Acute And Sub-Acute Reconstruction Of Periorbital Thermal Burns Involving The Anterior Lamella Of The Eyelid With Simultaneous Fullthickness Skin Grafting And Amniotic Membrane Grafting.

Thermal burns of the periorbital region offer a particular challenge to surgeons in terms of satisfactory operative reconstruction. Several approaches currently exist, including full-thickness skin grafting following scar excision. The authors present a novel technique for the reconstruction of periorbital thermal burns involving the anterior lamella of the eyelid with simultaneous full-thickness skin grafting and amniotic membrane grafting. A retrospective review of six patients with periorbital thermal burn injury involving the anterior lamella of the eyelid(s) who underwent operative reconstruction utilizing this technique was performed. The operative and post-surgical outcomes were evaluated. In total, sixteen graft sites were examined. All graft sites demonstrated appropriate coverage and healthy re-epithelialization over the previously necrotic and burned areas. No evidence of cicatricial changes or lagophthalmos was noted. The postoperative outcomes demonstrate excellent functionality and cosmesis. By providing acute reconstruction, the risk of ophthalmic damage and vision loss secondary to cicatricial eyelid changes may be avoided. Additionally, utilizing only a minimal amount of tissue for reconstruction preserves vital skin. This analysis demonstrates the viability of simultaneous full-thickness skin grafting and amniotic membrane grafting for the acute and sub-acute reconstruction of periorbital thermal burn injury.

Les brûlures thermiques de la région périorbitaire représentent pour les chirurgiens un défi particulier pour obtenir une reconstruction de qualité. Plusieurs approches existent actuellement, dont la greffe de peau totale après excision de la cicatrice. Les auteurs présentent une nouvelle technique de reconstruction des brûlures thermiques périorbitaires impliquant la lamelle antérieure de la paupière avec greffe de peau totale et greffe de membrane amniotique simultanées. Une revue rétrospective a été réalisée sur six patients présentant une brûlure thermique périorbitaire impliquant la lamelle antérieure de la/les paupière(s), reconstruits en utilisant cette technique. Les résultats opératoires et post-chirurgicaux ont été évalués. Au total, seize sites de greffe ont été examinés. Tous les sites de greffe ont démontré une couverture appropriée et une réépithélialisation saine sur les zones auparavant nécrotiques et brûlées. Aucun signe de remaniements cicatriciels ou de lagophtalmie n'a été noté. Les résultats postopératoires sont excellents sur le plan fonctionnel et esthétique. En assurant une reconstruction à la phase aiguë, le risque de lésions ophtalmiques et de perte de vision secondaire à des modifications de la paupière cicatricielle peut être évité. De plus, l'utilisation d'une quantité minimale de tissu pour la reconstruction préserve le capital cutané. Cette analyse démontre la viabilité de la greffe de peau totale et la greffe de membrane amniotique simultanées, pour la reconstruction aiguë et subaiguë de brûlure thermique périorbitaire.

Aerosol exposure to the angola strain of marburg virus causes lethal viral hemorrhagic Fever in cynomolgus macaques.

Cynomolgus macaques were exposed to the Angola strain of Lake Victoria Marburg virus (MARV) by aerosol to examine disease course and lethality. Macaques became febrile 4 to 7 days postexposure; the peak febrile response was delayed 1 to 2 days in animals that received a lower dose; viremia coincided with the onset of fever. All 6 macaques succumbed to the infection, with the 3 macaques in the low-dose group becoming moribund on day 9, a day later than the macaques in the high-dose group. Gross pathologic lesions included maculopapular cutaneous rash; pulmonary congestion and edema; pericardial effusion; enlarged, congested, and/or hemorrhagic lymphoid tissues; enlarged friable fatty liver; and pyloric and duodenal congestion and/or hemorrhage. Fibrinous interstitial pneumonia was the most consistent pulmonary change. Lymphocytolysis and lymphoid depletion, as confirmed by TUNEL (terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end labeling), were observed in the mediastinal lymph nodes and spleen. MARV antigen was detected in the lungs, mediastinal lymph nodes, spleen, and liver of all animals examined. In infected macaques, nuclear expression of interleukin-33 was lost in pulmonary arteriolar and mediastinal lymph node high endothelial venule endothelial cells; interleukin-33-positive fibroblastic reticular cells in the mediastinal lymph node were consistently negative for MARV antigen. These macaques exhibited a number of features similar to those of human filovirus infections; as such, this model of aerosolized MARV-Angola might be useful in developing medical countermeasures under the Animal Rule.

Aerobiology and inhalation exposure to biological select agents and toxins.

Aerosol is the most likely route of dissemination of biological select agents and toxins in a bioterrorist attack, regardless of the natural route of exposure to the agent. The use of animal models for testing preventative and therapeutic countermeasures requires knowledge of the pathogenesis of disease after inhalation exposure. Factors that relate to outcome after respiratory exposure include the inherent infectivity and virulence and/or toxicity of the agent in the host under investigation, in addition to characteristics of the aerosol particle and host that affect the delivered dose of, and host response to, the inhaled material. This introductory article discusses the emerging science of aerobiology and the unique features of respiratory tract anatomy, physiology, and immunology that are relevant to the pathogenesis of aerosolized biothreat agents.

Rapid discovery and optimization of therapeutic antibodies against emerging infectious diseases.

Using a comprehensive set of discovery and optimization tools, antibodies were produced with the ability to neutralize SARS coronavirus (SARS-CoV) infection in Vero E6 cells and in animal models. These anti-SARS antibodies were discovered using a novel DNA display method, which can identify new antibodies within days. Once neutralizing antibodies were identified, a comprehensive and effective means of converting the mouse sequences to human frameworks was accomplished using HuFR (human framework reassembly) technology. The best variant (61G4) from this screen showed a 3.5-4-fold improvement in neutralization of SARS-CoV infection in vitro. Finally, using a complete site-saturation mutagenesis methodology focused on the CDR (complementarity determining regions), a single point mutation (51E7) was identified that improved the 80% plaque reduction neutralization of the virus by greater than 8-fold. These discovery and evolution strategies can be applied to any emerging pathogen or toxin where a causative agent is known.

Grading impairment in leprosy.

The aim of the paper is to discuss the concept of 'severity grading' in relation to impairment in leprosy, and to describe the use of an impairment sum score, the Eyes, Hands, Feet (EHF) score, as an indicator of the severity and the evolution of impairment over time. The use of an impairment sum score, the EHF score, is illustrated using data on impairment at diagnosis and after a 2-year interval from MB patients released from MDT in the Western Region of Nepal. The WHO 1988 'disability' grading scale (0-2, for both eyes, hands and feet--six sites) was used as a measure of impairment. For the analysis, the WHO grades for the six sites were summed to form an EHF score (minimum 0, maximum 12). The sensitivity to change over time of the EHF score was compared with that of the 'method of maximum grades'. Using the 'method of maximum grades', 509/706 patients (72%) appeared not to have changed in impairment status, compared with only 399 (56.5%) with the EHF score. Improvement or deterioration of impairment status was missed in 113 patients (16%). In 216/706 patients (30.6%), the changes detected with the EHF score were bigger than those revealed by the method of maximum grades. The six components of the WHO impairment grading may be added up to form a EHF sum score of impairment. This score can be used to monitor changes in impairment status in individuals or in groups. It should be recorded and reported at least at diagnosis and release from treatment. Reporting could be done as the 'proportion of patients with improved EHF score', 'stable EHF score' and 'EHF score worse', and 'proportion of patients without impairment', 'proportion with WHO grade 1' and 'proportion with WHO grade 2'. It is recommended that the concepts and terminology of the WHO International Classification of Impairments, Activities and Participation (ICIDH-2) be adopted in the field of leprosy, particularly for the areas of prevention of impairment and disability and rehabilitation. The 'WHO disability grade' should be renamed 'WHO impairment grade'.

Structural changes in the promoter region mediate transvection at the sn-glycerol-3-phosphate dehydrogenase gene of Drosophila melanogaster.

Trans effects at the sn-glycerol-3-phosphate dehydrogenase locus (Gpdh) of Drosophila melanogaster give rise to an increase in GPDH activity and mRNA from the wild-type allele in heterozygotes with some low-activity alleles. Either the low-activity alleles that induce the effect have a defective P-element inserted between the promoter and a downstream intronic enhancer element or the promoter region is deleted. The trans effect is pairing dependent, characteristic of transvection at some other loci. The defective P-elements that mediate transvection are located between the promoter and at least up to 6 bp downstream of the transcription start site. Transvection at Gpdh appears similar to the "enhancer action in trans" mode at the yellow locus.

Generation of mucosal cytotoxic T cells against soluble protein by tissue-specific environmental and costimulatory signals.

We compared peripheral and mucosal primary CD8 T cell responses to inflammatory and noninflammatory forms of antigen in a T cell-adoptive transfer system. Immunization with the soluble antigen, ovalbumin (ova), administered i.p. or orally without adjuvant, activated nonmucosal CD8 T cells but did not induce cytotoxic activity. However, after activation, the transferred cells entered the intestinal mucosa and became potent antigen-specific killers. Thus, exogenous intact soluble protein entered the major histocompatibility complex class I antigen presentation pathway and induced mucosal cytotoxic T lymphocytes. Moreover, distinct costimulatory requirements for activation of peripheral versus mucosal T cells were noted in that the CD28 ligand, B7-1, was critical for activated mucosal T cell generation but not for activation of peripheral CD8 T cells. The costimulator, B7-2, was required for optimum activation of both populations. Infection with a new recombinant vesicular stomatitis virus encoding ovalbumin induced lytic activity in mucosal as well as peripheral sites, demonstrating an adjuvant effect of inflammatory mediators produced during virus infection. Generation of antiviral cytotoxic T lymphocytes was also costimulation-dependent. The results indicated that induction of peripheral tolerance via antigen administration may not extend to mucosal sites because of distinct costimulatory and inflammatory signals in the mucosa.

Glycosyltransferase regulation mediated by pre-TCR signaling in early thymocyte development.

CT1 is a carbohydrate moiety of CD45 that is expressed on fetal thymocytes in vivo. Examination of CT1 expression on thymocyte subsets revealed that primarily pro-T cells (CD44+ CD25+) and pre-T cells (CD44- CD25+) expressed CT1. Interestingly, non-T-lineage committed lymphoid progenitors (CD44+ CD25-) lacked CT1 indicating temporal regulation of expression of this determinant in early T-lineage committed development. In addition, CT1 was expressed by the majority of thymocytes in RAG-2(-/-) mice where thymocyte development is blocked at the CD44- CD25+ stage. Since late pre-T cells (CD44- CD25-) lacked the CT1 epitope we tested whether pre-TCR triggering regulated CT1 expression. Injection of CD3epsilon-specific mAb into RAG-2(-/-) mice induces differentiation of immature thymocytes to the double-positive stage of thymocyte development. Using this system, we demonstrated that expression of CT1 by RAG-2(-/-) thymocytes was rapidly lost from pre-T cells following anti-CD3 mAb treatment. Furthermore, the decline in CT1 expression induced by CD3 signaling paralleled a loss of mRNA for the glycosyltransferase responsible for the addition of CT1 to CD45. Flow cytometric analysis also revealed that the loss of the CT1 epitope was inversely correlated with an increase in peanut agglutinin ligand expression, demonstrating a complex regulation of cell surface glycosylation at a critical juncture in thymocyte development.

Progress of impairment scores following commencement of chemotherapy in multibacillary leprosy patients.

STUDY AIM: To investigate the progress of impairment over time in multibacillary (MB) leprosy patients. STUDY DESIGN: Retrospective cohort study. STUDY POPULATION: One-thousand-eighty-two MB patients newly registered in nine field clinics in the Western Region of Nepal between 1980 and 1993. METHODS: Data on impairment at diagnosis and at yearly intervals afterward were collected from patient records of MB patients already released from multidrug therapy (MDT). The World Health Organization (WHO) 1988 "disability" grading scale (0-2, for both eyes, hands and feet--six sites) was used as a measure of impairment. For the analysis we summed the WHO grading for the six sites into an eyes-hands-feet (EHF) sum score (minimum 0, maximum 12). The EHF score at 2 years of follow up was used to compute the main outcome measures: impairment at 2 years, yes or no, and deterioration of impairment compared with diagnosis. The combined effect of age, sex, classification and impairment status at diagnosis on the outcome was examined with logistic regression. RESULTS: At diagnosis, 55.8% of the patients had some impairment. This proportion decreased over 2 years to 43.9%. Among patients without initial impairment, 31/310 (10%) developed impairment during the study period. This was 81/396 (20.5%) among patients with impairment at diagnosis. The adjusted odds ratio (OR) for developing impairment was 1.87 [95% confidence interval (CI) 1.06-3.32] for patients with initial sensory impairment (WHO grade 1). and 1.98 (95% CI 1.15-3.4) for those with initial visible deformity (WHO grade 2). Among patients with impairment at diagnosis, 195/396 (49.2%) had improved after 2 years. CONCLUSIONS: The proportion of patients with impairment after 2 years of antileprosy treatment was 12% less than at diagnosis. Among patients without initial impairment, 10% had developed some impairment after 2 years. The risk of developing impairment was almost double for those with sensory impairment or visible deformity at diagnosis. For purposes of monitoring, evaluation and planning, both the proportion of patients with sensory impairment (WHO grade 1) and the proportion with visible deformity (WHO grade 2) should be reported at diagnosis and at release from treatment.

Construction and characterization of a recombinant adenovirus directing expression of the MAGE-1 tumor-specific antigen.

The finding that many human melanomas express distinct antigens that can be recognised by specific cytolytic T lymphocytes (CTL) implies that immunotherapeutic strategies against this cancer might prove effective. The ex vivo delivery of a tumour-associated antigen to autologous cells and the subsequent re-administration of these cells to the patient might prove effective in boosting the T cell immune response. Recombinant human adenoviral vectors provide an efficient delivery system and have many advantages over other viral and non-viral delivery vehicles. Infection of a panel of human melanoma cell lines by AdCMVMAGE-1, a novel recombinant adenovirus which incorporates the full-length MAGE-1 cDNA, was shown to induce production of high levels of MAGE-1 protein. Incubation of transduced HLA-A1 expressing melanoma cell lines with 2 anti-MAGE-1.A1 CTL clones resulted in specific recognition and lysis of target cells, indicating that the exogenous MAGE-1 protein was processed and presented in a normal manner. Furthermore, quantitative analyses demonstrated a correlation between the efficiency of transduction and the proportion of cells lysed. Importantly for future clinical trials, stimulation of peripheral blood lymphocytes (PBLs) from a melanoma patient by AdCMVMAGE-1-transduced autologous cells resulted in the generation of specific CTLs against the MAGE-1 antigen. Together, our data emphasize the utility of adenoviruses as vaccination vehicles and highlight the potential efficacy of this approach for the treatment of melanoma.

Activation and migration of CD8 T cells in the intestinal mucosa.

The activation of CD8 T cells was studied using transgenic mice expressing an MHC class I-restricted OVA-specific TCR (OT-I). Immunization of OT-I mice resulted in activation of lymph node T cells and up-regulation of expression of B220, CD11a, and CD44 and caused a preferential loss of mucosal-type T cells from the periphery. Immunization induced dramatic changes in intestinal lymphocytes, including the appearance of a CD11a high population that resembled peripheral T cells and induced a primary cytolytic response. Activation of adoptively transferred OT-I cells induced migration of CD8 T cells into the lamina propria and intraepithelial lymphocyte compartments. The results suggest that Ag presentation in the intestinal epithelium occurs, and that CD8 cells activated in the periphery can readily enter the mucosa during an ongoing immune response.

Molecular heterogeneity of naturally occurring sn-glycerol-3-phosphate dehydrogenase low-activity variants in Drosophila melanogaster.

Northern analyses of two low-activity sn-glycerol-3-phosphate dehydrogenase (Gpdh) alleles extracted from natural populations of Drosophila melanogaster showed that one of them, GpdhACyg22, produced wild-type levels of a normal sized (1.7-kb) mRNA but the other, GpdhAMB5, had very low levels of a 1.7-kb mRNA together with low levels of a transcript 200 bp larger. The two variant genes were cloned and sequenced. Compared with normal activity alleles, there were two nucleotide differences in the DNA sequence of GpdhACyg22 which were in first-codon positions and would be expected to give rise to Asn-13-->Tyr and Arg-272-->Cys substitutions. The second of these changes is most likely to account for the altered properties of the enzyme. In contrast, none of the nucleotide differences in GpdhAMB5 would give rise to amino acid substitutions, but a 76-bp deletion in the 5' region removed the normal TATA box and there was a 20-bp insertion in the same region. One of the two transcripts was derived from the use of a substitute TATA box sequence in the insertion, but the 1.9-kb transcript had heterogeneous 5' ends that were not associated with substitute TATA box sequences. The two transcripts either are produced at a lower rate or are less stable than the normal mRNA.

Defective P element insertions affect the expression of sn-glycerol-3-phosphate dehydrogenase alleles in natural populations of Drosophila melanogaster.

A distinctive and geographically widespread category of low-activity variant at the Gpdh locus in Drosophila melanogaster is shown to have defective P elements inserted between the TATA box and the transcription start site. In four examples the insertion was a single 1.15 kilobase (kb) KP element, whereas in another variant there were two KP elements in tandem. A sixth example contained a 0.61 kb defective P element. The target site for all of the insertions is GTGCAAAC. There was no sequence variation either between the insertions or in comparison with two other KP elements previously described from natural populations. The insertions cause a reduction in GPDH mRNA, and are the most likely cause of the low GPDH activity.

Condom use with regular and casual partners among women attending family planning clinics.

A survey of 16,632 women attending family planning clinics in Pennsylvania found that only 13 percent of the sample used condoms. Moreover, 67 percent of the women with regular partners never used condoms with those partners, and 72 percent of women who had casual partners never used them with those partners. Levels of condom use with both regular and casual partners were higher among women younger than 20, those who were nonwhite, those with multiple partners and those who reported a previous STD infection. However, women who had sex partners who used intravenous drugs were less likely to use condoms with regular or casual partners than were women who did not have drug-using partners.

Risk behaviors for HIV infection in clients of Pennsylvania family planning clinics.

Responses from a statewide survey of risk behaviors among clients of Planned Parenthood clinics in Pennsylvania indicate that a sizable proportion of these women are at intermediate or high risk for exposure to the human immunodeficiency virus (HIV). Overall, nearly five percent of the 15,499 women in the sample reported personal behavior, or a partner's behavior, that placed them at high risk, while an additional 20 percent were at intermediate risk. Among all women surveyed, 13 percent had had three or more partners in the past year, 12 percent had had a sexually transmitted disease, four percent had partners who were intravenous (IV) drug users, two percent were either current or past IV drug users, one percent had bisexual partners and less than one percent had partners who had hemophilia or who were infected with the AIDS virus.

PIP: Responses from a statewide survey of risk behaviors among clients of Planned Parenthood clinics in Pennsylvania indicate that a sizeable proportion of these women are at intermediate or high risk for exposure to the human immunodeficiency virus (HIV). Overall, nearly 5% of the 15,499 women in the sample reported personal behavior or a partner's behavior, that places them at high risk, while an additional 20% were at intermediate risk. Among all women surveyed, 13% had had 3 or more partners in the past year, 12% had had a sexually transmitted disease, 4% had partners who were intravenous (IV) drug users, 2% were either current or past IV drug users, 1% had bisexual partners and 1% had partners who had hemophilia or who were infected with the AIDS virus. Of the 16,553 clinic patients who supplied demographic information, 86% were white and 78% were aged 15-24. Information on marital status and education was compiled for women aged 20 and older. Of these 70% had never married and 91% had completed at least a high school education.

Assessing behavioral risk for HIV infection in family-planning and STD clinics: similarities and differences.

Four hundred fifty-five women in family-planning and sexually transmitted disease (STD) clinics were surveyed to determine the degree of participation in behaviors known to be associated with increased risk of human immunodeficiency virus (HIV) infection. A previous history of STD (20%) and multiple sexual partners (73%) were shown to be the high-risk behaviors most prevalent in these populations. Sexual intercourse with persons in high-risk groups (6%) and intravenous (IV) drug use (3.7%) were less prevalent. Differences between the populations from each type of clinic and between races were noted. Overall, almost one third of family-planning clinic clients and nearly half of STD clinic clients reported participation in at least one risk behavior, emphasizing the need for educational efforts toward disease prevention in these settings.