MiR-448-5p inhibits TGF-ß1-induced epithelial-mesenchymal transition and pulmonary fibrosis by targeting Six1 in asthma.

MicroRNAs (miRNAs) are small yet versatile gene tuners that regulate a variety of cellular processes, including cell growth and proliferation. The aim of this study was to explore how miR-448-5p affects airway remodeling and transforming growth factor-ß1 (TGF-ß1)-stimulated epithelial-mesenchymal transition (EMT) by targeting Sine oculis homeobox homolog 1 (Six1) in asthma. Asthmatic mice models with airway remodeling were induced with ovalbumin solution. MiRNA expression was evaluated using quantitative real-time polymerase chain reaction. Transfection studies of bronchial epithelial cells were performed to determine the target genes. A luciferase reporter assay system was applied to identify whether Six1 is a target gene of miR-448-5p. In the current study, we found that miR-448-5p was dramatically decreased in lung tissues of asthmatic mice and TGF-ß1-stimulated bronchial epithelial cells. In addition, the decreased level of miR-448-5p was closely associated with the increased expression of Six1. Overexpression of miR-448-5p decreased Six1 expression and, in turn, suppressed TGF-ß1-mediated EMT and fibrosis. Next, we predicted that Six1 was a potential target gene of miR-448-5p and demonstrated that miR-448-5p could directly target Six1. An SiRNA targeting Six1 was sufficient to suppress TGF-ß1-induced EMT and fibrosis in 16HBE cells. Furthermore, the overexpression of Six1 partially reversed the protective effect of miR-448-5p on TGF-ß1-mediated EMT and fibrosis in bronchial epithelial cells. Taken together, the miR-448-5p/TGF-ß1/Six1 link may play roles in the progression of EMT and pulmonary fibrosis in asthma.

A Review of Three Chinese Cases of Acromicric/Geleophysic Dysplasia with FBN1 Mutations.

OBJECTIVE: This study aims to explore the clinical features and molecular diagnosis of FBN1-related acromelic dysplasia in Chinese patients. METHODS: The clinical and genetic features of three FBN1-related acromicric dysplasia (AD)/geleophysic dysplasia (GD) Chinese patients from two families were reviewed, and comprehensive medical evaluations were performed. Targeted next-generation sequencing was used to detect genetic mutations associated with short statures, including FBN1. Sanger sequencing was used to determine the de novo mutation origin. RESULTS: Patient 1 presented with short stature, short and stubby hands and feet, mild facial dysmorphism, hepatomegaly, delayed bone age and beak-like femoral heads. Patient 2 and this patient's father merely presented with short stature, wide and short hands, and beak-like femoral heads. One novel mutation, c.5272G>T(p.D1758Y), and one known mutation, c.5183C>T(p.A1728V), were identified in these patients. CONCLUSION: The clinical features varied among these patients. The variant c.5272G>T(p.D1758Y) is a novel mutation.

[Recombinant adeno-associated virus conducted NgR(DN) enhances axonal regeneration of optic nerve after trauma: experiment with rats].

OBJECTIVE: To investigate the effect of recombinant adeno-associated virus conducted NgRDN on the axonal regeneration of optic nerve after trauma. METHODS: Two kinds of adeno-associated virus (AAV), AAV-NgRDN-EGFP containing dominant negative form of Nogo receptor and enhanced green fluorescent protein (EGFP) and rAAV-NgR-EGFP containing Nogo-66 receptor (NgR) and EGFP, were constructed. 45 adult Wistar male rats were randomly divided into three equal groups, all with both eyes as experimental eyes: Groups A, B, and C to undergo injection of rAAV-EGFP, rAAV-NgR-EGFP, and rAAV-NgRDN-EGFP respectively into the vitreous; and each group was subdivided into 3 equal subgroups: subgroups 1 underwent injection of rAAV only, subgroups 2 underwent injection of rAAV and lens trauma, and subgroups 3 underwent injection of rAAV and zymosan. The rats in the Subgroups A2, B2, and C2 underwent. Crush of the optic nerve 2 mm behind the eyeball with optic nerve forceps 3 weeks after the injection. Four days after the crush the right eyes were taken out and the retinal explants were cultured in 2 kinds of culture fluid: with or without myelin. The growth of axons at the edge of retinal explants was observed by immunofluorescent staining with betaIII tubulin. Two weeks after the crush the other eyes were taken out to isolate the optic nerves. Immunofluorescence assay was used to detect the expression of growth associated protein-43 (GAP-43) of optic nerve. The axonal regeneration of optic nerve was observed. RESULTS: betaIII tubulin staining showed that on the condition of culture fluid without myelin both rAAV-NgR-EGFP and rAAV-NgRDN-EGFP showed no effects on the axonal regeneration of retinal ganglion cells (RGCs). However, on the condition of culture fluid with myelin the count of axonal regeneration and the length of regenerated axons of Group B were (13+/-4) and (36 microm+/-4 microm), both significantly lower than those of Group A [(21+/-4) and (83 microm+/-11 microm) respectively, both P<0.01]. There were not significant differences in count of axonal regeneration and length of regenerated axons between Subgroups C1 and A1. The count of axonal regeneration and length of regenerated axons of Subgroups C2 were (317+/-45) and (508 microm+/-44 microm), both significantly higher than those of Subgroup C3 [(238+/-30) and (365 microm+/-48 microm) respectively, both P<0.01], and the values of both Subgroups C2 and C3 were significantly higher than those of Subgroups A2 and A3. The GAP43-positive area in the optic nerve of Group C was significantly larger than that of Group A (P<0.01), and that of Group B was significantly smaller than that of Group A (P<0.01). The GAP43-positive area in the optic nerve of Subgroup A2 was (18.71+/-1.72)x100 microm2, significantly larger than that of Subgroup A3 [(12.75+/-1.02)x100 microm2, P<0.01], and that of Subgroup A3 was significantly larger than that of Subgroup A1 (P<0.01). There were not significant differences in the GAP43-positive area among the subgroups in Group B. CONCLUSION: Transfection of rAAV-NgRDN-EGFP into RGC in an activated status enhances axonal regeneration of optic nerve. NgRDN AAV can inhibit effectively the role of NgR.

Targeted inhibition of Six1 attenuates allergic airway inflammation and remodeling in asthmatic mice.

Asthma is an inflammatory disease of the airways, characterized by lung eosinophilia, mucus hypersecretion by goblet cells and airway hyperresponsiveness to inhaled allergens. The purpose of this study was to evaluate the effects of Six1 on airway inflammation and remodeling and the underlying mechanisms in a murine model of chronic asthma. Female BALB/c mice were randomly divided into four groups: phosphate-buffered saline control, ovalbumin (OVA)-induced asthma group, OVA+siNC and OVA+siSix1. In this mice model, Six1 expression level was significantly elevated in OVA-induced asthma of mice. Additionally, downregulation of Six1 dramatically decreased OVA-challenged inflammation, infiltration, and mucus production. Moreover, silencing of Six1 resulted in decreased levels of immunoglobulin E and inflammatory mediators and reduced inflammatory cell accumulation, as well as inhibiting the expression of important mediators including matrix metalloproteinase MMP-2 and MMP-9, which is related to airway remodeling. Further analysis indicated that silencing of Six1 can significantly inhibit NF-kB pathway activation in the lungs. .In conclusion, these findings indicated that the downregulation of Six1 effectively inhibited airway inflammation and reversed airway remodeling, which suggest that Six1 represents a promising therapeutic strategy for human allergic asthma.

[Cluster analysis in micrangium detection in malignant nasal and paranasal sinus tumor].

OBJECTIVE: To study the application of cluster analysis in micrangium detection in malignant nasal and paranasal sinus tumor. METHODS: Microvessel density (MVD) counting and cluster analysis were used to detect the micrangium in patients with malignant nasal and paranasal sinus tumor to assess the association between the malignancy and MVD. RESULTS: According to cluster analysis, the MVD counting could be clustered into two groups, and the MVD showed significant differences between the tumor tissues, adjacent normal tissue and the control group (P<0.01), a result consistent with that by analysis of variance of the MVD. CONCLUSION: Cluster analysis can be used in clustering of MVD counting in malignant nasal and paranasal sinus tumor to simplify MVD counting, and offers an important analytic method for micrangium analysis in tumors.