Successful repair of ascending aortic pseudoaneurysm using autograft patch from fascia lata and saphenous vein

An 80-year-old man developed a pseudoaneurysm in the ascending aorta due to mediastinitis following cardiac surgery. We successfully repaired the pseudoaneurysm with an autograft patch harvested from fascia lata and the saphenous vein. The repair, which was carried out in two layers, can be expected to be durable.     

Pharmacokinetic analysis of transcellular transport of levofloxacin across LLC-PK1 and Caco-2 cell monolayers

To characterize the membrane transport responsible for the renal excretion and intestinal absorption of levofloxacin, we performed pharmacokinetic analysis of transcellular transport across LLC-PK(1) and Caco-2 cell monolayers. Transcellular transport of levofloxacin in LLC-PK(1) cells was greater in the basolateral-to-apical direction than in the opposite direction. Pharmacokinetic analysis indicated that basolateral uptake was the direction-determining step for the transcellular transport of levofloxacin in LLC-PK(1) cells. The apical efflux clearance of levofloxacin in LLC-PK(1) cells was increased at the medium pH 6 as compared with at pH 8, suggesting that membrane transport characteristics of levofloxacin are apparently similar to those of a prototypical organic cation, tetraethylammonium. On the other hand, transcellular transport of levofloxacin in Caco-2 cells was only slightly greater in the basolateral-to-apical direction than in the opposite direction. The apical efflux clearance of levofloxacin in Caco-2 cells was greater than basolateral efflux clearance, and apical influx clearance was greater than any other membrane transport clearance. In addition, the apical uptake of levofloxacin as well as quinidine in Caco-2 cells was inhibited significantly by nicotine and imipramine. The findings indicated that some transporters are responsible not only for the efflux but also for the influx of levofloxacin at the apical membrane of Caco-2 cells.     

Creation of novel cell-penetrating peptides for intracellular drug delivery using systematic phage display technology originated from Tat transduction domain

Many biologically active proteins need to be delivered intracellularly to exert their therapeutic action inside the cytoplasm. Cell penetrating peptides (CPPs) have been developed to efficiently deliver a wide variety of cargo in a fully biological active form into a range of cell types for the treatment of multiple preclinical disease models. To further develop this methodology, we established a systematic approach to identify novel CPPs using phage display technology. Firstly, we screened a phage peptide library for peptides that bound to the cell membrane. Secondly, to assess functionality as intracellular carriers, we recombined cDNAs of binding peptides with protein synthesis inhibitory factor (PSIF) to create fusion proteins. Randomly chosen clones were cultured and expression of peptide-PSIF fusion proteins induced, followed by screening of protein synthesis activity in cells. Using this systematic approach, novel and effective CPPs were rapidly identified. We suggest that these novel cell-penetrating peptides can utilized as drug delivery tools for protein therapy or an analytical tool to study mechanisms of protein transduction into the cytoplasm.     

Enhancement of peripheral nerve regeneration using bioabsorbable polymer tubes packed with fibrin gel

Nerve conduit tubes were developed using bioabsorbable polymer membranes, and the effects of tube shape--straight or bellows-shaped tubes--and the fibrin gel filling were investigated. The mechanical properties of the tubes were examined by in vitro tests, and their effectiveness for peripheral nerve regeneration was determined by grafting into experimentally transected sciatic nerves of rats. The bellows tube showed mechanically superior properties, and when used with the fibrin gel, it induced superior tissue formation of myelinated nerve fibers as compared to other tube types. The total area of myelinated axons regenerated in the fibrin-filled bellows tube was comparable to that of an isograft control, whereas those of the other tubes demonstrated inferior regeneration. This result suggests that the mechanically superior bellows tube filled with fibrin gel is an effective graft alternative for peripheral nerve regeneration.     

Hepatocellular adenoma in a male with familial adenomatous polyposis coli

Hepatocellular adenoma (HA) is a benign liver tumor most frequently occurring in young women using oral contraceptives. We report a rare case of HA in a 27-year-old male patient with familial adenomatous polyposis (FAP). The patient underwent a total colectomy and ileo-rectal anastomosis for FAP in 2003. A preoperative computed tomography scan of the abdomen disclosed a tumor in the left-lobe of the liver, 5.8 cm in diameter. Pathologic examination of a needle biopsy disclosed HA, but he had never used anabolic steroids or other known inducers of HA. The size of the liver mass gradually increased to 8.5 cm during a follow-up period of 38 months, and a left hepatectomy was performed in 2006. Pathology of the resected specimen confirmed the diagnosis of HA. Although FAP is known to be complicated with neoplasia in various extracolonic organs, only five reported cases of HA have developed in patients with FAP, including this case. This is the first report of HA to develop in a male FAP patient.     

Effect of ER-β gene disruption on estrogenic regulation of anxiety in female mice

It has been shown that long-term estrogen treatment in gonadectomized female mice increases anxiety levels. On the other hand, a recent study has reported that estrogen may down-regulate the levels of anxiety by acting through estrogen receptor (ER) β. In the present study, we investigated the role of ER-β in the regulation of anxiety levels in female mice after long-term estrogen treatment. Gonadectomized ER-β knockout (βERKO) female mice and their wild type (βWT) littermates were implanted several different doses (experiment 1: 2.0 μg/day, experiment 2: 1.0, 0.4, 0.2 or 0.1 μg/day) of an estradiol benzoate (EB) or placebo pellet. Ten days after pellet implant, behavioral tests commenced to measure the anxiety levels (experiment 1: light-dark transition test (LDT), experiment 2: LDT, elevated plus maze test (EPM) and social investigation test (SIT)). We found that, at higher-doses, long-term treatment of EB had anxiogenic effects in both βWT and βERKO mice as indicated by a decrease of the time spent in the light side and the number of transitions between two sides during LDT. In contrast, several behavioral measurements indicated that the lower-doses treatment of EB might reduce the anxiety levels possibly through ER-β. Particularly, the anxiolytic effects of EB in the SIT were more pronounced in βWT mice than βERKO mice. Together, the findings in the present study suggest that estrogen may have both anxiolytic and anxiogenic effects in female mice, and that ER-β gene disruption did not affect anxiogenic regulation by estrogen in female mice, but partially affected anxiolytic regulation.     

Weakness in intercellular association of keratinocytes in severely brittle nails

The brittle fingernail is a common complaint, but the features of the cellular structure of the nail plate remain unclear. In this study, clipped nailplates from two persons with severely brittle nails, one female aged 26 years and one male aged 82 years, were observed by light and electron microscopy and compared with normal nail plates. Numerous cracks were observed in clipped brittle nails, but not in normal nails, on light microscopy. When the deep areas of nail plates of the clipped normal nails were observed by electron microscopy, intercellular boundaries appeared intermingled, and two thin, electron-dense layers were observed in a narrow intercellular gap. In contrast, in brittle nails, marked dilatation of intercellular spaces was frequently observed and electron-dense layers were either not seen or were disrupted. When clipped normal nails were dehydrated in a desiccation chamber, similar dilatations - though not so severe -were observed, without evident cracks. These results suggest that dilatation of the intercellular space between nail keratinocytes is correlated with brittle nails and that dehydration may result in such intercellular dilatation.     

Galactose-PEG dual conjugation of beta-(1-->3)-D-glucan schizophyllan for antisense oligonucleotides delivery to enhance the cellular uptake

Antisense oligonucleotides (AS ODNs) are applied to silence a particular gene, and this approach is one of the potential gene therapies. However, naked oligonucleotides are easy to be degraded or absorbed in biological condition. Therefore, we need a carrier to deliver AS ODNs. This paper presents galactose moieties that were conjugated to the side chain of SPG to enhance cellular ingestion through endocytosis mediated by asialoglycoprotein receptor specifically located on parenchymal liver cells. We introduced galactose with two types of chemical bonds; amide and amine, and the amine connection showed lower ingestion and more toxicity than the amide one. Since PEG was known to induce endocytosis escape, we combined PEG and galactose aiming to provide both cellular up-take and subsequent endocytosis escape. We designed lactose or galactose moieties to attach to the end of the PEG chain that connects to the SPG side chain. When the PEG had the molecular weight of 5000-6000, the antisense effect reached the maximum. We believe that this new type of galactose and PEG dual conjugation broaden the horizon in antisense delivery.     

The function of connexin 43 on the differentiation of rat bone marrow cells in culture

Connexin (Cx) 43-mediated gap-junctional intercellular communication (GJC) mainly regulates the osteoblastic differentiation, but much of the function of Cx43 on the differentiation of bone marrow cells is unclear. This study is aimed to clarify relationship between the differentiation of rat bone marrow cells and the function of Cx43. Bone marrow cells derived from four-week-old Wistar strain rats were grown in the presence and absence of 18-alpha-glycyrrhetinic acid (AGA, 100 muM) to inhibit Cx43-mediated GJC. Expression of Cx43 gene and protein, and the level of intracellular cyclic adenosine monophosphate (cAMP) were determined as the assessment of the function in Cx43-mediated GJC, and alkaline phosphatase (ALP) activity and mineralization were measured as the assessment of osteoblastic differentiation. The Cx43 gene expression was first observed at 2 days, but under the condition in which rat bone marrow cells were treated with AGA, there was no significant effect on the Cx43 gene expression. By administrating AGA to rat bone marrow cells, all parameters of maturation but the Cx43 gene expression significantly decreased. The results of this experiment suggest that Cx43-mediated GJC plays a critical role in rat bone marrow cells, progress toward maturation.