SummaryXenograft transplantation of human tumor cells into immunodeficient mice is an important method to clarify the roles of specific molecules or chemicals in vivo. Recently, this method has been reported as a definitive examination to identify tumor stem cells. In this study, the authors compared the morphology and the quality and quantity of ribonucleic acid (RNA) and protein in paraffin-embedded tissues of nude mice implanted with human uterine cervical cancer cells, followed by fixation with commonly used fixatives, including 4% paraformaldehyde (PFA), 10% neutral buffered formalin (NBF), 20% NBF, and 99% ethanol (EtOH). The quality of the isolated RNA from PFA-and NBF-fixed paraffin-embedded tissues was high, while EtOH-fixed tissues showed degradation of RNA. NBF-fixed tissues showed excellent quality of morphology, but EtOH-fixed tissues showed contraction of cells. Immunohistochemical results showed differences depending on fixations. The 99% EtOH-fixed samples showed decreases of Ki-67 and VEGF-A immunoreactivities, but improved cytokeratin immunoreactivity. This study indicated that formalin fixation is better than alcohol fixation for RNA preservation in paraffinembedded cancer cell implantation models. Immunohistochemical results differed markedly depending on fixation materials and antibodies; therefore, suitable fixations are needed to quantify and compare the results of immunohistochemical staining on cancer cell implanted nude mice tissues. (J Histochem Cytochem 59:68-75, 2011)

Colorectal cancer (CRC) is one of the most common cancers worldwide, and many patients are already at an advanced stage when they are diagnosed. Therefore, novel biomarkers for early detection of colorectal cancer are required. In this study, we performed a global shotgun proteomic analysis using formalin-fixed and paraffin-embedded (FFPE) CRC tissue. We identified 84 candidate proteins whose expression levels were differentially expressed in cancer and non-cancer regions. A label-free semiquantitative method based on spectral counting and gene ontology (GO) analysis led to a total of 21 candidate proteins that could potentially be detected in blood. Validation studies revealed cyclophilin A, annexin A2, and aldolase A mRNA and protein expression levels were significantly higher in cancer regions than in non-cancer regions. Moreover, an in vitro study showed that secretion of aldolase A into the culture medium was clearly suppressed in CRC cells compared to normal colon epithelium. These findings suggest that decreased aldolase A in blood may be a novel biomarker for the early detection of CRC.

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Abstract. The keratinocyte growth factor receptor (KGFR), also known as FGFR2 IIIb, is mainly localized in epithelial cells and is activated by the keratinocyte growth factor (KGF) that is predominantly synthesized by mesenchymal cells. In this study, we examined the roles of KGFR and KGF in human esophageal cancer (EC). In noncancerous esophageal tissues, KGFR was localized in epithelial cells from the basal region of the epithelium to the lower one-third of the epithelium, and KGF was weakly localized in the basal to parabasal epithelial cells. On the other hand, Ki-67 was localized in the parabasal cells. In EC tissues, KGFR and KGF were expressed in cancer cells in 22 and 37 of 54 patients, respectively. The coexpression of KGFR and KGF in cancer cells was detected in 14 of 54 (26%) patients. Clinicopathologically, KGFR expression correlated with the well-differentiated cell type of EC (p<0.001), and KGF expression correlated with lymphatic invasion and lymph node metastasis (p=0.004 and 0.021, respectively). The coexpression of KGFR and KGF in cancer cells correlated with the well-differentiated cell type of EC (p=0.001). KGFR-positive, KGF-positive and KGFR/KGF coexpression patients tended to have shorter survival rates, but the survival rates were not statistically significantly different (p=0.44, 0.059 and 0.112, respectively). In human EC cell lines (TE-1, TE-8 and TE-11), KGFR mRNA was expressed but no KGF mRNA was detected. The KGFR mRNA level was highest in TE-1 cells, derived from well-differentiated SCC and lowest in TE-8 cells. KGFR was detected in the cancer cell lines by Western blot analysis. Recombinant human KGF significantly stimulated the growth of TE-8 and -11 cells, derived from moderately differentiated SCC, but had no effect on TE-1 cell growth. These results suggest that KGFR expression correlates with the differentiation of a normal esophageal epithelium and the well-differentiated cell type of EC. On the other hand, KGF may induce the growth of some EC cells in a paracrine manner and closely correlates with lymphatic invasion and lymph node metastasis.

In the histopathological diagnosis of cutaneous tumors, the differential diagnosis of squamous cell carcinoma (SCC) with crateriform architecture and keratoacanthoma (KA) is often difficult so an accurate understanding of the biological features and the identification of reliable markers of SCC and KA are crucial issues. Insulin-like growth factor 2 mRNA-binding protein-3 (IGF2BP3, also known as IMP3) is thought of as a bona fide oncofetal protein, which is overexpressed and is involved in cell proliferation, migration, and invasion in several kinds of tumors. However, the role of IMP3 in cutaneous SCC and KA has not been well studied. Therefore, we focused on studying the biological functions of IMP3 in SCC and KA. In human skin SCC cell lines, HSC-1 and HSC-5, and the human keratinocyte cell line, HaCaT, IMP3 mRNA levels were significantly higher than that of normal human skin. The knockdown of IMP3 expression reduced the proliferation of HSC-1, and significantly reduced invasion by HSC-1 and HSC-5. In contrast, the knockdown of IMP3 did not significantly affect invasion by HaCaT cells. In immunohistochemical studies of SCC and KA tissues, the Ki-67 labeling index (LI) of the suprabasal cell layer was significantly higher in SCC, compared with KA tissues and the tumor-free margin (TFM) adjacent to SCC and KA. Most SCC tissues stained strongly positive for IMP3, but KA tissues and TFM were mostly negative for IMP3. The Ki-67 LI of the IMP3-positive group was significantly higher than that of the IMP3-negative group in the suprabasal cell layer of SCC. These results suggest that IMP3 plays an important role in proliferation and, more significantly, in the invasion of SCC, and may be a suitable marker for the histopathological diagnosis of SCC with a crateriform architecture and KA. Furthermore, IMP3 may potentially be a new therapeutic target for SCC.