The infectivity of progeny adenovirus in the presence of neutralizing antibody.

Human adenoviruses (Ads), common pathogens that cause upper respiratory and gastrointestinal infections, are blocked by neutralizing antibodies (nAbs). However, Ads are not fully eliminated even in hosts with nAbs. In this study, we assessed the infectivity of progeny Ad serotype 5 (Ad5) in the presence of nAb. The infectivity of Ad5 was evaluated according to the expression of the Ad genome and reporter gene. Infection by wild-type Ad5 and Ad5 vector continued to increase until 3 days after infection even in the presence of nAb. We established an assay for determining the infection levels of progeny Ad5 using a sorting system with magnetic beads and observed little difference in progeny Ad5 counts in the presence and absence of nAb 1 day after infection. Moreover, progeny Ad5 in the presence of nAb more effectively infected coxsackievirus and adenovirus receptor (CAR)-positive cells than CAR-negative cells. We investigated the function of fiber proteins, which are the binding partners of CAR, during secondary infection, observing that fibre proteins spread from infected cells to adjacent cells in a CAR-dependent manner. In conclusion, this study revealed that progeny Ad5 could infect cells even in the presence of nAb, differing from the common features of the Ad5 infection cycle. Our findings may be useful for developing new therapeutic agents against Ad infection.

Intestinal Perforation in a Patient with Colon Cancer during Treatment with Regorafenib: A Case Report and Review of the Literature.

The multikinase inhibitor, regorafenib, is known to exert its antitumor effects by targeting several kinases, inhibiting interstitial intracellular signaling and suppressing tumor cell proliferation. Regorafenib causes gastrointestinal perforation and gastrointestinal fistula as adverse events, and discontinuation is recommended if these adverse events occur during administration. However, there are no prescribed standards for re-administration after discontinuation and for administration in patients with a history of gastrointestinal perforation. Herein, we report a case of gastrointestinal perforation in a patient, with a history of gastrointestinal microperforation, undergoing bevacizumab therapy, within a few days of starting regorafenib; this had a significant effect on the prognosis. The site of gastrointestinal perforation was consistent with previously reported sites around the tumor and at the anastomotic site. Based on a review of literature and our experience with the case presented here, we recommend that administration of regorafenib to patients with a history of gastrointestinal perforation should be avoided to the extent possible. Moreover, in case of prior administration of a drug reported to cause gastrointestinal perforation, such as an anti-VEGFR drug, the risk of gastrointestinal perforation should be considered during the administration of regorafenib. In the event of complaints, such as abdominal pain, gastrointestinal perforation should be considered as a differential diagnosis and appropriate tests and treatments should be initiated at an early stage.

Characteristics of an Emulsion Obtained Using Hydrophobic Hydroxypropyl Methylcellulose as an Emulsifier and a High-Pressure Homogenizer.

Hydrophobically modified hydroxypropyl methylcellulose (HM-HPMC), a polymer in which a small amount of HPMC is stearoxyl substituted, was used as an emulsifier of emulsion-type lotion. A high-pressure homogenizer (microfluidizer) was used. The viscosity of the 1% HM-HPMC aqueous gel decreased after passing through the microfluidizer from 5.5 to 2.7 Pa·s. When liquid paraffin (LP) was used as the oil phase, a stable emulsion was obtained with an LP ratio of 1-40%. The apparent viscosity decreased with LP ratios up to 20%, and then increased with increasing LP concentration. The emulsions with an LP ratio <20% presented a pseudo-viscous flow, similar to that of the diluted polymer solution. HM-HPMC likely adsorbed onto the oil with a stearoxyl group; thus, the interaction between the stearoxyl group, which explained the high viscosity of HM-HPMC, decreased, reducing the viscosity of the emulsion. The LP ratio was 40%, and the emulsion presented a plastic flow, which is typical of concentrated emulsions. The size of the droplet in the emulsion was approximately 1 µm regardless of the LP ratio. When low-viscosity LPs or monoester-type oils such as isopropyl myristate were used, some of the emulsions presented creaming. An emulsion using HM-HPMC as an emulsifier and an appropriate oil homogenized with a microfluidizer is stable, has low viscosity, and can be easily spread on skin.

Characteristics of dynamic standing balance with and without an insole in patients with spastic diplegia cerebral palsy.

[Purpose] This study examined characteristics of dynamic standing balance, with an without an insole, in patients with spastic diplegia cerebral palsy (CP). [Participants and Methods] This cross-sectional study used a crossover design. Eleven patients with spastic diplegia CP and gross motor levels between I and III with spastic diplegia CP (according to the Gross Motor Function Classification System expanded and revised version) were randomly allocated to either the barefoot or insole groups. The Index of postural stability (IPS) was evaluated while each patient was barefoot and while wearing insoles. The Pediatric Evaluation of Disability Inventory (PEDI) was used to measure functional self-care and mobility domains. [Results] While wearing the insoles, the center movement distance between right and left positions was significantly higher. While barefoot, IPS and area of postural sway correlated with the PEDI subscales for mobility and self-care. [Conclusion] Insoles promote standing balance and dynamic balance to move the center of pressure within the base of support. Such improvements may enhance activities of daily living in patients with spastic diplegia CP.

Influence of Characteristics of Oily Vehicle on Skin Penetration of Ufenamate.

Skin penetration amounts of a highly lipophilic drug, ufenamate, prepared in four oily vehicles, including white petrolatum (WP), liquid paraffin (LP), isopropyl myristate (IPM), and isocetyl stearate (ICS), were compared. Ufenamate was mixed in each vehicle at 5% and applied at a rate of 2 mg/cm2 to intact, stripped, and delipidized Yucatan micropig skin. The amounts of ufenamate and IPM in the stratum corneum (SC), epidermis, and dermis were determined. The skin penetration amounts of ufenamate from liquid oils were significantly higher than those from WP; the amounts of ufenamate were in the order WP

Identification of Adenovirus-Derived Cell-Penetrating Peptide.

Cell-penetrating peptides (CPPs) have been highly anticipated as an efficient delivery system due to their ability to cross biological membranes and transport various cargoes into cells. In the present study, we have identified adenovirus-derived CPPs using various capsid-mutant adenovirus (Ad) vectors. First, we examined the endocytosis-inducing ability of these vectors. A fiber-shaft substituted Ad vector, Ad type 5 vector with the fiber shaft domain replaced by that derived from Ad type 35, induced the highest fluorescein isothiocyanate (FITC)-dextran uptake into a human liver cell line, HepG2 cells. In contrast, the FITC-dextran uptake in HepG2 cells was not significantly different between coxsackievirus and adenovirus receptor (CAR)-binding-ablated Ad vector, integrin-binding-ablated Ad vector or conventional Ad vector. Next, we produced a recombinant Ad type 35 shaft protein using the Escherichia coli recombinant system. The recombinant Ad type 35 shaft protein retained the ability for FITC-dextran uptake and efficient gene delivery by plasmid transfection reagent. Furthermore, we identified 26 C-terminal amino acids in the Ad type 35 shaft protein as the cell membrane binding domain. The 26 amino-acid peptides also have the potential to be internalized into cultured cells. The internalization ability of the peptide was dependent on degree and was inhibited by an actin polymerization inhibitor (Latrunculin B) and by a lipid raft formation inhibitor (methyl-ß-cyclodextrin). The results of the present study indicate that Ad type 35-derived peptides induce endocytosis in cultured cells and have the ability to cross biological membranes. This report is the first paper to identify Ad-derived CPPs.

[A Case of Merkel Cell Carcinoma Complicated with Severe Thrombocytopenia Treated with Carboplatin/Etoposide Regimen after Surgery].

Thrombocytopenia is often caused by myelosuppression during chemotherapy. However, when platelet transfusions are required, pathological conditions such as idiopathic thrombocytopenic purpura(ITP)and thrombotic thrombocytopenic purpura( TTP)also occur. We report a case of Merkel cell carcinoma complicated with severe thrombocytopenia treated with carboplatin/etoposide regimen after surgery. The patient's platelet count did not increase in spite of platelet transfusions. However, the platelet count increased after steroid treatment was chosen under the diagnosis of ITP. Subsequent examinations revealed that the patient had HLA antibody, which caused the platelet transfusion refractoriness. When the platelet count does not increase in spite of platelet transfusions during chemotherapy, the possibility that the platelet transfusion refractoriness is due to the presence of HLA antibody should be considered.

Preparation and characterization of a powder containing an oily liquid drug with Eudragit EPO or L100 copolymer.

Oily liquid drugs are not convenient for oral administration. We developed a powder containing clofibrate (CF), a model of an oily drug, using aminoalkyl methacrylate copolymer (EPO) or methacrylic acid copolymer (L100). CF or a mixture of CF and soybean oil was emulsified with EPO or L100 aqueous solution. Using a high-pressure homogenizer, a stable emulsion was obtained, and a powder was then obtained by lyophilization of the emulsion. The content of CF in the powder depended on the formulation, with the highest contents being 24.6% and 27.1% for EPO and L100, respectively. The incorporation ratio of CF was higher for L100 than for EPO. The powder using EPO was sticky because of leaked CF and the low glass transition temperature of EPO. The powder using L100 was a typical powder obtained by lyophilization. The leakage of CF from the powder was <2%, lower than for EPO powder. The dissolution of CF from powder using EPO was fast, regardless of the pH of the medium, but the powder using L100 showed enteric-soluble characteristics, indicating that CF is well incorporated in L100.

Penetration of Ufenamate into Intact, Stripped, or Delipidized Skin Using Different Vehicles.

The purpose of this study was to clarify the effect of skin condition on skin penetration of the very high lipophilic drug, ufenamate (UF). UF was applied to stripped or delipidized skin using liquid paraffin (LP) or purified water containing polysorbate 80 at a dose of 2 µL/cm(2). We found that UF penetration into intact and stripped skin using a water vehicle was respectively 5 and 10 times higher than that using LP. UF is freely soluble in oil and insoluble in water; thus, activity in water is higher than that in LP. Therefore, it is useful to use a water-based vehicle for both intact sites and those with defective stratum corneum (SC). Conversely, we found that delipidization of SC decreased the penetration of UF significantly with both LP and water, and the amount measured in the epidermis was 1 µg/cm(2) with both vehicles. This indicates that UF is not suitable for so-called "dry skin." This study revealed clinically relevant differences in the penetration of UF into intact, stripped, or delipidized skin conditions.

Efficient Adenovirus Gene Transfer Methods in Human Colonic Caco-2 Epithelial Cells Using Capric Acid.

Adenovirus (Ad) vectors are widely used in gene therapy and in vitro/in vivo gene transfer. However, Ad-mediated gene transfer in epithelial cells shows low efficiency, because Ad fiber cannot bind to the primary receptor, the coxsackievirus and adenovirus receptor (CAR), present in tight junctions. Caco-2 monolayer cells cultured on Transwell-chamber plates for approximately 2 weeks are widely used for drug membrane permeation studies, but Ad-mediated gene transfer is difficult in Caco-2 monolayer cells. First, we examined the efficiency of gene transfer into Caco-2 monolayer cells. Luciferase production in cultured Caco-2 cells transduced with Ad vectors was 20-fold lower on day 12 than on day 1. In contrast, the expression of CAR protein in Caco-2 cells gradually increased along with the duration of culture. For efficient gene transfer into Caco-2 monolayer cells, the binding ability of Ad vectors with CAR was found to be important. Capric acid (C10), a medium-chain fatty acid is a tight-junction modulator used as a pharmaceutical agent. We found that a novel gene transfer method using transduction with Ad vectors in the presence of C10 led more efficiently to LacZ expression in Caco-2 monolayer cells than Ad vectors alone. The results of the present study indicate that C10 could be very useful for Ad-mediated gene transfer in human colonic Caco-2 epithelial cells.

Sec61ß regulates barrier functions of tight junction through expression of claudin-4 in Madin-Darby canine kidney cells.

Sec61ß is the ß subunit of the Sec61 translocon and is responsible for expression and delivery of basolateral membrane proteins, including claudins, major constituents of tight junction (TJ). In the present study, the effect of Sec61ß overexpression on TJ barrier functions in Madin-Darby canine kidney (MDCK) cells were investigated by monitoring transepithelial electrical resistance (TER) and the expression and distribution of claudins. We adopted the time required by TER to reach 50% (T1/2) as a measure of TJ modulation rate. Sec61ß overexpression increased TER by post-transcriptionally upregulating claudin-4 expression and resulted in increased TER. Sec61ß overexpression increased TJ modulation rates (lower T1/2), in conjunction with enhanced delivery of claudin-4 from and to plasma membranes. Marked co-distribution and indirect association of claudin-4 with Sec61ß were observed, contributing to the enhanced delivery of claudin-4. Thus, Sec61ß may be a novel TJ modulation target, including barrier function and modulation rates for drug delivery systems.

Effects of oils and emulsifiers on the skin penetration of stearyl glycyrrhetinate in oil-in-water emulsions.

We investigated whether an emulsifier or an emulsified oil affects the skin penetration of stearyl glycyrrhetinate (SG) when it is applied in an oil-in-water (O/W)-type emulsion under finite dose conditions in vitro. SG has a high molecular weight (MW: 723) and high lipophilicity (log P: 15.6). Skin penetration of SG applied with O/W emulsions was evaluated using 6 types of emulsifiers that are commonly used in cosmetics; however, no significant differences were observed between these emulsifiers. When applied with liquid paraffins in oil phase, SG skin penetration increased significantly as the molecular weight of the liquid paraffin decreased. The skin penetration of the fluorescent dye 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI; MW: 834, log P: 23.2) also increased with O/W-type emulsions containing liquid paraffins of lower molecular weights. These results indicate that use of O/W-type emulsions with an appropriate oil phase can improve SG skin penetration.

Effect of storage conditions on the recrystallization of drugs in solid dispersions with crospovidone.

The physical stability of amorphous solid dispersions (SDs) is influenced by their storage conditions. The goal of this work was to investigate the factors affecting the recrystallization of drugs in SDs after storage under conditions of high temperature and high humidity. SDs of three drugs (dipyridamole, nifedipine and indomethacin) with different functional groups (amino, carbonyl and hydroxyl) and onset times for crystallization of the amorphous state were prepared using crospovidone (CrosPVP). All of the drugs in the SDs remained in an amorphous state at 25 °C/50% relative humidity (RH) in closed glass bottles for at least six months. Under conditions of high temperature (40 °C/75%RH/closed and 60 °C/open), differences in interactions between the hydrogen bond donors of the drugs and the amide carbonyl group of CrosPVP are essential factors for recrystallization of the drugs in the SDs. On the other hand, under condition of high humidity (40 °C/75%RH/open), in addition to the difference in the interaction between the drug and CrosPVP, the rate of increase in moisture content affects their recrystallization in SDs.

Percolation analysis in electrical conductivity of Madin-Darby canine kidney and Caco-2 cells by permeation-enhancing agents.

The control of permeability through the paracellular route has been paid great attention to for enhanced bioavailability of macromolecular and hydrophilic drugs. The paracellular permeability is controlled by tight junctions (TJ), and claudins are the major constituents of TJ. Despite numerous studies on TJ modulation, the dynamics is not well understood, although it could be crucial for clinical applications. Here, we studied the time (t) course of electrical conductivity (Σ) in a monolayer of Madin-Darby canine kidney (MDCK) and Caco-2 cells upon treatment with modulators, the C-terminus fragments of Clostridium perfringens enterotoxin (C-CPE) and sodium caprate (C10). For C-CPE treatment, Σ remains approximately constant, then starts increasing at t=tc (percolation threshold). For C10, on the other hand, Σ increases to 1.6-2.0 fold of the initial value, stays constant, and then starts increasing again for both MDCK and Caco-2 cells at t=tc. We find that this behavior can be explained within a framework of percolation, where Σ shows a logarithmic dependence on t-tc with the power of µ; µ denotes the critical exponent. We obtain µ=1.1-1.2 regardless of cell type or modulator. Notably, µ depends only on the dimensionality (d) of the system, and these values correspond to those for d=2. Percolation is thus the operative mechanism for the increase in Σ through TJ modulation. The findings provide fundamental knowledge, not only on controlled drug delivery, but also on bio-nanotechnologies including the fabrication of biological devices.

Mental arithmetic modulates temporal variabilities of finger-tapping tasks in a tempo-dependent manner.

Background: Several psychiatric diseases impair temporal processing. Temporal processing is thought to be based on two domains: supra-second intervals and sub-second intervals. Studies show that temporal processing in sub-second intervals is mainly an automated process. However, the brain functions involved in temporal processing at each time scale remain unclear. We hypothesized that temporal processing in supra-second intervals requires several brain areas, such as the ventrolateral prefrontal cortex, intraparietal sulcus (IPS), and inferior parietal lobe, corresponding to various cognitions in a time scale-dependent manner. We focused on a dual-task paradigm (DTP) involving simultaneous performance of cognitive and motor tasks, which is an effective method for screening psychomotor functions; we then designed a DTP comprising finger tapping at various tempi as the temporal processing task and two cognitive tasks (mental arithmetic and reading) that might affect temporal processing. We hoped to determine whether task-dependent interferences on temporal processing in supra-second intervals differed depending on the cognitive tasks involved. Methods: The study included 30 participants with no history of neuromuscular disorders. Participants were asked to perform a DTP involving right index finger tapping at varying tempi (0.33, 0.5, 1, 2, 3, and 4 s inter-tapping intervals). Cognitive tasks comprised mental arithmetic (MA) involving three-digit addition, mental reading (MR) of three- to four-digit numbers, and a control (CTL) task without any cognitive loading. For comparison between tasks, we calculated the SDs of the inter-tapping intervals. Participants' MA abilities in the three-digit addition task were evaluated. Results: The MA and MR tasks significantly increased the SDs of the inter-tapping intervals compared to those of the CTL task in 2-3 s and 3-4 s for the MA and MR tasks, respectively. Furthermore, SD peaks in the finger-tapping tasks involving MA were normalized by those in the CTL task, which were moderately correlated with the participants' MA ability (r = 0.462, P = 0.010). Discussion: Our results established that DTP involving the temporal coordination of finger-tapping and cognitive tasks increased temporal variability in a task- and tempo-dependent manner. Based on the behavioral aspects, we believe that these modulations of temporal variability might result from the interaction between finger function, arithmetic processing, and temporal processing, especially during the "pre-semantic period". Our findings may help in understanding the temporal processing deficits in various disorders such as dementia, Parkinson's disease, and autism.

The relationship between spatiotemporal gait parameters and cognitive function in healthy adults: protocol for a cross-sectional study.

BACKGROUND: Motor dysfunctions, such as slower walking speed, precede the occurrence of dementia and mild cognitive impairment, suggesting that walking parameters are effective biomarkers for detecting early sub-clinical cognitive risk. It is often also concurrent with self-complained cognitive dysfunction, called motoric cognitive risk (MCR) syndrome. Our preliminary study found several walking parameters, obtained by a three-dimensional motion capture system, to be correlated with computer-based assessments of various cognitive function modalities, although the sample size was small. The Cognitive-Gait (CoGait) Database Project, described in the current protocol, aims to establish a database of multi-dimensional walking and cognitive performance data, collected from a large sample of healthy participants, crucial for detecting early sub-clinical cognitive risk. METHODS: We will recruit healthy volunteers, 20 years or older, without any neurological musculoskeletal or psychiatric disorders. The estimated sample size is 450 participants, including a 10% attrition rate. Using computer-based cognitive assessments, participants will perform six tasks: (i) the simple reaction time task, (ii) Go/No-Go task, (iii) Stroop Color-Word Test, (iv) N-back test, (v) Trail Making Test, and (vi) digit span test. We will also conduct paper-based cognitive assessments such as the Mini-Mental State Examination, Montreal Cognitive Assessment, and the Geriatric Depression Scale-15 for assessing MCR. Gait will be measured through joint kinematics and global positioning in participants' lower legs while walking at a comfortable and faster pace, using pants with an inertial measurement unit-based three-dimensional motion capture system. Finally, we will establish a prediction model for various cognitive performance modalities based on walking performance. DISCUSSION: This will be the first study to reveal the relationship between walking and cognitive performance using multi-dimensional data collected from a large sample of healthy adults, from the general population. Despite certain methodological limitations such as the accuracy of measurements, the CoGait database is expected to be the standard value for both walking and cognitive functions, supporting the evaluation of psychomotor function in early sub-clinical cognitive risk identification, including motoric-cognitive risk syndrome.

Effect of particle size of drug on conversion of crystals to an amorphous state in a solid dispersion with crospovidone.

The effect of particle size on amorphization of drugs in a solid dispersion (SD) was investigated for two drugs, indomethacin (IM) and nifedipine (NP). The SD of drugs were prepared in a mixture with crospovidone by a variety of mechanical methods, and their properties investigated by particle sizing, thermal analysis, and powder X-ray diffraction. IM, which had an initial particle size of 1 µm and tends to aggregate, was forced through a sieve to break up the particles. NP, which had a large initial particle size, was jet-milled. In both cases, reduction of the particle size of the drugs enabled transition to an amorphous state below the melting point of the drug. The reduction in particle size is considered to enable increased contact between the crospovidone and drug particles, increasing interactions between the two compounds.