Biomedical subjects

A Bosch

Publications and source records attributed to A Bosch.

At least 19 recordsLinked to original sources

PubMed1999

Group D adenoviruses infect primary central nervous system cells more efficiently than those from group C.

Group C adenovirus-mediated gene transfer to central nervous system cells is inefficient. We found that wild-type group D viruses, or recombinant adenovirus type 2 (Ad2) (group C) modified to contain Ad17 (group D) fiber, were more efficient in infecting primary cultures of neurons. Together with studies on primary vascular endothelial cells and tissue culture cell lines, our results indicate that there is not a universally applicable adenovirus serotype for use as a gene transfer vector.

Adenoviruses, Human↗

PubMed1998

Enhancement of the immunogenicity of a synthetic peptide bearing a VP3 epitope of hepatitis A virus.

The immune responses elicited in mice by different forms of the VP3(110-121) B-epitope of the hepatitis A virus (HAV) were studied. Different forms of incorporation in liposomes were tested, encapsulation, rather than surface exposure, being the best antigenic preparation. Three larger peptides of the VP3 epitope, two of them containing a hepatitis B virus T-epitope, and a third containing a putative T-epitope of HAV (VP3(102-121)) were assayed. While this latter T-epitope induced an enhancement of the response against the VP3 B-epitope, the artificially coupled T-epitopes failed to induce a significant increase. The administration of two multiple antigenic peptide (MAP) constructs, the first containing the VP3(110-121) and VP1(11-25) HAV sequences and the second only the VP1(11-25) sequence, also suggested the presence of a T-epitope, since the response against the VP1 peptide was increased in the first construct.

Animals↗

PubMed1998

Effects of keratinocyte and hepatocyte growth factor in vivo: implications for retrovirus-mediated gene transfer to liver.

We have previously shown that intravenous administration of keratinocyte growth factor (KGF) induces hepatocyte proliferation, allowing for efficient and noninvasive in vivo gene transfer with high-titer retroviral vectors in mice. The distinctive periportal distribution of transduced cells led us to investigate the ability of virus-sized particles to perfuse the liver adequately after growth factor treatment. We found that perfusion was adequate, and that transduction was limited to the periportal region because only those cells were stimulated to divide. Cells in this region also showed increased expression of Ram-1, the receptor for the murine Moloney leukemia virus (MoMLV) amphotropic envelope, after KGF treatment. In further studies we found that recombinant hepatocyte growth factor (HGF) induces a different population of hepatocytes to divide and upregulate Ram-1. The differential pattern of induction suggested that combining KGF and HGF would improve gene transfer efficiency further. Indeed, simultaneous delivery of both growth factors leads to an overall increase in the number of proliferating cells. Importantly, when coupled with MoMLV delivery, efficiency of gene transfer increased. These results confirm the utility of growth factors for noninvasive hepatic gene transfer in mice, and demonstrate how experiments to define the mechanism of transduction can be taken advantage of to develop improved gene transfer protocols.

Animals↗

PubMed1998

A new continuous epitope of hepatitis A virus.

A new continuous epitope of hepatitis A virus (HAV) was defined in the VP3 protein. Convalescent sera recognised the synthetic peptide 3110-3121 (FWRGDLVFDFQV). The replacement of the arginine, glycine, or aspartic acid at positions 112, 113, or 114, respectively by other amino acids induced the loss of synthetic peptide recognition by human convalescent sera, thereby confirming the presence of an epitope in the original VP3(110-121) sequence. Shorter VP3 peptides such as VP3(110-119). VP3(110-117), and VP3(110-116) and a tandem repeat of VP3(111-116) failed to react with convalescent sera, indicating the importance of the entire peptide in the epitope structure. The maximum inhibition of human convalescent binding to HAV by the VP3(110-121) peptide was around 60%, and 50% inhibition was achieved at a peptide concentration of 2.3-2.4 micrograms/ml. Antibodies generated by this peptide bound to intact HAV and neutralised its infectivity. Antipeptide antibodies inhibited convalescent serum binding to HAV. Monoclonal antibodies H7C27 and MAK-4E7 inhibited completely binding of the antipeptide antibodies to HAV.

Antibodies, Monoclonal↗

PubMed1998

Blood levels of polychlorinated dibenzodioxins, polychlorinated dibenzofurans and polychlorinated biphenyls in the general population of a Spanish Mediterranean city.

Polychlorinated dibenzodioxins (PCDDs), Polychlorinated dibenzofurans (PCDFs) and Polychlorinated biphenyls (PCBs) are among the most toxic environmental pollutants. We determined blood levels of these compounds in a population sample of the city of Mataró, Spain. Blood samples were drawn from a randomly selected sample of 198 subjects, of both genders, aged 18 to 69 years. These samples were pooled into 10 groups for laboratory analysis. For males, total level of PCDDs was 505.7 ppt, of PCDFs was 26.7 ppt, and the international toxic equivalent (I-TEQ) was 12.5. For females the levels were 739 ppt, 28.8 ppt and 14.71-TEQ, respectively. The most important contributors to the total I-TEQ were HexaCDD, PentaCDD and PentaCDF. The blood concentration of total PCBs was 2.02 mg/l in males, and 1.58 mg/l in females. Levels of PCDDs, PCDFs and PCBs increased by age in both sexes. The levels of PCDDs and PCDFs in residents of this Mediterranean city are among the lowest observed in industrialized countries.

Adolescent↗

PubMed1998

Flow cytometry detection of infectious rotaviruses in environmental and clinical samples.

A method for the detection of infectious human rotaviruses based on infection of CaCo-2 cells and detection of infected cells by indirect immunofluorescence and flow cytometry (IIF-FC) has been developed. The technique was validated by performing a seminested reverse transcription-PCR assay with sorted cell populations. The efficiency of the procedure has been compared with that of the standard method of infection of MA104 cells and ulterior detection by IIF and optical microscopy (IIF-OM) and with that of infection of MA104 cells and detection by IIF-FC. The limit of sensitivity for the detection of the cell-adapted strain Ito(r) P13, expressed as the most probable number of cytopathogenic units, was established as 200 and 2 for MA104 and CaCo-2 cells, respectively, by the IIF-FC method. The ratio of infectious virus particles to total virus particles for a wild-type rotavirus was determined to be 1/2 x 10(6) and 1/2 x 10(4) for IIF-OM with MA104 cells and IIF-FC with CaCo-2 cells, respectively. The use of IIF-FC with CaCo-2 cells was tested with fecal and water samples and proved to be more effective than the standard procedure for rotavirus detection.

Caco-2 Cells↗

PubMed1998

Interleukin-10 modulates the severity of hypersensitivity pneumonitis in mice.

Hypersensitivity pneumonitis (HP) is an inflammatory lung disease characterized by granuloma formation. We recently showed that interferon-gamma (IFN-gamma) is essential for inflammation and granuloma formation in HP. Interleukin-10 (IL-10) counteracts many of the biologic effects of IFN-gamma, suggesting that IL-10 modulates inflammation and granuloma formation in HP. We compared the expression of HP in C57BL/6 mice that lack IL-10 (IL-10 knockout [KO]) with that in wild-type (WT) littermates. IL-10 KO and WT mice were exposed to the thermophilic bacteria Saccharopolyspora rectivirgula or to saline alone for 3 wk. The IL-10 KO mice had higher cell counts in their bronchoalveolar lavage fluid (2.85 +/- 0. 43 x 10(6)) than did WT mice (1.4 +/- 0.3 x 10(6)/ml; P < 0.03), with a more prominent neutrophil response. They also had greater inflammation after antigen exposure than did the WT mice (P < 0. 0001). There was increased upregulation of IFN-gamma, IL-1, and tumor necrosis factor-alpha (TNF-alpha) mRNAs in the lungs of IL-10 KO mice. Adenovirus-mediated gene transfer of IL-10 to the liver of IL-10 KO mice reduced the inflammation from that seen in WT mice. These studies show that IL-10 has important anti-inflammatory properties in HP, and that lack of this cytokine leads to a more severe granulomatous inflammatory response.

Adenoviridae↗

PubMed1997

Disinfection of human enteric viruses on fomites.

The virucidal action of several commercially available disinfectant preparations was assayed against hepatitis A virus and human rotavirus dried on polystyrene. Overall, the level of virus disinfection achieved was very poor, usually inducing less than 3 log titre reduction. Suspension tests performed with the same disinfectants showed different virus inactivation rates, thus failing to provide a reliable indication of the actual virus disinfection on fomites. In our studies, bacteriophages of Bacteroides fragilis proved to be a simple, cheap and reliable screening tool for the evaluation of virus disinfection on non-porous surfaces. The same conclusion cannot be drawn for poliovirus.

Bacteroides fragilis↗

PubMed1997

Astrovirus survival in drinking water.

A method based on infection of CaCo-2 cultured cell monolayers (CC) and reverse transcription-PCR (RT-PCR) was developed for the specific detection of infectious astrovirus. The procedure was validated by titrating poliovirus stocks in parallel in CaCo-2 cells by determining the most probable number of cytopathogenic units and by cell culture and subsequent RT-PCR (CC-RT-PCR). CC-RT-PCR was then employed to measure the persistence of astrovirus suspended in dechlorinated tap water. After 60 days, the decay of astrovirus infectivity was 2 log units at 4 +/- 1 degrees C and 3.2 log units at 20 +/- 1 degrees C, while after 90 days, the titer reduction was 3.3 and 5 log units at 4 +/- 1 degrees C and 20 +/- 1 degrees C, respectively. Astrovirus decay in the presence of free chlorine (FC) was monitored by CC-RT-PCR. Residual infectivity was found after 2 h in the presence of 1 mg of FC/liter. Under these conditions, astrovirus shows a log titer reduction (LTR) or 4, while 0.5 mg of FC/liter induced an LTR of 2.4. The possibility of acquiring data on the survival of fastidious viruses in the environment opens new perspectives on the epidemiology of some significant infections transmitted by the fecal-oral route.

Astroviridae Infections↗

PubMed1996

Proliferation induced by keratinocyte growth factor enhances in vivo retroviral-mediated gene transfer to mouse hepatocytes.

Retroviral gene transfer to liver without prior injury has not yet been accomplished. We hypothesized that recombinant human keratinocyte growth factor would stimulate proliferation of hepatocytes and allow for efficient in vivo gene transfer with high titer murine Moloney retroviral vectors. This report shows that 48 h after intravenous injection of keratinocyte growth factor, hepatocyte proliferation increased approximately 40-fold compared to non-stimulated livers. When keratinocyte growth factor treatment was followed by intravenous injection of high titer (1 x 10(8) colony forming units/ml) retrovirus coding for the Escherichia Coli beta-galactosidase gene, there was a 600-fold increase in beta-galactosidase expression, with 2% of hepatocytes transduced. Thus, by exploiting the mitogenic properties of keratinocyte growth factor, retrovirus-mediated gene transfer to liver may be accomplished in vivo without the use of partial hepatectomy or pretreatment with other toxins to induce hepatocyte cell division.

Animals↗

PubMed1996

Hypoxanthine-guanine phosphoribosyltransferase (HPRT) expression in the central nervous system of HPRT-deficient mice following adenoviral-mediated gene transfer.

In this study we show that recombinant adenovirus can augment hypoxanthine-guanine phosphoribosyltransferase (HPRT) levels in the central nervous system (CNS) of HPRT-deficient mice. Recombinant adenovirus containing the cDNA for rat HPRT (rHPRT) expressed from the Rous sarcoma virus LTR (RSV LTR) was constructed (AdRSVrHPRT). AdRSVrHPRT was injected into the right caudate nucleus of 7-week-old HPRT-deficient mice. Brains were analyzed for gene transfer, transgene expression and function by DNA PCR, in situ RNA hybridization, and enzyme bioactivity. The results show that rHPRT cDNA delivered by an adenoviral vector can augment HPRT levels in brain tissue and documents the utility of gene transfer to restore HPRT activity in an HPRT-deficient CNS.

Adenoviridae↗

PubMed1996

Non isotopic automatable molecular procedures for the detection of enteroviruses.

Five microwell non isotopic hybridization assays, based on colorimetric immunoenzymatic reading, were developed and evaluated for the rapid and automatable detection of enteroviruses. Virus nucleic acids and/or capture probes were covalently bound to microtiter wells, and digoxigenin-11-dUTP was used as label for the detection of hybridized material. Among these procedures, a reverse transcriptase polymerase chain reaction (RT-PCR) hybridization assay was the most sensitive, enabling the detection of 10 MPNCU of poliovirus, and offering detection specificity for other enteroviruses, such as coxsackieviruses and echoviruses. The second most sensitive method was a complementary hybridization assay, simultaneously using three detection probes, one from the 5' end and two from the 3' end of poliovirus genome, offering a sensitivity for poliovirus detection of 5 x 10(3) MPNCU.

Automation↗

PubMed1996

Late presentation of congenital muscular torticollis: a non-dystonic cause of torticollis.

We describe three patients with a late presentation of congenital muscular torticollis to emphasize that this non-dystonic type of torticollis has to be considered in the differential diagnosis of cervical dystonia, even in adult patients. Surgery is the treatment of choice. Postoperatively, two of our patients had a transient lesion of the accessory nerve. We assume that in our patients the accessory nerve was embedded in the fibrous tissue around the affected muscle. To prevent damage to the accessory nerve, the incision has to be made distally at about 1 cm above the clavicle.

Adolescent↗

PubMed1996

Integration of 30 CA-repeat markers into the cytogenetic, genetic and YAC maps of human chromosome 21.

The number of polymorphic DNA markers developed for the whole human genome during the last 2 years has been vastly increased. For this reason, the genetic map is continuously improving, but the cytogenetic and physical maps are not progressing at the same speed. Therefore, there is a need to integrate genetic, cytogenetic and physical mapping data. We have developed and localized on the breakpoint map of human chromosome 21 thirty microsatellite markers. Twenty of them have been used in the construction of a genetic map of chromosome 21, which contains a total of 44 markers. This map has 39 uniquely placed loci at 23 anchor points, ordered with odds of at least 1,000:1. The sex average length of the map is 64.4 cM, with the male and female lengths being 49.4 and 79.2 cM, respectively. Twenty-six of these newly developed markers have been localised on the CEPH/Généthon and Joint YAC Screening Effort YACs. Although these microsatellites were found uniformly spread along chromosome 21, the detection of various markers in the same or adjacent YACs suggests that CA-repeat microsatellites are clustered in several regions. The localization of these markers on the cytogenetic, genetic and YAC maps has provided a refined location for them and is a step further towards the construction of an integrated map of HC21.

Chromosome Mapping↗

PubMed1996

[Organizational models for the hematology areas of the district hospitals of Catalonia].

PURPOSE: The haematological assistance in Catalonia is based upon the district hospitals, in the first step, and the stage III hospitals located usually in higher population nuclei, in the second step. The purpose of this work was to analyse the resources of the "primary haematological assistance" network provided by the district hospitals, to evaluate them and to propose a model for their organisation. MATERIAL AND METHODS: An enquiry was carried out to all members of the Grup de Treball d'Hospitals Comarcals de Catalunya (Catalonia's District Hospitals Task Force). The evaluable data included demographic figures of the population assisted, personnel of each haematological area, organising structure, clinical activity, cytomorphology, blood banks, laboratories and continuous formation activities. RESULTS: The enquiry was answered by 15 of the 21 district hospitals (71.4%) with haematologists in Catalonia. The population assisted in those hospitals is 2,100,000 (ranging between 55,000 an 450,000). All centres are integrated in the National Health network. Eleven of the hospitals analysed have only one haematologist (73.3%). If his dedication is 100% of the time, this would represent a doctor for 105,000 people. The time devoted to work is 690 hours a week for all the population, with a mean of 3,043. Four patients assisted per hour. The total number of hospital beds is 3,353 (50-450), with a mean number of 1 haematologist for every 167.6 beds. The number of patients hospitalized due to blood diseases ranges between 3 and 13 per month. Six of the 15 centres are adjunct to the outpatient clinic. Two centres have a blood bank and 7 have developed an autotransfusion programme. All the centres but one perform oral anticoagulant treatment follow-up, the number of patients assisted ranging from 20 to 210 per week. None of the hospitals has a separate Haematology Service; in most of them haematology is structurally and functionally dependent from Laboratories and in some there is a mixed Laboratory/Internal Medicine functional organisation, depending of the Medical Direction. No haematologist is ever on call specifically for his specialty. Continuous formation activities are carried out in 9 of the 15 centres (60%). COMMENT: Several measures are proposed to improve the haematological assistance, acting on different levels: continuous formation, patient flows and circuits, resident doctors training, anticoagulant treatment network, organisation models, credit cards from the Spanish Association of Haematology.

Adult↗

PubMed1995

Simultaneous on-line DNA sequencing on both strands with two fluorescent dyes.

We describe an automated DNA-sequencing technique which allows both the simultaneous sequencing from the two strands of double-stranded templates and the subsequent detection of the sequencing products online and in parallel. The technique is based on hardware technology also used in the ALF DNA sequencer (Pharmacia, Uppsala). A helium-neon laser was mounted into the sequencing device additionally to the standard argon laser. Two different primers, labeled with either fluorescein or Texas red, are used in a single sequencing reaction resulting in an output of two sequences. Both sequencing products are then analyzed on-line in the same lanes of a gel. This technique is especially useful for the complete sequencing of DNA fragments up to 1 kb. High accuracy sequencing of PCR products in double-stranded form can now be accomplished in only one sequencing reaction.

Fluorescent Dyes↗

PubMed1995

Characterisation of three microsatellite polymorphisms (D21S1262, D21S1419 and D21S1421) from band 21q22.1.

We have isolated three clones, containing highly polymorphic CA-repeat sequences, from a human chromosome 21 phage library (LA21NS01). These clones have been localised to band q22.1 by using a chromosome 21 somatic cell hybrid panel. D21S1262 is located between breakpoints 6918-8a1 and 3x2S, and D21S1419 and D21S1421 are localised between breakpoints JC6-A and MRC2G. Their observed heterozygosities range between 0.75 and 0.85 as shown by unrelated reference parents from the Centre d'Etude du Polymorphisme Humain. These highly polymorphic markers should be useful for improving the analysis of this region of chromosome 21, which contains important genes such as SOD1, GART and IFNAR.

Base Sequence↗

PubMed1995

Five new microsatellite polymorphisms at the q21 region of human chromosome 21.

Five clones, containing polymorphic CA-repeat sequences, have been isolated from a specific human chromosome 21 phage library and have been localised to band q21 of chromosome 21 using a somatic cell hybrid panel. These highly repetitive sequences (D21S1263, D21S1264, D21S1415, D21S1417 and D21S1420) have been characterised in the CEPH reference parents and have heterozygosities ranging from 0.30 to 0.81 and an average polymorphism information content (PIC) of 0.62. The relative order of these markers, based on the somatic cell hybrid panel, is cen-D21S1417, D21S1420-D21S1263, D21S1415-D21S1264-tel. The most polymorphic marker (D21S1264) has been included in the chromosome 21 genetic map. They have also been localised in the CEPH/Généthon YAC panel, providing a refined localisation of these polymorphic sequences. These five CA-repeat markers should provide a better characterisation of the q21 region of chromosome 21.

Animals↗