RESUMO
BACKGROUND: Opportunities to improve emergency surgery outcomes exist through guided better practice and reduced variability. Few attempts have been made to define optimal care in emergency surgery, and few clinically derived key performance indicators (KPIs) have been published. A summit was therefore convened to look at resources for optimal care of emergency surgery. The aim of the Donegal Summit was to set a platform in place to develop guidelines and KPIs in emergency surgery. METHODS: The project had multidisciplinary global involvement in producing consensus statements regarding emergency surgery care in key areas, and to assess feasibility of producing KPIs that could be used to monitor process and outcome of care in the future. RESULTS: Forty-four key opinion leaders in emergency surgery, across 7 disciplines from 17 countries, composed evidence-based position papers on 14 key areas of emergency surgery and 112 KPIs in 20 acute conditions or emergency systems. CONCLUSIONS: The summit was successful in achieving position papers and KPIs in emergency surgery. While position papers were limited by non-graded evidence and non-validated KPIs, the process set a foundation for the future advancement of emergency surgery.
Assuntos
Lesões Encefálicas Traumáticas/cirurgia , Pediatria/métodos , Acidentes por Quedas/mortalidade , Acidentes por Quedas/estatística & dados numéricos , Acidentes de Trânsito/mortalidade , Acidentes de Trânsito/estatística & dados numéricos , Adolescente , Mundo Árabe , Lesões Encefálicas Traumáticas/epidemiologia , Criança , Pré-Escolar , Técnica Delphi , Feminino , Humanos , Lactente , Masculino , Oriente Médio/epidemiologia , Pediatria/tendências , Estudos Retrospectivos , Centros de Traumatologia/organização & administração , Centros de Traumatologia/estatística & dados numéricos , Resultado do TratamentoRESUMO
BACKGROUND: Ultrasound (US) is often the imaging modality of choice in women with acute right iliac fossa (RIF) pain, identifying the appendix in up to 99% of patients. The literature, however, lacks clear guidelines on how ultrasonography should be performed to maximise sensitivity and specificity in such patients. Many centres perform untargeted abdomino-pelvic scans, including organs such as the liver and spleen, which unlikely contribute to the presenting complaint. AIMS: We aimed to evaluate the clinical utility of unfocussed abdominal and pelvic US in women of reproductive age with acute RIF pain. METHODS: This multicentre study describes 501 women between the ages of 12 and 50, over a 3-year period from three institutions, presenting acutely with RIF pain and investigated with US abdomen and pelvis. RESULTS: 5.9% of cases confirmed appendicitis sonographically. A normal appendix was visualised in 0.2%. Over 10% identified gynaecological pathology, 41% relating to the right ovary. 10.4% incidental extra-pelvic findings were unrelated to the acute clinical presentation. 0.8% of patients had extra-pelvic findings meriting further clinical assessment. CONCLUSION: The results herein reflect findings from high volume emergency surgical departments, demonstrating that unfocussed abdominal and pelvic ultrasounds are not an appropriate use of resources in reproductive women with RIF pain. Clinically relevant extra-pelvic US findings account for less than 1%, rarely contributing to the acute presentation. The appendix was only visualised in 6% of patients, suggesting that a focussed appendiceal and pelvic US would better assist diagnosis with a higher yield and increased sensitivity and specificity.
Assuntos
Abdome/diagnóstico por imagem , Dor Abdominal/etiologia , Apendicite/diagnóstico por imagem , Pelve/diagnóstico por imagem , Ultrassonografia/métodos , Dor Abdominal/patologia , Adolescente , Adulto , Apendicite/patologia , Criança , Estudos de Coortes , Feminino , Humanos , Pessoa de Meia-Idade , Estudos Retrospectivos , Adulto JovemRESUMO
Preventing virally induced liver disease begins with an understanding of the host factors that define susceptibility to infection. Hepatitis C virus (HCV) is a global health issue, with an estimated 170 million infected individuals at risk of developing liver disease including fibrosis and hepatocellular carcinoma. The liver is the major reservoir supporting HCV replication and this hepatocellular tropism is defined by HCV engagement of cellular entry receptors. Hepatocytes are polarized in vivo and this barrier function limits HCV entry. We previously reported that activated macrophages promote HCV entry into polarized hepatocytes via a TNF-α-dependent process; however, the underlying mechanism was not defined. In this study, we show that several TNF superfamily members, including TNF-α, TNF-ß, TWEAK and LIGHT, promote HCV entry via NF-κB-mediated activation of myosin light chain kinase (MLCK) and disruption of tight junctions. These observations support a model where HCV hijacks an inflammatory immune response to stimulate infection and uncovers a role for NF-κB-MLCK signalling in maintaining hepatocellular tight junctions.
Assuntos
Hepacivirus/fisiologia , Hepatite C/virologia , Fígado/virologia , Quinase de Cadeia Leve de Miosina/metabolismo , NF-kappa B/metabolismo , Fatores de Necrose Tumoral/metabolismo , Internalização do Vírus , Carcinoma Hepatocelular/virologia , Ativação Enzimática , Hepatite C/metabolismo , Hepatócitos/virologia , Humanos , Fígado/metabolismo , Cirrose Hepática/virologia , Neoplasias Hepáticas/virologia , Transdução de Sinais , Junções Íntimas/metabolismo , Junções Íntimas/virologia , Fator de Transcrição RelA/metabolismoRESUMO
BACKGROUND: Undergoing an endoscopy is a stressful experience for patients. AIMS: To audit the endoscopy pathway to improve patient satisfaction. METHODS: A prospective survey of endoscopy patients to identify system improvements that were then implemented. RESULTS: The survey was performed before (N = 71) and after (N = 60) process improvements identified by the initial survey. Information provision and staff communication skills were identified for optimisation. Patient anxiety at home was significantly reduced (median 2 vs. 1, p < 0.01). Education of endoscopy staff significantly improved the quality of information provided before and after the procedure with regard to sedation (median 4 vs. 5, p < 0.01), discomfort (median 4 vs. 5, p < 0.01), complications (28 vs. 82 %, p < 0.01), findings (89 vs. 100 %, p < 0.01) and follow-up (73 vs. 90 %, p = 0.015). Gloucester Comfort Scores during endoscopy improved (median 1 vs. 0, p < 0.01) without increasing sedation levels. Patient feelings of invasion/trauma significantly decreased. Overall 95 % of patients were satisfied. CONCLUSION: Structured information leaflets and improved staff communication skills reduce anxiety and enhance patients' experiences. They are now standard operating procedures.
Assuntos
Ansiedade/prevenção & controle , Ansiedade/psicologia , Endoscopia Gastrointestinal/psicologia , Educação em Saúde/estatística & dados numéricos , Satisfação do Paciente , Adulto , Idoso , Colonoscopia/psicologia , Colonoscopia/estatística & dados numéricos , Endoscopia Gastrointestinal/estatística & dados numéricos , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Medição da Dor , Estudos ProspectivosRESUMO
Hepatitis B virus (HBV) is one of the world's major unconquered infections, resulting in progressive liver disease, and current treatments rarely cure infection. A limitation to discovering new therapies is our limited knowledge of HBV entry and dissemination pathways that hinders the development of in vitro culture systems. To address this gap in our understanding we optimized the genesis of infectious lentiviral pseudoparticles (HBVpps). The recent discovery that the bile salt transporter sodium taurocholate co-transporting polypeptide (NTCP) acts as a receptor for HBV enabled us to assess the receptor dependency of HBVpp infection. HBVpps preferentially infect hepatoma cells expressing NTCP, whereas other non-liver cells engineered to express NTCP do not support infection, suggesting that additional hepatocyte-specific factors are required for HBVpp internalization. These results highlight the value of the HBVpp system to dissect the pathways of HBV entry and dissemination.
Assuntos
Vírus da Hepatite B/fisiologia , Hepatócitos/virologia , Interações Hospedeiro-Patógeno , Lentivirus/fisiologia , Transportadores de Ânions Orgânicos Dependentes de Sódio/metabolismo , Receptores Virais/metabolismo , Simportadores/metabolismo , Internalização do Vírus , Linhagem Celular , Vírus da Hepatite B/genética , Humanos , Lentivirus/genéticaRESUMO
INTRODUCTION: The aim of this study is to determine if simulated 3D vision improves the speed and accuracy of laparoscopic phantom tasks in laparoscopically naïve subjects. METHODS: Thirty laparoscopically naïve subjects were divided into matched groups according to age, sex, hand dominance and initial scores on a standardised visio-spatial test. Laprotrain(©) laparoscopic simulators were used, one attached to the standard 2D monitor and the other to a simulated 3D monitor and 3D glasses were worn by the subjects in this group. Five standardised laparoscopic tasks were developed and the subjects underwent testing on four separate occasions with more than 24 h between sessions. The subjects were timed for each task and errors were recorded by two independent observers. In the second part of the study, subjects switched to the opposite group and task times and errors were again recorded. Statistical differences between groups were calculated using student t-test and Fisher's exact test. RESULTS: There were fifteen subjects in each group with no significant difference in demographic or psychometric variables. The mean time to complete the tasks was faster in the 3D group compared with the 2D group. There was a lower rate of errors noted in the 3D group compared with the 2D group but this only reached statistical significance in two of the five laparoscopic tasks. In the crossover study, subjects who had trained on simulated 3D had better task times and fewer errors compared to those who had trained on 2D simulators. DISCUSSION & CONCLUSION: Training on a simulated 3D model (compared to standard 2D) allows trainees to reach proficiency sooner.
Assuntos
Competência Clínica , Percepção de Profundidade , Laparoscopia/educação , Desempenho Psicomotor , Adulto , Estudos Controlados Antes e Depois , Estudos Cross-Over , Feminino , Humanos , Imageamento Tridimensional , Laparoscopia/métodos , Masculino , Análise e Desempenho de Tarefas , Adulto JovemRESUMO
Many viruses target the polarized epithelial apex during host invasion. In contrast, hepatitis C virus (HCV) engages receptors at the basal surface of hepatocytes in the polarized liver parenchyma. Hepatocyte polarization limits HCV entry by undefined mechanism(s). Given the recent reports highlighting a role for receptor mobility in pathogen entry, we studied the effect(s) of hepatocyte polarization on viral receptor and HCV pseudoparticle (HCVpp) dynamics using real-time fluorescence recovery after photobleaching and single particle tracking. Hepatoma polarization reduced CD81 and HCVpp dynamics at the basal membrane. Since cell polarization is accompanied by changes in the actin cytoskeleton and CD81 links to actin via its C-terminus, we studied the dynamics of a mutant CD81 lacking a C-terminal tail (CD81(ΔC)) and its effect(s) on HCVpp mobility and infection. CD81(ΔC) showed an increased frequency of confined trajectories and a reduction of Brownian diffusing molecules compared to wild-type protein in non-polarized cells. However, these changes were notobserved in polarized cells. HCVpp showed a significant reduction in Brownian diffusion and infection of CD81(ΔC) expressing non-polarized cells. In summary, these data highlight the dynamic nature of CD81 and demonstrate a role for CD81 lateral diffusion to regulate HCV infection in a polarization-dependent manner.
Assuntos
Polaridade Celular , Hepacivirus/fisiologia , Hepatócitos/fisiologia , Receptores Virais/metabolismo , Tetraspanina 28/metabolismo , Internalização do Vírus , Células Hep G2 , Hepatócitos/imunologia , Hepatócitos/virologia , Humanos , Microscopia de FluorescênciaRESUMO
Laparoscopic appendicectomy has become standard in the treatment of acute appendicitis in most hospitals in Ireland. Studies have shown that it is a safe procedure for trainees to perform. However, these studies were conducted in university teaching hospitals whereas a significant proportion of training in Ireland takes place in peripheral hospitals which provide a different training environment. The aim of this study was to determine whether laparoscopic appendicectomy is a safe procedure for surgical trainees to perform in a peripheral hospital setting. A retrospective analysis was performed of appendicectomies carried out at a peripheral hospital over a 12 month period. Comparisons were made between consultant surgeons and trainees for a variety of outcomes. Of 155 appendicectomies, 129 (83.2%) were performed laparoscopically, of which 10 (7.75%) were converted to open. Consultants performed 99 (77%) laparoscopic appendicectomies. There were no statistically significant differences between consultants and trainees in complication rates (19 (19.2%) vs. 4 (13.3%), p = 0.46), mean length of hospital stay (4.7 +/- 4.0 vs. 3.4 +/- 3.3 days, p = 0.13), or rate of conversion to open operation (9 (9.1%) vs. 1 (3.3%), p = 0.45). For cases of complicated appendicitis there were no significant differences between consultants and trainees in complication rates (12 vs. 2, p = 0.40) or length of hospital stay (6.4 +/- 3.9 vs. 4.7 +/- 5.6 days, p = 0.27). We conclude that laparoscopic appendicectomy is a safe procedure for trainees to perform in the peripheral hospital setting and should be incorporated into surgical training programs at an early stage of training.
Assuntos
Apendicectomia/métodos , Cirurgia Geral/educação , Internato e Residência , Laparoscopia/educação , Adulto , Apendicectomia/estatística & dados numéricos , Feminino , Humanos , Irlanda , Laparoscopia/estatística & dados numéricos , Tempo de Internação/estatística & dados numéricos , Masculino , Complicações Pós-Operatórias , Estudos RetrospectivosRESUMO
INTRODUCTION: The management of gastric lymphoma is controversial and a wide variety of unimodality or multimodality approaches have been used. The aim of this report is to highlight the variety of treatment regimens deployed, the outcomes achieved and to present a modern management approach for this enigmatic tumour. RESULTS: 42 cases of primary gastric lymphoma managed at one centre over a 15-year period were reviewed. Weight loss (52%), pain (41%) and anorexia (33%) were the most common presenting symptoms. Most patients (86%) had high-grade lymphoma. Primary treatment modalities included surgery (36%), chemotherapy (40%), supportive care only (22%) and H. pylori eradication (2%). Adjuvant therapies included chemotherapy (17%), radiotherapy (7%) and combined chemoradiotherapy (5%). The overall median survival was 53 months, with a five year survival of 46%. In the curative group, the median survival was 75 months and five year survival 58%. Curative surgery or chemotherapy +/- radiotherapy were similarly effective for stage IE and IIE disease. CONCLUSIONS: The prognosis for gastric lymphoma is grade- and stage-dependent. With equivalent outcomes for cure in localised gastric lymphoma for surgery and chemotherapy, the latter is preferred in this unit because of gastric preservation, with surgery being reserved for failed medical management or presentations with haemorrhage, perforation or obstruction refractive to steroid therapy.
Assuntos
Linfoma/terapia , Neoplasias Gástricas/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Terapia Combinada , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Cuidados Paliativos , Taxa de Sobrevida , Resultado do TratamentoRESUMO
A second estrogen receptor, estrogen receptor-beta, was identified in 1996 and has led to an intensive re-evaluation of the role of estrogens in normal physiological and disease processes. While much has been learnt about this new receptor, there remain many outstanding questions, particularly regarding its prognostic significance and therapeutic implications.
Assuntos
Neoplasias da Mama/fisiopatologia , Receptor beta de Estrogênio/fisiologia , Neoplasias da Mama/terapia , Receptor beta de Estrogênio/genética , Feminino , Humanos , Prognóstico , Resultado do TratamentoRESUMO
We recently reported that retroviral pseudotypes bearing the hepatitis C virus (HCV) strain H and Con1 glycoproteins, genotype 1a and 1b, respectively, require CD81 as a coreceptor for virus-cell entry and infection. Soluble truncated E2 cloned from a number of diverse HCV genotypes fail to interact with CD81, suggesting that viruses of diverse origin may utilize different receptors and display altered cell tropism. We have used the pseudotyping system to study the tropism of viruses bearing diverse HCV glycoproteins. Viruses bearing these glycoproteins showed a 150-fold range in infectivity for hepatoma cells and failed to infect lymphoid cells. The level of glycoprotein incorporation into particles varied considerably between strains, generally reflecting the E2 expression level within transfected cells. However, differences in glycoprotein incorporation were not associated with virus infectivity, suggesting that infectivity is not limited by the absolute level of glycoprotein. All HCV pseudotypes failed to infect HepG2 cells and yet infected the same cells after transduction to express human CD81, confirming the critical role of CD81 in HCV infection. Interestingly, these HCV pseudotypes differed in their ability to infect HepG2 cells expressing a panel of CD81 variants, suggesting subtle differences in the interaction of CD81 residues with diverse viral glycoproteins. Our current model of HCV infection suggests that CD81, together with additional unknown liver specific receptor(s), mediate the virus-cell entry process.
Assuntos
Antígenos CD/metabolismo , Hepacivirus/classificação , Hepacivirus/patogenicidade , Hepatócitos/virologia , Proteínas do Envelope Viral/metabolismo , Sequência de Aminoácidos , Animais , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , HIV-1/genética , HIV-1/metabolismo , Hepacivirus/genética , Hepacivirus/fisiologia , Humanos , Fígado , Linfócitos/virologia , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Tetraspanina 28 , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genéticaRESUMO
Little is known about the role of Abs in determining the outcome of hepatitis C virus (HCV) infection. By using infectious retroviral pseudotypes bearing HCV glycoproteins, we measured neutralizing Ab (nAb) responses during acute and chronic HCV infection. In seven acutely infected health care workers, only two developed a nAb response that failed to associate with viral clearance. In contrast, the majority of chronically infected patients had nAbs. To determine the kinetics of strain-specific and crossreactive nAb emergence, we studied patient H, the source of the prototype genotype 1a H77 HCV strain. An early weak nAb response, specific for the autologous virus, was detected at seroconversion. However, neutralization of heterologous viruses was detected only between 33 and 111 weeks of infection. We also examined the development of nAbs in 10 chimpanzees infected with H77 clonal virus. No nAb responses were detected in three animals that cleared virus, whereas strain-specific nAbs were detected in six of the seven chronically infected animals after approximately 50 weeks of infection. The delayed appearance of high titer crossreactive nAbs in chronically infected patients suggests that selective mechanism(s) may operate to prevent the appearance of these Abs during acute infection. The long-term persistence of these nAbs in chronically infected patients may regulate viral replication.
Assuntos
Hepatite C Crônica/imunologia , Hepatite C/imunologia , Doença Aguda , Animais , Reações Cruzadas , Anticorpos Anti-Hepatite C/sangue , Humanos , Testes de Neutralização , Pan troglodytes , Especificidade da EspécieRESUMO
BACKGROUND: The identification of a second estrogen receptor (ER-beta) has significant implications for therapeutic strategy in breast cancer management arising from the potential agonist effect of Tamoxifen at estrogen receptor sites and as such, antiestrogen therapy may be inappropriate in patients with a dominance of ER-beta. METHODS: To determine the proportion of breast cancer patients who may be so at risk, we developed a novel multiplexed RT-PCR technique to establish the relative ER-alpha and ER-beta levels in 53 primary breast cancers, 11 normal breast tissues and six cell lines. We further assessed the prognostic significance of receptor status relative to the Nottingham prognostic index (NPI). The ER-alpha and ER-beta status was also determined by immunohistochemistry using previously published and 'in-house' scoring systems. RESULTS: Using RT-PCR analysis, 46 tumours were hormone receptor positive (ER+) with 42 displaying ER-alpha predominance. Comparison with immunohistochemistry demonstrated 44/53 (ER-alpha) and 27/50 (ER-beta) concordance rates. There was no significant difference in the NPI between ER-alpha and ER-beta predominant cohorts or between ER+ and ER- cohorts. CONCLUSION: This study identifies the existence of a subgroup of ER+ patients in whom Tamoxifen therapy may be inappropriate and has significant implications for adjuvant therapy of primary breast cancer.
Assuntos
Biomarcadores Tumorais/genética , Neoplasias da Mama/genética , Carcinoma Ductal de Mama/genética , Carcinoma Lobular/genética , Receptores de Estrogênio/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/epidemiologia , Carcinoma Ductal de Mama/epidemiologia , Carcinoma Lobular/epidemiologia , Receptor alfa de Estrogênio , Receptor beta de Estrogênio , Feminino , Perfilação da Expressão Gênica , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , Estudos Prospectivos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Risco , Saúde da MulherRESUMO
Removal of the V1-V3 loops from IIIB gp120 results in a protein, PR12, with altered immunogenicity compared to the full-length protein. Polyclonal immune sera raised in rats using PR12 as immunogen recognizes envelope glycoproteins of clades A, B, C, E, F and G and can neutralize chimeric human immunodeficiency virus type 1 (HIV-1) HXB2 viruses expressing envelopes from primary HIV-1 clades B, C, E and F. These data suggest that the immune response to PR12 is directed toward conserved epitopes expressed by viral glycoproteins of diverse genotypes. Five monoclonal antibodies (mAb) derived from PR12-immunized rats were unable to neutralize virus infectivity; hence the epitopes responsible for the induction of this cross-clade neutralizing activity remain to be elucidated. However, PR12 immune sera were able to compete with the human neutralizing mAb 2G12 for gp120 binding, implying that this epitope may be immunogenic when expressed in the context of this truncated protein.
Assuntos
Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , HIV-1/imunologia , Animais , Células CHO , Cricetinae , Anticorpos Anti-HIV/sangue , Proteína gp120 do Envelope de HIV/genética , HIV-1/classificação , HIV-1/genética , Humanos , Mutagênese , Testes de Neutralização , FilogeniaRESUMO
Between 1996 and 1999 thirteen cases of HIV infection were detected in Doncaster, a small town in the north of England (population approximately 250,000). A complex network of shared sexual histories involving local nightclubs linked these cases, with the only known risk factor being heterosexual intercourse. A series of frozen blood samples was collected in 1998-1999 and amplified by PCR to generate full-length gp120 clones. Sequencing demonstrated that all the transmission events in this heterosexual group involved the B subtype of HIV-1. When relationships between the samples were assessed it became clear that these 13 cases represented at least three separate strains of HIV-1, indicating that HIV is well established in this community. Eleven of the 13 cases were related, forming two distinct groups. Further investigation revealed that one group contained five patients whose general health was good and who were not receiving HAART. In contrast, the second group of six patients, including the putative index case, were symptomatic, receiving HAART, and may have been infected with a CXCR-4-utilizing virus. Several of the cases that were linked by genetic criteria were not linked by contact tracing, implying that further undiagnosed cases may exist in this community. To our knowledge, this is the largest outbreak of HIV studied within the heterosexual community in the United Kingdom to date, suggesting that this route of infection is becoming more common within the United Kingdom.
Assuntos
Surtos de Doenças , Infecções por HIV/epidemiologia , HIV-1/classificação , Feminino , Genoma Viral , Infecções por HIV/transmissão , Humanos , Masculino , Dados de Sequência Molecular , Filogenia , Receptores CXCR4 , Estudos Retrospectivos , Fatores Sexuais , Reino Unido/epidemiologiaRESUMO
To assess the antigenicity of envelope glycoproteins derived from primary human immunodeficiency virus type 1 populations, their interactions with the receptor CD4, and their coreceptor usage, we have cloned and expressed multiple gp120 proteins from a number of primary virus isolates. Characterization of these proteins showed a high degree of antigenic polymorphism both within the CD4 binding site and in defined neutralization epitopes, which may partially account for the general resistance of primary isolates to neutralizing agents. Furthermore, chimeric viruses expressing gp120 proteins with reduced CD4 binding abilities are viable, suggesting that primary viruses may require a less avid interaction with the receptor CD4 to initiate infection than do their laboratory-adapted counterparts. The coreceptor usage of chimeric viruses was related to the ability of the virus to bind CD4, with reduced CD4 binding correlating with preferential usage of CXCR4. Changes in coreceptor usage mapped to sequence changes in the C2 and V4 regions, with no changes seen in the V3 region.
Assuntos
Variação Antigênica , Antígenos CD4/metabolismo , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp120 do Envelope de HIV/metabolismo , Receptores de Quimiocinas/metabolismo , Sequência de Aminoácidos , Animais , Sítios de Ligação , Proteína gp120 do Envelope de HIV/química , Proteína gp120 do Envelope de HIV/genética , Infecções por HIV/virologia , HIV-1/imunologia , Humanos , Masculino , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Reação em Cadeia da Polimerase , Ratos , Proteínas Recombinantes de Fusão/metabolismo , Análise de Sequência de DNARESUMO
Nucleotide sequences of HIV-1 from plasma virus RNA and proviral DNA extracted from blood dendritic cells (DCs) and from T cells were analyzed to determine whether blood DCs may harbor a restricted population of virus variants. The sequence of the V3 loop and 51 bases from the 3' flanking region were determined in four patients not receiving antiviral therapy. There was no evidence of a unique or more restricted population of variants in DCs for any of the four patients studied. However, for one patient there was evidence of differences between plasma virus and virus in the T cell population, with virus in the plasma showing a closer relationship to DC-derived sequences.
Assuntos
Células Dendríticas/virologia , Proteína gp120 do Envelope de HIV/genética , Infecções por HIV/virologia , HIV-1/genética , Fragmentos de Peptídeos/genética , Linfócitos T/virologia , Sequência de Aminoácidos , DNA Viral/genética , Infecções por HIV/sangue , HIV-1/classificação , HIV-1/patogenicidade , Humanos , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , Provírus/genética , RNA Viral/genéticaRESUMO
Sequences from the nef/LTR overlap region of the human immunodeficiency virus type 2 (HIV-2) genome were amplified from uncultured peripheral blood mononuclear cells (PBMCs) from 40 HIV-2-infected individuals in The Gambia, West Africa. Additional sequences from the plasma of three blood donors were also derived. Analysis of HIV-2 U3 LTR transcription factor elements (PuB-1, p-ets, PuB-2, peri-kappa B, and NF-kappa B sites) indicated a relatively high level of conservation in vivo. The region immediately 3' of the nef termination codon, which exhibits clade-dependent specificity, was targeted by PCR to differentiate HIV-2 subtype A from subtype B infections, the two principal clinical HIV-2 subtypes. All clinical samples analyzed (n = 43) from The Gambia were identified as HIV-2 subtype A by a combination of LTR sequence analysis and subtype-specific amplification of subtypes A and B. Differential PCR amplification of the HIV-2 U3 LTR region represents a rapid means of differentiating subtype A from subtype B infections, the two dominant HIV-2 subtypes that are important in human disease.
Assuntos
Infecções por HIV/virologia , HIV-2/classificação , HIV-2/genética , Sequências Repetidas Terminais/genética , Sequência de Bases , Gâmbia/epidemiologia , Produtos do Gene nef/genética , Humanos , Leucócitos Mononucleares/virologia , Dados de Sequência Molecular , Análise de Sequência de DNA , Produtos do Gene nef do Vírus da Imunodeficiência HumanaRESUMO
Ideally HIV antibody tests have to be both extremely sensitive and able to recognize all known HIV subtypes. Three patients whose sera failed to react with a synthetic oligopeptide-based HIV antibody test are described in this report. The patients were a Pakistani male infected recently, an Australian male infected for several years, and a Ugandan woman with AIDS. The presence of anti-HIV antibodies was confirmed by means of a standard algorithm with different assay formats. All three sera failed to react in one antiglobulin enzyme-linked immunosorbent assay (ELISA) (Bioelisa HIV-1+2, Biokit SA). No single underlying reason could be identified for the assay failure in the three cases. The first patient, probably infected recently when first tested, was strongly positive by the same assay a year later, confirming the relative insensitivity of oligopeptide assays reported previously for detecting the early antibody response. The other two patients appear to have been infected for several years. Although unlikely to have been infected with a non-clade B virus, the sample from patient 2 lacked detectable antibody to the transmembrane glycoprotein (gp41), the site of the synthetic oligopeptides. Patient 3, of Ugandan origin, was found to be infected with a non-clade B virus. Although her serum reacted strongly to subtype B gp41 in Western blot, it failed to react in the antiglobulin ELISA. Since there appears to be no single common explanation for these three failures there is little opportunity to identify prospectively those situations where testing using assays employing synthetic oligopeptides on the solid phase is likely to fail.