determine differential osteopontin expression in HepG2 and Hep3B cells

Cancer progression depends on an accumulation of metastasis-supporting physiological changes which are regulated by cell signaling molecules. One such molecule, osteopontin (OPN), is a secreted phosphoprotein which mediates increased cellular migratory and invasive behavior, increased metastasis, protection from apoptosis, promotion of colony formation and 3D growth ability, induction of tumor-associated inflammatory cells, and induction of expression of angiogenic factors. Studies show that OPN expression is controlled by complex regulatory pathways at the transcriptional level in several cancers, but the molecular mechanisms which determine expression of OPN in HCC are largely unknown. In HepG2 and Hep3B tumor cell lines that differentially express OPN mRNA and protein, we identify elongation translation factor-1A1 (EF1A1) to be the trans-acting factor regulating differential OPN mRNA stability between HepG2 and Hep3B cell lines and characterize its interactions with G- and F-actin. EF1A1 binds to the OPN 5′-UTR to regulate OPN mRNA half-life. EF1A1 binds to actin in Hep3B cells. Pharmacologic manipulation to increase the G:F actin ratio in Hep3B increases OPN mRNA half-life and protein expression with simultaneous decrease in EF1A1 binding to OPN 5′-UTR. The converse findings were noted in HepG2 cells. Overall, our results suggest that EF1A1 regulation of OPN mRNA stability is actin dependent. EF1A1 has not been previously identified as a regulatory factor in OPN expression in cancer.

Osteopontin Expression According to Molecular Profile of Invasive Breast Cancer: A Clinicopathological and Immunohistochemical Study

Osteopontin (OPN) is a secreted, calcium-binding phosphorylated glycoprotein involved in several physiological and pathological events such as angiogenesis, apoptosis, inflammation, wound healing, vascular remodeling, calcification of mineralized tissues, and induction of cell proteases. There is growing interest in the role of OPN in breast cancer. In an attempt to obtain new insight into the pathogenesis of OPN-associated breast carcinomas, an immunohistochemical panel with 17 primary antibodies including cytokeratins and key regulators of the cell cycle was performed in 100 formalin-fixed paraffin-embedded samples of invasive breast carcinomas. OPN was expressed in 65% of tumors and was negatively correlated with estrogen (p=0.0350) and progesterone (p=0.0069) receptors, but not with the other markers and clinicopathological features evaluated including age, menstrual status, pathological grading, tumor size, and metastasis. There was no correlation between OPN expression and carcinomas of the basal-like phenotype (p=0.1615); however, OPN correlated positively with c-erbB-2 status (p=0.0286) and negatively with carcinomas of the luminal subtype (p=0.0353). It is well known that carcinomas overexpressing c-erbB-2 protein have a worse prognosis than luminal tumors. Here, we hypothesize that the differential expression of OPN in the first subtype of carcinomas may contribute to their more aggressive behavior.

High-resolution analysis of genomic profiles of hepatocellular carcinoma cells with differential osteopontin expression

Overexpression of osteopontin (OPN) could contribute to tumorigenesis and metastasis in hepatocellular carcinoma (HCC). Previous studies have shown that OPN-positive cancer cells are often localized in the periphery of cancer nodules adjacent to stromal cells. To identify the difference in intratumor genomic aberration pattern between OPN-positive and OPN-negative HCC cells, we adopted microarray-based comparative genomic hybridization (array-CGH) to achieve high-resolution analysis of genome-wide aberrations. Our present study indicates that, compared to OPN-negative cancer cells, OPN-positive cancer cells show much more amplification of chromosomal regions, including 4q13.1-q13.3, 4q21.23-q22.1 and 13q32.1-q32.3. Some candidate tumor related genes, such as SMR3B, MUC7, EPHA5, SPP1 and CLDN10, were detected with over 1.5-fold amplification. Together, the intratumor genomic heterogeneity may suggest that OPN-positive cancer cells play a more important role in the development of HCC malignancy than their OPN-negative counterparts.

Osteopontin expression according to molecular profile of invasive breast cancer: a clinicopathological and immunohistochemical study

Osteopontin (OPN) is a secreted, calcium-binding phosphorylated glycoprotein involved in several physiological and pathological events such as angiogenesis, apoptosis, inflammation, wound healing, vascular remodeling, calcification of mineralized tissues, and induction of cell proteases. There is growing interest in the role of OPN in breast cancer. In an attempt to obtain new insight into the pathogenesis of OPN-associated breast carcinomas, an immunohistochemical panel with 17 primary antibodies including cytokeratins and key regulators of the cell cycle was performed in 100 formalin-fixed paraffin-embedded samples of invasive breast carcinomas. OPN was expressed in 65% of tumors and was negatively correlated with estrogen (p=0.0350) and progesterone (p=0.0069) receptors, but not with the other markers and clinicopathological features evaluated including age, menstrual status, pathological grading, tumor size, and metastasis. There was no correlation between OPN expression and carcinomas of the basal-like phenotype (p=0.1615); however, OPN correlated positively with c-erbB-2 status (p=0.0286) and negatively with carcinomas of the luminal subtype (p=0.0353). It is well known that carcinomas overexpressing c-erbB-2 protein have a worse prognosis than luminal tumors. Here, we hypothesize that the differential expression of OPN in the first subtype of carcinomas may contribute to their more aggressive behavior.

Osteopontin Expression Correlates with Melanoma Invasion

Melanoma is one of the most aggressive cancers affecting humans. Although early melanomas are curable with surgical excision, metastatic melanomas are associated with high mortality. The mechanism of melanoma development, progression, and metastasis is largely unknown. In order to uncover genes unique to melanoma cells, we used high-density DNA microarrays to examine the gene expression profiles of metastatic melanoma nodules using benign nevi as controls. Over 190 genes were significantly overexpressed in metastatic melanomas compared with normal nevi by at least 2-fold. One of the most abundantly expressed genes in metastatic melanoma nodules is osteopontin (OPN). Immunohistochemistry staining on tissue microarrays and individual skin biopsies representing different stages of melanoma progression revealed that OPN expression is first acquired at the step of melanoma tissue invasion. In addition, blocking of OPN expression by RNA interference reduced melanoma cell numbers in vitro. Our observations suggest that OPN may be acquired early in melanoma development and progression, and may enhance tumor cell growth in invasive melanoma.

Differential expression of osteopontin and bax, bcl-2 by fluoride

To study the differential expression of bax, bcl-2 and osteopontin by fluoride in the renal tubular cells in vitro. The renal tubular cells were cultured and exposed to sodium fluoride (NaF) in 1, 5, 7.5, 12.5 mg F-/L level. The transcription level of bax, bcl-2 and osteopontin were investigated by reverse transcription - polymerase chain reaction (RT-PCR). The expression of bax mRNA in 7.5 and 12.5 mgF-/L groups (optical absorption ratio value was 2.37 +/- 0.18 and 2.64 +/- 0.19 respectively) was significantly increased (P < 0.01). On the contrary, the level of bcl-2 obviously decreased (5 mg F-/L group optical absorption ratio value, 0.80 +/- 0.22, P < 0.05; 7.5 mg F-/L group optical absorption ratio value 0.71 +/- 0.22, P < 0.01). The expression mRNA of osteopontin was significantly increased when cells were exposed to fluoride at 7.5 mg F-/L (optical absorption ratio value 2.01 +/- 0.40 P < 0.01), in that group the tubular cell apoptotic trend was obvious. NaF might induce tubular cell apoptosis via activation of bax expression and bcl-2 suppression. Osteopontin might protect the tubule against apoptosis in a lower fluoride level, but the function should be decreased in higher fluoride level.

Differential Osteopontin Expression in Phenotypically Distinct Subclones of Murine Breast Cancer Cells Mediates Metastatic Behavior

Cancer progression depends on an accumulation of metastasis-supporting cell signaling molecules, which target signal transduction pathways and, ultimately, gene expression. One such molecule, osteopontin (OPN), represents a key molecular signaling event in tumor progression and metastasis. However, the transcriptional regulatory mechanisms that underlie OPN expression in the setting of breast cancer have not been well studied. In this regard, we have examined the differential transcriptional regulation of OPN in the murine mammary epithelial tumor cell lines, 4T1 and 4T07, which are sublines derived from the parental population of 410.4 cells from Balb/cfC3H mice. These lines are phenotypically heterogeneous in their metastatic behavior. 4T1 hematogenously metastasizes to the lung, liver, bone, and brain, whereas 4T07 is highly tumorigenic but fails to metastasize. The tumor growth and metastatic spread of 4T1 cells closely mimics stage IV breast cancer. We demonstrate that a Ras-independent, phosphoinositide-3 kinase-dependent, c-Jun N-terminal kinase-dependent phosphorylation of c-Jun results in binding of an AP-1 c-Jun homodimer to the OPN promoter in 4T1 cells. This differential up-regulation of OPN gene transcription and protein expression in 4T1 cells conveys in vitro correlates of a metastatic phenotype. These results provide new insight into the transcriptional regulation of OPN as a key mediator of metastatic behavior in malignancy.

Differential osteopontin expression in lung cancer

Osteopontin (OPN) is a phosphorylated glycoprotein with diverse functions including cancer development, progression and metastasis. Its expression is induced by a variety of stimuli such as TNF-α and Ras proto-oncogene. However, differential OPN expression and its regulation in each histologic type of lung cancer are not well established. In this study, we assessed expression of OPN in lung cancer tissues with immunohistochemical analysis. OPN was predominantly expressed in tumor cells of non-small cell lung cancer (NSCLC) tissues: 11 of 16 cases (68.8%) of squamous cell carcinoma (SCC), five of 24 cases (20.8%) of adenocarcinoma (AD), but only two of 18 cases (11%) of small cell lung cancer (SCLC). Expectedly, OPN was principally expressed in NSCLC cell lines (H322 cells and HL460 cells) but not in SCLC cell line (H69 cells) by Western blotting and Northern blotting. Interestingly, Ras-p21 was specifically co-expressed with OPN staining in eight of eight cases with SCC (100%), whereas it was demonstrated in three of ten cases (30%) with AD and only one of 18 cases (5%) with SCLC. Collectively, these results suggest that OPN is mainly expressed in NSCLC, especially among SCC. OPN expression may be tightly regulated by Ras oncogene, and its concomitant induction with Ras activation may play a crucial role in the development of SCC.

Differential Expression of Osteopontin, PC4, and CEC5, a Novel mRNA Species, duringin VitroAngiogenesis

The formation of new capillaries from preexisting blood vessels, a process termed angiogenesis, plays a key role in many physiological and pathological conditions such as wound healing, embryogenesis, and tumor growth. The aim of this study was to identify changes in endothelial cell gene expression specifically associated with angiogenesis. Using anin vitromodel and the differential display strategy, we compared gene expression patterns of rat microvascular endothelial cells cultured in two (2D) and three-dimensional (3D) culture. In 2D culture, the cells express actin and proliferate, whereas in 3D culture actin expression is downregulated, and the cells are mitotically quiescent and reorganize into vascular tubes. We identified three differentially expressed genes,osteopontin, PC4, and CEC5,a novel mRNA species, with homology to calmodulin-dependent protein kinases. The expression patterns were confirmed by Northern blot analysis. In conclusion, the analysis of gene expression in endothelial cells in 2D and 3D culture allows the identification of genes differentially expressed during angiogenesis. These genes or proteins may serve as targets for therapeutic modulation of angiogenesis.

CAMP regulates the differential expression of osteopontin by cells in human endometrium.

cAMP regulates the differential expression of osteopontin by cells in human endometrium.