Background:The expression of Ltype amino-acid transporter 1 (LAT1) is tumour-specific and has been shown to have essential roles in cell growth and survival. However, little is known regarding the clinical significance of LAT1 expression in pancreatic cancer. This study was conducted to determine the prognostic significance of LAT1 expression.Methods:A total of 97 consecutive patients with surgically resected pathological stage I–IV pancreatic ductal adenocarcinoma were retrospectively reviewed. Tumour sections were stained by immunohistochemistry for LAT1, CD98, Ki-67 and vascular endothelial growth factor (VEGF), and microvessel density was determined by CD34 and p53.Results:L-type amino-acid transporter 1 and CD98 were highly expressed in 52.6% (51/97) and 56.7% (55/97) of cases, respectively (P=0.568). The expression of LAT1 within pancreatic cancer cells was significantly associated with disease stage, tumour size, Ki-67, VEGF, CD34, p53 and CD98. Ltype amino-acid transporter 1 expression was confirmed to be a significant prognostic factor for predicting poor outcome by multivariate analysis.Conclusion:Ltype amino-acid transporter 1 expression is a promising pathological marker for the prediction of outcome in patients with pancreatic cancer.

BackgroundThe expression of L-type amino acid transporter 1 (LAT1) has been described to play essential roles in tumor cell growth and survival. However, it remains unclear about the clinicopathological significance of LAT1 expression in biliary tract cancer. This study was conducted to determine biological significance of LAT1 expression and investigate whether LAT1 could be a prognostic biomarker for biliary tract cancer.MethodsA total of 139 consecutive patients with resected pathologic stage I-IV biliary tract adenocarcinoma were retrospectively reviewed. Tumor specimens were stained by immunohistochemistry for LAT1, Ki-67, microvessel density determined by CD34, and p53; and prognosis of patients was correlated. Biological significance of LAT1 expression was investigated by in vitro and in vivo experiments with LAT inhibitor, 2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid (BCH) using cholangiocarcinoma cell line.ResultsIn total patients, high LAT1 expressions were recognized in 64.0%. The expression of LAT1 was closely correlated with lymphatic metastases, cell proliferation and angiogenesis, and was a significant indicator for predicting poor outcome after surgery. LAT1 expression was a significant independent predictor by multivariate analysis. Both in vitro and in vivo preliminary experiments indicated that BCH significantly suppressed growth of the tumor and yielded an additive therapeutic efficacy to gemcitabine and 5-FU.ConclusionsHigh expression of LAT1 is a promising pathological marker to predict the outcome in patients with biliary tract adenocarcinoma. Inhibition of LAT1 may be an effective targeted therapy for this distressing disease.

S U M M A R YFading is one of the major obstacles to reliable observation in fluorescence microscopy. Using a confocal laser scanning microscope (CLSM) coupled to a computer, we quantitatively measured fading of fluorescence to formulate an equation, evaluated the anti-fading ability of several anti-fading media, and restored the faded images to the original level according to this equation. NIH 3T3 cells were stained with fluorescein isothiocyanate (FITC)-phalloidin, mounted with several commercial and homemade anti-fade media, and observed with CLSM under repeated illumination. With any mounting medium, attenuation of fluorescence intensity at a certain pixel occurred stepwise and the decrease was proportional to the intensity of the previous scan. From these results, we formulated an equation that has three coefficients: anti-fading factor ( A ), indicating the ability to retard fading; fluorescent intensity at the first scan ( EM 1 ); and background fluorescence ( B ). The fluorescent intensity at a certain point following n th scan is given as EM n ϭ EM 1 • A (n Ϫ 1) . This equation was available for restoring faded images to their original states, even after the image had faded to only 60% of its original intensity.

In order to confirm 14-3-3 sigma (sigma) protein distribution in human tissues, immunohistochemistry was performed using various paraffin-embedded human tissues. In normal human tissues, the strongest immunoreactivity for 14-3-3sigma protein was observed in squamous epithelia at various sites, followed by basal cells of the trachea, bronchus and basal or myoepithelial cells of various glands. Moderate to weak 14-3-3sigma immunoreactivity was seen in the epithelial cells of the alimentary tract, gall bladder, urinary tract and endometrium. In the lung, 14-3-3sigma immunoreactivity was also observed in hyperplastic type II alveolar cells and metaplastic squamous cells. Immunohistochemical study using non-small-cell lung cancers revealed that 14-3-3sigma immunoreactivity was stronger in squamous cell carcinomas than in adenocarcinomas. The present study revealed that 14-3-3sigma expression was exclusively present in various epithelial cells and had a tendency to be stronger in cells destined for squamous epithelium or differentiating toward squamous cells in human normal and neoplastic cells.

In polarized epithelial cells, agonists trigger Ca2+ waves and oscillations. These patterns may be caused by the compartmentalization of inositol 1,4,5-trisphosphate (IP3)-sensitive Ca2+ pools into specific regions. We have investigated the relationship between the distribution of IP3 receptors (IP3Rs) and the spatiotemporal pattern of Ca2+ signaling in the duct cells of the rat submandibular gland (SMG). Using immunofluorescence, although labeling was somewhat heterogeneous, the IP3Rs were colocalized to the apical pole of the duct cells. Immunoelectron microscopy identified small apical vesicles bearing IP3R2 in some types of duct cells. Real-time confocal imaging of intact ducts demonstrated that, after carbachol stimulation, an initial Ca2+ spike occurred in the apical region. Subsequently, repetitive Ca2+ spikes spread from the apical to the middle cytoplasm. These apical Ca2+ initiation sites were found only in some “pioneer cells,” rather than in all duct cells. We performed both Ca2+ imaging and immunofluorescence on the same ducts and detected the strongest immunosignals of IP3R2 in the Ca2+ initiation sites of the pioneer cells. The subcellular localization and expression level of IP3Rs correlated strongly with the spatiotemporal nature of the intracellular Ca2+ signal and distinct Ca2+ responses among the rat SMG duct cells.

ABSTRACT. We observed the dynamic changes in the localization of microfilaments during the exocytic secretion of rat parotid and submandibular gland acinar cells, and obtained results which led us to propose a new concept of microfilament function in exocytosis. With the electron microscopy , NBDPhallacidin (NBD-PL) fluorescence technique and immunohistochemistry for myosin, microfilaments consisting of F-actin and myosin were localized mainly underneath the luminal plasma membrane. Microfilaments were not detectable around the secretory granules which were stored in the cytoplasm, but were clearly observed around them whose membranes were continuous with the luminal plasma membrane. When viewed with NBD-PL and myosin fluorescence, the area of fused granule membranes revealed bright fluorescence in association with the luminal border, so that the luminal membrane undergoing exocytosis appeared like a 'bunch of grapes'. When excess exocytosis was stimulated by isoproterenol (IPR), the number of individual 'grapes' increased dramatically, indicating that the secretory granules are surrounded by microfilaments after the fusion with the luminal membrane. Microfilaments thus continuously undercoat the luminal membrane during exocytosis although the exocytic process involves the dilation and subsequent reduction of the luminal membrane due to the addition and removal of secretory granule membranes. This reduction of the dilated luminal membrane follwoing exocytosis was, however, inhibited when the microfilaments were disrupted by cytochalasin D. Following this treatment, the lumina was expanded extraordinarily and the secretory products remained in the enlarged lumina, showing that the release of secretory products is inhibited when the microfilament function is disturbed.These results indicate that 1) microfilaments are localized mainly underneath the luminal plasma membrane and act as an obstacle to exocytosis when cells are at the resting phase and 2) at the secretory phase microfilaments allow exocytosis by disorganizing their barrier system and then, by encircling the discharged secretory granule membranes, provide forces for the extrusion of secretory products through the action of the acto-myosin contractile system. Exocytosis is an ultimate step of packaged secretion that is recognized to involve the intracellular transport of secretory granules, the fusion of secretory granule membrane with plasma membrane, and the release of granule contents (10, 21) . It is the basis for the release of digestive enzymes, hormones, neuro-transmitters and

Background: We report herein a rare case of primary omental gastrointestinal stromal tumor (GIST).

Abstract. Upregulation of L-type amino acid transporter 1 (LAT1), a member of the system L amino acid transporter family, may be detected by immunohistochemical methods. Immunoreactive LAT1 expression in prostate cancer is considered to be a promising biomarker for high-grade malignancy. However, the mutual association between LAT1 and Gleason score, the most fixed indicator for grading the malignancy of prostate cancers, remains to be elucidated. The aim of this study was to clarify the correlations between LAT1 and other factors in prostate cancer, including the Gleason score. We evaluated 54 cases of primary prostate cancer, surgically resected without any neoadjuvant therapies and performed immunohistochemistry for LAT1, Ki-67, CD34 and vascular endothelial growth factor on the tissue sections. The Gleason score as well as the age, pathological stage (pStage) of prostate cancer and serum concentration of prostate-specific antigen (PSA) of each case were also assessed. Statistical analysis for the correlations between LAT1 expression and Gleason score and each of the other characteristics studied was performed. As a result, a strong significant correlation between immunoreactive LAT1 expression and Gleason score was identified (P<0.01). We concluded that immunoreactive LAT1 expression in tissue sections of prostate cancer may be useful as a biomarker for high-grade malignancy.