In vivo anti-influenza virus activity of Kampo (Japanese herbal) medicine “Sho-seiryu-to” and its mode of action

When BALB/c mice were treated with a Kampo (Japasese herbal) medicine “Sho-seiryu-to” (SST) (2 g/kg, 10 times) orally from 7 days before to 4 days after the infection and infected with mouse-adapted influenza virus A/PR/8/34 by nasal site-restricted infection, replication of the virus in the nasal cavity and spread of the virus to the lung were efficiently inhibited at 5 days after infection in comparison with water-treated mice. However, another Kampo medicine “Kakkon-to” showed no anti-influenza virus activity in the same condition. The antiviral IgA antibody in the nasal and broncho-alveolar washes of the SST treated mice increased significantly in comparison with that of water-treated control. Oral administration of SST (2 g/kg, 18 times) from 7 days before to 13 days after vaccination also significantly augmented serum hemagglutination-inhibiting antibody by nasal inoculation of influenza HA vaccine (5 μg/mouse) that was insufficient to induce antiviral antibody. SST did not inhibit the replication of mouse-adapted influenza virus A/PR/8/34 in Madin-Darby canine kidney cells. SST also did not inhibit the influenza virus sialidase activity against sodium p-nitrophenyl-N-acetyl-α-D-neuraminate and hemagglutination by mouse-adapted influenza virus A/PR/8/34. SST showed no influence on interferon production in nasal wash of mice at 5 days after the virus infection. These results suggest that SST confers better protection against influenza virus infection through augmentation of production of antiviral IgA antibody but not direct action to the virus, and can be used as an adjuvant to nasally inoculated influenza HA vaccine.     

In vitro and in vivo antibacterial activity of tigecycline against Vibrio vulnificus

Background/purpose

The aim of this study is to investigate the role of tigecycline in Vibrio vulnificus infection.

Utilization of frozen sections in the evaluation of chronic epilepsy–related cases

The role of frozen section consultation in the evaluation of chronic epilepsy–associated surgical excisions of brain tissue has not been previously examined. The study retrospectively reviews 335 cases in which a frozen section consultation was obtained in the setting of a resection for chronic epilepsy. In most cases (n = 323), 3 or fewer frozen sections were performed. The most commonly identified pathologies on final diagnosis included tumor or tumorlike lesions (79.1% of cases) and focal cortical dysplasia (20.9% of cases). Frozen section diagnoses discrepant with final diagnoses due to sampling error or misinterpretation were noted in 39 cases and most commonly involved a diagnosis of gliosis or tumor in the setting of a focal cortical dysplasia or diagnosis of gliosis in the setting of a low-grade tumor. In conclusion, frozen section consultation may be particularly useful in the evaluation of neoplasms arising in the setting of chronic epilepsy. Some epilepsy-associated pathology, such as focal cortical dysplasia, may be difficult to diagnose at the time of frozen section and such cases may not be an ideal target for intraoperative frozen section consultation.     

RNA interference inhibits yellow fever virus replication in vitro and in vivo

RNA interference (RNAi) is a process that is induced by double stranded RNA and involves the degradation of specific sequences of mRNA in the cytoplasm of the eukaryotic cells. It has been used as an antiviral tool against many viruses, including flaviviruses. The genus Flavivirus contains the most important arboviruses in the world, i.e., dengue (DENV) and yellow fever (YFV). In our study, we investigated the in vitro and in vivo effect of RNAi against YFV. Using stable cell lines that expressed RNAi against YFV, the cell lines were able to inhibit as much as 97% of the viral replication. Two constructions (one against NS1 and the other against E region of YFV genome) were able to protect the adult Balb/c mice against YFV challenge. The histopathologic analysis demonstrated an important protection of the central nervous system by RNAi after 10 days of viral challenge. Our data suggests that RNAi is a potential viable therapeutic weapon against yellow fever.

Overview of in vivo and ex vivo endpoints in murine food allergy models: Suitable for evaluation of the sensitizing capacity of novel proteins?

Significant efforts are necessary to introduce new dietary protein sources to feed a growing world population while maintaining food supply chain sustainability. Such a sustainable protein transition includes the use of highly modified proteins from side streams or the introduction of new protein sources that may lead to increased clinically relevant allergic sensitization. With food allergy being a major health problem of increasing concern, understanding the potential allergenicity of new or modified proteins is crucial to ensure public health protection. The best predictive risk assessment methods currently relied on are in vivo models, making the choice of endpoint parameters a key element in evaluating the sensitizing capacity of novel proteins. Here, we provide a comprehensive overview of the most frequently used in vivo and ex vivo endpoints in murine food allergy models, addressing their strengths and limitations for assessing sensitization risks. For optimal laboratory-to-laboratory reproducibility and reliable use of predictive tests for protein risk assessment, it is important that researchers maintain and apply the same relevant parameters and procedures. Thus, there is an urgent need for a consensus on key food allergy parameters to be applied in future food allergy research in synergy between both knowledge institutes and clinicians.     

Effects of fibrin pad hemostat on the wound healing process in vivo and in vitro

Fibrin Pad is a hemostatic pad designed to control surgical-related bleeding. It consists of a fully absorbable composite matrix scaffold coated with human-derived active biologics that immediately form a fibrin clot upon contact with targeted bleeding surfaces. Studies were conducted to investigate the effect of Fibrin Pad and its biologics-free composite matrix component (Matrix) on the wound healing process in in vitro and in vivo models. Fibrin Pad was evaluated in solid organ, soft tissue defects, and subcutaneous tissues. Immunocompromised rodents were used to avoid xeno-mediated responses. Extracts created from both materials were evaluated for biological activity using in vitro cell culture assays. Neither Fibrin Pad nor Matrix alone showed any inhibition of the wound healing of treated defect sites. An apparent accelerated healing was noted in the soft tissue and subcutaneous tissue defects with Fibrin Pad as compared to Matrix. Both materials showed desirable properties associated with tissue scaffolds. The in vitro study results show that Fibrin Pad extract can induce dose-dependent increases in fibroblast proliferation and migration. These studies confirm that the biologic components of Fibrin Pad can enhance wound healing processes in in vitro assays and fully support wound healing at the site of in vivo application.     

Indocyanine green-loaded biodegradable tumor targeting nanoprobes for in vitro and in vivo imaging

Indocyanine green (ICG) is a near-infrared (NIR) ?uorescence dye for extensive biological application, but limited by its poor aqueous stability in?vitro, concentration-dependent aggregation, rapid elimination from the body, and lack of target specificity. In this paper, to overcome these limitations, folate receptor-targeted, ICG dye-doped poly(d,l-lactide-co-glycolide) (PLGA) lipid nanoparticles (FA-ICG-PLGA-lipid NPs) were constructed by a single-step self-assemble and nanoprecipitation method. The prepared FA-ICG-PLGA-lipid NPs exhibited good biocompatibility, monodispersity, excellent NIR penetration ability, significant stability against photobleaching and long circulation time. The intracellular uptake experiment proved the targeting efficacy of the FA-ICG-PLGA-lipid NPs was more effective in folate receptor over-expressing MCF-7 cells than folate receptor negative A549 cells. Furthermore, the in?vivo experiments showed the FA-ICG-PLGA-lipid NPs were specifically targeted to the tumor, and its circulation time was much longer than free ICG. These biocompatible and biodegradable NIR-NPs prove a potential application in tumor diagnosis and targeted imaging due to its high aqueous stability, excellent NIR optical properties and significantly targeting property in?vivo.     

Ultrabright and ultrastable near-infrared dye nanoparticles for in vitro and in vivo bioimaging

We report a new strategy of using carrier-free pure near-infrared (NIR) dye nanoparticles (NPs) to achieve highly luminescent NIR fluorescent probes for in?vitro and in?vivo imaging. Bis(4-(N-(2-naphthyl)phenylamino) phenyl)-fumaronitrile (NPAPF) NPs are shown to exhibit favorable biocompatibility, wide-range pH stability (pH 4-10) and much more superior photostability than conventional dyes. Importantly, the combined merits of high dye loading content and aggregation-induced emission enhancement properties, endow the NIR probes with high brightness and a high quantum yield up to 14.9%. The NPAPF NPs can be readily conjugated with folic acid for targeted in?vitro cell imaging. Applications of the NPs probes in high efficiency in?vivo and ex?vivo imaging were successfully demonstrated. Intense fluorescent signals of NPAPF NPs can be distinctly, selectively and spatially resolved in tumor sites with ultrahigh sensitivity, even with 5?ms exposure time, due to the preferentially accumulation of NPs in tumor sites through passive enhanced permeability and retention effect. The totality of results clearly demonstrate the exciting potential of the functionalized NPAPF NPs as a NIR fluorescent probe for in?vitro and in?vivo imaging and diagnostics.     

Physical activity evaluation in Yugoslav Study of the Precursors of Atherosclerosis in School Children – YUSAD study

Introduction

It is observed that there is a lack of physical activity and exercise in children, stressing higher prevalence of childhood obesity. The purpose of the study was to evaluate duration of physical activity in a child population and correlation of dynamics in physical activity during 5 years of follow-up in the same population.

Material and methods

We evaluated 3243 school children from 12 regional centres across Serbia. The first examination was done when the children were 10 years old (baseline group), while the second examination was done on the same population when children were 15 years old. Physical activity was classified as recreational activity after school. We analysed 3 groups regarding physical activity: a group of children who were physically active less than 1 hour per day (group I), a second group active from 1 hour to < 3 hours per day (group II), and a third group active ≥ 3 hours per day (group III).

Results

In our study we have found on examination that the majority of children were physically active between 1 and 3 hours per day. Our results indicate that there is significant movement from groups I and III toward group II on the second examination regarding the proportion in the baseline group. There is a significant increase in the number of children in group I as they get older.

Conclusions

School children in Serbia are physically active predominantly between 1 and 3 hours per day at the age between 10 and 15 years.     

The novel HBx mutation F30V correlates with hepatocellular carcinoma in vivo,reduces hepatitis B virus replicative efficiency and enhances anti-apoptotic activity of HBx N terminus in vitro

ObjectiveWe aimed to investigate HBx genetic elements correlated with hepatitis B virus (HBV) -related hepatocellular carcinoma (HCC) and their impact on (a) HBV replicative efficiency, (b) HBx binding to circular covalently closed DNA (cccDNA), (c) apoptosis and cell-cycle progression, and (d) HBx structural stability.MethodsThis study included 123 individuals chronically infected with HBV: 27 with HCC (77.9% (21/27) genotype D; 22.1% (6/27) genotype A) and 96 without HCC (75% (72/96) genotype D; 25.0% (24/96) genotype A). HepG2 cells were transfected by wild-type or mutated linear HBV genome to assess pre-genomic RNA (pgRNA) and core-associated HBV-DNA levels, HBx-binding onto cccDNA by chromatin immunoprecipitation-based quantitative assay, and rate of apoptosis and cell-cycle progression by cytofluorimetry.ResultsF30V was the only HBx mutation correlated with HCC (18.5% (5/27) in HCC patients versus 1.0% (1/96) in non-HCC patients, p 0.002); a result confirmed by multivariate analysis. In vitro, F30V determined a 40% and 60% reduction in pgRNA and core-associated HBV-DNA compared with wild-type (p <0.05), in parallel with a significant decrease of HBx binding to cccDNA and decreased HBx stability. F30V also decreased the percentage of apoptotic cells compared with wild-type (14.8 ± 6.8% versus 19.1 ± 10.1%, p <0.01, without affecting cell-cycle progression) and increased the probability of HBx-Ser-31 being phosphorylated by PI3K-Akt kinase (known to promote anti-apoptotic activity).ConclusionsF30V was closely correlated with HBV-induced HCC in vivo, reduced HBV replicative efficiency by affecting HBx-binding to cccDNA and increased anti-apoptotic HBx activity in vitro. This suggests that F30V (although hampering HBV's replicative capacity) may promote hepatocyte survival, so potentially allowing persistent production of viral progeny and initiating HBV-driven hepatocarcinogenesis. Investigation of viral genetic markers associated with HCC is crucial to identify those patients at higher risk of HCC, who hence deserve intensive liver monitoring and/or early anti-HBV therapy.     

Viral agents causing lower respiratory tract infections in hospitalized children: evaluation of the Speed-Oligo? RSV assay for the detection of respiratory syncytial virus

Respiratory syncytial virus (RSV) is the viral agent which is more frequently involved in lower respiratory tract infections (LRTIs) in infants under 1 year of age in developed countries. A new oligochromatographic assay, Speed-Oligo? RSV, was designed and optimized for the specific detection and identification of RSV subtypes A and B. The test was evaluated in 289 clinical samples from 169 hospitalized children using an immunochromatography (IC) test, virus isolation by culture, and an in-house real-time polymerase chain reaction (RT-PCR). Other viruses causing LRTIs were investigated by cell culture or PCR-based tests. Sixty-two patients were infected by RSV (36.7%). In addition, adenovirus, influenza B, parainfluenza 2, and human metapneumovirus were detected in rates ranging from 5 to 8%. A proportion of 10.1% of the patients had mixed infections. The sensitivity, specificity, and positive and negative predictive values were, respectively, 94.9, 99.4, 98.9, and 97.4% for Speed-Oligo? RSV, 92.9, 96.3, 92.9, and 96.3% for RT-PCR/RSV, and 58.4, 98.1, 93.3, and 82.6% for IC. Our rates of viral detection and co-infection were similar to those of previously reported series. Finally, we find that Speed-Oligo? RSV is a rapid and easy-to-perform technique for the detection of RSV and the identification of subtypes A and B.     

Cost-effectiveness of rifampin for 4 months and isoniazid for 9 months in the treatment of tuberculosis infection

The purpose of this study was to evaluate the cost-effectiveness of the strategy of controlling the contacts of tuberculosis patients with latent tuberculosis infection by means of treatment with rifampin for 4 months or isoniazid for 9 months. The cost was the sum of the cost of treating latent tuberculosis infection in all contacts plus the cost of treating tuberculosis in whom the disease was not avoided. The effectiveness was expressed as cases avoided. The efficacy adopted was 90 % for rifampin for 4 months and 93 % for isoniazid for 9 months. We carried out a sensitivity analysis for efficacies of rifampin for 4 months of 80 %, 75 %, 69 % and 65 %. Of the 1,002 patients studied, 139 were treated with rifampin for 4 months and 863 were treated with isoniazid for 9 months. The cost-effectiveness was €436,842.83/50 cases avoided with rifampin for 4 months and €692,164.42/40 cases avoided with isoniazid for 9 months. Rifampin for 4 months was dominant. In the sensitivity analysis, rifampin for 4 months was dominant for efficacies of 75 % or greater. The cost-effectiveness analysis favoured the use of rifampin for 4 months when its efficacy was 75 % or greater.     

PEI protected aptamer molecular probes for contrast-enhanced in vivo cancer imaging

Aptamers have emerged as promising molecular probes for cancer diagnosis. However, their application for in?vivo cancer imaging remains limitation due to the poor stability in blood and the degradation by nucleases. In the present study, we generated PEI/aptamer molecular complexes for cancer imaging in?vivo by using deoxyribonuclease (DNase)-activatable fluorescence probes (DFProbes) to monitor DNA degradation. The results showed that the complexes with PEI at the N/P ratio from 3.8 to 15 effectively prevented the degradation of DFProbes both in?vitro and in?vivo. Moreover, PEI successfully protected TD05 aptamers from DNase degradation without affecting its specific recognition of Ramos cells. In tumor bearing mice, PEI/aptamer molecular complexes further demonstrated superior passive tumor targeting and extended circulation time as compared with free aptamer. Hence, the well-defined PEI/aptamer probe is a novel strategy to deliver targeted aptamer for tumor diagnosis and imaging in?vivo.