A new class of monoclonal Aß antibodies selectively targets and triggers deposition of Aß protofibrils.

Aggregation and accumulation of amyloid-ß peptide (Aß) are a critical trigger for the onset of Alzheimer's disease (AD). While the plaques are the most outstanding Aß pathological feature, much of the recent research emphasis has been on soluble Aß species because of their diffusible, proinflammatory, and toxic properties. The focus on soluble aggregated Aß species has also increased the interest in antibodies that are selective for different Aß conformations. In the current study, we developed and characterized a new class of monoclonal antibodies (referred to as mAbSL) that are selective for Aß protofibrils. Cloning and sequencing of the heavy and light chain variable regions for multiple antibodies identified sequence characteristics that may impart the conformational selectivity by the antibodies. Transfection of FreeStyle 293F cells with the plasmids permitted in-house expression and purification of mAbSL antibodies along with non-conformation-selective Aß monoclonal antibodies (Aß mAbs). Several of the purified mAbSL antibodies demonstrated significant affinity and selectivity for Aß42 protofibrils compared with Aß42 monomers and Aß42 fibrils. Competition ELISA assays assessing the best overall antibody, mAbSL 113, yielded affinity constants of 7 nM for the antibody-Aß42 protofibril interaction, while the affinity for either Aß42 monomers or Aß42 fibrils was roughly 80 times higher. mAbSL 113 significantly inhibited Aß42 monomer aggregation by a unique mechanism compared with the inhibition displayed by Aß mAb 513. Aß42 protofibril dynamics were also markedly altered in the presence of mAbSL 113, whereby insoluble complex formation and protofibril deposition were stimulated by the antibody at low substoichiometric molar ratios. As the field contemplates the therapeutic effectiveness of Aß conformation-selective antibodies, the findings presented here demonstrate new information on a monoclonal antibody that selectively targets Aß protofibrils and impacts Aß dynamics.

[Etiology of nonspecific chronic cough in children and relationship between TRPV1 gene polymorphisms and nonspecific chronic cough].

OBJECTIVE: To explore the causes of nonspecific chronic cough in children and relationship between transient receptor potential vanilloid 1 (TRPV1) gene polymorphisms and nonspecific chronic cough. METHODS: A total of 195 children with chronic cough were followed up half a month, one month and three months after their first visit to hospital. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to examine polymorphisms of the TRPV1 gene in the children. A total of 205 healthy or surgical children without chronic cough served as the control group. RESULTS: The etiologic distribution of the 195 children with chronic cough was as follows: 96 (49.2%) cases of cough variant asthma (CVA), 48 (24.6%) cases of CVA complicated by upper airway cough syndrome (UACS), 34 (17.4%) cases of post-infectious cough, and 17 (8.7%) cases of UACS. Three genotypes were identified in both groups at positions rs222747 (CC, GC and GG), rs222748 (CC, TC and TT) and rs8065080 (CC, TC and TT). The frequencies of genotype and allele at position rs222747 did not accord with the law of Hardy-Weinberg. There was no significant difference in frequencies of genotype and allele at positions rs222748 and rs8065080 between the two groups. CONCLUSIONS: CVA, UACS and post-infectious cough are common causes of nonspecific chronic cough in children. TRPV1 gene polymorphisms at positions rs222748 and rs8065080 may be unrelated to nonspecific chronic cough in children.

atToc159 is a selective transit peptide receptor for the import of nucleus-encoded chloroplast proteins.

The members of the Toc159 family of GTPases act as the primary receptors for the import of nucleus-encoded preproteins into plastids. Toc159, the most abundant member of this family in chloroplasts, is required for chloroplast biogenesis (Bauer, J., K. Chen, A. Hiltbunner, E. Wehrli, M. Eugster, D. Schnell, and F. Kessler. 2000. Nature. 403:203-207) and has been shown to covalently cross-link to bound preproteins at the chloroplast surface (Ma, Y., A. Kouranov, S. LaSala, and D.J. Schnell. 1996. J. Cell Biol. 134:1-13; Perry, S.E., and K. Keegstra. 1994. Plant Cell. 6:93-105). These reports led to the hypothesis that Toc159 functions as a selective import receptor for preproteins that are required for chloroplast development. In this report, we provide evidence that Toc159 is required for the import of several highly expressed photosynthetic preproteins in vivo. Furthermore, we demonstrate that the cytoplasmic and recombinant forms of soluble Toc159 bind directly and selectively to the transit peptides of these representative photosynthetic preproteins, but not representative constitutively expressed plastid preproteins. These data support the function of Toc159 as a selective import receptor for the targeting of a set of preproteins required for chloroplast biogenesis.

Members of the Toc159 import receptor family represent distinct pathways for protein targeting to plastids.

Plastids represent a diverse group of organelles that perform essential metabolic and signaling functions within all plant cells. The differentiation of specific plastid types relies on the import of selective sets of proteins from among the approximately 2500 nucleus-encoded plastid proteins. The Toc159 family of GTPases mediates the initial targeting of proteins to plastids. In Arabidopsis thaliana, the Toc159 family consists of four genes: atTOC159, atTOC132, atTOC120, and atTOC90. In vivo analysis of atToc159 function indicates that it is required specifically for the import of proteins necessary for chloroplast biogenesis. In this report, we demonstrate that atToc120 and atToc132 represent a structurally and functionally unique subclass of protein import receptors. Unlike atToc159, mutants lacking both atToc120 and atToc132 are inviable. Furthermore, atToc120 and atToc132 exhibit preprotein binding properties that are distinct from atToc159. These data indicate that the different members of the Toc159 family represent distinct pathways for protein targeting to plastids and are consistent with the hypothesis that separate pathways have evolved to ensure balanced import of essential proteins during plastid development.

[Proportion of incidence of etiological agents in children with non-specific chronic cough in Chongqing: a follow-up study].

OBJECTIVE: To investigate the proportion of incidence of children with non-specific chronic cough in Chongqing and analyze the characteristics of etiology during the follow-up. METHOD: Diagnostic criteria were defined for children with non-specific chronic cough according to the Guidelines of diagnosis and therapy for children with chronic cough that were formulated by the Subspecialty Group, Society of Pediatrics, Chinese Medical Association and Chinese Journal of Pediatrics in 2008. Totally 266 patients in whom cough was the main or the only symptom,lasting > 4 weeks, presenting to Asthma Center of Children's Hospital, Chongqing Medical University between June 2008 and April 2009 were recruited into this study. Based on the Guidelines, diagnosis was made after taking history, physical examination and assistant examination. After etiological treatment, the patients were followed up during the second week, the fourth week and the twelfth week. Etiological diagnosis was confirmed if cough was resolved after specific therapy. If cough was not resolved,the diagnosis was rechecked and a new therapy was applied. RESULT: Totally 125 (47.0%) patients received final diagnoses of cough variant asthma (CVA), 58 (21.8%) was CVA and upper airway cough syndrome (UACS), 44 (16.5%) was diagnosed postinfection cough, 35 (13.2%) of UACS. In different age groups, the proportion of incidence of etiological agents is statistically distinct. In the ≤ 3 years old group, 35 patients (70.0%) were diagnosed CVA, 10 (20.0%) was postinfection cough; in 3 - 6 years group, 71 patients (50.7%) had CVA; the incidence of UACS was significantly higher in ≥ 6 years group. CONCLUSION: It is concluded that CVA, CVA and UACS, post infection cough, and simple UACS were identified as the three top reasons for children with chronic cough in Chongqing. Children with chronic cough of different age groups had different etiology of cough. The characteristic of each etiology need further study.

[Analysis of 53 cases with bronchoscopically confirmed pediatric tracheobronchomalacia].

OBJECTIVE: Tracheobronchomalacia is one of the common respiratory tract dysplasia in children. Its symptoms are nonspecific, and routine methods are unreliable in the assessment of tracheobronchomalacia in children. In addition, many physicians are confused about its clinical characteristics, so tracheobronchomalacia is often underdiagnosed. The purpose of this study was to explore the clinical features of tracheobronchomalacia in children and to investigate the diagnostic value of flexible bronchoscopy for children with tracheobronchomalacia. METHOD: For diagnosis and treatment, 229 children out of 4725 patients hospitalized in the division of respiratory disorders were examined by Olympus BF3c-20 flexible bronchoscopy or by Olympus BF-P20 flexible bronchoscopy under general anesthesia with propofol, in Chongqing Children's Hosptial from April 2004 to April 2006. Fifty-three cases were confirmed to have tracheobronchomalacia by bronchoscopy, patients' data including airway lesion, age, sex, clinical characteristics, aided examinations, treatment, final outcomes, were collected and analyzed. RESULTS: (1) Of the 53 children with tracheobronchomalacia, 31 were not suspected for this diagnosis prior to bronchoscopy, who were instead misdiagnosed as refractory pneumonia, difficult-to-control asthma, bronchial foreign body, bronchopulmonary dysplasia and pulmonary atelectasis of unknown origin or bronchiolitis. (2) In the 53 children with tracheobronchomalacia aged one month to eight years, 41 were infants, 6 were younger than two years, 4 were younger than 3 years and the rest 2 cases were older than 3 years. The risk of tracheobronchomalacia related inversely with ages. Ten cases were girls and 43 were boys. (3) Eleven cases had tracheomalacia, 24 bronchomalacia, 18 tracheobronchomalacia; 12 cases had malacia on left lung, 11 on right lung, 19 on both sides; 21 children were mild cases, 25 moderate cases, 7 severe cases. (4) In the 53 children with tracheobronchomalacia, 28 had recurrent or prolonged wheezing, 16 chronic cough, 5 recurrent respiratory infections, 2 atelectasis of unknown origin, and 2 dyspnea. CONCLUSIONS: The infants and toddlers seem to be predisposed more to the bronchomalacia than the older children. Clinical features of children with airway malacia were variable and atypical, expiratory stridor and cough are the most commonly reported symptoms. Flexible bronchoscopy should be regarded as a "golden standard" method for diagnosing TBM.

[Role of Foxp3 expression and CD4+CD25+ regulatory T cells on the pathogenesis of childhood asthma].

OBJECTIVE: To explore the impact of Foxp3 expression and CD(4)(+)CD(25)(+) regulatory T cells on pathogenesis of childhood asthma. METHODS: Totally 15 patients with acute asthma exacerbation, 15 children with asthma remission and 10 children who were hospitalized for skeleton deformity without atopic disorders or history of allergic diseases or respiratory infections within a month as controls were recruited in this study from Sep. 2004 to Mar. 2005. The percentage of CD(4)(+)CD(25)(+) T cells were detected by 2-color flow cytometry. The levels of interleukin (IL)-4, IL-10, interferon (IFN)-gamma, transforming growth factor (TGF)-beta in plasma and supernatant were assayed by ELISA. Both the asthmatic children and the control children were selected to induce sputum by hypertonic saline. Sputum was processed for detecting the expression of Foxp3-mRNA. The expression of Foxp3-mRNA in both sputum and PBMC was detected by semi-quantitative RT-PCR with beta-actin as internal control. RESULTS: The percentage of CD(4)(+)CD(25)(+) regulatory T cells in exacerbation and remission asthmatic children was significantly lower than that of the control children both prestimulation [(10.1 +/- 2.1)% vs. (15.5 +/- 2.7)%, (11.7 +/- 2.5)% vs. (15.5 +/- 2.7)%, P < 0.05] and poststimulation with PHA [(12.4 +/- 2.3)% vs. (26.9 +/- 3.8)%, (17.3 +/- 3.2)% vs. (26.9 +/- 3.8)%, P < 0.05]. The percentage of CD(4)(+)CD(25)(+) regulatory T cells was significantly higher after PHA stimulation in normal children [(15.5 +/- 2.7)% vs. (26.9 +/- 3.8)%, P < 0.01]. The expression of Foxp3-mRNA (Foxp3/beta-actin) in asthmatic children was significantly lower than that in the control children in both PBMC and induced sputum. The expression of Foxp3-mRNA in PBMC was significantly higher after PHA stimulation in the control children (0.77 +/- 0.22 vs. 1.07 +/- 0.21, P < 0.05). However, there was no significant difference in Foxp3-mRNA expression in asthmatic children pre and post PHA stimulation. A significant positive correlation between the Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells was detected. The levels of IFN-gamma and TGF-beta were significantly lower in asthmatic children than those in the control children, and the levels of IFN-gamma and TGF-beta correlated positively with Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells. The level of IL-4 both in plasma and supernatant was higher in asthmatic children. The levels of IL-10 was higher only in exacerbation than in control children, the levels of IL-4 and IL-10 had no correlation with Foxp3-mRNA expression and the percentage of CD(4)(+)CD(25)(+) regulatory T cells. CONCLUSION: Insufficient secretion of TGF-beta, decreased Foxp3 expression, insufficient number of CD(4)(+)CD(25)(+) regulatory T cells and the defective ability of converting CD(4)(+)CD(25)(-) T cells to CD(4)(+)CD(25)(+) regulatory T cells might play an important role in pathogenesis of asthma.

Phytochrome control of the Arabidopsis transcriptome anticipates seedling exposure to light.

Phytochromes mediate a profound developmental shift when dark-grown seedlings are exposed to light. Here, we show that a subset of genes is upregulated in phytochrome B (phyB) mutants even before dark-grown Arabidopsis thaliana seedlings are exposed to light. Most of these genes bear the RY cis motif, which is a binding site of the transcription factor ABSCISIC ACID INSENSITIVE3 (ABI3), and the phyB mutation also enhances ABI3 expression. These changes in transcriptome have physiological consequences, because seedlings of the abi3 mutant showed enhanced responses to pulses of far-red light, whereas ABI3 overexpressers exhibited the opposite pattern. Seedlings of the wild type derived from seeds germinated in full darkness showed enhanced expression of genes bearing the RY cis motif and reduced responses to far-red light. We propose that, via changes in ABI3 expression, light, perceived mainly by phyB in the seed, generates a downstream transdevelopmental phase signal that preconditions the seedling to its most likely environment.