Secreted Phosphoprotein 1 Expression Research Articles

Association of SPP1 promoter variants with hip osteoarthritis susceptibility in Chinese population

Three genetic variants in the promoter of SPP1 (secreted phosphoprotein 1) gene have been reported to affect transcriptional activity of SPP1, thus conferring an increased risk for some diseases. To testify if these variants are associated with risk of hip osteoarthritis (OA) as well, we performed a case–control study including 389 hip OA patients and 315 healthy controls. Genotypes of SPP1 were determined by DNA sequencing, and differential expressions of SPP1 in relation with genotypes were evaluated by RT-PCR and ELISA. The results showed that rs17524488 (delG>insG) increased the risk of hip OA, with the adjusted OR 1.48 (95% CI 1.18–1.85, P<0.01) for risk allele insG, 1.90 (95% CI 1.35–2.66, P<0.01) for delG/insG and 2.04 (95% CI 1.20–3.49, P<0.01) for insG/insG respectively. However, as for rs11730582 (T>C), the adjusted ORs were 1.18 (95% CI 0.94–1.49, P=0.148) for allele C, 1.26 (95% CI 0.90–1.75, P=0.158) for TC, and 1.31 (95% CI 0.77–2.24, P=0.293) for CC, indicating no association of rs11730582 with hip OA risk. The variant rs28357094 was not observed in the tested subjects. Furthermore, the delG/insG and insG/insG genotypes of rs17524488 both correlated with higher levels of SPP1 expression in articular cartilage (P<0.01 for all comparisons) as well as in in synovial fluid (P<0.01 for all comparisons) compared with delG/delG, while rs11730582 had no effect on the SPP1 expression (P>0.05 for all comparisons). These results collectively indicate that the genetic variant rs17524488 in SPP1 promoter confers high risk for hip OA in a Chinese population, possibly through enhancing SPP1 expression.

Expression and regulation of secreted phosphoprotein 1 in the bovine corpus luteum and effects on T lymphocyte chemotaxis

Secreted phosphoprotein 1 (SPP1) in the bovine corpus luteum (CL) regulates cell function during the transitional periods of luteinization and luteal regression. The objectives were to i) characterize SPP1 expression in the CL throughout the estrous cycle, ii) determine factors that regulate SPP1 expression in luteal cells, and iii) examine the role of SPP1 in lymphocyte chemotaxis, proliferation, and function. SPP1 mRNA was greater in fully functional (d10) CL and late cycle (d18) CL compared with developing (d4) CL. Additionally, SPP1 mRNA increased within 1 h and remained elevated 4 and 8 h following induction of luteolysis with prostaglandin (PG)F2α. Expression of the SPP1 receptor, β3 integrin, was not different throughout the estrous cycle but decreased following induction of luteolysis. Expression of CD44 increased during the estrous cycle but did not change during luteal regression. In cultured luteal cells, SPP1 mRNA was upregulated by PGF2α and/or tumor necrosis factor α. Western blots revealed the presence of both full-length SPP1 and multiple cleavage products in cultured luteal cells and luteal tissue. Depletion of endogenous SPP1 did not hinder luteal cell-induced lymphocyte proliferation or lymphocyte phenotype but did inhibit lymphocyte migration toward luteal cells. Based on these data, it is concluded that SPP1 is initially activated to establish and maintain cellular interactions between steroidogenic and nonsteroidogenic cells during the development of the CL. Upon induction of luteolysis, SPP1 serves as a signaling molecule to recruit or activate immune cells to facilitate luteal regression and tissue degradation.

Differential Expression of Secreted Phosphoprotein 1 in the Motor Cortex among Primate Species and during Postnatal Development and Functional Recovery

We previously reported that secreted phosphoprotein 1 (SPP1) mRNA is expressed in neurons whose axons form the corticospinal tract (CST) of the rhesus macaque, but not in the corresponding neurons of the marmoset and rat. This suggests that SPP1 expression is involved in the functional or structural specialization of highly developed corticospinal systems in certain primate species. To further examine this hypothesis, we evaluated the expression of SPP1 mRNA in the motor cortex from three viewpoints: species differences, postnatal development, and functional/structural changes of the CST after a lesion of the lateral CST (l-CST) at the mid-cervical level. The density of SPP1-positive neurons in layer V of the primary motor cortex (M1) was much greater in species with highly developed corticospinal systems (i.e., rhesus macaque, capuchin monkey, and humans) than in those with less developed corticospinal systems (i.e., squirrel monkey, marmoset, and rat). SPP1-positive neurons in the macaque monkey M1 increased logarithmically in layer V during postnatal development, following a time course consistent with the increase in conduction velocity of the CST. After an l-CST lesion, SPP1-positive neurons increased in layer V of the ventral premotor cortex, in which compensatory changes in CST function/structure may occur, which positively correlated with the extent of finger dexterity recovery. These results further support the concept that the expression of SPP1 may reflect functional or structural specialization of highly developed corticospinal systems in certain primate species.

Abstract 395: Farnesyl diphosphate farnesyltransferase induces metastasis via upregulating secreted phosphoprotein 1 in non-small cell lung cancer.

Abstract Identifying cancer progression associated genes that predicting patient outcome might provide opportunities for developing better therapeutic strategies for lung cancer patients. In the present study, we investigated whether farnesyl diphosphate farnesyltransferase (FDFT1), an enzyme of mevalonate pathway, is a potential biomarker of lung cancer metastasis. We further investigated its prognostic value and functional role in lung cancer invasion and metastasis. Our results by immunohistochemistry (IHC) analysis showed the expression of FDFT1 was significantly higher in tumors compared to normal lung tissues. Overexpression of FDFT1 was significantly associated with tumor size (P &amp;lt; 0.001), advanced tumor stage (P &amp;lt;0.001), lymph node metastasis (P = 0.02), distal metastasis (P = 0.043), and poor prognosis (P &amp;lt;0.001). Loss of FDFT1 expression significantly suppressed invasion and migration potential of lung cancer cell lines, whereas FDFT1 overexpression promoted invasion and migration in vitro and in vivo. Microarray analysis further identified secreted phosphoprotein 1 (SPP1) as an important mediator for FDFT1-induced invasion and migration. Furthermore, patients with high FDFT1 and high SPP1 expression showed dismal outcomes as compared to patients with low FDFT1 and low SPP1 expression with good prognosis. In conclusion, we identified the FDFT1 in this study as a new lung cancer progression biomarker that is associated poor prognosis and its overexpression resulted in enhanced tumor invasion and metastasis through upregulation of SPP1. Our findings may implicate the possibility of targeting FDFT1 as a novel therapeutic strategy to inhibit lung cancer metastasis. Citation Format: Yi-Fang Yang, Yi-Hua Jan, Michael Hsiao. Farnesyl diphosphate farnesyltransferase induces metastasis via upregulating secreted phosphoprotein 1 in non-small cell lung cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 395. doi:10.1158/1538-7445.AM2013-395 Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.

Knockdown of prolyl‐4‐hydroxylase domain 2 inhibits tumor growth of human breast cancer MDA‐MB‐231 cells by affecting TGF‐β1 processing

The prolyl-4-hydroxylase domain 1-3 (PHD1-3) enzymes are regulating the protein stability of the α-subunit of the hypoxia-inducible factor-1 (HIF-1), which mediates oxygen-dependent gene expression. PHD2 is the main isoform regulating HIF-1α hydroxylation and thus stability in normoxia. In human cancers, HIF-1α is overexpressed as a result of intratumoral hypoxia which in turn promotes tumor progression. The role of PHD2 for tumor progression is in contrast far from being thoroughly understood. Therefore, we established PHD2 knockdown clones of MDA-MB-231 breast cancer cells and analyzed their tumor-forming potential in a SCID mouse model. Tumor progression was significantly impaired in the PHD2 knockdown MDA-MB-231 cells, which could be partially rescued by re-establishing PHD2 expression. In a RNA profile screen, we identified the secreted phosphoprotein 1 (SPP1) as one target, which is differentially regulated as a consequence of the PHD2 knockdown. Knockdown of PHD2 drastically reduced the SPP1 expression in MDA-MB-231 cells. A correlation of SPP1 and PHD2 expression was additionally verified in 294 invasive breast cancer biopsies. In subsequent analyses, we identified that PHD2 alters the processing of transforming growth factor (TGF)-β1, which is highly involved in SPP1 expression. The altered processing capacity was associated with a dislocation of the pro-protein convertase furin. Thus, our data demonstrate that in MDA-MB-231 cells PHD2 might affect tumor-relevant TGF-β1 target gene expression by altering the TGF-β1 processing capacity.

Differential expression of secreted phosphoprotein 1 in response to estradiol-17β and in ovarian tumors in chickens

Secreted phosphoprotein 1 (SPP1), a highly phosphorylated protein containing a polyaspartic acid sequence and a conserved RGD motif, plays important roles in physiological processes such as inflammatory responses, calcification, organ development, immune cell function and carcinogenesis. Results of the present study indicate expression of SPP1 mRNA in various organs such as oviduct, small intestine and kidney from chickens, particularly in the glandular epithelium (GE) of the shell gland and, to a lesser extent, in luminal epithelium (LE) of the infundibulum and magnum, and GE of the isthmus of the oviduct. We determined that DES (diethylstilbestrol, a synthetic nonsteroidal estrogen) decreases SPP1 expression in the oviduct and that SPP1 mRNA and protein are significantly more abundant in GE of ovarian endometrioid carcinoma, but not the other cancerous and normal ovaries of hens. Further, microRNA-140 was discovered to influence SPP1 expression via its 3′-UTR which suggests that post-transcriptional regulation influences SPP1 expression in chickens. Collectively, results of this study indicate that SPP1 is novel in that its expression is down-regulated by estrogen in epithelial cells of the chicken oviduct and that it is up-regulated in chicken ovarian endometrioid tumor that could be used for monitoring effects of therapies for this disease in laying hens.

Abstract 5005: Large-scale DNA methylation profiling in gastrointestinal stromal tumors (GIST) reveals epigenetic regulation of SPP1 as an independent prognostic factor

Abstract Aims Gastrointestinal stromal tumors (GISTs) have distinct gene expression patterns according to anatomical localisation and genotype, with potential impact on biological aggressiveness. DNA methylation of CpG dinucleotides in the promoter region of genes is an important mechanism for regulation of gene expression. We performed a large-scale analysis of DNA methylation in GISTs to identify methylation patterns associated with clinico-pathological parameters and prognosis. Methods Seventy-five GISTs with either KIT or PDGFRA mutations were analysed for DNA methylation of 1.505 CpG sites in the promoter regions of 807 cancer-related genes using a large-scale approach (Cancer Methylation Panel I, Illumina). Epigenetic regulation of Secreted Phosphoprotein 1 (SPP1) expression was analysed at mRNA and protein levels in GIST882 and GIST48b cells after treatment with a demethylating agent. Impacts on tumorigenesis-related signalling pathways were analysed by Western Blot after stimulation of GIST cell lines with SPP1. Prognostic impact of SPP1 at methylation and protein levels were investigated in 75 and 121 GIST tumor samples, respectively, and compared to classical prognostic parameters tumor size, mitotic counts and tumor localisation. Results Principal component analysis revealed distinct DNA methylation patterns according to anatomical localisation, genotype and mitotic counts. Hypomethylation of a CpG dinucleotide 647 base pairs upstream of the transcription start site of SPP1 was significantly correlated to SPP1 mRNA and protein level, and also to tumor progression (p=0.003). In vitro analyses confirmed epigenetic regulation of SPP1 expression, and demonstrated potential impact on proliferation and invasion. High protein level of SPP1 was significantly correlated to shorter disease-free survival in univariate (p=2x10−7) and multivariate (p&amp;lt;1x10−6) analyses. Conclusion The expression of SPP1 is regulated epigenetically in GISTs, and high expression of SPP1 is a novel and independent prognostic parameter in GISTs. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5005. doi:1538-7445.AM2012-5005

Molecular signatures and new candidates to target the pathogenesis of rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic, inflammatory joint disease of unknown etiology and pronounced interpatient heterogeneity. To characterize RA at the molecular level and to uncover pathomechanisms, we performed genome-wide gene expression analysis. We identified a set of 1,054 genes significantly deregulated in pair-wise comparisons between RA and osteoarthritis (OA) patients, RA and normal donors (ND), or OA and ND. Correlation analysis revealed gene sets regulated identically in all three groups. As a prominent example secreted phosphoprotein 1 (SPP1) was identified to be significantly upregulated in RA compared with both OA and ND. SPP1 expression was found to correlate with genes expressed during an inflammatory response, T-cell activation and apoptosis, suggesting common underlying regulatory networks. A subclassification of RA patients was achieved on the basis of proteoglycan 4 (PRG4) expression, distinguishing PRG4 high and low expressors and reflecting the heterogeneity of the disease. In addition, we found that low PRG4 expression was associated with a more aggressive disease stage, which is in accordance with PRG4 loss-of-function mutations causing camptodactyly-arthropathy-coxa vara-pericarditis syndrome. Altogether we provide evidence for molecular signatures of RA and RA subclasses, sets of new candidate genes as well as for candidate gene networks, which extend our understanding of disease mechanisms and may lead to an improved diagnosis.

The conceptus increases secreted phosphoprotein 1 gene expression in the mouse uterus during the progression of decidualization mainly due to its effects on uterine natural killer cells

Within the mouse endometrium, secreted phosphoprotein 1 (SPP1) gene expression is mainly expressed in the luminal epithelium and some macrophages around the onset of implantation. However, during the progression of decidualization, it is expressed mainly in the mesometrial decidua. To date, the precise cell types responsible for the expression in the mesometrial decidua has not been absolutely identified. The goal of the present study was to assess the expression of SPP1 in uteri of pregnant mice (decidua) during the progression of decidualization and compared it with those undergoing artificially induced decidualization (deciduoma). Significantly (P<0.05) greater steady-state levels of SPP1 mRNA were seen in the decidua when compared with deciduoma. Further, in the decidua, the majority of the SPP1 protein was localized within a subpopulation of granulated uterine natural killer (uNK) cells but not co-localized to their granules. However, in addition to being localized to uNK cells, SPP1 protein was also detected in another cell type(s) that were not epidermal growth factor-like containing mucin-like hormone receptor-like sequence 1 protein-positive immune cells that are known to be present in the uterus at this time. Finally, decidual SPP1 expression dramatically decreased in uteri of interleukin-15-deficient mice that lack uNK cells. In conclusion, SPP1 expression is greater in the mouse decidua when compared with the deciduoma after the onset of implantation during the progression of decidualization. Finally, uNK cells were found to be the major source of SPP1 in the pregnant uterus during decidualization. SPP1 might play a key role in uNK killer cell functions in the uterus during decidualization.