Plant-expressed Zika virus envelope protein elicited protective immunity against the Zika virus in immunocompetent mice.

Zika virus infection causes multiple clinical issues, including Guillain-Barré syndrome and neonatal malformation. Vaccination is considered as the only strategy for the prevention of ZIKV-induced clinical issues. This study developed a plant-based recombinant vaccine that transiently expressed the ZIKV envelope protein (ZikaEnv:aghFc) in Nicotiana benthamiana and evaluated the protective immunity afforded by it in immunocompetent mice. ZikaEnv:aghFc induced both humoral and cellular immunity at a low dose (1-5 µg). This immune-inducing potential was enhanced further when adjuvanted CIA09A. In addition, antigen-specific antibodies and neutralizing antibodies were vertically transferred from immunized females to their progeny and afforded both protective immunity to ZIKV and cross-protection to Dengue virus infection. These results suggest that our plant-based ZIKV vaccine provides a safe and efficient protective strategy with a competitive edge.

EGT022, an RGD-containing recombinant disintegrin, inhibits the VEGF-induced angiogenic process by targeting integrin ß3 in endothelial cells.

OBJECTIVES: EGT022, an RGD-containing recombinant disintegrin from human ADAM metallopeptidase domain 15 (ADAM15), has been reported to stimulate vascular maturation of retinal blood vessels with promotion of pericyte coverage through binding to integrin αIIbß3. Previous studies have reported that angiogenesis can be inhibited by several RGD motif-containing disintegrins; however, the effect of EGT022 on Vascular endothelial growth factor (VEGF)-induced angiogenesis has not yet been determined. This study was conducted in order to evaluate the anti-angiogenic function of EGT022 in VEGF-induced endothelial cells. METHODS: A proliferation and migration assay was performed using human umbilical vein endothelial cells (HUVEC) cells stimulated with VEGF to determine whether the angiogenic process was suppressed by EGT022. An in vitro trans-well assay and Mile's permeability assay were performed to determine the effect of EGT022 on permeability. Western blot was performed in order to further determine whether EGT022 can inhibit phosphorylation of VEGF receptor-2 (VEGFR2) and Phospholipase C gamma1 (PLC-γ1). An integrin binding assay and luciferase assay were performed for identification of the integrin target of EGT022. RESULTS: Angiogenesis including proliferation, migration, tube formation, and permeability was significantly inhibited by EGT022 in HUVEC cells. Our findings also demonstrated that EGT022 binds directly to integrin αvß3, induces dephosphorylation of integrin ß3, and inhibits phosphorylation of VEGFR2. In addition, phosphorylation of PLC-γ1 and activation of Nuclear Factor of Activated T-cell (NFAT), a downstream pathway of VEGF, are inhibited by EGT022 in HUVEC cells. CONCLUSION: These results clearly demonstrate the anti-angiogenic role played by EGT022 as a potent antagonist of integrin ß3 in endothelial cells.

Sex differences in seizure effects on social anxiety in persons with epilepsy.

PURPOSE: We investigated sex differences in the effect of seizures on social anxiety in persons with epilepsy. METHOD: In this cross-sectional multicenter study, social anxiety was measured using the short forms of the Social Phobia Scale (SPS-6) and Social Interaction Anxiety Scale (SIAS-6). SPS-6 scores ≥ 9 and SIAS-6 scores ≥ 12 were considered to indicate social phobia and social interaction anxiety, respectively. The Patient Health Questionnaire-9, Stigma Scale-Revised, and Family Adaptation-Partnership-Growth-Affection-Resolve scale were also completed. A logistic regression analysis with an interaction term was used to analyze the data. RESULTS: Out of 285 participants, a SPS-6 score ≥ 9 and a SIAS-6 score ≥ 12 were noted in 62 (21.8%) and 36 (12.6%) of participants, respectively. There was no difference in the prevalence of social anxiety between men and women. Intractable seizures and lack of seizure freedom were associated with a SPS-6 score ≥ 9 and a SIAS-6 score ≥ 12, but statistical significance was lost in the adjusted models. However, intractable seizures and lack of seizure freedom significantly interacted with sex for a SPS-6 score ≥ 9 (p = 0.018) and a SIAS-6 score ≥ 12 (p = 0.048) in both the separate and adjusted models. Specifically, intractable seizures tended to be positively associated with SPS-6 scores ≥ 9 than non-intractable seizures in men only (odds ratio = 2.602, p = 0.068), whereas lack of seizure freedom tended to be negatively associated with SIAS-6 scores ≥ 12 than seizure freedom in women only (odds ratio = 4.804, p = 0.053). CONCLUSION: We found significant sex differences in seizure effects on social anxiety. Intractable seizures were associated with social phobia in men, whereas lack of seizure freedom in the last year was associated with social interaction anxiety in women.

Nasopharyngeal electrodes in temporal lobe epilepsy: A reappraisal of their diagnostic utility.

OBJECTIVE: To compare electroencephalography (EEG) recordings with nasopharyngeal electrodes (NPEs) plus anterior temporal electrodes (ATEs) (NPE recordings) and those with only ATEs (non-NPE recordings) for the detection of interictal epileptiform discharges (IEDs) in patients with suspected temporal lobe epilepsy (TLE). METHODS: We retrospectively analyzed the initial EEGs of 229 patients that were recorded simultaneously with ATEs and NPEs in addition to the electrodes of the 10-20 system of electrode placement. Two data sets of NPE and non-NPE recordings were reviewed independently by three interpreters with differing degrees of experience. Discordant findings in the interpretation among the three interpreters were resolved by a consensus to yield final results. RESULTS: IEDs were detected in 76.4% of patients with NPE recordings compared to 55.5% with non-NPE recordings (p < 0.01). Bilateral independent IEDs were found in 26.2% and 11.4% of EEGs with NPE and non-NPE recordings (p < 0.01). The degree of agreement for the detection of IEDs among the three interpreters was higher with the NPE than with non-NPE recordings (κappa score, 0.70 vs. 0.54). The increased diagnostic yield of NPE recordings for the detection of IEDs was particularly prominent in patients with mesial and non-lesional TLEs. CONCLUSIONS: EEG recordings using NPEs are useful to improve the sensitivity and level of agreement among interpreters for the detection of IEDs in patients with TLE. SIGNIFICANCE: NPE recordings may be recommended in routine EEGs for the evaluation of patients with suspected TLE.

Cerebral Infarction Observed on Brain MRI in Unconscious Out-of-Hospital Cardiac Arrest Survivors: A Pilot Study.

BACKGROUND: Cumulative evidence regarding the use of brain magnetic resonance imaging (MRI) for predicting prognosis of unconscious out-of-hospital cardiac arrest (OHCA) survivors treated with targeted temperature management (TTM) is available. Theoretically, these patients are at a high risk of developing cerebral infarction. However, there is a paucity of reports regarding the characteristics of cerebral infarction in this population. Thus, we performed a pilot study to identify the characteristics and risk factors of cerebral infarction and to evaluate whether this infarction is associated with clinical outcomes. METHODS: A single-center, retrospective, registry-based cohort study was conducted at Severance Hospital, a tertiary center. Unconscious OHCA survivors were registered and treated with TTM between September 2011 and December 2015. We included patients who underwent brain MRI in the first week after the return of spontaneous circulation. We excluded patients who underwent any endovascular interventions to focus on "procedure-unrelated" cerebral infarctions. We assessed hypoxic-ischemic encephalopathy (HIE) and procedure-unrelated cerebral infarction separately on MRI. Patients were categorized into the following groups based on MRI findings: HIE (-)/infarction (-), infarction-only, and HIE (+) groups. Conventional vascular risk factors showing p < 0.05 in univariate analyses were entered into multivariate logistic regression. We also evaluated if the presence of this procedure-unrelated cerebral infarction lesion or HIE was associated with a poor clinical outcome at discharge, defined as a cerebral performance category of 3-5. RESULTS: Among 71 unconscious OHCA survivors who completed TTM, underwent MRI, and who did not undergo endovascular interventions, 14 (19.7%) patients had procedure-unrelated cerebral infarction based on MRI. Advancing age [odds ratio (OR) 1.11] and atrial fibrillation (OR 5.78) were independently associated with the occurrence of procedure-unrelated cerebral infarction (both p < 0.05). There were more patients with poor clinical outcomes at discharge in the HIE (+) group (88.1%) than in the infarction-only (30.0%) or HIE (-)/infarction (-) group (15.8%) (p < 0.001). HIE (+) (OR 38.69, p < 0.001) was independently associated with poor clinical outcomes at discharge, whereas infarction-only was not (p > 0.05), compared to HIE (-)/infarction (-). CONCLUSIONS: In this pilot study, procedure-unrelated cerebral infarction was noted in approximately one-fifth of unconscious OHCA survivors who were treated with TTM and underwent MRI. Older age and atrial fibrillation might be associated with the occurrence of procedure-unrelated cerebral infarction, and cerebral infarction was not considered to be associated with clinical outcomes at discharge. Considering that the strict exclusion criteria in this pilot study resulted in a highly selected sample with a relatively small size, further work is needed to verify our findings.

Family cohesion is differently associated with felt stigma depending on enacted stigma in adults with epilepsy.

PURPOSE: There have been little researches examining the role of family functioning on psychological outcomes in the field of adult epilepsy. We determined whether family functioning is correlated with felt stigma in adults with epilepsy. METHODS: In this cross-sectional study, adults with epilepsy and their caregivers were recruited. Data were collected using the Family Adaptability and Cohesion Evaluation Scale (FACES) III, the Family adaptation, partnership, growth, affection, and resolve (APGAR) questionnaire, the Stigma Scale for Epilepsy (SS-E), the modified questionnaire for episodes of discrimination, and the Beck Depression Inventory. Family functioning was measured by the caregivers. RESULTS: A total of 273 adult patients and their primary caregivers were included. Multivariate logistic analyses showed that family cohesion and excellent family functioning were negatively correlated with felt stigma after controlling for confounding variables. Enacted stigma, depressive symptoms, and university education were also significant. Interaction between enacted stigma and family cohesion on felt stigma was significant (p = 0.049). Family cohesion was negatively correlated with felt stigma only in the patients with enacted stigma (p = 0.011). CONCLUSIONS: Family functioning especially family cohesion may have protective effects against development of felt stigma in adults with epilepsy. Such protecting effects against felt stigma may be different according to enacted stigma. This understanding is helpful for developing effective psychosocial interventions to reduce felt stigma in patients with epilepsy.

Factors associated with stigma and depressive symptoms in family members of patients with epilepsy.

PURPOSE: Literature regarding family stigma related to epilepsy is scarce. This study investigated the prevalence of family stigma and depressive symptoms and the associated factors among the family members of patients with epilepsy. METHODS: In a cross-sectional study, Stigma Scale-Revised score ≥ 4 and Patient Health Questionnaire-9 score ≥ 10 were considered indicative of moderate-to-severe stigma and depressive symptoms, respectively. Stepwise logistic regression analyses were performed. RESULTS: Of the 482 family members, a mean age was 47.1 ±â€¯9.4 years, and 73.4% were female. Of the patients, a mean age was 25.5 ±â€¯16.7 years, and 45.0% were female. Idiopathic generalized epilepsy and focal epilepsy were noted in 22.4% and 65.6% of patients, respectively. Family stigma and depressive symptoms were noted in 10.0% and 11.2% of family members, respectively. Family stigma was significantly associated with high seizure frequency and being a sibling or offspring of a patient independent of their depressive symptoms. By contrast, depressive symptoms in family members were significantly associated with polytherapy, being parents of a patient, and neurological comorbidities independent of family stigma. In a subset of patients and their family, patients had higher proportion of stigma and depressive symptoms than their family. Depressive symptoms and stigma among patients were significantly correlated with those among parents, but not spouse. CONCLUSION: Family stigma is common in families with epilepsy and is closely related to depressive symptoms. Frequent seizures, polytherapy, neurological comorbidities, and the relationship to a patient may be factors that are independently associated with family stigma and depressive symptoms in family members.

Does the new Korean term for epilepsy reduce the stigma for Korean adults with epilepsy?

PURPOSE: The purpose of this study was to evaluate differences in stigma, disclosure management of epilepsy, and knowledge about epilepsy between patients with epilepsy who recognized and did not recognize the new Korean term for epilepsy. METHODS: This was a cross-sectional, multicenter study. The Stigma Scale-Revised, the Disclosure Management Scale, the Patient Health Questionnaire-9, and a questionnaire assessing knowledge about epilepsy were used. The set of questionnaires had two versions, using either the old or new name for epilepsy. Multivariate logistic regression analyses were used. RESULTS: A total of 341 patients with epilepsy and 509 family members were recruited. Approximately 62% of patients felt some degree of epilepsy-related stigma. Mild stigma, severe concealment of epilepsy diagnosis, and increased knowledge about epilepsy were independently identified as factors associated with recognition of the new term in patients. Recognition of the new term was more prevalent in patients and family members with higher education, female family members, and family members having patients with younger age at seizure onset and shorter duration of epilepsy. There were no significant differences between the two types of questionnaires. About 81% of patients and 93% of family members had a positive attitude about renaming epilepsy. CONCLUSION: The use of the new Korean term for epilepsy (cerebroelectric disorder) increased knowledge about epilepsy but did not reduce stigma and concealment of epilepsy diagnosis in Korean adults with epilepsy. Higher education may be an important factor for knowing the new term in patients and family members.

Factors associated with social anxiety in South Korean adults with epilepsy.

PURPOSE: The aim of this study was to examine social anxiety in South Korean adults with epilepsy and to identify associated factors. METHOD: This was a cross-sectional, multicenter study in South Korea. Social anxiety was assessed using short forms of the Social Phobia Scale (SPS-6) and Social Interaction Anxiety Scale (SIAS-6). The SPS-6 scores ≥9 and SIAS-6 scores ≥12 were considered indicative of social phobia and social interaction anxiety, respectively. The Patient Health Questionnaire-9 (PHQ-9); Stigma Scale-Revised (SS-R); Disclosure Management Scale; Family Adaptation, Partnership, Growth, Affection, Resolve (F-APGAR) scale; and a questionnaire assessing knowledge about epilepsy were also used. RESULTS: Of a total of 219 patients with epilepsy, 21% and 11% had SPS-6 scores ≥9 and SIAS-6 scores ≥12, respectively. In logistic regression analysis, SPS-6 scores ≥9 were independently associated with SS-R scores of 4-9 (odds ratio [OR]: 8.626, 95% confidence interval [CI]: 2.515-29.587, p = .001), SS-R scores 1-3 (OR: 5.496, 95% CI: 1.757-17.197, p = .003), and PHQ-9 scores ≥10 (OR: 4.092, 95% CI: 1.823-9.185, p = .001). In contrast, SIAS-6 scores ≥12 were related only to PHQ-9 scores ≥10 (OR: 8.740, 95% CI: 3.237-23.599, p < .001). Belonging to a dysfunctional family and lack of knowledge about epilepsy tended to be associated with social phobia (p = .071) and social interaction anxiety (p = .090), respectively. Epilepsy-related variables were not related to social anxiety. CONCLUSION: Social anxiety is not rare in patients with epilepsy. In this study, social phobia was associated with perceived stigma and depressive symptoms, whereas social interaction anxiety was related only to depressive symptoms in patients with epilepsy.

Continuous Positive Airway Pressure Therapy Restores Cardiac Mechanical Function in Patients With Severe Obstructive Sleep Apnea: A Randomized, Sham-Controlled Study.

BACKGROUND: Continuous positive airway pressure (CPAP) therapy might decrease left ventricular (LV) and right ventricular (RV) loads and improve cardiac mechanical function in patients with obstructive sleep apnea (OSA). However, the benefits of CPAP therapy for cardiac mechanical function in patients with OSA have not previously been proved in a randomized, sham-controlled clinical trial. This study therefore investigated the effects of CPAP therapy on LV and RV mechanical function in patients with severe OSA and compared them with the effects of a sham intervention. METHODS: In this randomized sham-controlled trial, we analyzed LV and RV function by conventional and speckle-tracking echocardiography before and after 3 months of treatment in 52 patients with severe OSA. Patients were randomly assigned (1:1) to receive either CPAP or sham treatment for 3 months. The main investigator and patients were masked to the trial randomization. RESULTS: After 3 months, CPAP treatment significantly improved LV global longitudinal strain (GLS) compared with the sham treatment (-20.0% ± 2.1% vs -18.0% ± 2.5%; P = .004), although there were no differences in LV dimension or ejection fraction. CPAP treatment reduced RV size and improved the fractional area change (51.3% ± 7.9% vs 46.9% ± 6.7%; P = .038) compared with the sham treatment. CPAP treatment did not ameliorate the RV GLS compared with the sham treatment. CONCLUSIONS: In patients with severe OSA, CPAP treatment for 3 months improved LV and RV function compared with sham treatment. LV mechanical function assessed by speckle-tracking echocardiography and RV fractional area change assessed by two-dimensional echocardiography were significantly improved by CPAP treatment.

Efficient induction of cell-mediated immunity to varicella-zoster virus glycoprotein E co-lyophilized with a cationic liposome-based adjuvant in mice.

Varicella zoster virus (VZV) is a neurotropic and lymphotropic alpha herpesvirus that causes varicella and herpes zoster (HZ). At a primary infection, VZV causes varicella in young children. Reactivation of latent VZV in sensory ganglia causes painful HZ in elderly people, occasionally leading to a serious complication, postherpetic neuralgia (PHN). A live attenuated VZV vaccine, the first vaccine licensed for the prevention of HZ and PHN is not very effective, while a recombinant subunit vaccine provides higher and longer protection against HZ. In the present study, we developed a new adjuvant system CIA09A, which is composed of cationic liposomes, the Toll-like receptor 4 (TLR4) agonist de-O-acylated lipooligosaccharide, and Quillaja saponin fraction QS-21. We then determined its adjuvant activity for recombinant VZV glycoprotein E (gE) in mice. Co-lyophilization of the liposomal adjuvant formulation with gE did not abolish the immune-stimulating activity. In fact, the CIA09A-adjuvanted gE vaccine was highly effective in eliciting both humoral and cellular immune responses to the recombinant gE protein and VZV in a VZV-primed mouse model. Furthermore, the frequency of gE-specific polyfunctional CD4+ T cells expressing interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and interleukin (IL)-2 was significantly increased in mice immunized with the adjuvanted vaccine. These data indicate that co-lyophilization of protein antigens with CIA09A enables development of a liposome-adjuvanted vaccine in a single vial to induce strong cell-mediated immunity required for vaccine efficacy. Thus, the CIA09A-adjuvanted gE vaccine warrants further development as a new prophylactic vaccine against HZ.

The effect of presymptomatic hypertension in posterior reversible encephalopathy syndrome.

OBJECTIVE: The effect of blood pressure (BP) on the lesion distribution of posterior reversible encephalopathy syndrome (PRES) is controversial. The aim of this study was to identify the relationship between brain lesion distribution patterns and BP. METHODS: Sixty-five patients with PRES were selected from the database. Data regarding brain MRI findings, clinical symptoms, medical conditions, and BP at the presymptomatic period (24 hr before the symptom onset) and at the symptom onset were collected. The brain lesion distribution degree was numerically calculated (lesion scoring point [LSP]) and compared with BP and medical conditions. RESULTS: Mean onset-MAP was higher than mean pre-MAP. Pre-MAP correlated with onset-MAP. The LSP was significantly correlated with pre-MAP (p = 0.009, correlation coefficient [cc] = 0.323), whereas no significant correlation was found between LSP and onset-MAP (p = 0.159, cc = 0.177). Similarly, when patients were grouped by mean MAP values, LSP was significantly higher in the patients with high MAP at the presymptomatic period (p = 0.004), whereas no difference was found in the LSP value between patients with low MAP and high MAP at the symptom onset (p = 0.272). CONCLUSION: The patient with PRES who has relatively higher BP in the presymptomatic period would be more likely to have wider lesion distribution than the patient with lower BP. BP elevation during presymptomatic period may be a heralding sign of impending PRES and a factor affecting the severity of PRES although BP was not investigated at earlier time points.

Increased Superoxide Dismutase 2 by Allopregnanolone Ameliorates ROS-Mediated Neuronal Death in Mice with Pilocarpine-Induced Status Epilepticus.

Excessive production of reactive oxygen species (ROS), along with dysfunction of the antioxidant defense system, such as that involving superoxide dismutase (SOD), may play a major role in neuronal death following status epilepticus (SE). Neurosteroids, which are allosteric modulators of the GABAA receptor in cerebral metabolism, have been suggested as being neuroprotective in various animal models; however, their effect to preventing ROS has not been examined. Herein, we investigate the neuroprotective role of allopregnanolone, the prototypical neurosteroid in the brain, in relation to the ROS-mediated neuronal injury. Adult male C57BL/6 mice were subjected to SE and treated with allopregnanolone. Hippocampal cell death was assessed by the terminal deoxynucleotidyl transferase dUTP nick end labeling assay, and ROS production was investigated by in situ detection of oxidized hydroethidine. SOD2 expression was analyzed by both western blot and immunofluorescent staining in the hippocampal subfields. In mice treated with allopregnanolone after SE, hippocampal cell death, DNA fragmentation, oxidative DNA damage, and ROS production were reduced significantly compared to mice subjected to vehicle treatment after SE. Hippocampal SOD2 expression was significantly increased by allopregnanolone. These finding suggest that allopregnanolone plays a neuroprotective role, with not only anticonvulsant but also antioxidant effects, by increasing SOD2 in pilocarpine-induced SE model.

Effects of continuous positive airway pressure therapy on left ventricular diastolic function: a randomised, sham-controlled clinical trial.

Continuous positive airway pressure (CPAP) therapy may decrease left ventricular (LV) loads and improve myocardial oxygenation. In this study, we investigated the effect of CPAP on LV diastolic function compared with sham treatment in patients with severe obstructive sleep apnoea (OSA).This 3-month prospective single-centre randomised sham-controlled trial analysed 52 patients with severe OSA. Patients were randomly assigned (1:1) to receive either CPAP or sham treatment for 3 months. The main investigator and patients were masked to the trial randomisation. The primary end-point was change of early diastolic mitral annular (e') velocity over the 3-month period. Secondary end-points were pulse wave velocity (PWV), 24-h ambulatory blood pressure (BP) and variables of ventricular-vascular coupling at 3 months.After 3 months of follow-up, CPAP treatment significantly increased the e' velocity, and was greater than the sham treatment (0.65±1.70 versus -0.61±1.85 cm·s-1, p=0.014). The PWV, 24-h mean diastolic BP, night-time diastolic BP, arterial elastance index and ventricular-vascular coupling index after 3 months of follow-up decreased significantly in the CPAP group.In patients with severe OSA, CPAP treatment for 3 months improved LV diastolic function more than sham treatment, and was accompanied by improvements in arterial stiffness and ventricular-vascular coupling.

Unblinded, randomized multicenter trial comparing lamotrigine and valproate combination with controlled-release carbamazepine monotherapy as initial drug regimen in untreated epilepsy.

PURPOSE: To compare controlled-release carbamazepine monotherapy (CBZ-CR) with lamotrigine and valproate combination therapy (LTG + VPA) in equivalent total drug load, as initial drug regimen in untreated patients with partial and/or generalized tonic-clonic seizures (GTCS). METHODS: This unblinded, randomized, 60-week superiority trial recruited patients having two or more unprovoked seizures with at least one seizure during previous three months. After randomization into CBZ-CR or LTG + VPA, patients entered into eight-week titration phase (TP), followed by 52-week maintenance phase (MP). Median doses of CBZ-CR and LTG + VPA were 600 mg/day and 75 mg/day + 500 mg/day, respectively. Primary outcome measure was completion rate (CR), a proportion of patients who have completed the 60-week study as planned. Secondary efficacy measures included seizure-free rate (SFR) for 52-week of MP and time to first seizure (TTFS) during MP. RESULTS: Among 207 randomized patients, 202 underwent outcome analysis (104 in CBZ-CR, 98 in LTG + VPA). CR was 62.5% in CBZ-CR and 65.3% in LTG + VPA (p = 0.678). SFR during MP was higher in LTG + VPA (64.1%) than CBZ-CR (47.8%) (P = 0.034). TTFS was shorter with CBZ-CR (p = 0.041). Incidence of adverse effects (AEs) were 57.7% in CBZ-CR and 60.2% in LTG + VPA and premature drug withdrawal rates due to AEs were 12.5% and 7.1%, respectively, which were not significantly different. CONCLUSION: CR was comparable between LTG + VPA and CBZ-CR, however, both SFR for 52-week MP and TTFS during MP were in favor of LTG + VPA than CBZ-CR. The study suggested that LTG + VPA can be an option as initial drug regimen for untreated patients with partial seizures and/or GTCS except for women of reproductive age.

Potentiation of Th1-Type Immune Responses to Mycobacterium tuberculosis Antigens in Mice by Cationic Liposomes Combined with De-O-Acylated Lipooligosaccharide.

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis. Bacillus Calmette-Guérin (BCG) vaccine is the only TB vaccine currently available, but it is not sufficiently effective in preventing active pulmonary TB or adult infection. With the purpose of developing an improved vaccine against TB that can overcome the limitations of the current BCG vaccine, we investigated whether adjuvant formulations containing de-O-acylated lipooligosaccharide (dLOS) are capable of enhancing the immunogenicity and protective efficacy of TB subunit vaccine. The results revealed that dLOS/dimethyl dioctadecyl ammonium bromide (DDA) adjuvant formulation significantly increased both humoral and Th1-type cellular responses to TB subunit vaccine that are composed of three antigens, Ag85A, ESAT-6, and HspX. The adjuvanted TB vaccine also effectively induced Th1-type response in a BCG-primed mouse model, suggesting a potential as a booster vaccine. Finally, dLOS/DDA-adjuvanted TB vaccine showed protective efficacy against M. tuberculosis infection in vitro and in vivo. These data indicate that dLOS/DDA adjuvant enhances the Th1-type immunity and protective efficacy of TB subunit vaccine suggesting that it would be a promising adjuvant candidate for development of a booster vaccine.

Trafficking patterns of NMDA and GABAA receptors in a Mg2+-free cultured hippocampal neuron model of status epilepticus.

An altered pattern of receptor trafficking is one of the pathophysiologic mechanisms of status epilepticus (SE). The gradual internalization of GABAA receptors (GABARs) occurs in both in vitro and in vivo models of SE and is thought to be a cause of decreased GABAergic inhibition. Unlike GABARs, little is known about alterations in NMDA receptor (NMDAR) trafficking during SE, even though increased activity of NMDARs is indispensable for the induction and maintenance of SE. Therefore, we aimed to simultaneously investigate the changes in the trafficking patterns of GABARs and NMDARs in an in vitro cultured hippocampal neuron model of SE. For induction of epileptiform discharges, hippocampal neurons were exposed to external medium without Mg2+. Biotinylation assay and immunofluorescence staining for GABAR ß2,3 and NMDAR NR1 subunits were performed to quantify and visualize surface GABARs and NMDARs, respectively. The frequency of spontaneous action potentials increased more than 4-fold after Mg2+-free induction. The level of surface GABARs decreased over time after Mg2+-free induction, dropping to approximately 50% of control levels an hour after Mg2+-free induction. By contrast, the trafficking of NMDARs to the surface was enhanced after a slight time lag, increasing by 30% of control levels an hour after Mg2+-free induction. Our data showed the changes of both NMDAR and GABAR trafficking during prolonged SE induced by a Mg2+-free extracellular environment and confirmed that this in vitro SE model is suitable for examining alterations in the receptor trafficking pattern by prolonged seizure activity. These results suggest that targeting of surface NMDAR could be a promising method in controlling benzodiazepine-resistant SE.

Increased Immunogenicity and Protective Efficacy of a P. aeruginosa Vaccine in Mice Using an Alum and De-O-Acylated Lipooligosaccharide Adjuvant System.

Pseudomonas aeruginosa (P. aeruginosa) is an opportunistic pathogen that commonly causes fatal infections in cystic fibrosis and burn patients as well as in patients who are hospitalized or have impaired immune systems. P. aeruginosa infections are difficult to treat owing to the high resistance of the pathogen to conventional antibiotics. Despite several efforts, no effective prophylactic vaccines against P. aeruginosa are currently available. In this study, we investigated the activity of the CIA06 adjuvant system, which is composed of alum and de-O-acylated lipooligosaccharide, on a P. aeruginosa outer membrane protein (OMP) antigen vaccine in mice. The results indicated that CIA06 significantly increased the antigen-specific IgG titers and opsonophagocytic activity of immune sera against P. aeruginosa. In addition, the antibodies induced by the CIA06-adjuvanted vaccine exhibited higher cross-reactivity with heterologous P. aeruginosa strains. Finally, mice immunized with the CIA06-adjuvanted vaccine were effectively protected from lethal P. aeruginosa challenge. Based on these data, we suggest that the CIA06 adjuvant system might be used to promote the immunogenicity and protective efficacy of the P. aeruginosa OMP vaccine.

Effects of exogenous recombinant human bone morphogenic protein-7 on the corneal epithelial mesenchymal transition and fibrosis.

AIM: To evaluate the effect of exogenous recombinant human bone morphogenic protein-7 (rhBMP-7) on transforming growth factor-ß (TGF-ß)-induced epithelial mesenchymal cell transition (EMT) and assessed its antifibrotic effect via topical application. METHODS: The cytotoxic effect of rhBMP-7 was evaluated and the EMT of human corneal epithelial cells (HECEs) was induced by TGF-ß. HECEs were then cultured in the presence of rhBMP-7 and/or hyaluronic acid (HA). EMT markers, fibronectin, E-cadherin, α-smooth muscle actin (α-SMA), and matrix metaloproteinase-9 (MMP-9), were evaluated. The level of corneal fibrosis and the reepithelization rate were evaluated using a rabbit keratectomy model. Expression of α-SMA in keratocytes were quantified following treatment with different concentrations of rhBMP-7. RESULTS: Treatment with rhBMP-7 attenuated TGF-ß-induced EMT in HECEs. It significantly attenuated fibronectin secretion (31.6%; P<0.05), the α-SMA protein level (72.2%; P<0.01), and MMP-9 expression (23.6%, P<0.05) in HECEs compared with cells grown in the presence of TGF-ß alone. E-cadherin expression was significantly enhanced (289.7%; P<0.01) in the presence of rhBMP-7. Topical application of rhBMP-7 combined with 0.1% HA significantly reduced the amount of α-SMA+ cells by 43.18% (P<0.05) at a concentration of 2.5 µg/mL and by 47.73% (P<0.05) at 25 µg/mL, compared with the control group, without disturbing corneal reepithelization. CONCLUSION: rhBMP-7 attenuates TGF-ß-induced EMT in vitro, and topical application of rhBMP-7 reduces keratocyte myodifferentiation during the early wound healing stages in vivo without hindering reepithelization. Topical rhBMP-7 application as biological eye drops seems to be feasible in diseases involving TGF-ß-related corneal fibrosis with corneal reepithelization disorders.