Cyr61, CTGF, and WISP-1 play important roles in cell proliferation, migration and differentiation. Their expression appears to be regulated differently in various types of tumors. To explore the correlation between expression of CCN genes and NSCLC, we used real-time PCR and immunohistochemistry to evaluate the mRNA and protein levels of these three genes in NSCLC and their matched normal lung tissues. Downregulation of Cyr61 and CTGF and upregulation of WISP-1 occurred in the NSCLC samples compared to their normal counterparts, suggesting that these molecules might be associated with tumor formation and progression in NSCLC.
Cyr61 is the first cloned member of the CCN family and its regulatory roles in tumor cells have been widely reported in many types of cancers. In breast cancers, Cyr61 is overexpressed and can stimulate tumor progression 
. A gastric adenocarcinoma cell line became more tumorigenic when the cells were genetically engineered to express high levels of Cyr61 
. Expression of Cyr61 was high in rhabdomyosarcomas and cell lines derived from malignant melanomas, colon adenocarcinomas, and bladder papillomas 
. Malignant gliomas often have high levels of Cyr61 associated enhanced tumorigenicity mediated through the integrin-linked kinase signaling pathway 
. Upregulation of Cyr61 expression was recently identified in peritoneal metastases from human pancreatic cancer 
. Paradoxically, Cyr61 appears to have the opposite role in lung cancer. We have reported that Cyr61 was downregulated in four of 5 samples of lung cancer, and lung cancer cells stably transfected with a Cyr61 expression vector were less tumorigenic than the vector alone transfected control cells 
. Our further studies showed that forced expression of Cyr61 in lung cancer cells resulted in their cell-cycle arrest in G1 phase mediated by p53 
. Here, we found that expression of Cyr61
is decreased in NSCLC samples compared to their matched controls, which strongly supports our former hypothesis. Cyr61 was also reported to inhibit growth of prostate cancer 
, endometrial cancer 
and leiomyomas 
. Taken together, the data suggested that Cyr61 might behave as a tumor suppressor under certain circumstances in several tissue types, including NSCLC.
CTGF was identified as a mitogen found in the conditioned medium of human umbilical vein endothelial cells 
. It encodes a protein of 349 amino acids with 43% sequence identity to Cyr61, and all 38 cysteines in CTGF and Cyr61 are completely conserved. CTGF was found to be overexpressed in mammary tumors 
, melanomas 
, pancreatic cancers 
, sarcomas including chondrosarcomas 
; while an inverse correlation has been reported between the malignant phenotype and the level of CTGF expression in fibroblasts and endothelial cell tumors 
. In our experiments, we found that the level of expression of CTGF
in NSCLCs was lower than in the matched normal lung samples, implying its potential tumor-suppressing function. Consistent with our finding, a recent study by Chien et al. 
showed that CTGF suppressed lung cancer cell growth by induction of p53, as well as, by inhibition of insulin-like growth factor-I dependent Akt phosphorylation and epidermal growth factor-dependent extracellular signal-regulated kinase 1/2 phosphorylation.
On the other hand, we found that WISP-1
was overexpressed in NSCLC samples compared to their normal lung tissue counterparts, suggesting that WISP-1 might act as an oncoprotein in NSCLC. WISP-1 was identified as a gene that was upregulated in Wnt-1 transformed C57 MG mouse mammary epithelial cells; it has complete conservation of all 38 cysteine residues with those of Cyr61 and CTGF 
. WISP-1 has been associated with either enhancing or inhibiting growth of tumors. For example, it is strongly expressed in human breast and colon cancers 
. Forced overexpression of WISP-1 in normal rat kidney fibroblasts (NRK-49F) was sufficient to induce their transformation 
. In contrast, WISP-1 expression was inversely correlated with proliferation, metastasis and growth of melanoma cells 
. Additionally, our result that upregulation of WISP-1 was positively correlated with lung cancer metastasis was consistent with findings from a mouse model 
. Paradoxically, Soon et al.
found that in vitro
overexpression of WISP-1 decreased motility of lung cancer cells 
. Since WISP-1 is a secreted protein and functions mainly by interaction with ECM, the difference in findings might reflect the difference in ECM in vivo
and in vitro
Univariate statistical analysis further disclosed that low levels of either Cyr61 or CTGF were related with the progression of NSCLC, and the downregulation of Cyr61 and CTGF was more notable in patients with family history than those without family history. These data implied that Cyr61 and CTGF might be tumor suppressor genes in lung cancer. Through multiple linear regression analyses, moreover, we found that clinical features are closely associated with levels of expression of Cyr61, CTGF and WISP-1. Expression level of Cyr61 in 71% lung cancer samples was determined by eight independent variables, mainly by AC, SC and age. Levels of CTGF in NSCLCs were also markedly associated with clinical features, but with some differences. Gender seemed to play a role in expression of CTGF, but not Cyr61. Sex hormone might regulate expression of CTGF, but more work is needed to confirm this impression. Statistical analysis also revealed that levels of these molecules were correlated with one another, suggesting that their regulation may relate to similar signaling pathways. Furthermore, levels of Cyr61, CTGF, and WISP-1 were tightly related to pivotal clinical and prognostic features of NSCLC. Perhaps, examining the levels of these three molecules either in the primary tumor or in the malignant cells from sputa of patients may help guide therapy.
In summary, our study examined the three CCN molecules (Cyr61, CTGF, and WISP-1) and found correlations between their levels and clinical features of NSCLCs. Although the detailed mechanism remains to be investigated, our results might provide new parameters for diagnosis and prognosis of NSCLC.