FIZZ2/RELM-ß induction and role in pulmonary fibrosis.

Found in inflammatory zone (FIZZ) 2, also known as resistin-like molecule (RELM)-ß, belongs to a novel cysteine-rich secreted protein family named FIZZ/RELM. Its function is unclear, but a closely related family member, FIZZ1, has profibrotic activities. The human ortholog of rodent FIZZ1 has not been identified, but human FIZZ2 has significant sequence homology to both rodent FIZZ2 (59%) and FIZZ1 (50%). Given the greater homology to rodent FIZZ2, analyzing the role of FIZZ2 in a rodent model of bleomycin-induced pulmonary fibrosis would be of greater potential relevance to human fibrotic lung disease. The results showed that FIZZ2 was highly induced in lungs of rodents with bleomycin-induced pulmonary fibrosis and of human patients with idiopathic pulmonary fibrosis. FIZZ2 expression was induced in rodent and human lung epithelial cells by Th2 cytokines, which was mediated via STAT6 signaling. The FIZZ2 induction in murine lungs was found to be essential for pulmonary fibrosis, as FIZZ2 deficiency significantly suppressed pulmonary fibrosis and associated enhanced extracellular matrix and cytokine gene expression. In vitro analysis indicated that FIZZ2 could stimulate type I collagen and α-smooth muscle actin expression in lung fibroblasts. Furthermore, FIZZ2 was shown to have chemoattractant activity for bone marrow (BM) cells, especially BM-derived CD11c(+) dendritic cells. Notably, lung recruitment of BM-derived cells was impaired in FIZZ2 knockout mice. These findings suggest that FIZZ2 is a Th2-associated multifunctional mediator with potentially important roles in the pathogenesis of fibrotic lung diseases.

Involvement of endoplasmic reticulum stress in myofibroblastic differentiation of lung fibroblasts.

Stress that impairs endoplasmic reticulum (ER) function leads to an accumulation of unfolded or misfolded proteins in the ER (ER stress) and triggers the unfolded protein response (UPR). Recent studies suggest that ER stress is involved in idiopathic pulmonary fibrosis (IPF). The present study was undertaken to determine the role of ER stress on myofibroblastic differentiation of fibroblasts. Fibroblasts in fibroblastic foci of IPF showed immunoreactivity for GRP78. To determine the role of ER stress on α-smooth muscle actin (α-SMA) and collagen type I expression in fibroblasts, mouse and human lung fibroblasts were treated with TGF-ß1, and expression of ER stress-related proteins, α-SMA, and collagen type I was analyzed by Western blotting. TGF-ß1 significantly increased expression of GRP78, XBP-1, and ATF6α, which was accompanied by increases in α-SMA and collagen type I expression in mouse and human fibroblasts. A chemical chaperone, 4-PBA, suppressed TGF-ß1-induced UPR and α-SMA and collagen type I induction. We also showed that TGF-ß1-induced UPR was mediated through the reactive oxygen species generation. Our study provides the first evidence implicating the UPR in myofibroblastic differentiation during fibrosis. These findings of the role of ER stress and chemical chaperones in pulmonary fibrosis may improve our understanding of the pathogenesis of IPF.

Data mining of micrornas in breast carcinogenesis which may be a potential target for cancer prevention.

The microRNAs (miRNAs) negatively regulate the stability and translation of target messenger RNAs by selectively binding. It has been implicated in diverse processes such as cellular differentiation, cell-cycle control, apoptosis, and carcinogenesis. Examination of tumor-specific miRNA expression profiles has revealed wide spread dysregulation of these molecules in diverse cancers. The available genomic bulk evidences were extracted from The Cancer Genome Atlas by using IluminaGA_miRNASeq platform in human breast cancer samples. After mining collected data, group of each miRNA ID was analyzed through five D/Bs (mirWalk, miranda, mirDB, RNA22, and TargetScan) on predicted and validated miRNA targets. Oncogenes known to have a high correlation with breast cancer (C-myc, HER2, cyclin D-1, N-RAS, FGF-4, FGF-3, BRCA1, and BRCA2) are subject in this study to select their relevant miRNAs. Function of miRNA regulation will be essential to achieve a complete understanding of carcinogenesis and these miRNAs would be potential target for breast cancer prevention.

Expression of SIRT1 and cortactin is associated with progression of non-small cell lung cancer.

Cortactin is an F-actin binding protein involved in cell migration and tumor metastasis. Recent reports suggest that silent mating-type information regulation 2 homologue 1 (sirtuin1; SIRT1) enhances the function of cortactin and promotes cell migration. We investigated SIRT1 and cortactin expression in 144 invasive non-small cell lung cancers (NSCLC) and 19 adenocarcinomas in situ (AIS) by immunohistochemistry and evaluated their clinicopathological significance in NSCLC. Positive SIRT1 and cortactin expression was observed in 67% (96 of 144) and 58% (84 of 144) of patients with invasive NSCLC, respectively. SIRT1 and cortactin expression was significantly associated with unfavorable clinicopathological factors, including high pathological T stage, lymph node metastasis, and advanced tumor invasion (AIS vs. invasive adenocarcinoma). Cortactin was significantly associated with high pathological T stage and lymph node metastasis in SIRT1-positive tumors. Cytoplasmic SIRT1 was significantly associated with high pathological T stage and large tumor size compared to that of nuclear SIRT1. Large tumor size, high pathological T stage, lymph node metastasis, and cytoplasmic SIRT1 expression were significantly associated with shorter overall survival in a univariate analysis. Our findings suggest that SIRT1 and cortactin may play a role in the progression of NSCLC and may cooperate during tumor progression in NSCLC.

Effects of delay in the snap freezing of colorectal cancer tissues on the quality of DNA and RNA.

PURPOSE: The success of basic molecular research using biospecimens strongly depends on the quality of the specimen. In this study, we evaluated the effects of delayed freezing time on the stability of DNA and RNA in fresh frozen tissue from patients with colorectal cancer. METHODS: Tissues were frozen at 10, 30, 60, and 90 minutes after extirpation of colorectal cancer in 20 cases. Absorbance ratio of 260 to 280 nm (A(260)/A(280)) and agarose gel electrophoresis were evaluated. In addition, the RNA integrity number (RIN) was assayed for the analysis of the RNA integrity. RESULTS: Regardless of delayed freezing time, all DNA and RNA samples revealed A(260)/A(280) ratios of more than 1.9, and all DNA samples showed a discrete, high-molecular-weight band on agarose gel electrophoresis. The RINs were 7.53 ± 2.04, 6.70 ± 1.88, 6.47 ± 2.58, and 4.22 ± 2.34 at 10, 30, 60, and 90 minutes, respectively. Though the concentration of RNA was not affected by delayed freezing, the RNA integrity was decreased with increasing delayed freezing time. CONCLUSION: According to the RIN results, we recommend that the collection of colorectal cancer tissue should be done within 10 minutes for studies requiring RNA of high quality and within 30 minutes for usual RNA studies.

Discoidin domain receptor 1 is associated with poor prognosis of non-small cell lung carcinomas.

Discoidin domain receptors (DDRs) are a novel class of receptor tyrosine kinases that bind to several collagens and facilitate cell adhesion. DDR1 is involved in cancer invasion. However, very limited data are available on the aspect of clinical significance of DDR1 expression in cancer. The aim of this study was to investigate prognostic impact of DDR1 expression in non-small cell lung carcinoma (NSCLC). Tumor tissues from 171 NSCLC, including 86 squamous cell carcinomas, 69 adenocarcinomas, and 16 pure bronchioloalveolar carcinomas (BAC), were analyzed for expression of DDR1 using immunohistochemical staining. In addition, two lung adenocarcinoma cell lines (A549, H358) were transfected with two isoforms of DDR1, DDR1a and DDR1b, and then migration and invasion assays were carried out. DDR1 was expressed in 6 of 16 BAC (38%) and 95 of 155 invasive NSCLC (61%, p=0.065). DDR1 up-regulation tend to be more frequently observed in invasive adenocarcinoma (64%) compared to DDR1 expression in BAC (38%) (p=0.056). In invasive NSCLC, DDR1 expression was significantly correlated with lymph node metastasis (p=0.001). Overexpression of DDR1 in lung cancer cells resulted in a significant increase of cell motility and invasiveness (p<0.001) and the interaction of DDR1 with collagen facilitates the invasiveness of NSCLC cells. DDR1 overexpression produced an activation of MMP-9 in H358 cells. In conclusion, these findings indicate that up-regulation of DDR1 may contribute to the progression and poor prognosis of NSCLC and this effect may be associated with increased invasiveness.