Transient early neurotrophin release and delayed inflammatory cytokine release by microglia in response to PAR-2 stimulation.

Activated microglia exerts both beneficial and deleterious effects on neurons, but the signaling mechanism controlling these distinct responses remain unclear. We demonstrated that treatment of microglial cultures with the PAR-2 agonist, 2-Furoyl-LIGRLO-NH2, evoked early transient release of BDNF, while sustained PAR-2 stimulation evoked the delayed release of inflammatory cytokines (IL-1 ß and TNF-α) and nitric oxide. Culture medium harvested during the early phase (at 1 h) of microglial activation induced by 2-Furoyl-LIGRLO-NH2 (microglial conditioned medium, MCM) had no deleterious effects on cultured neurons, while MCM harvested during the late phase (at 72 h) promoted DNA fragmentation and apoptosis as indicated by TUNEL and annexin/PI staining. Blockade of PAR-1 during the early phase of PAR-2 stimulation enhanced BDNF release (by 11%, small but significant) while a PAR-1 agonist added during the late phase (24 h after 2-Furoyl-LIGRLO-NH2 addition) suppressed the release of cytokines and NO. The neuroprotective and neurotoxic effects of activated microglial exhibit distinct temporal profiles that are regulated by PAR-1 and PAR-2 stimulation. It may be possible to facilitate neuronal recovery and repair by appropriately timed stimulation and inhibition of microglial PAR-1 and PAR-2 receptors.

Microwave-assisted derivatization combined with coacervative extraction for determining glutathione in biomatrix samples, followed by capillary liquid chromatography.

In this study, an ecofriendly analytical method was developed for determining glutathione (GSH) levels in biomatrix samples. 9-(bromomethyl)acridine was used for the first time as a derivatization reagent in GSH analysis. Microwave-assisted derivatization reduced the reaction time to 1 min. After derivatization, coacervative extraction was employed to extract GSH derivative from the complex biomatrix and to increase sensitivity. Because the negatively charged group of the GSH derivative was neutralized by the extracting agent Aliquat 336, aggregates formed without any coacervating agents. Furthermore, capillary liquid chromatography coupled with ultraviolet detection was applied to decrease waste generation and increase selectivity. This method successfully quantified GSH levels in various biomatrices, including erythrocytes, HaCaT cells, BALB/3T3 cells, and 3T3-L1 fibroblasts. This method only required a low sample volume (≤10 µL). A standard addition method was utilized to spike the biomatrix samples with 0-4.8 nmol GSH to construct calibration curves. The proposed method performed well, with a determination coefficient of 0.999 and relative standard deviations of less than 6.59% for the slope and the intercept, as determined by linear regression analysis. The limit of detection of GSH in the standard solution was 800 nM or 0.4 pmol. Compared to non-derivatized GSH, the proposed method for detecting derivatized GSH provides 750-fold greater sensitivity.

Determination of parabens using two microextraction methods coupled with capillary liquid chromatography-UV detection.

Parabens are common preservatives and environmental hormones. As such, possible detrimental health effects could be amplified through their widespread use in foods, cosmetics, and pharmaceutical products. Thus, the determination of parabens in such products is of particular importance. This study explored vortex-assisted dispersive liquid-liquid microextraction techniques based on the solidification of a floating organic drop (VA-DLLME-SFO) and salt-assisted cloud point extraction (SA-CPE) for paraben extraction. Microanalysis was performed using a capillary liquid chromatography-ultraviolet detection system. These techniques were modified successfully to determine four parabens in 19 commercial products. The regression equations of these parabens exhibited good linearity (r2=0.998, 0.1-10µg/mL), good precision (RSD<5%) and accuracy (RE<5%), reduced reagent consumption and reaction times (<6min), and excellent sample versatility. VA-DLLME-SFO was also particularly convenient due to the use of a solidified extract. Thus, the VA-DLLME-SFO technique was better suited to the extraction of parabens from complex matrices.

Clinical features of fatal asthma.

To characterize the clinical features of fatal asthma, we retrospectively analyzed the clinical characteristics of patients who died of an acute asthma attack in our hospital during a 15-year period from 1989 to 2003. Twelve patients had fatal asthma during this period, including eight who were dead on arrival in the emergency room (ER) and three who died within 1 hour of admission to the ER. Patients were categorized into three groups according to the clinical presentations during the fatal attack: (1) rapid (< 3 hours) decompensation in four patients; (2) gradual development of respiratory failure over several days in two patients; and (3) acute deterioration after unstable asthma lasting several days in six patients. All patients in groups 1 and 2 had reported previous near-fatal attacks. The proportion of young patients was highest in group 3, with half of them (3/6) younger than 35 years of age. Only one patient in group 3 had had a previous near-fatal attack. Five of the seven patients, with previous near-fatal attacks, had a pattern of decompensation during their fatal attack that was similar to their previous attacks. In conclusion, nearly all patients with fatal asthma in this study died outside of the hospital or within 1 hour after admission to the ER. Patients had patterns of decompensation during the fatal attack that were similar to those of their previous attacks. Early detection of warning signs, early admission to the ER, adequate treatment, and extremely close observation of patients, especially within 1 hour after ER arrival, may prevent or decrease the incidence of fatal asthmatic attack.

Protective Effect of Vanillic Acid against Hyperinsulinemia, Hyperglycemia and Hyperlipidemia via Alleviating Hepatic Insulin Resistance and Inflammation in High-Fat Diet (HFD)-Fed Rats.

Excess free fatty acid accumulation from abnormal lipid metabolism results in the insulin resistance in peripheral cells, subsequently causing hyperinsulinemia, hyperglycemia and/or hyperlipidemia in diabetes mellitus (DM) patients. Herein, we investigated the effect of phenolic acids on glucose uptake in an insulin-resistant cell-culture model and on hepatic insulin resistance and inflammation in rats fed a high-fat diet (HFD). The results show that vanillic acid (VA) demonstrated the highest glucose uptake ability among all tested phenolic acids in insulin-resistant FL83B mouse hepatocytes. Furthermore, rats fed HFD for 16 weeks were orally administered with VA daily (30 mg/kg body weight) at weeks 13-16. The results show that levels of serum insulin, glucose, triglyceride, and free fatty acid were significantly decreased in VA-treated HFD rats (p < 0.05), indicating the protective effects of VA against hyperinsulinemia, hyperglycemia and hyperlipidemia in HFD rats. Moreover, VA significantly reduced values of area under the curve for glucose (AUCglucose) in oral glucose tolerance test and homeostasis model assessment-insulin resistance (HOMA-IR) index, suggesting the improving effect on glucose tolerance and insulin resistance in HFD rats. The Western blot analysis revealed that VA significantly up-regulated expression of hepatic insulin-signaling and lipid metabolism-related protein, including insulin receptor, phosphatidylinositol-3 kinase, glucose transporter 2, and phosphorylated acetyl CoA carboxylase in HFD rats. VA also significantly down-regulated hepatic inflammation-related proteins, including cyclooxygenase-2 and monocyte chemoattractant protein-1 expressions in HFD rats. These results indicate that VA might ameliorate insulin resistance via improving hepatic insulin signaling and alleviating inflammation pathways in HFD rats. These findings also suggest the potential of VA in preventing the progression of DM.