Visual detection of white spot syndrome virus using DNA-functionalized gold nanoparticles as probes combined with loop-mediated isothermal amplification

The integration of loop-mediated isothermal amplification (LAMP) and DNA-functionalized AuNPs as visual detection probes (LAMP–AuNPs) was developed and applied for the detection of white spot syndrome virus (WSSV) from Penaeid shrimp in this study. The principle of this combination assay relies on the basis of stability characteristics of the DNA-functionalized AuNPs upon hybridization with the complementary target DNA toward salt-induced aggregation. If the detected target DNA is not complementary to the ssDNA probes, the DNA-functionalized AuNPs will be aggregated due to the screening effect of salt, resulting in the change of solution color from red to blue/gray and shift of the surface plasmon peak to longer wavelength. While the DNA-functionalized AuNPs are perfectly matched to the detected target DNA, the color of solution still remains red in color and no surface plasmon spectral shift. This assay provides simply technique, time-saving and its detection results could be achieved qualitatively and quantitatively by visualization using the naked eye due to the colorimetric change and by measurement using the UV–vis spectroscopy due to the surface plasmon spectral shift, respectively. In this study, LAMP–AuNPs assay was successfully developed with the detection of WSSV-LAMP generated product at 0.03 μg/reaction, and showed the sensitivity of 2 × 10² copies WSSV plasmid DNA, that is comparable to the most sensitive method reported to date. The LAMP–AuNPs assay described in this study revealed a highly sensitive, rapid and reliable diagnostic protocol for detection of WSSV. This technique has a potential as a routine method for assessing the infectious diseases in Penaeid shrimp not only for WSSV, but also for other shrimp pathogens, and can be useful tool in field conditions for the diagnosis or surveillance programs.     

Mixed lineage kinase domain-like is a key receptor interacting protein 3 downstream component of TNF-induced necrosis

Tumor necrosis factor (TNF) is an important inflammatory cytokine and induces many cellular responses, including inflammation, cell proliferation, apoptosis, and necrosis. It is known that receptor interacting protein (RIP) kinases, RIP1 and RIP3, are key effectors of TNF-induced necrosis, but little is known about how these two RIP kinases mediate this process, although reactive oxygen species (ROS) generation and JNK activation have been suggested to be two downstream events of RIP kinases. Here we report the identification of mixed lineage kinase domain-like, MLKL, as a key RIP3 downstream component of TNF-induced necrosis. Through screening a kinase/phosphatase shRNA library in human colon adenocarcinoma HT-29 cells, we found that knockdown of MLKL blocked TNF-induced necrosis. Our data suggest that MLKL functions downstream of RIP1 and RIP3 and is recruited to the necrosome through its interaction with RIP3. Finally, we found that MLKL is required for the generation of ROS and the late-phase activation of JNK during TNF-induced necrosis. However, because these two events are not involved in TNF-induced necrosis in HT-29 cells, the target of MLKL during TNF-induced necrosis remains elusive. Taken together, our study suggests that MLKL is a key RIP3 downstream component of TNF-induced necrotic cell death.     

Protective effects of garlic extract on cardiac function,heart rate variability,and cardiac mitochondria in obese insulin-resistant rats

Purpose

Garlic has been shown to exhibit antioxidant effects and cardioprotective properties. However, the effects of garlic extract on the heart in insulin resistance induced by long-term high-fat-diet consumption are not well defined. Therefore, we sought to determine the effects of garlic extract in the obese insulin-resistant rats.

Methods

Male Wistar rats (180–200 g) were divided into two groups: normal-diet or high-fat-diet (n = 24/group) fed for 12 weeks. Rats in each groups were divided into three subgroups (n = 8 each): vehicle or garlic extract (250 or 500 mg/kg/day, respectively) treated for 28 days. At the end of the treatment, the metabolic parameters, heart rate variability (HRV), cardiac function, and cardiac mitochondrial function were determined.

Results

Rats that received a high-fat-diet for 12 weeks had increased body weight, visceral fat, plasma insulin levels, total cholesterol, oxidative stress levels, depressed HRV, and cardiac mitochondrial dysfunction. Garlic extract at both concentrations significantly decreased the plasma insulin, total cholesterol, homeostasis model assessment index, and oxidative stress levels. Furthermore, garlic extract at both doses restored the HRV, cardiac function, and cardiac mitochondrial function.

Conclusion

We concluded that garlic extract at both concentrations exerted cardioprotective effects against cardiac dysfunction and mitochondrial dysfunction in obese insulin-resistant rats.     

The effects of acetylcholinesterase inhibitors on the heart in acute myocardial infarction and heart failure: From cells to patient reports

Cardiovascular diseases remain a major cause of morbidity and mortality worldwide. Cardiovascular diseases such as acute myocardial infarction, ischaemia/reperfusion injury and heart failure are associated with cardiac autonomic imbalance characterized by sympathetic overactivity and parasympathetic withdrawal from the heart. Increased parasympathetic activity by electrical vagal nerve stimulation has been shown to provide beneficial effects in the case of cardiovascular diseases in both animals and patients by improving autonomic function, cardiac remodelling and mitochondrial function. However, clinical limitations for electrical vagal nerve stimulation exist because of its invasive nature, costly equipment and limited clinical validation. Therefore, novel therapeutic approaches which moderate parasympathetic activities could be beneficial for in the case of cardiovascular disease. Acetylcholinesterase inhibitors inhibit acetylcholinesterase and hence increase cholinergic transmission. Recent studies have reported that acetylcholinesterase inhibitors improve autonomic function and cardiac function in cardiovascular disease models. Despite its potential clinical benefits for cardiovascular disease patients, the role of acetylcholinesterase inhibitors in acute myocardial infarction and heart failure remediation remains unclear. This article comprehensively reviews the effects of acetylcholinesterase inhibitors on the heart in acute myocardial infarction and heart failure scenarios from in vitro and in vivo studies to clinical reports. The mechanisms involved are also discussed in this review.     

Chitin biomass-nifedipine amorphous solid dispersion for enhancement of hydrophobic drug dissolution in aqueous media

Solid dispersions of poorly-water soluble nifedipine using microparticles fabricated from chitin biomass were prepared to improve the drug dissolution in an aqueous medium. After the drug loading and solvent evaporation, the drug loading capacity of 1, 6, and 19 %w/w was obtained. In the microstructure, the drug was dispersed on the porous chitin carrier with minor agglomeration leading to reduced crystallization and improved dissolution, nearly 100%, as compared to the pure nifedipine powder and the physical mixture with chitin. There was an interaction between functional groups of the drug and polymer, consequently the release prolonged for the entire 6 h with the maximum drug solubility of about 300 μg/ml. An in vitro release study showed that nifedipine effectively released from chitin into a simulated gastric releasing medium (pH 1.2). In this case, the release mechanism was best fitted with a Zero-order model based on the Fick's Law in the first 2 h. This research indicated that employing microparticle obtained from chitin biomass in the solid dispersion system lowered the drug recrystallization, facilitated drug release, and provided the sustainability in pharmaceutical science.     

Anesthetic management for small bowel enteroscopy in a World Gastroenterology Organization Endoscopy Training Center

AIM:To study the anesthetic management of patients undergoing small bowel enteroscopy in the World Gastroenterology Organization(WGO) Endoscopy Training Center in Thailand.METHODS:Patients who underwent small bowel enteroscopy during the period of March 2005 to March 2011 in Siriraj Gastrointestinal Endoscopy Center were retrospectively analyzed.The patients’ characteristics,pre-anesthetic problems,anesthetic techniques,anesthetic agents,anesthetic time,type and route of procedure and anesthesia-related complications were assessed.RESULTS:One hundred and forty-four patients underwent this procedure during the study period.The mean age of the patients was 57.6 ± 17.2 years,andmost were American Society of Anesthesiologists(ASA) class Ⅱ(53.2%).Indications for this procedure were gastrointestinal bleeding(59.7%),chronic diarrhea(14.3%),protein losing enteropathy(2.6%) and others(23.4%).Hematologic disease,hypertension,heart disease and electrolyte imbalance were the most common pre-anesthetic problems.General anesthesia with endotracheal tube was the anesthetic technique mainly employed(50.6%).The main anesthetic agents administered were fentanyl,propofol and midazolam.The mean anesthetic time was 94.0 ± 50.5 min.Single balloon and oral(antegrade) intubation was the most common type and route of enteroscopy.The anesthesia-related complication rate was relatively high.The overall and cardiovascular-related complication rates including hypotension in the older patient group(aged ≥ 60 years old) were significantly higher than those in the younger group.CONCLUSION:During anesthetic management for small bowel enteroscopy,special techniques and drugs are not routinely required.However,for safety reasons anesthetic personnel need to optimize the patient’s condition.     

Dengue Virus Infection of Mice: Morphology and Morphogenesis of Dengue Type-2 Virus in Suckling Mouse Neurones

In dengue type-2 virus-infected neurones of suckling mice, formation of single-membrane vesicles is observed in the distended cisternae of the endoplasmic reticulum mostly of the perinuclear zone around 72 h after inoculation. Electron-dense 50-nm virus particles are arranged in chains in these distended cisternae; some form small crystalloid aggregates. Aberrant particles of different shapes are also seen in the distended cisternae about the same time that the virus particles appear. Parallel filamentous structures are occasionally observed in the cisternae that contain very few virions, either characteristic or aberrant. Increasing cytopathic changes are present after 75 to 96 h. There is an intense vesicular formation. Large numbers of virions and aberrant particles are seen either in the endoplasmic reticulum cisternae or smooth membrane vesicles. They are spread throughout the neurocytoplasm, extending into the dendrites. Dengue virions which are enclosed in fairly intact membrane-bound vesicles are released during cytolysis of the neurones. Morphogenesis of dengue virus type 2 is discussed.     

Impact of mutations in DNA gyrase genes on quinolone resistance in Campylobacter jejuni

Amino acid substitutions providing quinolone resistance to Campyloabcter jejuni have been found in the quinolone resistance‐determining region of protein DNA gyrase subunit A (GyrA), with the highest frequency at position 86 followed by position 90. In this study, wild‐type and mutant recombinant DNA gyrase subunits were expressed in Escherichia coli and purified using Ni‐NTA agarose column chromatography. Soluble 97 kDa GyrA and 87 kDa DNA gyrase subunit B were shown to reconstitute ATP‐dependent DNA supercoiling activity. A quinolone‐inhibited supercoiling assay demonstrated the roles of Thr86Ile, Thr86Ala, Thr86Lys, Asp90Asn, and Asp90Tyr amino acid substitutions in reducing sensitivity to quinolones. The marked effect of Thr86Ile on all examined quinolones suggested the advantage of this substitution in concordance with recurring isolation of quinolone‐resistant C. jejuni. An analysis of the structure‐activity relationship showed the importance of the substituent at position 8 in quinolones to overcome the effect of Thr86Ile. Sitafloxacin (SIT), which has a fluorinate cyclopropyl ring at R‐1 and a chloride substituent at R‐8, a characteristic not found in other quinolones, showed the highest inhibitory activity against all mutant C. jejuni gyrases including ciprofloxacin‐resistant mutants. The results suggest SIT as a promising drug for the treatment of campylobacteriosis caused by CIP‐resistant C. jejuni. Copyright © 2016 John Wiley & Sons, Ltd.