Use of Computed Tomography Scans for Cochlear Implants

While 3-dimensional (3D) imaging by computed tomography has long been desirable for research and treatment of cochlear-implant patients, technical challenges have limited its wide application. Recent developments in scanner hardware and image processing techniques now allow image quality improvements that make clinical applications feasible. Validation experiments were performed to characterize a new methodology and its imaging performance.     

The role of the microtubules in tumor necrosis factor-alpha-induced endothelial cell permeability

Tumor necrosis factor (TNF)-alpha, a major proinflammatory cytokine, triggers endothelial cell activation and barrier dysfunction which are implicated in the pathogenesis of pulmonary edema associated with acute lung injury syndromes. The mechanisms of TNF-alpha-induced vascular permeability are not completely understood. Our initial experiments demonstrated that TNF-alpha-induced decreases in transendothelial electrical resistance across human pulmonary artery endothelial cells are independent of myosin light chain phosphorylation catalyzed by either myosin light chain kinase or Rho kinase. We next assessed the involvement of another cytoskeletal component, the tubulin-based microtubule network, and found TNF-alpha to induce a decrease in stable tubulin content and partial dissolution of peripheral microtubule network as evidenced by anti-acetylated tubulin and anti-beta-tubulin immunofluorescent staining, respectively. Microtubule-stabilizing agents, paclitaxel and epothilone B, significantly attenuated TNF-alpha-induced decreases in transendothelial electrical resistance, inhibited the cytokine-induced increases in actin stress fibers, formation of intercellular gap, and restored the TNF-alpha-compromised vascular endothelial (VE)-cadherin-based cell-cell junctions. Importantly, neither TNF-alpha nor paclitaxel treatment was associated with endothelial cell apoptosis. Inhibition of p38 mitogen-activated protein kinase by SB203580 significantly attenuated TNF-alpha-induced microtubule destabilization, actin rearrangement, and endothelial barrier dysfunction. These results strongly suggest the involvement of microtubule rearrangement in TNF-alpha-induced endothelial cell permeability via p38 mitogen-activated protein kinase activation.     

The DQA1*0104 allele is carried by DRB1*1001- and DRB1*1401-positive haplotypes in Caucasians, Africans and Orientals

Abstract: The DQA1*0104 allele is known to differ from DQA1*0101 by a single nucleotide in the sequenced part of the first exon. DQA1*0104 has a guanine in the second position of the second expressed codon, whereas DQA1*0101 and all other sequenced DQA1 alleles have an adenine in that position, changing aspartic acid to glycine. The DQA1*0104 allele was originally described in African Americans with the DRB1*12, DRB3*0101, DQA1*0104, DQB1*0501, DRB1*12, DRB3*0202, DQA1*0104, DQB1*0605 or DRB1*14, DQA1*0104, DQB1*0503 haplotypes. When developing DQA1 typing by PCR amplification with sequence-specific primers (PCR-SSP), we observed that all DR10- and DR14-positive samples carried the DQA1*0104 allele, wheres all DRB1*01 -positive DNAs carried the closely related DQA1*0101 allele. In the present study, samples representing the major ethnic groups with DR-DQ haplotypes known to carry the DQA1*0104 allele or the very similar DQA1*0101 allele were investigated by Taq I RFLP analysis, PCR-SSP typing and nucleotide sequencing. The DQA1*0104 allele was found to differ from DQA1*0101 not only in the second expressed codon, but also by a productive mutation in the signal peptide. All investigated DRB1*1001 -(n = 24) and DRB1*1401 -positive (n = 25) haplotypes, defined by homozygosity or association, of Caucasian, African or Oriental origin carried the DQA1*0104 allele, whereas the DQA1*0101 allele was found on all DRB1*01 -positive (n = 32) haplotypes. These findings demonstrate that in the assignment of HLA class II alleles, polymorphism outside the second exon sometimes must be considered. The maintenance of the DQA1*0104 allele on a few distinct haplotypes indicates that the allele is old and might also be compatible with a functional difference between the DQA1*0101 and DQA1*0104 alleles.     

Expression and function of NKRP1A molecule on human monocytes and dendritic cells

In this study, we analyzed the expression and function of the lymphocyte surface lectin NKRP1A on peripheral blood monocytes (Mo) or Mo and dendritic cells (DC) derived from thymic and bone marrow precursors. De novo expression of NKRP1A and CD14 molecules was detected upon culture of CD2^? CD3^? CD14^? CD16^? CD1a^? NKRP1A^? immature thymic precursors for 7 days in the presence of granulocyte-macrophage colony-stimulating factor (GM-CSF). Under these culture conditions, by day 21, a fraction of cells had lost CD14 and acquired both CD80 (B7.1) and CD86 (B7.2) molecules. These cells displayed a DC-like morphology and were surface NKRP1A positive. CD34⁺ NKRP1A^? CD14^? precursors, isolated from bone marrow and cultured in the presence of GM-CSF, also expressed both NKRP1A and CD14: these antigens were newly expressed on about one third of cells which had lost the CD34 precursor marker. In addition, NKRP1A was constitutively present on resting CD14⁺ peripheral blood Mo. When these cells were cultured in the presence of GM-CSF, the resulting DC population retained the expression of NKRP1A and acquired CD80, while they lost the CD14 antigen. Functional analysis revealed that the engagement of NKRP1A molecule leads to a strong intracellular calcium ([Ca²⁺]_i) increase both in resting peripheral blood Mo and in vitro-derived DC. [Ca²⁺]_i increase was mainly due to extracellular calcium influx, as it was completely abrogated by the addition of EGTA. More importantly, the engagement of the NKRP1A molecule induced interleukin (IL)-1β and IL-12 production by resting Mo and DC, respectively. Altogether these data indicate that NKRP1A lectin is present at the surface of Mo and DC and may play a relevant role in the activation and function of both cell types.     

Ex vivo expansion and prophylactic infusion of CMV-pp65 peptide-specific cytotoxic T-lymphocytes following allogeneic hematopoietic stem cell transplantation.

Cytomegalovirus reactivation and infection post-allogeneic hematopoietic stem cell transplant continue to cause morbidity and mortality. Current pharmacologic therapies are limited by side effects. Adoptive transfer of ex vivo generated cytomegalovirus-specific T cells has the potential to restore immunity, prevent cytomegalovirus, and circumvent the need for pharmacologic therapies. We have generated donor-derived cytomegalovirus-specific cytotoxic T cells using dendritic cells pulsed with the HLA-A2 restricted nonapeptide NLVPMVATV (NLV) derived from the cytomegalovirus-pp65 protein. These cytotoxic T cells have been given prophylactically to 9 recipients aged 4 to 65 years on or after day 28 post-allogeneic hematopoietic stem cell transplant. Only 2 of 9 recipients received T cell depletion in vivo or in vitro. There were no immediate adverse reactions to the infusions. During 97-798 days of follow-up, 2 recipients developed cytomegalovirus reactivation; neither developed cytomegalovirus disease or required pharmacotherapy. Three recipients developed acute graft versus host disease after infusion. Two recipients died, 1 from thrombotic thrombocytopenia purpura secondary to cyclosporine, 1 from complications of graft versus host disease. A transient increase in numbers of cytomegalovirus-specific T cells demonstrated by NLV-tetramer binding was seen in 6 recipients. Prophylactic adoptive transfer of NLV-specific T cells is safe and may be effective in preventing cytomegalovirus reactivation.     

Effects of Creep and Cyclic Loading on the Mechanical Properties and Failure of Human Achilles Tendons

The Achilles tendon is one of the most frequently injured tendons in humans, and yet the mechanisms underlying its injury are not well understood. This study examines the ex vivo mechanical behavior of excised human Achilles tendons to elucidate the relationships between mechanical loading and Achilles tendon injury. Eighteen tendons underwent creep testing at constant stresses from 35 to 75 MPa. Another 25 tendons underwent sinusoidal cyclic loading at 1 Hz between a minimum stress of 10 MPa and maximum stresses of 30–80 MPa. For the creep specimens, there was no significant relationship between applied stress and time to failure, but time to failure decreased exponentially with increasing initial strain (strain when target stress is first reached) and decreasing failure strain. For the cyclically loaded specimens, secant modulus decreased and cyclic energy dissipation increased over time. Time and cycles to failure decreased exponentially with increasing applied stress, increasing initial strain (peak strain from first loading cycle), and decreasing failure strain. For both creep and cyclic loading, initial strain was the best predictor of time or cycles to failure, supporting the hypothesis that strain is the primary mechanical parameter governing tendon damage accumulation and injury. The cyclically loaded specimens failed faster than would be expected if only time-dependent damage occurred, suggesting that repetitive loading also contributes to Achilles tendon injuries. © 2003 Biomedical Engineering Society. PAC2003: 8719Rr     

Antibody response after RSV infection in children younger than 1 year of age living in a rural area of Mozambique

Serological responses have been studied in respiratory syncytial virus (RSV) infected children < 1 year of age attending the outpatient department of the Manhiça District Hospital (Mozambique). Molecular characterization of viral RNA in nasopharyngeal aspirates from the infected children indicated a high level of genetic uniformity among the infecting viruses, all of which belonged to a single genotype of RSV group A. A representative virus strain, Moz00, was isolated from one of the infants and was used, together with the group A strain A2 and the group B strain 8/60, as antigens in the quantification of infant antibody responses. In this study, 97.5% (39/40) and 96.4% (27/28) of infected children produced an antibody response against Moz00 detected by the membrane fluorescent antibody test (MFAT) and the neutralization test (NT), respectively. Seroconversion rates decreased when the A2 and 8/60 strains were used as antigen in MFAT (95.4% and 88.2%, respectively) or NT (81.8% and 54.5%, respectively), indicating that antibody responses had both group‐ and strain‐specific components. Antibodies in convalescent sera of infected children were compared with maternally derived antibodies detected in a group of children also < 1 year of age, but with no evidence of RSV infection. The convalescent sera exhibited reduced neutralizing capacity when the 8/60 strain was used as antigen (P = 0.028), suggesting that the infant antibody response lacks neutralizing capacity against strains of the heterologous virus group. Restricted cross‐reactivity and neutralizing capacity of antibodies generated by young children might be expected to induce only moderate protection in subsequent epidemics against genetically distant strains. J. Med. Virol. 69:579–587, 2003. © 2003 Wiley‐Liss, Inc.     

Frequent transmission of enterococcal strains between mechanically ventilated patients treated at an intensive care unit

The objectives of this investigation were to study the respiratory tract colonization and transmission of enterococci between 20 patients treated with mechanical ventilation at an intensive care unit (ICU), to compare genotyping with phenotyping, and to determine the antibiotic susceptibilities of the isolated enterococci. Samples were collected from the oropharynx, stomach, subglottic space, and trachea within 24 h of intubation, every third day until day 18, and thereafter every fifth day until day 33. Enterococcal isolates (n = 170) were analyzed by pulsed-field gel electrophoresis and with the PhenePlate (PhP) system. The antimicrobial susceptibilities to five agents were determined. Seventeen of the 20 subjects were colonized with enterococci in the respiratory tract; 12 were colonized in the lower respiratory tract. Genotype analyses suggested that 13 patients were involved in a transmission event, including all patients intubated more than 12 days. In conclusion, colonization of resistant enterococci in the respiratory tract of intubated patients treated at an ICU was common. Transmission of enterococci between patients occurred frequently. Prolonged intubation period seems to be a risk factor for enterococcal cross-transmission.     

Expression of Intercellular Adhesion Molecule-1 (ICAM-1) on Cultured Human Endometrial Stromal Cells and Its Role in the Interaction With Natural Killers

PROBLEM: Recent evidence emphasizes the role of natural killer cells (NKs) as potential effectors of peritoneal immune surveillance directed against the outgrowth of endometrial cells, refluxed with menstrual debris, in ectopic sites. This NK-mediated cytotoxicity toward autologous endometrial antigens seems to be significantly decreased in endometriosis patients. METHOD: We set up experiments to clarify which molecules are involved in NK-endome-trial cell interaction. In particular, we evaluated the surface expression and functional activity of intercellular adhesion molecule-1 (ICAM-1), a cell surface glycoprotein that has been identified as one of the ligands for lymphocyte function-associated antigen-1 (LFA-1), present on almost all leucocyte cell types. Immunofluorescence flow cytometry was used to assess ICAM-1 expression on resting and IL 1β-activated endometrial stromal cells in culture. Dermal fibroblasts were used as control cells. Cytotoxicity and binding assays by ⁵¹Cr release in presence and absence of a specific monoclonal antibody (mAb) against ICAM-1 were then performed in order to determine the effect of this molecule on NK-mediated cytotoxic and binding activity toward endometrial stromal cells. RESULTS: The results of this study indicated that ICAM-1 expression on endometrial stromal cells seems to be constitutively higher than on dermal fibroblasts and can be up-regulated upon exposure to IL 1β. Furthermore, a mAb against ICAM-1 strongly inhibits the binding but not the cytotoxicity of NKs toward endometrial cells. No difference in the expression of this molecule was observed throughout the cycle. CONCLUSIONS: The presence of ICAM-1 on human endometrium might relate to the action of the immunocompetent cells in human specific reproductive events.