The rs662 polymorphism of paraoxonase 1 affects the difference in the inhibition of butyrylcholinesterase activity by organophosphorus pesticides in human blood.

OBJECTIVES: Organophosphorus pesticides (OPs) are a human health hazard. OPs inhibit acetylcholinesterase (AChE) at nerve endings and accumulate acetylcholine (ACh) at these sites. High levels of ACh and long exposure promote cholinergic crisis. The hydrolysis of OPs by serum paraoxonase 1 (PON1) plays a role in cholinergic crisis in humans. Human serum PON1 can break down organophosphate before binding to ChE. We investigated the effect of PON1 polymorphisms on AChE activity after OP treatment. METHODS: 50 healthy volunteers were randomly recruited with informed consent. We investigated butyrylcholinesterase (BuChE) activity changes in plasma as a biomarker of AChE after OP treatment in human blood samples immediately following blood sampling. After the standardization of BuChE activity in human blood, we correlated changes in BuChE activity with changes in blood pH. We analyzed the PON1 polymorphisms (rs854560 and rs662) of 50 participants to retrospectively investigate the interindividual variability of changes in BuChE activity. RESULTS: Changes in BuChE activity are strongly correlated with pH changes after OP treatment (R2 = 0.913). We used changes in pH as a surrogate marker for BuChE inhibition after OP treatment. OP treatment significantly decreased BuChE activity by 56.4 ± 5.1% (p < 0.001). The degree of BuChE inhibition was significantly different in the PON1 rs662 genotype (56.10 ± 4.74% vs. 57.96 ± 5.67% vs. 52.34 ± 1.51%; GG vs. GA vs. AA, respectively). CONCLUSION: Changes in pH can be used as a surrogate marker for the detection of BuChE inhibition after OP exposure. The rs662 polymorphism of PON1 may explain the inter-individual variability in BuChE inhibition.

Physiological properties of Scomber japonicus meat hydrolysate prepared by subcritical water hydrolysis.

The health-beneficial biological activities, including antioxidant and tyrosinase inhibitory activities, of Scomber japonicus muscle protein hydrolysates prepared by subcritical water hydrolysis were investigated. After 5 min of subcritical hydrolysis at 140 degrees C, 59.76% of S. japonicus muscle protein was hydrolyzed, the highest degree of hydrolysis in all the groups were tested. According to the response surface methodology results, as the reaction temperature and reaction time became lower and shorter, the yield became higher. The highest 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity (90.63%) occurred in hydrolysates treated at 140 degrees C for 5 min, and the highest tyrosinase inhibitory activity (65.54%) was identified in hydrolysates treated at 200 degreesC for 15 min. Changes in the molecular weight distribution of S. japonicus muscle proteins after subcritical water hydrolysis were observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Subcritical water hydrolysis is a suitable technique for obtaining S.japonicus muscle protein hydrolysates with useful biological activities, within a short time (5-15 min).

Pitavastatin Regulates Ang II Induced Proliferation and Migration via IGFBP-5 in VSMC.

Angiotensin II (Ang II), a key mediator of hypertensive, causes structural changes in the arteries (vascular remodeling), which involve alterations in cell growth, vascular smooth muscle cell (VSMC) hypertrophy. Ang II promotes fibrotic factor like IGFBP5, which mediates the profibrotic effects of Ang II in the heart and kidneys, lung and so on. The purpose of this study was to identify the signaling pathway of IGFBP5 on cell proliferation and migration of Ang II-stimulated VSMC. We have been interested in Ang II-induced IGFBP5 and were curious to determine whether a Pitavastatin would ameliorate the effects. Herein, we investigated the question of whether Ang II induced the levels of IGFBP5 protein followed by proliferation and migration in VSMC. Pretreatment with the specific Angiotensin receptor type 1 (AT1) inhibitor (Losartan), Angiotensin receptor type 2 (AT2) inhibitor (PD123319), MAPK inhibitor (U0126), ERK1/2 inhibitor (PD98059), P38 inhibitor (SB600125) and PI3K inhibitor (LY294002) resulted in significantly inhibited IGFBP5 production, proliferation, and migration in Ang II-stimulated VSMC. In addition, IGFBP5 knockdown resulted in modulation of Ang II induced proliferation and migration via IGFBP5 induction. In addition, Pitavastatin modulated Ang II induced proliferation and migration in VSMC. Taken together, our results indicated that Ang II induces IGFBP5 through AT1, ERK1/2, P38, and PI3K signaling pathways, which were inhibited by Pitavastatin. These findings may suggest that Pitavastatin has an effect on vascular disease including hypertension.

Valsartan independent of AT1 receptor inhibits tissue factor, TLR-2 and -4 expression by regulation of Egr-1 through activation of AMPK in diabetic conditions.

Patients suffering from diabetes mellitus (DM) are at a severe risk of atherothrombosis. Early growth response (Egr)-1 is well characterized as a central mediator in vascular pathophysiology. We tested whether valsartan independent of Ang II type 1 receptor (AT1R) can reduce tissue factor (TF) and toll-like receptor (TLR)-2 and -4 by regulating Egr-1 in THP-1 cells and aorta in streptozotocin-induced diabetic mice. High glucose (HG, 15 mM) increased expressions of Egr-1, TF, TLR-2 and -4 which were significantly reduced by valsartan. HG increased Egr-1 expression by activation of PKC and ERK1/2 in THP-1 cells. Valsartan increased AMPK phosphorylation in a concentration and time-dependent manner via activation of LKB1. Valsartan inhibited Egr-1 without activation of PKC or ERK1/2. The reduced expression of Egr-1 by valsartan was reversed by either silencing Egr-1, or compound C, or DN-AMPK-transfected cells. Valsartan inhibited binding of NF-κB and Egr-1 to TF promoter in HG condition. Furthermore, valsartan reduced inflammatory cytokine (TNF-α, IL-6 and IL-1ß) production and NF-κB activity in HG-activated THP-1 cells. Interestingly, these effects of valsartan were not affected by either silencing AT1R in THP-1 cells or CHO cells, which were devoid of AT1R. Importantly, administration of valsartan (20 mg/kg, i.p) for 8 weeks significantly reduced plasma TF activity, expression of Egr-1, TLR-2, -4 and TF in thoracic aorta and improved glucose tolerance of streptozotocin-induced diabetic mice. Taken together, we concluded that valsartan may reduce atherothrombosis in diabetic conditions through AMPK/Egr-1 regulation.

Inhibitory effects of seaweed extracts on the growth of the vaginal bacterium Gardnerella vaginalis.

Of 44 species of seaweed screened for potential anti-Gardnerella vaginalis activity, 27 (61.4%) showed antimicrobial activity by the agar disk-diffusion method. Among them, the strongest activities against the pathogen were exhibited by Chlorophyta, with Ulva pertusa producing an 11.3-mm zone of inhibition at 5 mg disk⁻¹. The MIC values of U. pertusa extracts against both G. vaginalis KCTC 5096 and KCTC 5097, the main cause of vaginosis, were 312 µg ml⁻¹, while the MIC values against both Candida albicans KCTC 7270 and KCTC 7965, the main cause of candidiasis, were 2.5 mg ml⁻¹. Against Lactobacillus gasseri KCTC 3173 and Lactobacillus jensenii KCTC 5194, members of the normal vaginal microflora, no inhibitory effect was seen even at 10 mg ml⁻¹. To identify the primary active compounds, a U. pertusa powder was successively fractionated according to polarity, and the main active agents against G. vaginalis KCTC 5096 were determined to be nitrogenous compounds (156 µg ml⁻¹ of the MIC value). According to these results, it was suggested that extracts of the seaweed U. pertusa are valuable for the development of natural therapeutic agents for treating women with bacterial vaginosis.

Seasonal variation of antibacterial activities in the green alga Ulva pertusa Kjellman.

The present study was performed to screen out the extracts of algae and assess the seasonal variation in antimicrobial activity of Ulva pertusa against Gardnerella vaginalis. Seasonal variation in antibacterial activity was observed, with the extracts showing no activity during summer and autumn, and showing antibacterial activity from early winter (December) to middle spring (April). The maximum value of antimicrobial activity (6.5 mm inhibition zone at 5 mg disk(-1)) of U. pertusa against G. vaginalis was observed in April. Otherwise, for both chlorophyll a and b, the highest content (2.87 mg g(-1) and 1.37 mg g(-1)) was observed in March 2009. These results may reflect variation in cellular chemical compositions such as secondary metabolite(s) rather than chlorophyll and biological activities according to season.

High glucose induces connective tissue growth factor expression and extracellular matrix accumulation in rat aorta vascular smooth muscle cells via extracellular signal-regulated kinase 1/2.

Connective tissue growth factor (CTGF) is a potent pro-fibrotic factor, which is implicated in fibrosis through extracellular matrix (ECM) induction in diabetic cardiovascular complications. It is an important downstream mediator in the fibrotic action of transforming growth factor ß (TGFß) and is potentially induced by hyperglycemia in human vascular smooth muscle cells (VSMCs). Therefore, the goal of this study is to identify the signaling pathways of CTGF effects on ECM accumulation and cell proliferation in VSMCs under hyperglycemia. We found that high glucose stimulated the levels of CTGF mRNA and protein and followed by VSMC proliferation and ECM components accumulation such as collagen type 1, collagen type 3 and fibronectin. By depleting endogenous CTGF we showed that CTGF is indispensable for the cell proliferation and ECM components accumulation in high glucose-stimulated VSMCs. In addition, pretreatment with the MEK1/2 specific inhibitors, PD98059 or U0126 potently inhibited the CTGF production and ECM components accumulation in high glucose-stimulated VSMCs. Furthermore, knockdown with ERK1/2 MAPK siRNA resulted in significantly down regulated of CTGF production, ECM components accumulation and cell proliferation in high glucose-stimulated VSMCs. Finally, ERK1/2 signaling regulated Egr-1 protein expression and treatment with recombinant CTGF reversed the Egr-1 expression in high glucose-induced VSMCs. It is conceivable that ERK1/2 MAPK signaling pathway plays an important role in regulating CTGF expression and suggests that blockade of CTGF through ERK1/2 MAPK signaling may be beneficial for therapeutic target of diabetic cardiovascular complication such as atherosclerosis.

Single oral dose toxicity test of polycalcium, a mixed composition of polycan and calcium lactate-gluconate 1:9 (G/G) in SD rat.

The object of this study was to obtain acute oral toxicity information of Polycalcium, a mixed composition of Polycan and Calcium lactate-gluconate 1:9 (g/g), in Sprague-Dawely (SD) rats. In order to investigate the toxicity and identify target organs, Polycalcium were once orally administered to female and male SD rats at dose levels of 2000, 1000, 500 and 0 (control) mg/kg body weights. The mortality, changes on body weight and clinical signs were monitored during 14 days after treatment with gross observation, changes on the organ weights and histopathology of principle organs and treatment sites based on the recommendation of KFDA Guidelines [2009-116, 2009]. As the results of single oral treatment of Polycalcium, no treatment related mortalities were observed within 14 days after end of treatment up to 2000 mg/kg, the limited dosage of rodents in the both genders. In addition, no Polycalcium treatment related changes on the body and organ weights, clinical signs, necropsy and histopathological findings were detected. The results obtained in this study suggest that the Polycalcium is non-toxic in rats. The LD50 and approximate LD in rats after single oral dose of Polycalcium were considered over 2000 mg/kg in both female and male, respectively.

Higenamine reduces HMGB1 during hypoxia-induced brain injury by induction of heme oxygenase-1 through PI3K/Akt/Nrf-2 signal pathways.

Growing lines of evidence suggests that high mobility group box-1 (HMGB1) plays an important role for promoting inflammation and apoptosis in brain ischemia. Previously, we demonstrated that inducers of heme oxygenase-1 (HO-1) significantly reduce HMGB1 release in inflammatory conditions in vitro and in vivo. Thus, we tested our hypothesis that higenamine protects brain injury by inhibition of middle cerebral artery occlusion (MCAO)-mediated HMGB1 release in vivo, and glucose/glucose oxidase (GOX)-induced apoptosis in C6 cells in vitro due to HO-1 induction. Higenamine increased HO-1 expression in C6 cells in both hypoxia and normoxia, in which the former was much more significant than the latter. Higenamine increased Nrf-2 luciferase activity, translocated Nrf-2 to nucleus, and increased phosphorylation of Akt in C6 cells. Consistent with this, LY 294002, a PI3K inhibitor, inhibited HO-1 induction by higenamine and apoptosis induced by glucose/GOX in C6 cells was prevented by higenamine, which effect was reversed by LY 294002. Importantly, administration of higenamine (i.p) significantly reduced brain infarct size, mortality rate, MPO activity and tissue expression of HMGB1 in MCAO rats. In addition, recombinant high mobility group box 1 induced apoptosis in C6 cells by increasing ratio of Bax/bcl-2 and cleaved caspase c, which was inhibited by higenamine, and all of these effects were reversed by co-treatment with ZnPPIX. Therefore, we conclude that higenamine, at least in part, protects brain cells against hypoxic damages by up-regulation of HO-1. Thus, higenamine may be beneficial for the use of ischemic injuries such as stroke.

Inhibition of oral pathogens and collagenase activity by seaweed extracts.

Fifty-seven species of common seaweed from the Coast of Korea were screened for antimicrobial (i.e. inhibition of Prevotella intermedia and Porphyromonas gingivalis growth) activity. As a source of bioactive compounds, seaweeds can produce many secondary metabolites with a variety of activities. Using the agar diffusion method, only 17 species (29.8%) showed inhibitory activity. Of these, methanol extracts of Enteromorpha linza, Sargassum sagamianum, and Ulva pertusa showed strong inhibitory effects against both P. intermedia and P. gingivalis. The MIC values of E. linza, S. sagamianum, and U. pertusa extracts against P. intermedia were 625, 78 and 625 microg ml(-1) and those against P. gingivalis were 312, 156 and 625 microg ml(-1), respectively. When these three species' extracts were separated into five fractions according to their polarity, the main active agents were determined to be phenolic compounds. We then compared the antimicrobial activities of these phenolic compounds against various periodontal pathogens using a MIC test. Phenolic compound containing extracts at concentrations of 10 to 100 microg ml(-1) showed a moderate to significant inhibitory effect on collagenase 1,2 and 3 activity.

Easy, Fast Self-Heating Polyurethane Nanocomposite with the Introduction of Thermally Annealed Carbon Nanotubes Using Near-Infrared Lased Irradiation.

In this study, high-crystallinity single walled carbon nanotubes (H-SWNTs) were prepared by high-temperature thermal annealing at 1800 °C and a self-heating shape memory polyurethane nanocomposite with excellent self-heating characteristics was developed within a few seconds by irradiation with near-infrared rays. With a simple method (heat treatment), impurities at the surface of H-SWNTs were removed and at the same time the amorphous structure converted into a crystalline structure, improving crystallinity. Therefore, high conductivity (electric, thermal) and interfacial affinity with PU were increased, resulting in improved mechanical, thermal and electric properties. The electrical conductivity of neat polyurethane was enhanced from ~10-11 S/cm to 4.72 × 10-8 S/cm, 1.07 × 10-6 and 4.66 × 10-6 S/cm, while the thermal conductivity was enhanced up to 60% from 0.21 W/mK, 0.265 W/mK and 0.338 W/mK for the composites of 1, 3 and 5 wt%, respectively. Further, to achieve an effective photothermal effect, H-SWNTs were selected as nanofillers to reduce energy loss while increasing light-absorption efficiency. Thereafter, near-infrared rays of 818 nm were directly irradiated onto the nanocomposite film to induce photothermal properties arising from the local surface plasmon resonance effect on the CNT surface. A self-heating shape memory composite material that rapidly heated to 270 °C within 1 min was developed, even when only 3 wt.% of H-SWNTs were added. The results of this study can be used to guide the development of heat-generating coating materials and de-icing materials for the wing and body structures of automobiles or airplanes, depending on the molding method.

Artificial Intelligence, Wearables and Remote Monitoring for Heart Failure: Current and Future Applications.

Substantial milestones have been attained in the field of heart failure (HF) diagnostics and therapeutics in the past several years that have translated into decreased mortality but a paradoxical increase in HF-related hospitalizations. With increasing data digitalization and access, remote monitoring via wearables and implantables have the potential to transform ambulatory care workflow, with a particular focus on reducing HF hospitalizations. Additionally, artificial intelligence and machine learning (AI/ML) have been increasingly employed at multiple stages of healthcare due to their power in assimilating and integrating multidimensional multimodal data and the creation of accurate prediction models. With the ever-increasing troves of data, the implementation of AI/ML algorithms could help improve workflow and outcomes of HF patients, especially time series data collected via remote monitoring. In this review, we sought to describe the basics of AI/ML algorithms with a focus on time series forecasting and the current state of AI/ML within the context of wearable technology in HF, followed by a discussion of the present limitations, including data integration, privacy, and challenges specific to AI/ML application within healthcare.

Rapid and Local Self-Healing Ability of Polyurethane Nanocomposites Using Photothermal Polydopamine-Coated Graphene Oxide Triggered by Near-Infrared Laser.

In this study, we report the self-healing ability of polyurethane (PU) nanocomposites based on the photothermal effect of polydopamine-coated graphene oxide (PDA-rGO). Polydopamine (PDA) was coated on the graphene oxide (GO) surface, while simultaneously reducing GO by the oxidation of dopamine hydrochloride in an alkaline aqueous solution. The PDA-rGO was characterized by Fourier-transform infrared spectroscopy, X-ray diffraction, Raman spectroscopy, thermogravimetric analysis, and scanning electron microscopy-energy-dispersive X-ray analysis. PDA-rGO/PU nanocomposites with nanofiller contents of 0.1, 0.5 and 1 wt% were prepared by ex situ mixing method. The photothermal effect of the PDA-rGO in the PU matrix was investigated at 0.1 W/cm2 using an 808 nm near-infrared (NIR) laser. The photothermal properties of the PDA-rGO/PU nanocomposites were superior to those of the GO/PU nanocomposites, owing to an increase in the local surface plasmon resonance effect by coating with PDA. Subsequently, the self-healing efficiency was confirmed by recovering the tensile stress of the damaged nanocomposites using the thermal energy generated by the NIR laser.

Changes in Gait Parameters and Gait Variability in Young Adults during a Cognitive Task while Slope and Flat Walking.

This study investigates the effects of a cognitive task while walking on a slope or a flat surface on gait parameters and gait variability in young adults. The participants consisted of thirty healthy young subjects. They were instructed to walk on a slope or on a flat surface while performing or not performing a cognitive task, which involved speaking a four-syllable word in reverse. A wearable inertia measurement unit (IMU) system was used to measure spatiotemporal parameters and gait variability. Flat gait (FG) while performing the cognitive task (FGC) and uphill gait (UG) while performing the cognitive task (UGC) significantly altered stride times, gait speeds, and cadence as compared with FG and UG, respectively. Downhill gait (DG) while performing the cognitive task (DGC) caused no significant difference as compared with DG. Gait variability comparisons showed no significant difference between UGC and UG or between FGC and FG, respectively. On the other hand, variabilities of stride times and gait speeds were significantly greater for DGC than DG. FGC and UGC induce natural changes in spatiotemporal gait parameters that enable the cognitive task to be performed safely. DGC should be regarded as high complexity tasks involving greater gait variability to reduce fall risk.

EP2 receptor activation by prostaglandin E2 leads to induction of HO-1 via PKA and PI3K pathways in C6 cells.

Recently we proposed that COX-2 induction precedes expression of HO-1 in ischemic preconditioned rat brain. In the current study, we investigated the molecular mechanism by which prostaglandin E(2), one of COX-2 metabolites, induces HO-1 in rat C6 brain cells. We demonstrated that concentration of PGE(2) increased HO-1 expression in C6 cells in vitro. The effects of PGE(2) were mimicked by PGE(2) receptor EP(2) agonists, 11-deoxy PGE(2), and cAMP analog, dibutyl-cAMP. HO-1 expression by PGE(2) was inhibited by LY294002, PI3K inhibitor and H89, PKA inhibitor. The EP(2)-specific antagonist, AH8006 also inhibited PGE(2)-mediated HO-1 expression in a concentration-dependent manner. Finally, PGE(2) inhibited GOX-induced apoptosis as assayed by FACS analysis or DNA strand breaks assay, and this cell death was reversed by ZnPPIX, HO-1 inhibitor. In addition to HO-1 induction, PGE(2) also increased phosphorylation of Bad by PKA- and PI3K-depednent manner. Taken together, we conclude that PGE(2) induces HO-1 protein expression through PKA and PI3K signaling pathways via EP(2) receptor in C6 cells. The induction of HO-1 along with increase of p-Bad by PGE(2) is responsible for anti-apoptosis against oxidant stress.

Propofol limits rat myocardial ischemia and reperfusion injury with an associated reduction in apoptotic cell death in vivo.

Propofol, a rapidly acting, short duration, intravenous hypnotic anesthetic induction agent, is often used in clinical situations where myocardial ischemia/ reperfusion (I/R) injury is a threat. The aim of the present study was to evaluate the protective effect of propofol on myocardial I/R injury in rat due to apoptosis. Myocardial I/R injury were induced by occluding the left anterior descending (LAD) coronary artery for 25 min followed by either 2 h or 6 h reperfusion. Apoptosis was evaluated by Western blot analysis (Bcl-2, Bax expression), DNA strand breaks, TUNEL analysis and measuring myocardial caspase-3 activity. Propofol significantly reduced infarct size and improved I/R-induced myocardial contractile dysfunction by improving left ventricular diastolic pressure and positive and negative maximal values of the first derivative (+dp/dt) of left ventricular pressure. Propofol increased Bcl-2/Bax expression ratio and decreased caspase-3 activity in I/R rat hearts, which resulted in reduction of myocardial apoptosis as evidenced by TUNEL analysis and DNA laddering experiments. In an in vitro study, propofol increased H9c2 cell viability against oxidative stress induced by glucose oxidase (GOX) in a dose-dependent manner. These data suggest propofol limits I/R injury with an associated reduction in apoptotic cell death in vivo.

Anti-apoptotic effect of magnolol in myocardial ischemia and reperfusion injury requires extracellular signal-regulated kinase1/2 pathways in rat in vivo.

Magnolol, an active component extracted from Magnolia officinalis, has been reported to have protective effect on ischemia and reperfusion (I/R)-induced injury in experimental animals. The aim of the present investigation was to further evaluate the mechanism(s) by which magnolol reduces I/R-induced myocardial injury in rats in vivo. Under anesthesia, left anterior descending (LAD) coronary artery was occluded for 30 min followed by reperfusion for 24 h (for infarct size and cardiac function analysis). In some experiments, reperfusion was limited to 1 h or 6 h for analysis of biochemical and molecular events. Magnolol and DMSO solution (vehicle) were injected intra-peritoneally 1 h prior to I/R insult. The infarct size was measured by TTC technique and heart function was monitored by Millar Catheter. Apoptosis related events such as p-ERK, p-Bad, Bcl-xl and cytochrome c expression were evaluated by Western blot analysis and myocardial caspase-3 activity was also measured. Magnolol (10 mg/kg) reduced infarct size by 50% (P < 0.01 versus vehicle), and also improved I/R-induced myocardial dysfunction. Left ventricular systolic pressure and positive and negative maximal values of the first derivative of left ventricular pressure (dP/dt) were significantly improved in magnolol-treated rats. Magnolol increased the expression of phosphor ERK and Bad which resulted in inhibition of myocardial apoptosis as evidenced by TUNEL analysis and DNA laddering experiments. Application of PD 98059, a selective MEK1/2 inhibitor, strongly antagonized the effect of magnolol. Taken together, we concluded that magnolol inhibits apoptosis through enhancing the activation of ERK1/2 and modulation of the Bcl-xl proteins which brings about reduction of infarct size and improvement of cardiac function in I/R-induced injury.

Vaccination of shrimp (Penaeus chinensis) against white spot syndrome virus (WSSV).

Two structural protein genes, VP19 and VP466, of white spot syndrome virus (WSSV) were cloned and expressed in Sf21 insect cells using a baculovirus expression system for the development of injection and oral feeding vaccines against WSSV for shrimps. The cumulative mortalities of the shrimps vaccinated by the injection of rVP19 and rVP466 at 15 days after the challenge with WSSV were 50.2% and 51.8%, respectively. For the vaccination by oral feeding of rVP19 and rVP466, the cumulative mortalities were 49.2% and 89.2%, respectively. These results show that protection against WSSV can be generated in the shrimp, using the viral structural protein as a protein vaccine.

Neutralization of white spot syndrome virus (WSSV) for Penaeus chinensis by antiserum raised against recombinant VP19.

This study was carried out to neutralize the WSSV one of the most virulent pathogen causing large economic damage in shrimp culture industry using the antiserum produced against recombinant WSSV envelope protein VP19 (rVP19) as a tool to evaluate WSSV infection mechanism. A fragment of VP19 was expressed in Sf21 insect cell using baculovirus expression system as fusion protein with 6 His-tag. Then, polyclonal antiserum against rVP19 was raised in white rabbit. A constant amount of WSSV (at 10(4) diluted stock) was incubated with various antiserum concentrations and injected into shrimp, Penaeus chinensis, for the neutralization challenge. At 9 days post injection, the shrimp in the positive control injected with WSSVshowed 100% mortality The shrimps injected with WSSV preincubated with preimmune serum showed 83.3% mortality at 15 days post injection. The shrimps injected with the WSSV preincubated with 1 microl, 5 microl or 10 microl r VP19 antiserum and shrimp mortalities showed 66.6%, 40.0% and 26.6% at 15 days post injection, respectively The high concentration of antiserum group showed lower mortality than those of the low concentration of antiserum group. This indicates that the WSSV can be neutralized by the rVP19 antiserum in a dose-dependent manner. The neutralization challenge result suggested that VP19 might play an important role in WSSV infection to shrimp.

Production of polyclonal antibody against recombinant ORF 112 L of rock bream (Oplegnathus fasciatus) iridovirus (RBIV) and in vitro neutralization.

Rock bream iridovirus (RBIV) is a causative agent of epizootics among cultured rock bream (Oplegnathus fasciatus) in Korea. In this study, the immunogenic property of ankyrin repeats gene (ORF 112L) from RBIV was evaluated to develop vaccines against RBIV. ORF 112L of RBIV was cloned into expression vector of pGEX-4T-1. The recombinant protein was successfully expressed using E. coli BL21 (DE3). The soluble recombinant RBIV protein was applied to affinity column for the purification of the protein. Mice were immunized by the injection of purified recombinant protein to produce polyclonal antibodies. EUSA was carried out to identify the immune reaction abilities of polyclonal antibody to recombinant protein. The antigenic property of this protein was evaluated by using in vitro neutralization with BF-2 cells. In neutralization test, BF-2 cells infected with the mixture of RBIV and antisera containing anti-GST-ORF 112L polyclonal antibody were healthy showing few cytopathic effect (CPE) similar with the negative control (without RBIV). These studies suggest that the protein from the ankyrin repeats gene, ORF 112L of RBIV may play an important role in the mechanism of infection. Also, it can be possible to develop protein or gene vaccines using ORF 112L against RBIV.