We have cloned a novel hematopoietic granulocyte colony-stimulating factor (G-CSF)-induced olfactomedin-related glycoprotein, termed hGC-1 (human G-CSF-stimulated clone-1). mRNA differential display was used in conjunction with a modified two-phase liquid culture system. The hGC-1 gene encodes a 510-amino acid glycoprotein whose exact in vivo localization and function still remains elusive. A recent study showed that hGC-1 mRNA expression is upregulated in gastric caner. To further explore a potential relationship between hGC-1 and gastric carcinoma, we investigated the expression pattern of hGC-1 protein in 173 cases of gastric carcinoma. Using immunohistochemistry, we demonstrated that hGC-1 is expressed in the esophagus, stomach, small intestine and colon. We demonstrated a striking correlation between hGC-1 expression and the histological type and differentiation grades of gastric carcinoma. Enhanced hGC-1 expression is more frequently seen in intestinal- type adenocarcinoma, while loss of expression tends to occur in the diffuse-type. hGC-1 is highly expressed in well or moderately differentiated cancer tissues and remarkably reduced or lost in poorly differentiated or undifferentiated tissues. We have subsequently examined the hGC-1 expression in colon carcinoma and explore the relationship between hGC-1 expression and clinico-pathological features of colon cancer patients. The expression of hGC-1 in colon adenocarcinoma tissues was examined by dot-blot analysis, in situ hybridization and immunohistochemistry. Compared to normal colon mucosa, upregulation of hGC-1 was more frequently detected in more differentiated colon cancers, while downregulation or no expression was associated with poorly differentiated colon cancers. Interestingly, hGC-1 downregulation was also found in late tumor-node-metastasis (TNM) stage, metastasis, and patients with shorter survival. To investigate the involvement of hGC-1 in colon cancer progression, human colon carcinoma (HT-29) cells overexpressing hGC-1 were established. The morphology and cortical actin distribution of HT-29 cells were altered by hGC-1 over-expression. However, this did not change cell proliferation, but decreased cell adhesion and migration. Our findings indicate that hGC-1 is involved in colon cancer adhesion and metastasis, and that hGC-1 may be a useful marker for tumor differentiation and progression of human colon carcinoma.These investigations define for the first time the expression pattern of hGC-1 in the normal human gastrointestinal tract and provide a novel and sensitive marker for the differentiation of gastric carcinoma. The human olfactomedin 4 gene (OLFM4, also known as hGC-1, GW112) is thought to be a useful marker for early myeloid development. To understand the molecular mechanisms underlying granulocyte colony-stimulating factor (G-CSF)-stimulated OLFM4 expression, we characterized the promoter region of OLFM4. The 35-bp region (-101 to -66) of the proximal promoter regulated reporter gene expression, and mutation of the nuclear factor (NF)-kappaB binding site within the promoter abolished the binding of the transcription factor and the ability to regulate OLFM4 expression. G-CSF increased reactive oxygen species (ROS) production in human CD34(+) cells, which was abrogated by inhibition of phosphatidylinositol 3-kinase (PI3K) or NADPH oxidase. Phosphorylation of ERK1/2 mitogen-activated protein kinase (MAPK) induced by G-CSF inhibited by the antioxidant N-acetyl-L-cysteine (NAC), ERK1/2 inhibitor PD98059, or U0126. However, phosphorylation of signal transducer and activator of transcription (STAT)3 was only partially inhibited by NAC, but not by PD98059 or U0126. Inhibition of the ERK pathway remarkably decreased OLFM4 expression and this inhibition required NF-kappaB transcription factor. Inhibition of ROS or the ERK pathway remarkably decreased G-CSF-induced OLFM4 expression. Our results suggest that G-CSF-induced expression of OLFM4 is regulated by the transcription factor NF-kappaB, and that this induction is mediated by the ERK1/2 MAPK signaling pathway through PI3K-driven ROS production. In further studies of the role of hGC-1 in myelopoiesis, we found that hGC-1 is overexpressed in myeloid leukemia patients compared with normal individuals in peripheral blood leukocytes (p<0.01) and its expression in the accelerated phase of chronic myeloid leukemia (CML) patients was significantly higher than that in chronic phase (p<0.01) using a dot blot and quantitative RT-PCR analysis. Hypomethylation of CpG sites in the promoter of hGC-1 gene were observed in CML patients by pyrosequence and 5-aza-2-deoxycytidine induced hGC-1 expression in myeloid leukemia cells, suggesting that promoter CpG methylation status affects the expression of hGC-1 gene. All-trans-retinoic acid (ATRA) and interferons (IFNs) are active anti-leukemia agents. ATRA and IFNs have shown synergistic interactions in various experimental conditions and represent a potentially useful therapeutic combination in the treatment of various types of leukemia. However, the target genes and molecular basis of these interactions still needs to be further elucidated. Here, we identified that hGC-1 was a target gene of RA in myeloblastic leukemia cells. Treatment with ATRA induced hGC-1 expression in HL-60 cells and enhanced hGC-1 expression in AML-193 and GDM-1 cells. Deletion analysis led to the identification of a positive retinoic acid response element (DR5) and a negative response element (DR1) within hGC-1 promoter. Furthermore, electrophoretic mobility-shift assays demonstrated that RARα/RXRαbinds to the DR5 site. Transfection study in COS-7 cells revealed RARα/RXRαmediated the RA induced transactivation of hGC-1 promoter. We also found that hGC-1 was an early responsive gene of IFN αand β in myeloid leukemia cells (HL-60, AML-193 and GDM-1). Αn effective interferon-stimulated response element (ISRE) was identified in the promoter of hGC-1 gene by examining the deletion mutants in luciferase reporter gene assay. Combined application of ATRA and IFNαenhanced hGC-1 expression synergistically. Taken together, hGC-1 is identified as a novel target gene of methylation modification, RA and IFNs in myeloid leukemia cells. Because these findings implicate a role of hGC-1 in tumorgenesis, and because hGC-1 is located on chromosome 13q14.1, near a site of a putative prostate cancer tumor suppressor gene, we examined the role of hGC-1 in prostate carcinogenesis. Higher grade and metastasis of prostate cancer are mainly cause death of prostate cancer patients. RNA expression of hGC-1 was detected in the normal prostate however the expression of hGC-1 protein have not examined. We have done an immunohistochemistry staining with hGC-1 antibody and stained 18 normal prostate specimens and 159 different grades of prostate cancer specimens. We have found that lower or lost expression of hGC-1 are significantly associated with higher Gleason score of prostate cancer specimens. hGC-1 was not detected in PC-3, LNCap and Du145 cells that derived from metastasized prostate cancer. Furthermore, we compared the expression of hGC-1 to prostate specific antigen (PSA) and E-cadherin by immunohistochemistry staining with series cutting prostate cancer tissue arrays containing 112 specimens. The results indicated that expressed pattern of hGC-1 are most correlated with the expression of E-cadherin (P<0.001) and less correlated with PSA (P<0.004).

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Liu, Wenli; Liu, Yueqin; Li, Hongzhen et al. (2018) Olfactomedin 4 contributes to hydrogen peroxide-induced NADPH oxidase activation and apoptosis in mouse neutrophils. Am J Physiol Cell Physiol 315:C494-C501
Liu, Wenli; Rodgers, Griffin P (2016) Olfactomedin 4 expression and functions in innate immunity, inflammation, and cancer. Cancer Metastasis Rev 35:201-12
Liu, W; Li, H; Hong, S-H et al. (2016) Olfactomedin 4 deletion induces colon adenocarcinoma in Apc(Min/+) mice. Oncogene :
Liu, Wenli; Yan, Ming; Sugui, Janyce A et al. (2013) Olfm4 deletion enhances defense against Staphylococcus aureus in chronic granulomatous disease. J Clin Invest 123:3751-5
Liu, Wenli; Yan, Ming; Liu, Yueqin et al. (2012) Olfactomedin 4 inhibits cathepsin C-mediated protease activities, thereby modulating neutrophil killing of Staphylococcus aureus and Escherichia coli in mice. J Immunol 189:2460-7
Liu, Wenli; Lee, Hyun Woo; Liu, Yueqin et al. (2010) Olfactomedin 4 is a novel target gene of retinoic acids and 5-aza-2'-deoxycytidine involved in human myeloid leukemia cell growth, differentiation, and apoptosis. Blood :
Liu, Wenli; Yan, Ming; Liu, Yueqin et al. (2010) Olfactomedin 4 down-regulates innate immunity against Helicobacter pylori infection. Proc Natl Acad Sci U S A 107:11056-61
Liu, Wenli; Liu, Yueqin; Zhu, Jianqiong et al. (2008) Reduced hGC-1 protein expression is associated with malignant progression of colon carcinoma. Clin Cancer Res 14:1041-9
Chin, Kyung L; Aerbajinai, Wulin; Zhu, Jiangqiong et al. (2008) The regulation of OLFM4 expression in myeloid precursor cells relies on NF-kappaB transcription factor. Br J Haematol :