Biliary stenting as alternative therapy to stone clearance in elderly patients with bile duct stones. .

BACKGROUND AND STUDY AIMS: Endoscopic retrograde cholangio- pancreatography with stone retrieval following endoscopic sphinc- terotomy (ES) is the standard method for the management of cho- ledocholithiasis. However, biliary stenting is used to treat patients with endoscopically irretrievable bile duct stones, especially elderly and high-risk patients. The aim of this study was to evaluate the benefits and risks of biliary stenting versus stone clearance follow- ing ES in the management of choledocholithiasis. PATIENTS AND METHODS: Between January 2010 and December 2012, 165 patients with common bile duct stones who underwent biliary stenting or stone clearance following ES were enrolled. One 7 Fr. double-pigtail plastic stent was placed without ES or stone extraction. The procedure time, hospitalization period, adverse events, additional endoscopic interventions required and one-year mortality were evaluated retrospectively. RESULTS: Ninety-nine and 66 patients were included in stenting group and in stone clearance group, respectively. Except for age, number of stones, and use of antithrombotic agents in the stent group, there were no statistically significant difference between groups. The average procedure time and hospitalization period in the stenting group were significantly shorter than those in stone clearance group (mean 21 min vs. 43.9 min, P < 0.0001; 3.8 days vs. 6.5 days, P < 0.0001). No significant differences were seen in ad- verse events and additional endoscopic interventions required be- tween both groups for at least a 1.5-year follow-up. No one-year mortality occurred. CONCLUSIONS: Biliary stenting using a double-pigtail stent proved to be a useful alternative therapy to stone clearance following ES in the management of choledocholithiasis in elderly patients.

Localization of the invadopodia-related proteins actinin-1 and cortactin to matrix-contact-side cytoplasm of cancer cells in surgically resected lung adenocarcinomas.

OBJECTIVES: Actin-associated proteins at cell-matrix-contact sites form invadopodia in cancer cells and participate in migration, matrix degradation and invasion. We investigated an alteration of subcellular localization of invadopodia-related actin-associated proteins, actinin-1 and cortactin, in lung adenocarcinomas, its clinical significance, and its possible regulatory factors. METHODS: Invadopodia-related proteins, actinin-1 and cortactin, were immunohistochemically examined in 90 cases of lung adenocarcinomas. Expression of invadopodia-associated proteins and their possible regulators in lung adenocarcinomas were examined by real-time RT-PCR, database search, and immunohistochemistry. RESULTS: Actinin-1 and cortactin showed matrix-contact-side localization in adenocarcinoma cells, but rarely in normal bronchiolar epithelial cells, alveolar cells, or precursor lesion atypical adenomatous hyperplasia cells. Immunoelectron-microscopic examination of adenocarcinoma cells revealed actinin-1 localization to matrix-contact-side cytoplasm with cytoplasmic protrusions. Matrix-contact-side localization of actinin-1 and cortactin was correlated with tumor stages, lymph node metastasis, vascular permeation, and loss of basement membrane. The tumor-specific survival rate was worse for the group in which matrix-contact-side localization of cortactin was high than for the low group. mRNA of the Rho guanine exchange factor epithelial cell transforming sequence-2 (Ect2) tended to be overexpressed in lung adenocarcinomas and cytoplasmic expression of Ect2 tended to be correlated with matrix-contact-side localization of actinin-1. CONCLUSION: Matrix-contact-side localization of invadopodia-related proteins in the lung adenocarcinoma cells were correlated with invasion, metastasis, and poor prognosis. Ect2 was a possible regulator of matrix-contact-side localization of invadopodia-related proteins.

Ras-association domain family protein 6 induces apoptosis via both caspase-dependent and caspase-independent pathways.

The Ras-association domain family (RASSF) comprises six members (RASSF1-6) that each harbors a RalGDS/AF-6 (RA) and Sav/RASSF/Hippo (SARAH) domain. The RASSF proteins are known as putative tumor suppressors but RASSF6 has not yet been studied. We have here characterized human RASSF6. Although RASSF6 has RA domain, it does not bind Ki-Ras, Ha-Ras, N-Ras, M-Ras, or TC21 under the condition that Nore1 (RASSF5) binds these Ras proteins. The message of RASSF6 is detected by RT-PCR in several cell lines including HeLa, MCF-7, U373, A549, and HepG2 cells, but the protein expression is low. The enhanced expression of RASSF6 causes apoptosis in HeLa cells. RASSF6 activates Bax and induces cytochrome C release. Caspase-3 activation is also induced, but the caspase inhibitor, Z-VAD-FMK, does not block RASSF6-mediated apoptosis. Apoptosis-inducing factor and endonuclease G are released from the mitochondria upon expression of RASSF6 and their releases are not blocked by Z-VAD-FMK. The knock down of RASSF6 partially blocks tumor necrosis factor-alpha-induced cell death in HeLa cells. These findings indicate that RASSF6 is implicated in apoptosis in HeLa cells and that it triggers both caspase-dependent and caspase-independent pathways.

Monoclonal antibody 7F9 recognizes rat protein homologous to human carboxypeptidase-M in developing and adult rat lung.

BACKGROUND AND OBJECTIVE: The purpose of this study was to obtain an antibody that would be useful for investigating the yet unclear molecular mechanism underlying the differentiation of lung alveolar type I and II cells. METHODS: Monoclonal antibodies were raised against membrane proteins from embryonal day 18.5 rat lungs and characterized by immunoblotting on rat lung lysates at various developmental stages to select an appropriate clone. The antigen of the selected antibody was purified by serial column chromatography and immunoprecipitation and identified by mass spectrometry. RESULTS: 7F9 antibody recognizes a 65-kDa protein that is expressed most prominently from embryonal day 20.5 to postnatal day 1. This protein was identified as a rat protein that is similar to 5730456K23Rik protein. The protein is homologous to human carboxypeptidase-M. Although human carboxypeptidase-M is known as a marker of type I cells, the expression of this rat protein was detected in columnar epithelial cells expressing type II cell markers, SP-C and a lamellar body protein ABCA3, in developing lung. Its expression was detected in alveolar cells lacking T1alpha, a type I cell marker protein, in adult lung. It was also expressed in RLE-6TN cells derived from type II cells. The expression in RLE-6TN cells was down-regulated by transforming growth factor-beta1 and up-regulated by Wnt3a. CONCLUSIONS: 7F9 antibody detects a protein in rat lung cells expressing type II markers. The antibody is a useful tool for studying signalling triggered by transforming growth factor-beta1 and Wnt3a in rat type II cells.

CIN85 is localized at synapses and forms a complex with S-SCAM via dendrin.

Membrane-associated guanylate kinase inverted (MAGI)-1 plays a role as a scaffold at cell junctions in non-neuronal cells, while S-SCAM, its neuronal isoform, is involved in the organization of synapses. A search for MAGI-1-interacting proteins by yeast two-hybrid screening of a kidney cDNA library yielded dendrin. As dendrin was originally reported as a brain-specific postsynaptic protein, we tested the interaction between dendrin and S-SCAM and revealed that dendrin binds to the WW domains of S-SCAM. Dendrin is known to be dendritically translated but its function is largely unknown. To gain insights into the physiological meaning of the interaction, we performed a second yeast two-hybrid screening using dendrin as a bait. We identified CIN85, an endocytic scaffold protein, as a putative dendrin-interactor. Immunocytochemistry and subcellular fractionation analysis supported the synaptic localization of CIN85. The first SH3 domain and the C-terminal region of CIN85 bind to the proline-rich region and the N-terminal region of dendrin, respectively. In vitro experiments suggest that dendrin forms a ternary complex with CIN85 and S-SCAM and that this complex formation facilitates the recruitment of dendrin and S-SCAM to vesicle-like structures where CIN85 is accumulated.

Receptor for advanced glycation end-products is a marker of type I lung alveolar cells.

Lung alveolar epithelial cells are comprised of type I (ATI) and type II (ATII) cells. ATI cells are polarized, although they have very flat morphology. The identification of marker proteins for apical and basolateral membranes of ATI cells is important to investigate into the differentiation of ATI cells. In this paper, we characterized receptor for advanced glycation end-products (RAGE) as a marker for ATI cells. RAGE was localized on basolateral membranes of ATI cells in the immunoelectron microscopy and its expression was enhanced in a parallel manner to the differentiation of ATI cells in vivo and in primary cultures of ATII cells. RAGE and T1 alpha, a well-known ATI marker protein, were targeted to basolateral and apical membranes, respectively, when expressed in polarized Madine Darby canine kidney cells. Moreover, RAGE was expressed in ATI cells after T1 alpha in vivo and in ex in vivo organ cultures. In conclusion, RAGE is a marker for basolateral membranes of well-differentiated ATI cells. ATI cells require some signal provided by the in vivo environment to express RAGE.