Genetic analysis and functional study of novel CFTR variants in Chinese children with cystic fibrosis.

OBJECTIVES: To describe the clinical characteristics of Chinese cystic fibrosis (CF) patients and to investigate the variants of CFTR and their potential pathogenicity. STUDY DESIGN: Chinese patients with potential CF diagnosis were studied. Clinical data were reviewed retrospectively from medical records. Whole exome sequencing and genetic evaluation were conducted to explore potential gene variants. The disruption of the variants to protein structure and function was explored and validated using in vitro experiments and in silico analysis. RESULTS: Four patients were recruited to the study, three of them were diagnosed as CF, and one was diagnosed as CFTR-related disorder. The age at symptom onset for the patients in this study ranged from newborn to 6 years, while the age at diagnosis varied from 3 to 11 years. All four patients exhibited bilateral diffuse bronchiectasis with Pseudomonas aeruginosa infections, and three of them had malnutrition. Finger clubbing was observed in three patients, two of whom displayed mixed ventilatory dysfunction. The CFTR variants spectrum of Chinese children with CF differs from that of Caucasian. A total of six variants were identified, two of which were first reported (c.1219G > T [p.Glu407*] and c.1367delT [p.Ala457Leufs*12]). The nonsense variants c.1219G > T, c.1657C > T and c.2551C > T and the frameshift variant c.1367delT were predicted to introduce premature stop codon and produce shorten CFTR protein, which was also first validated by in vitro truncation assay in this study. The missense variant c.1810A > C was predicted to disrupt the function of the nucleotide-binding domain 1 (NBD1) in the CFTR protein. The splicing variant c.1766 + 5G > T caused skipping of exon 13 and damaged the integrity of CFTR protein. CONCLUSIONS: Our study expands the spectrum of phenotypes and genotypes for CF of Chinese origin, which differs significantly from that of Caucasian. Genetic analysis and counseling are crucial and deserve extensive popularization for the diagnosis ofCF in patients of Chinese origin.

Facile Preparation of Polyacrylonitrile-Based Activated Carbon Fiber Felts for Effective Adsorption of Dipropyl Sulfide.

Activated carbon fibers (ACFs) derived from various polymeric fibers with the characteristics of a high specific surface area, developed pore structure, and good flexibility are promising for the new generation of chemical protection clothing. In this paper, a polyacrylonitrile-based ACF felt was prepared via the process of liquid phase pre-oxidation, along with a one-step carbonization and chemical activation method. The obtained ACF felt exhibited a large specific surface area of 2219.48 m2/g and pore volume of 1.168 cm3/g, as well as abundant polar groups on the surface. Owing to the developed pore structure and elaborated surface chemical property, the ACF felt possessed an intriguing adsorption performance for a chemical warfare agent simulant dipropyl sulfide (DPS), with the highest adsorption capacity being 202.38 mg/g. The effects of the initial concentration of DPS and temperature on the adsorption performance of ACF felt were investigated. Meanwhile, a plausible adsorption mechanism was proposed based on the kinetic analysis and fitting of different adsorption isotherm models. The results demonstrated that the adsorption process of DPS onto ACF felt could be well fitted with a pseudo-second-order equation, indicating a synergistic effect of chemical adsorption and physical adsorption. We anticipate that this work could be helpful to the design and development of advanced ACF felts for the application of breathable chemical protection clothing.

Pulse pressure and its association with body composition among Chinese men and women without diagnosed hypertension: the China Kadoorie Biobank.

BACKGROUND: Wide brachial pulse pressure (PP) has been associated with cardiovascular events, while its population distribution and association with body composition were poorly characterized in large populations. METHODS: We evaluated the age and sex distributions of PP and its associations with body composition using baseline data from the China Kadoorie Biobank. A total of 434 200 participants without diagnosed hypertension were included in the analysis. Wide PP was defined as PP above 65 mmHg. Body composition variables, including BMI, waist circumference, waist-to-hip ratio (WHR), fat mass index (FMI), fat-free mass index (FFMI), and body fat percentage (BF%), were obtained from bioelectrical impedance analysis. RESULTS: Overall, 14.3% of the participants had wide PP. Older age was consistently associated with wider PP in women but only after the andropause stage in men. The independent associations of BMI with wide PP were stronger than other body composition measures. The adjusted differences (men/women, mmHg) in PP per standard deviation (SD) increase in BMI (1.55/1.47) were higher than other body composition (BF%: 0.32/0.64, waist circumference: 0.33/0.39; WHR: 0.49/0.42). In addition, sex differences were observed. In men, the per SD difference in PP was higher for FFMI than for FMI (0.91 vs. 0.67, P  < 0.05), whereas in women, it was higher for FMI than for FFMI (1.01 vs. 0.72, P  < 0.05). CONCLUSION: Our nationwide population-based study presented the sex-specific distribution of PP over age and identified differential associations of PP with fat and fat-free mass in men and women.

Differential Gut Microbiota, Dietary Intakes in Constipation Patients with or without Hypertension.

SCOPE: Diet and gut microbiota are involved in blood pressure regulations, but few studies have focused on the constipation patients. The study seeks to identify differences in gut microbiota between hypertensive and normotensive subjects in constipation patients, analyzes the relationship between dietary patterns and blood pressure, and explores mediation effects of gut microbiota. METHODS AND RESULTS: Gut microbial genera and dietary information of 186 functional constipation participants are characterized by 16S rRNA sequencing and a food frequency questionnaire. The hypertensive subjects shows lower α-diversity and ß-diversity of gut microbiota than normotensive (p < 0.05) and 17 differential microbial genera. The dried-beans intake frequency inversely correlated with systolic and diastolic blood pressure after multivariate adjustment (r = -0.273, p-FDR < 0.01; r = -0.251, p-FDR = 0.026, respectively). Logistic regression indicates that the individuals often consumed dried-beans have a lower hypertension risk than those never consumed [OR = 0.137, 95% CI: (0.022, 0.689), p = 0.022]. A marginal mediating effect of the genus Monoglobus is observed for the association between high-fiber dietary pattern and hypertension. CONCLUSION: In patients with functional constipation, hypertension-related gut microbial differences are identified. Dried-beans intake is inversely associated with blood pressure, and a genus may potentially mediate the association between high-fiber dietary pattern and hypertension.

Effects of dietary fibers or probiotics on functional constipation symptoms and roles of gut microbiota: a double-blinded randomized placebo trial.

Dietary fibers/probiotics may relieve constipation via optimizing gut microbiome, yet with limited trial-based evidences. We aimed to evaluate the effects of formulas with dietary fibers or probiotics on functional constipation symptoms, and to identify modulations of gut microbiota of relevance. We conducted a 4-week double-blinded randomized placebo-controlled trial in 250 adults with functional constipation. Intervention: A: polydextrose; B: psyllium husk; C: wheat bran + psyllium husk; D: Bifidobacterium animalis subsp. lactis HN019 + Lacticaseibacillus rhamnosus HN001; Placebo: maltodextrin. Oligosaccharides were also included in group A to D. 16S rRNA sequencing was used to assess the gut microbiota at weeks 0, 2, and 4. A total of 242 participants completed the study. No time-by-group effect was observed for bowel movement frequency (BMF), Bristol stool scale score (BSS), and degree of defecation straining (DDS), while BSS showed mean increases of 0.95-1.05 in group A to D (all P < 0.05), but not significantly changed in placebo (P = 0.170), and 4-week change of BSS showed similarly superior effects of the interventions as compared placebo. Group D showed a marginal reduction in plasma 5-hydroxytryptamine. Group A resulted in a higher Bifidobacterium abundance than placebo at week 2 and 4. Fourteen genera showed intervention-specific increasing or decreasing trends continuously, among which Anaerostipes showed increasing trends in groups B and C, associated with BMF increase. Random forest models identified specific baseline microbial genera panels predicting intervention responders. In conclusion, we found that the dietary fibers or probiotics may relieve hard stool, with intervention-specific changes in gut microbiota relevant to constipation relief. Baseline gut microbiota may predispose the intervention responsiveness. ClincialTrials.gov number, NCT04667884.

What is the context?Supplementation of dietary fibers, such as psyllium husk or wheat bran (10 ~ 15 g/day) may relieve constipation symptoms, but bloating and flatulence are major concerns on a high fiber intake.Functional constipation patients had alternated gut microbiota profiles, while meta-analysis suggested that multispecies probiotics may increase bowel movement frequency and relieve hard stool in functional constipation.Dietary fibers or probiotics may lead to before-after changes of gut microbiota in patients with functional constipation, but time-series continued changes of gut microbiota during the intervention are unknown.Elevation of 5-hydroxytryptamine synthesis in enterochromaffin cells may affect bowel movement. And the elevated plasma 5-hydroxytryptamine was observed in functional constipation patients.What is new? Daily supplement of three prebiotic formulas with dietary fibers (polydextrose, psyllium husk, wheat bran, together with oligosaccharides), or a probiotic formula with Bifidobacterium animalis subsp. lactis HN019 + Lacticaseibacillus rhamnosus HN001 effectively relieved hard stool in functional constipation patients after 4 weeks intervention.We identified continued increasing or decreasing gut microbial genera over the intervention. Dietary fiber ­ gut microbiota (Anaerostipes)­constipation relieve (bowel movement frequency) evidence axis was identified in this human trial.Probiotic supplementation marginally reduced plasma 5-hydroxytryptamine, possibly associated with changes in BMF-related gut microbial genera.Intervention-specific baseline gut microbiota well predicted the responsiveness of constipation symptom relief.What is the impact? We provided references for the dosage and duration of dietary fiber/probiotics recommendations for adults with functional constipation, and advanced the microbial genera evidences of the fibers/probiotics-microbiota-laxation theory in humans.

Gamma-Muricholic Acid Inhibits Nonalcoholic Steatohepatitis: Abolishment of Steatosis-Dependent Peroxidative Impairment by FXR/SHP/LXRα/FASN Signaling.

Nonalcoholic steatohepatitis (NASH) reflects the outcome of steatosis-based peroxidative impairment. Here, the effect and mechanism of γ-muricholic acid (γ-MCA) on NASH were investigated on the basis of its actions in hepatic steatosis, lipid peroxidation, peroxidative injury, hepatocyte apoptosis, and its NAFLD activity score (NAS). The agonist action of γ-MCA on farnesoid X receptor (FXR) upregulated the small heterodimer partner (SHP) expression of hepatocytes. An increase in SHP attenuated the triglyceride-dominated hepatic steatosis which was induced in vivo by a high-fat high-cholesterol (HFHC) diet and in vitro by free fatty acids depending on the inhibition of liver X receptor α (LXRα) and fatty acid synthase (FASN). In contrast, FXR knockdown abrogated the γ-MCA-dependent lipogenic inactivation. When compared to their excessive production in HFHC diet-induced rodent NASH, products of lipid peroxidation (MDA and 4-HNE) exhibited significant reductions upon γ-MCA treatment. Moreover, the decreased levels of serum alanine aminotransferases and aspartate aminotransferases demonstrated an improvement in the peroxidative injury of hepatocytes. By TUNEL assay, injurious amelioration protected the γ-MCA-treated mice against hepatic apoptosis. The abolishment of apoptosis prevented lobular inflammation, which downregulated the incidence of NASH by lowering NAS. Collectively, γ-MCA inhibits steatosis-induced peroxidative injury to ameliorate NASH by targeting FXR/SHP/LXRα/FASN signaling.

Profile analysis and functional modeling identify circular RNAs in nonalcoholic fatty liver disease as regulators of hepatic lipid metabolism.

Nonalcoholic fatty liver disease (NAFLD) is the leading cause of chronic liver disease, associated with an outcome of hepatic fibrosis/cirrhosis and hepatocellular carcinoma. However, limited exploration of the underlying mechanisms hinders its prevention and treatment. To investigate the mechanisms of epigenetic regulation in NAFLD, the expression profile of circular RNA (circRNA) of rodents in which NAFLD was induced by a high-fat, high-cholesterol (HFHC) diet was studied. Modeling of the circRNA-microRNA (miRNA) -mRNA regulatory network revealed the functional characteristics of NAFLD-specific circRNAs. The targets and effects in the liver of such NAFLD-specific circRNAs were further assessed. Our results uncovered that the downregulation of 28 annotated circRNAs characterizes HFHC diet-induced NAFLD. Among the downregulated circRNAs, long intergenic non-protein coding RNA, P53 induced transcript (LNCPINT) -derived circRNAs (circ_0001452, circ_0001453, and circ_0001454) targeted both miR-466i-3p and miR-669c-3p. Their deficiency in NAFLD abrogated the circRNA-based inhibitory effect on both miRNAs, which further inactivated the AMPK signaling pathway via AMPK-α1 suppression. Inhibition of the AMPK signaling pathway promotes hepatic steatosis, depending on the transcriptional and translational upregulation of lipogenic genes, such as those encoding sterol regulatory element-binding protein 1 (SREBP1) and fatty acid synthase (FASN) in hepatocytes. The levels of LNCPINT-derived circRNAs displayed a negative association with hepatic triglyceride (TG) concentration. These findings suggest that loss of LNCPINT-derived circRNAs may underlie NAFLD via miR-466i-3p- and miR-669c-3p-dependent inactivation of the AMPK signaling pathway.

Metabolomic Changes Upon Conjugated Linoleic Acid Supplementation and Predictions of Body Composition Responsiveness.

CONTEXT: Conjugated linoleic acid (CLA) may optimize body composition, yet mechanisms underlining its benefits are not clear in humans. OBJECTIVE: We aimed to reveal the CLA-induced changes in the plasma metabolome associated with body composition improvement and the predictive performance of baseline metabolome on intervention responsiveness. METHODS: Plasma metabolome from overnight fasted samples at pre- and post-intervention of 65 participants in a 12-week randomized, placebo-controlled trial (3.2 g/day CLA vs 3.2 g/day sunflower oil) were analyzed using untargeted LC-MS metabolomics. Mixed linear model and machine learning were applied to assess differential metabolites between treatments, and to identify optimal panel (based on baseline conventional variables vs metabolites) predicting responders of CLA-derived body composition improvement (increased muscle variables or decreased adiposity variables) based on dual-energy x-ray absorptiometry. RESULTS: Compared with placebo, CLA altered 57 metabolites (P < 0.10) enriched in lipids/lipid-like molecules including glycerophospholipids (n = 7), fatty acyls (n = 6), and sphingolipids (n = 3). CLA-upregulated cholic acid (or downregulated aminopyrrolnitrin) was inversely correlated with changes in muscle and adiposity variables. Inter-individual variability in response to CLA-derived body composition change. The areas under the curves of optimal metabolite panels were higher than those of optimal conventional panels in predicting favorable response of waist circumference (0.93 [0.82-1.00] vs 0.64 [0.43-0.85]), visceral adiposity index (0.95 [0.88-1.00] vs 0.58 [0.35-0.80]), total fat mass (0.94 [0.86-1.00] vs 0.69 [0.51-0.88]) and appendicular fat mass (0.97 [0.92-1.00] vs 0.73 [0.55-0.91]) upon CLA supplementation (all FDR P < 0.05). CONCLUSION: Post-intervention metabolite alterations were identified, involving in lipid/energy metabolism, associated with body composition changes. Baseline metabolite profiling enhanced the prediction accuracy for responsiveness of CLA-induced body composition benefits.

Differential effects of formaldehyde exposure on airway inflammation and bronchial hyperresponsiveness in BALB/c and C57BL/6 mice.

Epidemiological evidence suggests that formaldehyde (FA) exposure may influence the prevalence and severity of allergic asthma. However, the role of genetic background in FA-induced asthma-like responses is poorly understood. In the present study, we investigated the nature and severity of asthma-like responses triggered by exposure to different doses of FA together with or without ovalbumin (OVA) in two genetically different mouse strains-BALB/c and C57BL/6. Both mouse strains were divided into two main groups: the non-sensitized group and the OVA-sensitized group. All the groups were exposed to 0, 0.5 or 3.0 mg/m3 FA for 6 h/day over 25 consecutive days. At 24 h after the final FA exposure, the pulmonary parameters were evaluated. We found that FA exposure induced Th2-type allergic responses in non-sensitized BALB/c and C57BL/6 mice. In addition, FA-induced allergic responses were significantly more prominent in BALB/c mice than in C57BL/6 mice. In sensitized BALB/c mice, however, FA exposure suppressed the development of OVA-induced allergic responses. Exposure to 3.0 mg/m3 FA in sensitized C57BL/6 mice also led to suppressed allergic responses, whereas exposure to 0.5 mg/m3 FA resulted in exacerbated allergic responses to OVA. Our findings suggest that FA exposure can induce differential airway inflammation and bronchial hyperresponsiveness in BALB/c and C57BL/6 mice.

IL-13 R110Q, a Naturally Occurring IL-13 Polymorphism, Confers Enhanced Functional Activity in Cultured Human Bronchial Smooth Muscle Cells.

PURPOSE: Interleukin (IL)-13, a Th2-type cytokine, plays a pivotal role in the pathogenesis of asthma through its direct effects on airway smooth muscles. A naturally occurring IL-13 polymorphism, R110Q, is strongly associated with increased total serum IgE levels and asthma. In the present study, we aimed to determine whether the IL-13 R110Q variant would display different biochemical properties or altered functions in comparison with wild-type (WT) IL-13 in cultured human bronchial smooth muscle cells (hBSMCs). METHODS: Culture supernatants and cell proteins were collected from cultured hBSMCs that were treated with 50 ng/mL IL-13 or IL-13 R110Q for 24 hours. Eotaxin released into hBSMC culture medium was determined by ELISA. The expression levels of the high-affinity IgE receptor (FcεRI) α-chain, smooth muscle-specific actin alpha chain (α-SMA), smooth muscle myosin heavy chain (SmMHC), and calreticulin in the cells were measured on Western blots. RESULTS: Compared with WT IL-13, treatment with the IL-13 R110Q variant resulted in a significant increase in eotaxin release as well as significant, although modest, increases in the expression levels of α-SMA, SmMHC, calreticulin, and FcεRI α-chain. CONCLUSIONS: The results of the present study suggenst that the IL-13 R110Q variant may enhance enhanced functional activities in hBSMCs.

Diphenyleneiodonium protects preoligodendrocytes against endotoxin-activated microglial NADPH oxidase-generated peroxynitrite in a neonatal rat model of periventricular leukomalacia.

The contribution of microglial activation to preoligodendroglial (preOL) damage in the central nervous system (CNS) is considered to be one of the principal causes of periventricular leukomalacia (PVL) pathogenesis. The present study explores the effect of diphenyleneiodonium (DPI), a NADPH oxidase (NOX) inhibitor, on protection of preOLs from bacterial lipopolysaccharide (LPS)-induced microglial toxicity in vivo and in vitro. In vitro, preOLs co-cultured with microglia exhibited increased preOL apoptosis, accompanied by overproduction of superoxide anion (O(2)(-)) and the formation of peroxynitrite (ONOO(-)) after LPS exposure. LPS also significantly up-regulated accumulation of activated microglial NOX subunits p67-phox and gp91-phox in the plasma membrane. Diphenyleneiodonium (DPI) (10µm) was found to significantly attenuate up-regulation of this NOX activity. In vivo, DPI was administered (1mg/kg/day) by subcutaneous injection for 3 days to two-day-old neonatal Sprague-Dawley rats subjected to intracerebral injection of LPS. Treatment with DPI within 24h of LPS injection significantly ameliorated white matter injury, decreasing preOL loss, O(2)(-) generation, and ONOO(-) formation, and inhibiting p67-phox, gp91-phox synthesis and p67phox membrane translocation in microglia. These results indicated that LPS-induced preOL apoptosis may have been mediated by microglia-derived ONOO(-). DPI prevented this LPS-induced brain injury, most likely by inhibiting ONOO(-) formation via NOX, thereby preventing preOL loss and immature white matter injury.

[Effects of glial cell line-derived neurotrophic factor and memantine on long-term prognosis in neonatal rats with periventricular leukomalacia].

OBJECTIVE: To evaluate the effects of glial cell line-derived neurotrophic factor (GDNF) and memantine on the long-term prognosis in neonatal rats with ischemia-induced periventricular leukomalacia (PVL). METHODS: Thirty-two 5-day-old neonatal rats were randomly divided into 4 groups: sham-operated, PVL, GDNF-treated and memantine-treated. PVL was induced by right carotid artery ligation and hypoxia in the PVL, GDNF-treated and memantine-treated groups. GDNF (100 µg/kg) or memantine (20 mg/kg) was injected in the two treatment groups immediately after PVL inducement. The weight of the rats was measured immediately before and after hypoxia ischemia (HI). Both of Morris water maze test and Rivlin inclined plane test were performed at 26 days old (21 days after HI). The values of the escape latency (EL) and swimming distance, and the maximum inclined plane degree which the rats could stand at least 5 seconds were compared among the four groups. RESULTS: The lower weight, the prolonged mean values of EL and swimming distance and the reduced maximum inclined plane degree were observed in the PVL group compared to those in the sham-operated, GDNF-treated and memantine-treated groups. There were no significant differences in the weight, the values of EI and swimming distance and the maximum inclined plane degree between the two treatment groups and the sham-operated group. CONCLUSIONS: The administration of either GDNF or memantine can markedly increase the abilities of spatial discrimination,learning and memory, and motor coordination, promote weight gain, and improve long-term prognosis in rats with PVL.

Effect of 1400W on blocking lipopolysaccharide-induced microglial toxicity to preoligodendrocytes.

BACKGROUND: The maternal-fetal infection/inflammation is believed to be the mechanism in the pathogenesis of periventricular leukomalacia (PVL). The activation of microglias (MGs) may contribute to preoligodendroglial damage. The present study was undertaken to explore the effect of N-[3-(aminomethyl) benzyl] acetamidine (1400W), a selective inhibitor of inducible nitric oxide synthase (iNOS), on the blockage of lipopolysaccharide (LPS)-induced microglial toxicity to preoligodendrocytes (preOLs). METHODS: The co-cultural MGs and preOLs obtained from two-day-old Sprague-Dawley (SD) neonatal rats were divided into three groups: co-culture control group, coculture LPS group, and co-culture LPS plus 1400W group. The concentration of nitric oxide (NO) was measured by nitric acid-deoxidize-colorimetry, the level of peroxynitrite (ONOO(-)) determined by immunocytochemistry, the synthetic level of inducible nitric oxide synthase (iNOS) detected by western blotting, and the apoptotic rate of preOLs assessed by flow cytometry after the co-cultural cells were induced by LPS (100 ng/ml) for 48 hours. RESULTS: Compared with those in the co-culture control group, the levels of NO (82.27+/-3.41 micromol/L vs. 167.86+/-9.87 micromol/L, P<0.01), ONOO(-) (6.14+/-1.27 vs. 34.38+/-7.75, P<0.01), and iNOS (0.18+/-0.027 vs. 0.79+/-0.068, P<0.01) induced by LPS increased remarkably in the co-culture LPS group, with a higher apoptotic rate of preOLs (6.73+/-1.39% vs. 24.77+/-2.05%, P<0.01). The levels of NO (69.55+/-5.07 micromol/L, P<0.01), ONOO(-) (10.33+/-3.47, P<0.01) and iNOS (0.35+/-0.042, P<0.01) were decreased significantly using 1400W at a dose of 10 micromol/L in the co-culture LPS plus 1400W group, and the apoptotic rate of preOLs (11.80+/-2.06% vs. 24.77+/-2.05%, P<0.01) also decreased compared with the co-culture LPS group. CONCLUSION: 1400W can block effectively the LPS-induced microglial toxicity to preOLs by inhibiting iNOS specifically, resulting in a significant reduction of toxicity parameters investigated and a marked increase of the survival preOLs.

[1400W blocks death pathway of LPS-induced activated-microglia to preOLs].

OBJECTIVE: To explore the efficacy of inductible nitric oxide synthase (iNOS) inhibitor 1400W in vivo in blocking the death pathway of lipopolysaccharide (LPS)-induced activated-microglia to preoligodendrocytes (preOLs) in neonatal rats with infective-type periventricular leukomalacia (PVL) induced by LPS. METHODS: Two-day-old neonatal rats were randomly divided into: a sham-operated group, an untreated PVL group, and four 1400W-treated PVL groups that were subcutaneously administrated with 20 mg/kg of 1400W at 0 h, 8 hrs, 16 hrs, and 24 hrs after LPS induction, respectively. The brain specimens were obtained 5 days after LPS induction. The pathological assessment of cerebral white matter was performed under a light microscope. Concentrations of nitric oxide (NO) were measured by nitric acid-deoxidize colorimetry. Synthesis of iNOS was determined by Western blot analysis. Peroxynitrite (ONOO(-)) level and the amount of preOLs were determined by immunocytochemistry. RETHODS: The obvious injuries of periventricular white matter, massive loss of positive O4-labelled preOLs, and increased levels of NO, ONOO(-) and iNOS were observed in neonatal rats with PVL. Compared to the untreated PVL group, the use of 1400W at 0 h, 8 hrs and 16 hrs after LPS induction significantly improved white matter injuries, reduced the levels of NO, ONOO(-) and iNOS, and increased the amount of O4-labelled preOLs. However, the use of 1400W at 24 hrs after LPS induction did not result in the improvements. CONCLUSIONS: iNOS inhibitor 1400W can effectively block the toxicity of LPS-activated microglia to preOLs and protect cerebral white matter through inhibiting iNOS and reducing the production of NO and ONOO(-). The use of 1400W within 16 hrs after LPS induction may provide cerebral protections in neonatal rats with PVL.

[Effect of 1400W, an inhibitor of inducible nitric oxide synthetase, on blocking the toxicity of lipopolysaccharide-induced activated microglia to preoligodendrocytes].

OBJECTIVE: To explore the toxicity of LPS-induced activated microglia to preoligodendrocytes (preOLs) and the effect of 1400W, a selective inhibitor of inducible nitric oxide synthetase (iNOS), on the blockage of the toxicity. METHODS: Co-cultured microglia and preOLs obtained from two-day-old Sprague-Dawley (SD) rats were divided into three groups: co-culture control group, co-culture LPS group and co-culture LPS plus 1400W group. After cultured cells were induced by LPS (100 ng/ml) for 48 hours, the concentration of nitric oxide (NO) was measured by nitric acid-oeoxidize-colorimetry, the level of peroxynitrite (ONOO(-)) was determined by immunocytochemistry, and the synthetic level of iNOS was detected by Western blotting, respectively. The morphologic observation of apoptotic preOLs stained with Hoechst 33342/PI and the apoptotic rate of preOLs detected by flow cytometry were processed simultaneously. Data were analyzed with SPSS 11.0 software. RESULTS: Compared to co-culture control group, there was significant increase in levels of NO [(82.27+/-3.41) micromol/L vs. (167.86+/-9.87) micromol/L, t=8.593, P<0.01], ONOO(-)[(6.14+/-1.27) x 10(7)/L vs. (34.38+/-7.75) x 10(7)/L, t=5.892, P<0.01], and iNOS [(0.18+/-0.027) vs. (0.79+/-0.068), t=9.26, P<0.01] induced by LPS in co-culture LPS group, and with a higher apoptotic rate of preOLs [(6.73+/-1.39)% vs. (24.77+/-2.05)%, t=12.619, P<0.01]. However, all levels of NO [(69.55+/-5.07) micromol/L, t=8.896, P<0.01], ONOO(-) [(10.33+/-3.47) x 10(7)/L, t=14.96, P<0.01] and iNOS (0.35+/-0.042, t=5.506, P<0.01) decreased significantly with the use of 1400W at a dose of 10 micromol/L in co-culture LPS plus 1400W group, and the apoptotic rate of preOLs [(11.8+/-2.06)%, t=7.715, P<0.01] was also reduced evidently. CONCLUSIONS: NO, ONOO(-) and iNOS, etc. play important roles in the death pathway of preOLs induced by LPS. 1400W can block effectively the toxicity of LPS-activated microglia toxicity to preOLs through inhibiting iNOS selectively and reducing the production of NO and ONOO(-), and improve the survival rate of preOLs.