Inhibition of KIR2.1 decreases pulmonary artery smooth muscle cell proliferation and migration.

The investigation of effective therapeutic drugs for pulmonary hypertension (PH) is critical. KIR2.1 plays crucial roles in regulating cell proliferation and migration, and vascular remodeling. However, researchers have not yet clearly determined whether KIR2.1 participates in the proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) and its role in pulmonary vascular remodeling (PVR) also remains elusive. The present study aimed to examine whether KIR2.1 alters PASMC proliferation and migration, and participates in PVR, as well as to explore its mechanisms of action. For the in vivo experiment, a PH model was established by intraperitoneally injecting Sprague­Dawley rats monocrotaline (MCT). Hematoxylin and eosin staining revealed evidence of PVR in the rats with PH. Immunofluorescence staining and western blot analysis revealed increased levels of the KIR2.1, osteopontin (OPN) and proliferating cell nuclear antigen (PCNA) proteins in pulmonary blood vessels and lung tissues following exposure to MCT, and the TGF­ß1/SMAD2/3 signaling pathway was activated. For the in vitro experiments, the KIR2.1 inhibitor, ML133, or the TGF­ß1/SMAD2/3 signaling pathway blocker, SB431542, were used to pre­treat human PASMCs (HPASMCs) for 24 h, and the cells were then treated with platelet­derived growth factor (PDGF)­BB for 24 h. Scratch and Transwell assays revealed that PDGF­BB promoted cell proliferation and migration. Immunofluorescence staining and western blot analysis demonstrated that PDGF­BB upregulated OPN and PCNA expression, and activated the TGF­ß1/SMAD2/3 signaling pathway. ML133 reversed the proliferation and migration induced by PDGF­BB, inhibited the expression of OPN and PCNA, inhibited the TGF­ß1/SMAD2/3 signaling pathway, and reduced the proliferation and migration of HPASMCs. SB431542 pre­treatment also reduced cell proliferation and migration; however, it did not affect KIR2.1 expression. On the whole, the results of the present study demonstrate that KIR2.1 regulates the TGF­ß1/SMAD2/3 signaling pathway and the expression of OPN and PCNA proteins, thereby regulating the proliferation and migration of PASMCs and participating in PVR.

The promoting role of Cx43 on the proliferation and migration of arterial smooth muscle cells for angiotensin II-dependent hypertension.

BACKGROUND: Recent studies have shown that endothelin-1 and angiotensin II (AngII) can increase gap junctional intercellular communication (GJIC) by activating Mitogen-activated protein kinases (MAPKs) pathway. However, not only the precise interaction of AngII with Connexin43(Cx43) and the associated functions remain unclear, but also the regulatory role of Cx43 on the AngII-mediated promotion proliferation and migration of VSMCs is poorly understood. MATERIAL AND METHODS: Our research applicated pressure myography measurements, immunofluorescence and Western blot analyses to investigate the changes in physiological indicators in spontaneously hypertensive rats (SHRs) and AngII-stimulated proliferation and migration of A7r5 SMCs(Rat vascular smooth muscle cells). The aim was to elucidate the role of CX43 in hypertension induced by AngII. RESULTS: Chronic ramipril (angiotensin converting enzyme inhibitor) management for SHRs significantly attenuated blood pressure and blood vessel wall thickness, also reduced contraction rate in the cerebral artery. The cerebral artery contraction rates, mRNA and protein expression of Cx43, osteopontin (OPN) and proliferating cell nuclear antigen (PCNA) protein expression in the SHR + ramipril and SHR + ramipril + carbenoxolone (CBX, Cx43 specific blocker) groups were significantly lower than those in the SHR group. Cx43 protein expression and Ser368 phosphorylated Cx43 protein levels increased significantly in AngII-stimulated A7r5 cells. However, the levels of phosphorylated Cx43 decreased after pre-treatment with candesartan (AT1 receptor blocker), GF109203X (protein kinase C (PKC) blocker) and U0126 (mitogen-activated protein kinases/extracellular signal-regulated kinase1/2(MEK/ERK1/2)-specific blocker) in AngII-stimulated A7r5 cells. Cx43 was widely distributed in the cell membrane, nucleus, and cytoplasm of the SMCs. Furthermore, pre-treatment of the AngII- stimulated A7r5 cells with Gap26 (Cx43 blocker) significantly inhibited cell migration and decreased the expression levels of MEK1/2, ERK1/2, P-MEK1/2, and P-ERK1/2. CONCLUSION: Our research confirms that Cx43 plays an important role in the regulation of proliferation and migration of VSMCs via MEK/ERK and PKC signal pathway in AngII-dependent hypertension.

[Expression of ANO1 during cardiac fibroblasts differentiation and its electrophysiological characteristics as calcium-activated chloride channel protein].

OBJECTIVE: To investigate the expression and electrophysiological characteristics of calcium-activated chlorine channel anoctamin-1 (ANO1) protein during the differentiation of cardiac fibroblasts (CFs) into myofibroblasts (MFs), and to elucidate the role of ANO1 in myocardial fibrosis. METHODS: The primary CFs from neonatal rats were isolated and the cells differentiated into MFs by subculture. The Ca2+-activated Cl- current (ICl(Ca)) in CFs and MFs were measured by whole-cell patch clamp, and the expressions of ANO1, α-smooth muscle actin(α-SMA)and vimentin in CFs and MFs were detected by immunofluorescence assay and Western blot, respectively. RESULTS: The current density in the early adherent CFs was stronger than that in MFs. ANO1 was expressed preferentially within and around the nuclei, and a small amount of ANO1 was expressed on the cell membrane. Moreover, ANO1 expression was weak in the early adherent CFs and displayed stronger expression in the MFs with proliferation tendency. CONCLUSION: The expression of ANO1 is closely related to the differentiation of MFs and it may be involved in modulation myocardial fibrosis.

Age­associated variation in the expression and function of TMEM16A calcium­activated chloride channels in the cochlear stria vascularis of guinea pigs.

The present study was designed to investigate the expression and function of transmembrane protein 16 (TMEM16A), a calcium­activated chloride channel (CaCC), in the stria vascularis (SV) of the cochlea of guinea pigs at different ages, and to understand the role of CaCCs in the pathogenesis of presbycusis (age­related hearing loss), the most common type of sensorineural hearing loss that occurs with natural aging. Guinea pigs were divided into the following groups: 2 weeks (young group), 3 months (youth group), 1 year (adult group), D­galactose intervention (D­gal group; aging model induced by subcutaneous injection of D­galactose) and T16Ainh­A01 (intraperitoneal injection of 50 µg/kg/day TMEM16A inhibitor T16Ainh­A01 for 2 weeks). Differences in the hearing of guinea pigs between the various age groups were analyzed using auditory brainstem response (ABR), and immunofluorescence staining was performed to detect TMEM16A expression in the SV and determine the distribution. Reverse transcription­quantitative PCR and western blot analyses were conducted to detect the mRNA and protein levels of TMEM16A in SV in the different age groups. Morris water maze behavior analysis demonstrated that spatial learning ability and memory were damaged in the D­gal group. Superoxide dismutase activity and malondialdehyde content assays indicated that there was oxidative stress damage in the D­gal group. The ABR thresholds gradually increased with age, and the increase in the T16Ainh­A01 group was pronounced. Immunofluorescence analysis in the cochlear SV of guinea pigs in different groups revealed that expression of TMEM16A increased with increasing age (2 weeks to 1 year); fluorescence intensity was reduced in the D­gal model of aging. As the guinea pigs continued to mature, the protein and mRNA contents of TMEM16A in the cochlea SV increased gradually, but were decreased in the D­gal group. The findings indicated that CaCCs in the cochlear SV of guinea pigs were associated with the development of hearing in guinea pigs, and that downregulation of TMEM16A may be associated with age­associated hearing loss.

Mosapride combined with probiotics on gastrointestinal function and growth in premature infants.

The aim of this study was to investigate the effects of mosapride combined with probiotics on gastrointestinal function and growth and development in premature infants. A total of 240 premature infants treated at Weifang People's Hospital between June 2012 and May 2015 who matched our criteria were randomly divided into three groups of 80 cases each. Group A received routine treatment, group B received routine treatment combined with live B. subtilis and E. faecium granules with multivitamins (Medilac-Vita), and group C received routine treatment and Saccharomyces boulardii sachets (Bioflor). Mosapride was administered to patients in groups B and C to promote intestinal peristalsis. Gastrin and bilirubin levels, as well as jaundice fade time, were recorded. Growth and development condition (i.e., head circumference and weight), duration and incidence of feeding intolerance (FI), as well as other symptoms were also analyzed. By day 14, gastrin concentrations in groups B and C were significantly higher than those in group A (P<0.05). Serum bilirubin levels in groups B and C showed a progressive decline from day 7 to day 14, and jaundice duration in group A was significantly longer (P<0.05). Furthermore, at 2 weeks, the average weight growth rate and head circumference were significantly greater in groups B and C, weight loss recovery time was shorter, and EUGR incidence was lower (P<0.05). The incidence rate of gastric retention and FI were lower in groups B and C than group A (P<0.05), and neonatal hyperbilirubinemia, parenteral nutrition-associated cholestasis, necrotizing enterocolitis, and neonatal sepsis incidence was significantly lower in groups B and C (P<0.05). Mosapride combined with probiotics can effectively reduce FI incidence in premature infants, shorten enteral feeding time, promote the absorption of required nutrients, and promote the development and recovery of early physiological weight loss in preterm infants.

[Diversity of uncultured actinomycetes in saline-alkali soil from Jiuquan area of Hexi Corridor].

ABSTRACT: In order to more accurately understand community structure and diversity of actinomycetes in saline-alkali soil from Jiuquan area of Hexi Corridor, the community structure and diversity from three kinds of soil samples (primary, secondary saline alkali soil and farmland soil) were analyzed using uncultured methods. The results showed that the 16S rDNA clone library of actinomycetales from the primary saline-alkali soil belonged to 19 OTUs, Micrococcineae, Propionibacterineae, Corynebacterineae, Frankineae, Pseudonocardineae and unknown groups of Actinomycetales; the 16S r DNA clone library of actinomycetales from the secondary saline-alkali soil belonged to 14 OTUs, Micrococcineae, Propionibacterineae, Corynebacterineae, Frankineae, Pseudonocardineae and unknown groups of Actinomycetales; the 16S rDNA clone library of farmland soil belonged to 7 OTUs, Micrococcineae, Propionibacterineae, Corynebacterineae, Frankineae, Pseudonocardineae and unknown groups of Actinomycetales; Micrococcineae was the common population in the three soils, and also was the dominant population in primary saline alkali soil and farmland soil. The diversity index and rarefaction curves analysis showed that actinomycetes species richness was in order of primary saline-alkali soil > secondary saline-alkali soil > farmland soil. The dilution curves of primary saline-alkali soil and secondary saline-alkali soil were not leveled off, which indicated the actinomycetes diversity in saline-alkali soil was more enriched than the actual. The rich and diverse actinomycetes resources in saline-alkali soil from Jiuquan area of Hexi Corridor provide important data on the actinomycetes ecology distribution research, exploitation and utilization in saline-alkali soil.

Phylogeny and virulence divergency analyses of Toxoplasma gondii isolates from China.

BACKGROUND: Toxoplasma gondii (T. gondii) is a very successful parasite that can infect virtually all warm blooded animals with a worldwide distribution. It causes a large range of clinical manifestations in both humans and domesticated animals. In addition, marked biological differences exist among T. gondii strains in the pathogenicity and geographical distribution. Molecular epidemiology studies primarily based on restriction fragment length polymorphism (RFLP) method revealed that three main types are predominant in North America and Europe, whereas other diverse genotypes are found in other parts of the world. Microsatellite (MS) as a type of genetic marker has been widely used in many organisms. Limited MS genotyping, however, to fingerprint T. gondii isolates has been reported and little is known about the MS data of the strains predominantly prevalent in China. METHODS: Genotyping of twenty-eight Chinese T. gondii isolates were performed using 15 MS markers located on 12 different chromosomes. Results were analyzed in terms of population structure by a Bayesian statistical approach. Phylogenetic analysis was obtained from a Neighbor-Net phylogenetic network. The virulence analyses of some representative isolates were determined by inoculation of mice and cell invasion assays. The gene expressions of some virulence-associated factors (VFs) were performed by quantitative real-time PCR (qRT- PCR). RESULTS: Three haplogroups were clustered among the 28 isolates although minor genetic differences were found within haplogroups. The majority of strains belong to one haplogroup corresponding to the previously described Chinese 1 type (ToxoDB#9). Phylogenetic networks uncovered a limited diversity of T. gondii strains and the virulence differs in the strains sharing the same genotype. No remarkable difference, however, was noted in the tested VFs except for dense granule protein3 (GRA3), which was found to have a higher expression in low virulent TgCtwh6 (Wh6) strain than that in high virulent TgCtwh3 (Wh3) strain. CONCLUSION: The profile of microsatellite typing data from Chinese T. gondii strains revealed a limited genetic diversity and the selected VFs and phylogenetic network analyses displayed less divergence, although the strain virulence differs in the Chinese 1 type of T. gondii predominantly prevalent in China.

Preventive effect of pidotimod on reactivated toxoplasmosis in mice.

As one of food-borne parasitic diseases, toxoplasmosis entails the risk of developing reactivation in immunocompromised patients. The synthetic dipeptide pidotimod is a potent immunostimulating agent that improves the immunodefenses in immunodepression. To investigate the efficacy of pidotimod as a preventive treatment, we used a murine model of reactivated toxoplasmosis with cyclophosphamide (CY)-induced immunosuppression. Pidotimod administration significantly restored the body weight and spleen organ index, increased survival time (from 70 to 90%), and decreased the parasitemia (from 80 to 35%) of CY-induced mice with reactivated toxoplasmosis. Cytokine profiles and CD4(+) T cells subpopulation analyses by Cytometric Bead Array and flow cytometry demonstrated that pidotimod treatment resulted in a significant upregulation of pro-inflammatory cytokines (IFN-γ, TNF-α, and IL-2) and Th1 cells (from 3.73 ± 0.39 to 5.88 ± 0.46%) after CY induction in infected mice. Additionally, histological findings and parasite DNA quantification revealed that mice administered with pidotimod had a remarkable reduction of parasite burden (two-log) and amelioration of histopathology in the brains. The in vitro studies showed that pidotimod significantly restored concanavalin A-induced splenocyte proliferation and pro-inflammatory cytokines in the supernatants of splenocyte culture. It could be concluded that the administration of pidotimod in immunocompromised mice significantly increases the Th1-biased immune response, prolongs survival time, and ameliorates the load of parasites in the blood. This is the first report of the preventive effect of pidotimod on reactivated toxoplasmosis.

Silencing of miR155 promotes the production of inflammatory mediators in Guillain-Barré syndrome in vitro.

MicroRNA-155 (miR155) has been demonstrated as a central regulator of immune responses induced by inflammatory mediators. Previous studies suggest that miR155 may play adverse effects in various diseases. We hereby explored the roles of miR155 in the pathogenesis of Guillain-Barré syndrome (GBS). Peripheral blood mononuclear cells (PBMCs) were separated from GBS patients and healthy controls. Expression of miR155 in PBMCs was detected by quantitative PCR. An inhibitor of miR155 was transfected into the cultured PBMCs and the GBS-related cytokines were detected. Significantly, our study demonstrated that miR155 was downregulated in PBMCs from GBS patients and silencing of miR155 profoundly promoted the production of Th1-type cytokines in vitro. Our data effectively demonstrate a protective role of miR155 in GBS, which suggests that miR155 may be a promising target for the therapy of the disease.

Comparative studies of macrophage-biased responses in mice to infection with Toxoplasma gondii ToxoDB #9 strains of different virulence isolated from China.

BACKGROUND: Different from three clonal lineages of Toxoplasma gondii in North America and Europe, the genotype China 1 is predominantly prevalent in China. However, there are different virulent isolates within China 1, such as virulent TgCtwh3 and avirulent TgCtwh6, and little is known about differences in macrophage activation between them. The objective of this study focused on cytokine production, phenotype and markers of activated macrophages, and correlated signaling pathway induced by the two isolates. METHODS: Adherent peritoneal macrophages (termed Wh3-Mφ and Wh6-Mφ, respectively) harvested from infected mice were cultured for detection of Nitric Oxide and arginase activity, and activated markers on Wh3-Mφ/Wh6-Mφ were determined by flow cytometry. In in vitro experiments, the levels of IL-12p40 and TNF-α were measured using ELISA kits, and mRNA expressions of IL-12p40, TNF-α, iNOS, Arg-1 and Ym1 were assayed by real-time PCR. To confirm the activation state of NF-kB p65 in infected cells stained by IF, protein levels of iNOS, Arg-1, Ym1, nuclear NF-κB p65, and phosphorylation of STAT6/STAT3/IκBα were evaluated by Western Blotting. A one-way ANOVA test was used to compare differences among multiple groups. RESULTS: The result revealed that contrary to the virulent TgCtwh3, the less virulent TgCtwh6 isolate induced a significant increase in IL-12p40 and TNF-α. Although both isolates down-regulated CD80, CD86 and MHCII molecule expression on macrophages, TgCtwh3 promoted up-regulation of PD-L2 and CD206. Wh6-Mφ generated a high level of NO whereas Wh3-Mφ up-regulated Ym1 and arginase expression at transcriptional and protein levels. In terms of signaling pathway, TgCtwh3 induced phospho-STAT6, conversely, TgCtWh6 led to NF-κB p65 activation. CONCLUSIONS: The virulent TgCtwh3 isolate induced macrophages to polarize toward alternatively activated cells with STAT6 phosphorylation, whereas the less virulent TgCtwh6 elicited the development of classically activated macrophages with nuclear translocation of NF-κB p65. This discrepancy suggests that it is necessary to thoroughly analyze the genotype of TgCtwh3 and TgCtwh6, and to further study other effector molecules that contribute to the macrophage polarization in T. gondii.

Changes of Treg-associated molecules on CD4+CD25 +Treg cells in myasthenia gravis and effects of immunosuppressants.

OBJECTIVE: Myasthenia gravis (MG) is a CD4(+) T cell-dependent autoimmune disease, and close attention has been paid to the role of CD4(+)CD25(+)Treg cells (Tregs). Previous results regarding Tregs in MG patients have been conflicting. The discrepancy was partly ascribed to selecting different Treg-associated molecules in defining Tregs. Therefore, we considered it necessary to find a reliable index for assessing the immunologic state in MG patients and explore the effect of IS on them. METHODS: We adopted flow cytometric techniques to measure the numbers and frequencies of Tregs in peripheral blood taken from 57 patients and 91 age-matched healthy donors, and we also analyzed FOXP3 mean fluorescence intensity on Tregs. RESULTS: The number and frequency of Tregs in peripheral blood of MG patients significantly decreased, together with down-regulation of FOXP3 expression. There was dynamic change of Treg cell level and the inverse relationship with clinical symptom, suggesting that the immunologic disorder in MG patients was related to peripheral Tregs population. Meanwhile, CD4(+)CD25(+)FOXP3(+)Helios(+)T cells might be activated Tregs, rather than nTregs. Moreover, the number and frequency of CD4(+)CD25(+)FOXP3(+)Helios(+)T cells significantly decreased in MG patients, indicating that the reduction of the activated Tregs population might be a critical contributor to the pathogenesis of MG. CONCLUSIONS: The significant reduction of the peripheral Tregs population in MG patients might be responsible for the immunologic disorders in MG patients. IS such as GC took its effect possible by increasing the population size, and the underlying mechanism should be further investigated.

[Isolation and identification of Japanese encephalitis virus in Tanghe County, Henan Province].

OBJECTIVE: To isolate Japanese encephalitis virus (JEV) from mosquitoes collected in Tanghe County, Henan province and analyze the genotype of the newly isolated JEV strains and the characteristics of amino acid in the E gene. METHODS: Viruses were isolated from mosquitoes collected in 2004 and identified by biological, serological and molecular biologic methods. PrM and E segments of the newly isolated JEV were amplified by RT-PCR, the PCR products were purified and sequenced. Multiple alignment, phylogenetic and amino acid (AA) analysis were carried out by Clustal X (1.8) program, MEGA 3.1 and GENEDOS (3.2). RESULTS: Totally 3722 mosquitoes were collected including Culex, Armigeres, Aedes, Anopheline. Three new JEV strains isolated from Culex belonged to genotype 1. The homologue of nucleotide and amino acid of E gene between new JEV strains and live attenuated vaccine strain SA14-14-2 was 86.9-87.7% and 95.2%-97.0%, respectively. Totally there were 12 common sites of amino acid differences in E gene between them. CONCLUSION: Newly isolated viruses in Henan province belonged to JEV genotype 1. It suggests that the vaccine strain SA14-14-2 currently used for preventing JE is able to protect people from JEV infection, although there are some amino acid differences between them.

[Clinical significance of detection of T-cell subgroups in patients with aplastic anemia].

The study was aimed to investigate the changes of T-cell subgroups in the peripheral blood (PB) of patients with aplastic anemia (AA) and the relationships between these changes and the pathogenesis of AA and the immunosuppressive therapeutic effects in AA, in order to provide a basis for selecting rational therapy of AA patients. T-cell subtype and the ratio of CD4+/CD8+ cell in the PB of 88 AA patients which had been diagnosed clearly and given conventional therapy or conventional therapy combined with immunotherapy were analyzed by tri-colour fluorescence-labeled monoclonal antibody and using multiparameter flow cytometry. The patients with AA were divided into normal type of ratio, inverted type of ratio, hypernormal type of ratio according to the ratio of CD4+/CD8+ cell in normal group, and then the relations of these subtype with patients' conditions and therapeutic effects were investigated. The results showed that the percentage of normal type of ratio in all patients was 39.8%, the percentage of inverted type of ratio in all patients was 44.3%, The percentage of hypernormal type of ratio in all patients was 15.9%. In the conventional therapy alone, there was no significant difference on therapeutic effects among these three immunological subtypes. In combined immunotherapy, total therapeutic efficacy of AA patients with inverted type of ratio and AA patients with immunologic abnormality (inverted type + hypernormal type) was 84.2% and 82.6% respectively, which were more than that in conventional therapy (45.5% and 42.8%) (p < 0.05). Total therapeutic efficacy in these patients was better than that in AA patients with normal type. It is concluded that significant abnormal ratios of CD4+/CD8+ exist in the majority of AA patients, abnormal ratios of CD4+/CD8+ both may be showed as increase or decrease, immunologic abnormality may play a role in pathogenesis of the patients with AA. The detection of PB T-cell subtype in patients with aplastic anemia contributes to evaluation of patients' condition and choice of rational treatment prescription, and enhancement of diagnostic level and therapeutic efficacy significantly, which is an important indicator for therapeutic strategy also.

[Volt-ampere characteristics of Yuan points of three yin channels of hand in women before, during and after menstruation].

OBJECTIVE: To observe the changes of volt-ampere characteristics of Yuan-Primary acupoints in women before, during and after menstruation, and to find out the relationships between Qi-blood in human body and volt-ampere characteristics of acupoints, so as to develop a new method for quantitative analysis on changes of Qi-blood. METHODS: A high-sensitive detection system of volt-ampere characteristics of acupoint was applied to detecting the volt-ampere characteristics of Taiyuan (LU 9), Daling (PC 7) and Shenmen (HT 7) in healthy women before, during and after menstruation. RESULTS: The resistances on LU 9, PC 7 and HT 7 with some certain scan currents were quite different among the women before, during and after menstruation. For such changes, PC 7 was of the most obvious change, LU 9 was of the less and HT 7 was of the least. The total ratio of the scanning spots with significantly increasing changes of resistances on these three acupoints along with the menstrual process was 70.6%. CONCLUSION: The volt-ampere characteristics of acupoints vary with the menstrual process, which is of the acupoint specificity. The resistances on the acupoints increase with the menstrual process, and such results may be due to the blood loss during menstruation. The volt-ampere characteristics can be used as a quantitative index to study on the change of Qi and blood.

[Genetics subtypes and distribution of Seoul virus in Henan].

OBJECTIVE: To investigate the sub-genotypes and distribution ot Seoul virus in Henan. METHODS: Rodents were collected in the major epidemic areas and rats lungs were studied by indirect immunofluorescence assay. Partial M and S segments were amplified with nested reverse transcription-polymerase chain reaction using Hantavirus genotype-specific primers, sequenced, analyzed and compared with other known sequences. RESULTS: The Hantavirus carried by Rattus norvegicus, Rattus flavipectus and Mus musculus were all belonged to Seoul virus in the main epidemic areas of Henan. We constructed two phylogenetic tree based on the partial M and S segment sequences while phylogenetic analysis distinguished three genetic subtypes (S1, S2 and S3). S1 and S3 were found main subtypes in Henan. CONCLUSION: The results indicated that the genetic subtypes of Hantavirus were complicated and widely distributed in Henan.

[Inhibitory kinetic-synchronous fluorimetric determination of trace aniline].

A novel kinetic-synchronous fluorimetric method is reported for the determination of trace aniline. The method is based on the inhibitory effect of aniline on the oxidation of fluorescein by potassium bromate in the presence of surfactant -CTMAB in the acid medium. Fluorescein's oxidized product produces strong fluorescence. But its excitation and emission spectra overlap seriously. So this article adopted synchronous fluorimetric spectra to diminish spectra overlap and obtained very good sensitivity and selectivity. The detection limit for aniline is 0.3 microg x L(-1), and the linear range is 0-10 microg x L(-1). The proposed method has been applied to the determination of trace aniline in waste water and laboratory water with satisfactory results.

[A novel mutation of KCNQ2 gene in a Chinese family with benign familial neonatal convulsions].

OBJECTIVE: To diagnose a Chinese benign familial neonatal convulsions (BFNC) family at the level of gene and investigate its molecular pathogenesis. METHODS: All family members were studied by clinical examinations and linkage analysis. Mutation analysis of KCNQ2 gene was made by means of polymerase chain reaction (PCR)-direct sequencing and PCR-single strand conformation polymorphism (SSCP) in the proband, 16 family members and 72 unrelated normal individuals. RESULTS: Linkage analysis hinted the linkage of BFNC to KCNQ2, while the linkage to KCNQ3 was excluded. Mutation 1931delG of KCNQ2 gene was found in the proband by DNA-direct sequencing. The same SSCP variant as the proband's was showed in the rest affected members of this family but not in the unaffected members of this family and all of the 72 unrelated normal individuals. CONCLUSION: 1931delG of KCNQ2 gene can cause BFNC in China and is novel mutation. The combination of linkage analysis and gene analysis is useful for gene diagnosis.