Mycoplasma pneumoniae Induces Chronic Respiratory Infection, Airway Hyperreactivity, and Pulmonary Inflammation: a Murine Model of Infection-Associated Chronic Reactive Airway Disease

Because chronic Mycoplasma pneumoniae respiratory infection is hypothesized to play a role in asthma, the potential of M. pneumoniae to establish chronic respiratory infection with associated pulmonary disease was investigated in a murine model. BALB/c mice were intranasally inoculated once with M. pneumoniae and examined at 109, 150, 245, 368, and 530 days postinoculation. M. pneumoniae was detected in bronchoalveolar lavage fluid by culture or PCR in 70 and 22% of mice at 109 and 530 days postinoculation, respectively. Lung histopathology was normal up to 368 days postinoculation. At 530 days, however, 78% of the mice inoculated with M. pneumoniae demonstrated abnormal histopathology characterized by peribronchial and perivascular mononuclear infiltrates. A mean histopathologic score (HPS) at 530 days of 5.1 was significantly greater (P < 0.01) than that for controls (HPS score of 0). Serum anti-M. pneumoniae immunoglobulin G was detectable in all of the mice inoculated with M. pneumoniae and was inversely correlated with HPS (r = -0.95, P = 0.01) at 530 days postinoculation. Unrestrained whole-body plethysmography measurement of enhanced pause revealed significantly elevated airway methacholine reactivity in M. pneumoniae-inoculated mice compared with that in controls at 245 days (P = 0.03) and increased airway obstruction at 530 days (P = 0.01). Murine M. pneumoniae respiratory infection can lead to chronic pulmonary disease characterized by airway hyperreactivity, airway obstruction, and histologic inflammation.     

A novel autosomal recessive non-syndromic deafness locus (DFNB35) maps to 14q24.1-14q24.3 in large consanguineous kindred from Pakistan

Autosomal recessive nonsyndromic deafness is one of the most frequent forms of inherited hearing impairment. Over 30 autosomal recessive nonsyndromic hearing loss loci have been mapped, and 15 genes have been isolated. Of the over 30 reported autosomal recessive nonsyndromic hearing loss (NSHL) loci, the typical phenotype is prelingual non-progressive severe to profound hearing loss with the exception of DFNB8, which displays postlingual onset and DFNB13, which is progressive. In this report we describe a large inbred kindred from a remote area of Pakistan, comprising six generations and segregating autosomal recessive nonsyndromic prelingual deafness. DNA samples from 24 individuals were used for genome wide screen and fine mapping. Linkage analysis indicates that in this family the NSHL locus, (DFNB35) maps to a 17.54 cM region on chromosome 14 flanked by markers D14S57 and D14S59. Examination of haplotypes reveals a region that is homozygous for 11.75 cM spanning between markers D14S588 and D14S59. A maximum two-point LOD score of 5.3 and multipoint LOD score of 7.6 was obtained at marker D14S53. The interval for DFNB35 does not overlap with the regions for DFNA9, DFNA23 or DFNB5.     

Improving the teaching skills of residents as tutors/facilitators and addressing the shortage of faculty facilitators for PBL modules

Background  

Residents play an important role in teaching of medical undergraduate students. Despite their importance in teaching undergraduates they are not involved in any formal training in teaching and leadership skills. We aimed to compare the teaching skills of residents with faculty in facilitating small group Problem Based Learning (PBL) sessions.     

DFNB39, a recessive form of sensorineural hearing impairment, maps to chromosome 7q11.22-q21.12

This article describes the identification of a novel locus (DFNB39) responsible for an autosomal recessive form of hearing loss segregating in a Pakistani consanguineous family. The hearing impaired members of this family present with profound prelingual sensorineural hearing impairment and use sign language for communications. Linkage was established to microsatellite markers located on chromosome 7q with a maximum multipoint lod score of 3.8. The region of homozygosity spans a 19 cM region that is bounded by markers D7S3046 and D7S644.     

Effect of clarithromycin on cytokines and chemokines in children with an acute exacerbation of recurrent wheezing: a double-blind, randomized, placebo-controlled trial.

BACKGROUND: Clarithromycin is postulated to possess immunomodulatory properties in addition to its antimicrobial activity. OBJECTIVE: To evaluate the effect of clarithromycin on serum and nasopharyngeal cytokine and chemokine concentrations in children with an acute exacerbation of recurrent wheezing. METHODS: Children with a history of recurrent wheezing or asthma and who presented with an acute exacerbation of wheezing were enrolled in a double-blind, randomized trial of clarithromycin vs placebo. Concentrations of tumor necrosis factor alpha (TNF-alpha), interferon-gamma (IFN-gamma), interleukin-1beta (IL-1beta), IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, granulocyte-macrophage colony-stimulating factor, RANTES, eotaxin, macrophage inflammatory protein 1alpha, macrophage inflammatory protein 1beta, and monocyte chemoattractant protein 1 were measured in serum and/or nasopharyngeal aspirates before, during, and after therapy. Mycoplasma pneumoniae and Chlamydophila pneumoniae infection were evaluated for by polymerase chain reaction and serologic testing. RESULTS: Nasopharyngeal concentrations of TNF-alpha, IL-1beta, and IL-10 were significantly and persistently lower in children treated with clarithromycin compared with placebo. There tended to be a greater effect of clarithromycin on nasopharyngeal cytokine concentrations in patients with evidence of M. pneumoniae or C. pneumoniae infection. No significant differences were detected in serum cytokines for children treated with clarithromycin compared with placebo. CONCLUSION: Clarithromycin therapy reduces mucosal TNF-alpha, IL-1beta, and IL-10 concentrations in children with an acute exacerbation of recurrent wheezing.     

Novel sequence variants in the TMC1 gene in Pakistani families with autosomal recessive hearing impairment

Though many hearing impairment genes have been identified, only a few of these genes have been screened in population studies. For this study, 168 Pakistani families with autosomal recessive hearing impairment not due to mutations in the GJB2 (Cx26) gene underwent a genome scan. Two-point and multipoint parametric linkage analyses were carried out. Twelve families had two-point or multipoint LOD scores of 1.4 or greater within the transmembrane cochlear expressed gene 1 (TMC1) region and were subjected to further screening with direct DNA sequencing. Five novel putatively functional non-synonymous sequence variants, c.830A>G (p.Y277C), c.1114G>A (p.V372M), c.1334G>A (p.R445H), c.2004T>G (p.S668R), and c.2035G>A (p.E679K), were found to segregate within seven families, but were not observed in 234 Pakistani control chromosomes. The variants c.830A>G (p.Y277C), c.1114G>A (p.V372M), and c.1334G>A (p.R445H) occurred at highly conserved regions and were predicted to lie within hydrophobic transmembrane domains, while non-synonymous variants c.2004T>G (p.S668R) and c.2035G>A (p.E679K) occurred in extracellular regions that were not highly conserved. There is evidence that the c.2004T>G (p.S668R) variant may have occurred at a phosphorylation site. One family has the known splice site mutation c.536 -8T>A. The prevalence of non-syndromic hearing impairment due to TMC1 in this Pakistani population is 4.4% (95%CI: 1.9, 8.6%). The TMC1 protein might have an important function in K(+) channels of inner hair cells, which would be consistent with the hypothetical structure of protein domains in which sequence variants were identified.     

Purification of alkaline phosphatase from extracellular vesicles of fracture callus cartilage

Summary Extracellular matrix vesicles from fracture callus cartilage were isolated by differential centrifugation and resolved by equilibrium centrifugation on a discontinuous sucrose gradient into two bands. The phosphohydrolytic activity towards p-nitrophenyl phosphate, tetrasodium pyrophosphate and adenosine triphosphate was distributed similarly after differential and equilibrium centrifugation suggesting the association of this activity with the matrix vesicles. The two bands isolated by equilibrium centrifugation of the partially purified vesicular preparation demonstrated high levels of alkaline phosphatase activity. Observed with an electron microscope, the 1.07–1.14 g/cm³ band from the gradient was enriched in electron luscent matrix vesicles while the 1.27 g/cm³ band contained electron dense matrix vesicles. Enzymatic analysis of the 1.27 g/cm³ band indicated a slight contamination due to the presence of mitochondria and lysosomes while the 1.07–1.14 g/cm³ band gave no enzymatic indication of subcellular contamination. A phosphohydrolytic enzyme active towards p-nitrophenyl phosphate, tetrasodium pyrophosphate and adenosine triphosphate was purified from the 1.07–1.14 g/cm³ fraction by DEAE-cellulose column chromatography. Electron micrographs of callus cartilage sections demonstrated densification of the plasma membrane and matrix vesicles following substrate incubation with-glycerophosphate or tetrasodium pyrophosphate. The histochemical and biochemical data indicate that a phosphatase, with multiple substrate specificity, is a component of fracture callus cartilage matrix vesicles.     

Blood-Brain Barrier Permeability of Asiaticoside,Madecassoside and Asiatic Acid in Porcine Brain Endothelial Cell Model

Neurotherapeutic potentials of Centella asiatica and its reputation to boost memory, prevent cognitive deficits and improve brain functions are widely acknowledged. The plant's bioactive compounds, i.e. asiaticoside, madecassoside and asiatic acid were reported to have central nervous system (CNS) actions, particularly in protecting the brain against neurodegenerative disorders. Hence, it is important for these compounds to cross the blood-brain barrier (BBB) to be clinically effective therapeutics. This study aimed to explore the capability of asiaticoside, madecassoside and asiatic acid to cross the BBB using in vitro BBB model from primary porcine brain endothelial cells (PBECs). Our findings showed that asiaticoside, madecassoside and asiatic acid are highly BBB permeable with apparent permeability (P_app) of 70.61 ± 6.60, 53.31 ± 12.55 and 50.94 ± 10.91 × 10^?6 cm/s respectively. No evidence of cytotoxicity and tight junction disruption of the PBECs were observed in the presence of these compounds. Asiatic acid showed cytoprotective effect towards the PBECs against oxidative stress. This study reported for the first time that Centella asiatica compounds demonstrated high capability to cross the BBB, comparable to central nervous system drugs, and therefore warrant further development as therapeutics for the treatment of neurodegenerative diseases.