DFNA54, a third locus for low-frequency hearing loss

Nonsyndromic hereditary hearing impairment (NSHHI) is a highly heterogeneous disorder with more than 90 loci mapped, of which nearly one-half of the responsible genes are identified. In dominant NSSHI hearing loss is typically biased towards the high frequencies while low-frequency hearing loss is unusual. Only two NSHHI loci, DFNA1 and DFNA6/14/38, are associated with predominantly low- frequency loss. We mapped the loci harboring the gene responsible for autosomal dominant low-frequency hearing loss in a multigenerational family. The pedigree of a Swiss family with low-frequency hearing loss was established. Using genomic DNA, DFNA1 and DFNA6/14/38 were excluded by linkage analysis or by direct sequencing of the responsible gene. Genome-wide linkage analysis was performed using commercially available microsatellite markers. Two-point linkage analysis demonstrated linkage to chromosome 5q31, the locus for DFNA15, with a lod score of 6.32 at recombination fraction =0 for marker D5S436. Critical recombinations were seen at markers D5S1972 and D5S410. Sequencing of the corresponding gene POU4F3 yielded no pathogenic mutation segregating with the affected members. In addition to Wolfram syndrome gene 1 (DFNA6/14/38) and diaphanous (DFNA1) there is evidence for a third gene involved in low-frequency hearing loss located at DFNA15. Because of the differences in auditory phenotype and the absence of pathogenic mutation in the coding region of POU4F3 it is likely that there is a second gene in 5q31, designated DFNA54, associated with NSHHI.     

The lipid raft microdomain-associated protein reggie-1/flotillin-2 is expressed in human B cells and localized at the plasma membrane and centrosome in PBMCs

Reggie-1/flotillin-2 is a plasma membrane-associated cytoplasmic protein, which defines non-caveolar raft microdomains. Reggie-1/flotillin-2 is enriched in detergent insoluble (TX100) membrane fractions (DIG), co-localizes with activated GPI-linked proteins and the fyn-kinase in neurons and T cells, and thus apparently participates in the assembly of protein complexes essential for signal transduction. In T cells activated by crosslinking the GPI-linked protein Thy-1 or by crosslinking the ganglioside GM1, reggie-1/flotillin-2 co-localizes with the T cell receptor. To determine whether reggie-1/flotillin-2 is also expressed in B cells, primary B cells from human blood and cell lines representing the developmental stages of pro, pre, mature and plasma B cells were analyzed by Western blotting, RT-PCR and immunofluorescence. Here, we show that reggie-1/flotillin-2 is expressed throughout B cell development, as well as in primary B cells, purified by cell sorting. On non-activated mature B cell Raji cell line we found reggie-1/flotillin-2 are exclusively in the detergent (TX100) insoluble membrane fractions that are staining positive for the raft marker GM1. Immunofluorescence microscopy showed that reggie-1/flotillin-2 is localized at the plasma membrane and marks intracellular spots in PBMCs. Confocal co-localization studies showed that reggie-1/flotillin-2 is associated with the plasma membrane, and the centrosomes (microtubule organizing centers) in these PBMCs. Comparison of reggie-1/flotillin-2 cDNA sequences with the genomic sequence database allowed us to determine the exon/intron structures in mouse and human. The gene organizations are highly conserved suggesting an important function of reggie-1/flotillin-2. Since reggie/flotillin proteins co-cluster with the T cell receptor and fyn kinases upon T cell stimulation, our findings of reggie-1/flotillin-2 in B cells suggest a similar role in B cell function.     

Ultraviolet light-induced modification of crosslinked hyaluronan gels

Hyaluronan (HA) gels (hylans) crosslinked with divinyl sulfone (DVS) are highly biocompatible and can be structurally modified to obtain desired mechanical properties that are attractive for their use as tissue-engineering scaffolds. However, unmodified hylan gels are not good substrates for cell attachment or infiltration, likely as a result of their smooth surface and the highly anionic nature of HA. This study investigated whether the cell-adhering characteristics of hylan gels could be enhanced by irradiation with ultraviolet (UV) light, with or without prior dehydration. The attachment and proliferation of neonatal rat smooth muscle cells atop these gels was compared with that on unmodified (control; C) or dehydrated (D) gels. UV-induced changes to gel structure and chemistry were characterized by confocal and electron microscopy, and fluorphore-assisted carbohydrate electrophoresis (FACE). Cell attachment was sparse on both unmodified (C) and dehydrated (D) gels. Significantly higher levels of cell attachment were observed on the surface of irradiated (UV) and dehydrated-irradiated (DUV) gels, likely because of texturing of the gel surface by UV light. In addition, dehydration of gels before UV irradiation created irregular pore-like structures through which cells appeared to migrate into the interior. FACE assays demonstrated that UV-irradiation alters the chemistry of HA, causing limited breakdown of HA chains and DVS crosslinks within gel and possibly creating new crosslinks that have not yet been identified. Because the hylan gels are altered structurally and chemically, binding of cells to the material is likely to be more permanent than possible by other approaches, such as coating of cell-adhesive matrix factors on the gel surface, described previously. The significance of this work is that we have developed a technique for the modification of DVS-crosslinked HA (hylans) to enhance their performance as a cellular scaffold for tissue-engineering applications.     

Risk factors for HIV infection in injection drug users and evidence for onward transmission of HIV to their sexual partners in Chennai, India

OBJECTIVES: Determining HIV prevalence in injection drug users (IDUs) and their regular sex partners in Chennai, India. METHODS: A total of 226 IDUs and their regular sex partners were enrolled during April-July 2003. After informed consent was obtained, a semistructured questionnaire was administered and serum was tested for HIV antibody. RESULTS: The HIV seroprevalence was 30% (68/226) in IDUs and 5% in their regular sex partners (11/226). While in 25% of couples only the male partner was HIV positive, 5% of the couples were concordant for HIV infection and 70% were HIV negative. Fifty-seven percent of the HIV-positive IDUs and 45% of the HIV-infected women thought that they had "no chance" or "very little chance" of getting HIV, reflecting low HIV risk perception. More than 20% IDUs reported borrowing or lending of injection equipment. In univariate analyses "sex" and "condom use" with sex workers had no bearing but "more than twice a day injecting frequency," "history of incarceration," "tattoos," "recruitment from northern part of the city," and ever-injecting drugs in drug-selling places had significant association with HIV infection in IDUs. In an adjusted model, the odds of HIV infection were 2 times higher among IDUs who had ever injected drugs in drug-selling places and 6 times higher in those who were recruited from the northern part of central Chennai. CONCLUSION: Reducing sharing of injection equipment and unsafe tattooing through targeted and environmental interventions, increasing HIV risk perception, and promoting safer sex practices among IDUs and their sex partners are urgent program needs.     

DADOS-Survey: an open-source application for CHERRIES-compliant Web surveys

Background  

The Internet has been increasingly utilized in biomedical research. From online searching for literature to data sharing, the Internet has emerged as a primary means of research for many physicians and scientists. As a result, Web-based surveys have been employed as an alternative to traditional, paper-based surveys. We describe DADOS-Survey, an open-source Web-survey application developed at our institution that, to the best of our knowledge, is the first to be compliant with the Checklist for Reporting Results of Internet E-Surveys (CHERRIES). DADOS-Survey was designed with usability as a priority, allowing investigators to design and execute their own studies with minimal technical difficulties in doing so.     

Infection and transformation of Fv-2rr erythroprogenitor cells with Friend virus

Friend virus was used to infect and transform Fv-2rr erythroprogenitor cells in vivo. The RB (Fv-2rr) cell line was characteristic of Friend virus-induced cell lines in Fv-2ss mice, i.e., it produced infectious Friend virus and synthesized hemoglogin. The RB (Fv-2rr) cell line expressed the envelope protein of the spleen focus-forming virus (gp52) and a novel, related envelope protein (gp48). The results demonstrate that Fv-2rr erythroprogenitor cells can be infected and transformed in vivo.     

Dengue disease spectrum among infants in the 2001 dengue epidemic in Chennai,Tamil Nadu,India

An investigation in a referral pediatric hospital has indicated that during a recent dengue outbreak in Chennai, Tamil Nadu, India, dengue in infancy constituted 20% of total dengue virus infections with low mortality rates in this hospital. In developing countries, strengthening of dengue management capabilities at hospitals can prevent dengue-related deaths in infants.     

Tumoural cysticercosis

The unusual occurrence of very large cysticerci presenting as tumour-like lesions prompted this search of the pathology records of a south Indian hospital, to find similar cases. The maximum diameter of 187 consecutively diagnosed extraneural cysticerci, was recorded and those cases exceeding the mean by two standard deviations reviewed. Six biopsies showing cysticerci with a maximum diameter ranging from 4 to 11 cm, were identified, and hospital records and all pathological material available, were reexamined. The clinical differential diagnosis included benign and malignant tumours, hydatid cyst and haematoma. All the cysts were confirmed to be cysticerci, but species diagnosis was not possible by examining archival material. In an appropriate geographical setting, large cysticerci should form part of the differential diagnosis of cystic tumour-like lesions. An attemp can then be made, while the specimen is still fresh, to identify the species.     

Distribution of galanin immunoreactivity in the central nervous system and the responses of galanin-containing neuronal pathways to injury

Radioimmunoassay and immunocytochemistry were used to study the distribution of galanin, a novel 29 amino acid porcine intestinal peptide, in the central nervous system of the rat and pig. The pattern of distribution was similar in the two species, with the highest concentrations of galanin-like immunoreactivity found in the neurohypophysis, hypothalamus and sacral spinal cord. Immunocytochemical studies of these regions localized galanin-like immunoreactivity to cell bodies in the paraventricular and supraoptic nuclei of the hypothalamus, to fibres in the pars nervosa and to numerous cell bodies and fibres in the dorsal horn of the spinal cord. On both gel and high pressure liquid chromatography, galanin-like immunoreactivity in rat and pig nervous tissue eluted as a single peak in a position similar to purified procine intestinal galanin standard. Surgical and pharmacological manipulations in the rat suggest the presence of galanin in afferent fibres. An increase of galanin-like immunoreactivity was observed in the sacral spinal cord of the rat following thoracic spinal cord transection. Thus galanin-like immunoreactivity in the brain is mainly localized in the hypothalamopituitary region. The decrease of galanin-like immunoreactivity in the dorsal horn of the spinal cord, following dorsal rhizotomy and pre-treatment of rats with capsaicin, indicates that many of the fibres, which are of small diameter, may well be derived from spinal sensory neurones.