RESUMO
BACKGROUND: Preoperative assessment of C2 pedicle morphology is critical to safe pedicle screw placement. To avoid iatrogenic injury, complex digital templating software has been introduced; however, this technology may not be available in many centers. We report a technique for preoperative assessment of C2 pedicle screw placement safety based upon 2-dimensional sagittal computed tomography (CT) scan images and verify its utility in clinical practice. METHODS: A total of 46 consecutive patients underwent cervical spine CT scans between 2005 and 2011. The C2 pedicle morphology was assessed on sagittal CT imaging by 5 independent reviewers to determine the feasibility and risk associated with pedicle screw placement. Thirty consecutive patients underwent C2 pedicle screw placement and were followed clinically for a minimum of 2 years. The ability to place a screw was noted, and accuracy of screw placement was assessed postoperatively by CT scan. RESULTS: The CT scan analysis demonstrated that 11% (5/46) of patients had sufficient pedicle size bilaterally to allow safe placement of long pedicle screws with a low risk of vertebral artery injury, whereas 15% (7/46) were considered a high risk bilaterally. Screw placement was deemed low risk in 28%, moderate risk in 38%, and high risk in 34%. Excellent intraobserver reliability and good interobserver reliability was observed. Clinically, 18 of 20 (90%) low-risk and 21 of 24 (88%) moderate-risk pedicle screws were placed safely versus 5 of 16 (31%) high-risk pedicle screws (P < .001). CONCLUSIONS: Using the described technique for evaluating the C2 pedicle via sagittal CT scan images allows for safe and reliable pedicle screw placement without relying upon complex digital templating software, which may have limited availability. LEVEL OF EVIDENCE: II. CLINICAL RELEVANCE: This study aids in the surgical decision-making behind the placement of C2 pedicle screws using CT scans without reliance upon complex digital templating software.
RESUMO
Except during acutely lethal infection, macaques infected with SIVmac251 produce antibodies that neutralize the virus in CEMx174 cells, macaque PBMC and macrophage cultures. In a previous report, we had shown that whereas neutralization of the SIVmac251 was complete in lymphocyte cultures, "protected" macrophages had actually become latently infected, and remained viral DNA-positive, but the infection was nonproductive as long as antibodies were maintained in the medium. Removal of the antibodies as long as 1 week later, resulted in resurgence of virus replication. In the present study, we compared neutralization of SIVmac239 with that of neutralization of SHIV and HIV-1, and sought to determine whether the failure to prevent infection in macrophages was also typical of neutralization of SHIV and HIV-1 in macaque and human macrophage cultures, respectively. The results showed that similar to SIVmac251, neutralizing antibodies did not block SIVmac239 infection in macaque macrophages, although they blocked infection of the virus in T cells. The data from neutralization of SHIV using anti-SHIV antibodies and for neutralization of HIV-1 (89.6 and Bal) using anti-HIV IgG in both T cells and macrophages, however, can be summarized with a single statement: neutralization of SHIV and HIV-1 was complete in all of the cultures, with no evidence of establishment of latent infection in or resurgence of virus replication after antibodies were removed from macrophage cultures. The non-neutralizability of SIVmac (251 and 239) in macrophages is therefore unique to the SIVmac and not relevant to neutralization of HIV-1.
Assuntos
HIV-1/imunologia , Macrófagos/imunologia , Vírus Reordenados/imunologia , Vírus da Imunodeficiência Símia/imunologia , Animais , Anticorpos Antivirais/imunologia , Sequência de Bases , Células Cultivadas , Primers do DNA , Humanos , Linfócitos/virologia , Macaca mulatta , Testes de Neutralização , Vírus Reordenados/isolamento & purificaçãoRESUMO
We examined the renal pathology and viral genetic changes following inoculation of six rhesus macaques with lymphocyte-tropic SIVmac239. Portions of the renal cortex were sieved into glomerular and tubulointerstitial (TI) fractions and examined for SIVmac sequences by PCR and for p27 core antigen. SIVmac sequences were detected in renal tissue from five of six macaques (three of five glomerular and five of five TI fractions were positive for SIV by PCR). Glomerulosclerosis (segmental and global) was evident in two macaques that were positive for env sequences in the glomerular fractions. Diffuse mesangial hyperplasia and matrix expansion were present in all three animals with glomerular SIV, as was an increase in glomerular collagen I and collagen IV. Tubulointerstitial inflammation was evident in all virus-inoculated macaques. The TI infiltration of CD68+ cells was most pronounced in the animals with SIVmac present in the glomerulus. All SIVmac-infected macaques exhibited increased glomerular deposition of IgM and to a lesser extent IgG, but no C3 or IgA was evident. Sequence analyses of the viral env gene (gp120) isolated from the glomerular and TI fractions of a macaque that developed glomerulopathy revealed the presence of specific viral variants in glomerular and TI fractions. In addition, chimeric viruses constructed with glomerular but not tubulointerstitial gp120 sequences were converted to a macrophage-tropic phenotype. These results indicate that infection by lymphocyte-tropic SIVmac239 is primarily associated with immunoglobulin deposition in the glomerulus and suggests that when glomerulosclerosis develops there is selection of viral variants that are macrophage tropic in nature.
Assuntos
Glomerulonefrite/imunologia , Glomerulonefrite/virologia , Rim/patologia , Rim/virologia , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/isolamento & purificação , Animais , Antígenos Virais/análise , Antígenos Virais/genética , Linfócitos T CD4-Positivos/imunologia , Colágeno/análise , Produtos do Gene gag , Genes env , Imuno-Histoquímica , Testes de Função Renal , Glomérulos Renais/ultraestrutura , Leucócitos Mononucleares/imunologia , Tecido Linfoide/virologia , Macaca mulatta , Dados de Sequência Molecular , Análise de Sequência de DNA , Síndrome de Imunodeficiência Adquirida dos Símios/complicações , Vírus da Imunodeficiência Símia/crescimento & desenvolvimento , Vírus da Imunodeficiência Símia/patogenicidade , Proteínas do Envelope ViralRESUMO
Macaques inoculated orally, vaginally, or parenterally with SHIV(KU-1) develop severe systemic infection, acute loss of CD4+ T cells, and AIDS. We showed in a previous report that passive immunization with neutralizing serum protected macaques against infection with parenterally inoculated pathogenic SHIV given 24 hr later. In the study reported here we asked whether the identical passive immunization protocol would protect macaques against infection with pathogenic SHIV following oral inoculation of the virus. Ten pigtail macaques were inoculated orally with one animal infectious dose of SHIV(KU-1). Four of the 10 had been given pooled anti-SHIV plasma (15 ml/kg) 24 hr earlier, 4 others were given the same dose of anti-SHIV plasma 2 hr after virus challenge, and the 2 remaining animals were used as controls. The neutralizing antibodies failed to protect macaques against infection after mucosal challenge with SHIV(KU-1).
Assuntos
Vacinas contra a AIDS/imunologia , Anticorpos Anti-HIV/imunologia , Infecções por HIV/prevenção & controle , Imunização Passiva , Vacinas contra a SAIDS/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/prevenção & controle , Vacinas contra a AIDS/administração & dosagem , Animais , HIV/imunologia , Anticorpos Anti-HIV/administração & dosagem , Humanos , Macaca nemestrina , Mucosa , Testes de Neutralização , Vacinas contra a SAIDS/administração & dosagem , Vírus da Imunodeficiência Símia/imunologiaRESUMO
By animal-to-animal passage in macaques we derived a pathogenic chimeric simian-human immunodeficiency virus (SHIV) that caused CD4+ T cell loss and AIDS in pigtail macaques and used it to inoculate 20 rhesus and pigtail macaques by the intravaginal and intravenous routes. On the basis of the outcome of infection and patterns of CD4+ T cell loss and viral load, disease was classified into four patterns: acute, subacute, chronic, and nonprogressive infection. During the study period, 15 of the 20 animals developed fatal disease, including AIDS, encephalitis, pneumonia, and severe anemia. Opportunistic pathogens identified in these animals included Pneumocystis, cytomegalovirus, Cryptosporidium, Toxoplasma, and Candida. No single parameter by itself predicted outcome, although a combination of low CD4+ T cell counts in blood, high plasma virus levels, and presence of autoantibodies to red blood cells reliably predicted a fatal outcome. Five animals (25%) died within 3 months of inoculation and constituted the group with acute disease, whereas the nine animals (45%) with subacute disease died between 3 and 8 months postinoculation. This 70% mortality within 8 months is significantly shorter than in HIV-1-infected human beings, of whom 70% develop fatal disease a decade after infection. SHIV infection in macaques provides a useful model with which to evaluate antiviral strategies, combining all the advantages of the SIVmac system, yet using a virus bearing the envelope gene of HIV-1.
Assuntos
Síndrome da Imunodeficiência Adquirida/etiologia , Modelos Animais de Doenças , HIV-1/genética , Macaca mulatta/virologia , Macaca nemestrina/virologia , Vírus Reordenados/genética , Vírus da Imunodeficiência Símia/genética , Infecções Oportunistas Relacionadas com a AIDS/imunologia , Infecções Oportunistas Relacionadas com a AIDS/patologia , Anemia Hemolítica Autoimune/sangue , Anemia Hemolítica Autoimune/imunologia , Anemia Hemolítica Autoimune/metabolismo , Animais , Anticorpos Antivirais/sangue , Linfócitos T CD4-Positivos , Teste de Coombs , Anticorpos Anti-HIV/sangue , HIV-1/imunologia , Humanos , Contagem de Linfócitos , Macaca mulatta/genética , Macaca mulatta/imunologia , Macaca nemestrina/genética , Macaca nemestrina/imunologia , Vírus Reordenados/imunologia , Vacinas contra a SAIDS/administração & dosagem , Vacinas contra a SAIDS/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/classificação , Síndrome de Imunodeficiência Adquirida dos Símios/mortalidade , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/imunologia , Viremia/sangueRESUMO
Previous reports from our lab had shown that sera obtained from SIV(mac)-infected animals neutralized SIV(mac) infectivity in CD4(+) T cells but failed to protect monkey primary macrophages from infection with the virus. However, the antibodies could inhibit completion of the viral life cycle in the macrophages at the postentry stage(s). In this report we examined the mechanisms of the late effect of the antibodies. Using monoclonal antibodies (MAbs), we demonstrated that only antibodies to the SIV envelope protein (KK17 and KK42) but not antibody to the viral core protein (FA2) had the same inhibitory effect as that of the anti-SIV sera. To identify the stage of the viral replication cycle that was inhibited by anti-SIV antibodies in macrophages, we used various PCR techniques to study viral entry/reverse transcription (by amplifying the viral gag gene), viral genome nuclear transport (by amplifying 2-LTR circular forms), viral integration (by Alu-PCR assay), and viral protein expression (by RIPA). We found that in macrophage cultures inoculated with SIV(mac)251 that were preincubated with antienvelope MAbs, viral DNA was detected at 8 h postinoculation but the 2-LTR circular forms and integrated viral DNAs were undetectable, and viral proteins were not expressed in these infected macrophages. These results strongly suggested that anti-SIV antibodies inhibited SIV(mac) replication in macrophages by blocking nuclear transport of viral genomes since viral DNA could not be detected in the nuclei of treated cultures. Furthermore, we showed that although viral replication in macrophages was interrupted by the antibodies, when cocultured with permissive T cells, the viral genomes presented in the cytoplasm of the macrophages could readily transfer to T cells during cell-cell contact. Importantly, this transfer could not be prevented by the antibodies. These results might explain the failure of passive antibody immunization against SIV(mac)251--a critical obstacle in AIDS vaccine development.
Assuntos
Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Anticorpos Antivirais/farmacologia , DNA Viral/metabolismo , Estágios do Ciclo de Vida/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Vírus da Imunodeficiência Símia/imunologia , Animais , Anticorpos Monoclonais/farmacologia , Ácido Aspártico Endopeptidases/fisiologia , Comunicação Celular , Células Cultivadas , Genoma Viral , Humanos , Macaca mulatta , Macrófagos/virologia , Testes de Neutralização , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/fisiologia , Linfócitos T/virologia , Proteínas do Envelope Viral/imunologia , Replicação Viral/efeitos dos fármacosRESUMO
We examined plasma from macaques infected with three different phenotypes of SIVmac for their ability to neutralize the infectivity of homologous and heterologous virus in lymphocyte (CEMx174 cells or normal rhesus macaque peripheral blood lymphocytes) or normal rhesus macaque macrophage (Mphi) cultures. Similar to previous findings, we observed that some plasmas failed to neutralize or poorly neutralized the infectivity of SIVmac239 and SIVmac251(<1:20 plasma dilution) in lymphocyte cultures. In contrast, when primary rhesus Mphi cultures were used as the indicator cells, the same plasmas neutralized both viruses at high dilutions (1:200 to 1:20,000). Neutralization of virus infectivity by the various plasmas was confirmed by SIV core antigen capture assays. We excluded the possibility that this differential neutralization in Mphi was related to the differences in the ability of the virus strain to replicate in these two cell types by demonstrating that the replication efficiency of SIVmac251 in CEMx174 cells, PBMC, and Mphi cultures was very similar. The role of Fc receptors on the Mphi surface in the clearance of the virus-antibody complexes was also excluded since similar neutralizing results were obtained using whole plasmas, purified IgG antibodies, and purified Fab fragments derived from the IgG fraction of these plasmas. The mechanism of virus neutralization in Mphi does not appear to involve blocking of virus entry into the cells since radiolabeled virus reacted with anti-SIV antibodies was taken up by rhesus Mphi as efficiently as virus reacted with normal antibody. DNA of the neutralized virus was identified in the Mphi cultures, but virus replication, as evidenced by accumulation of viral protein products, was not detectable so long as the antibodies were present in the medium. Removal of the antibodies resulted in a resumption of virus replication in the Mphi. These results indicate that virus infectivity can be efficiently neutralized by antibodies in Mphi cultures by a mechanism that is fundamentally different from that in lymphocyte cultures.
Assuntos
Anticorpos Antivirais/imunologia , Linfócitos/virologia , Macrófagos/virologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Vírus da Imunodeficiência Símia/imunologia , Animais , Anticorpos Antivirais/sangue , Complexo Antígeno-Anticorpo/imunologia , Complexo Antígeno-Anticorpo/metabolismo , Linhagem Celular , Meios de Cultura , Humanos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologia , Linfócitos/imunologia , Macaca , Macrófagos/imunologia , Testes de Neutralização , Receptores Fc/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/sangue , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/crescimento & desenvolvimento , Especificidade da Espécie , Linfócitos T/imunologia , Linfócitos T/virologia , Replicação ViralRESUMO
We have examined both the sequence changes in the LTR, gag, vif, vpr, vpx, tat, rev, vpu, env, and nef genes and the cell tropism of a cell-free stock of chimeric simian-human immunodeficiency virus (SHIV) isolated from the cerebrospinal fluid of a pig-tailed macaque (PNb) that developed AIDS. This virus (SHIVKU-1) is highly pathogenic when inoculated into other macaques. DNA sequence analysis of PCR-amplified products revealed a total of 5 nucleotide changes in the LTR while vif had 2 consensus amino acid changes. The gag, vif, and vpx had no consensus amino acid substitutions, whereas vpr had 1 consensus substitution. The tat and rev genes of the HXB2 region of SHIVKU-1 had 2 and 1 consensus amino acid changes, respectively. The vpu gene of the HXB2 region of SHIV, which originally had an ACG at the beginning of the gene, reverted to an initiation ATG codon and in addition contained a consensus amino acid substitution at position 69 of this protein. As expected, the majority of the nucleotide substitutions were found in the env and nef genes. Thirteen and 5 amino acid changes were predicted for the corresponding Env and Nef proteins, respectively. In addition, one-third of the env gene clones isolated from the SHIVKU-1 stock had a 5-amino-acid deletion in the V4 region. Using three independent assays, we determined that the changes in the SHIVKU-1 were associated with an increase in the efficiency of replication in macrophages. The strikingly few consensus changes in the virus suggest that conversion of this virus to one capable of causing AIDS in pig-tailed macaques was associated with relatively few changes in the viral envelope and/or accessory genes. These results will provide the basis for the development of a pathogenic, molecular clone of SHIV capable of causing AIDS in pig-tailed macaques.
Assuntos
HIV-1/genética , Síndrome de Imunodeficiência Adquirida dos Símios/microbiologia , Vírus da Imunodeficiência Símia/genética , Síndrome da Imunodeficiência Adquirida/microbiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Quimera , Clonagem Molecular , Genes Virais , HIV-1/patogenicidade , Macaca nemestrina/microbiologia , Macrófagos/microbiologia , Dados de Sequência Molecular , Vírus da Imunodeficiência Símia/patogenicidade , Especificidade da Espécie , Linfócitos T/microbiologia , Proteínas Estruturais Virais/genética , Replicação ViralRESUMO
SIVsmmPBj14 is a highly pathogenic lentivirus which causes acute diarrhea, rash, massive lymphocyte proliferation predominantly in the gastrointestinal tract, and death within 7 to 14 days. In cell culture, the virus has mitogenic effects on resting macaque T lymphocytes. In contrast, SIVmac239 causes AIDS in rhesus macaques, generally within 2 years after inoculation. In a previous study, replacement of amino acid residues 17 and 18 of the Nef protein of SIVmac239 with the corresponding amino acid residues of the Nef protein of SIVsmmPBj14 yielded a PBj-like virus that caused extensive activation of resting T lymphocytes in cultures and acute PBj-like disease when inoculated into pig-tailed macaques. This study suggested that nef played a major role in both processes. In this study, we replaced the nef/long terminal repeat (LTR) region of a nonpathogenic simian-human immunodeficiency virus (SHIV), SHIVPPc, with the corresponding region from SIVsmmPBj14 and examined the biological properties of the resultant virus. Like SIVsmmPBj14, SHIVPPcPBjnef caused massive stimulation of resting peripheral blood mononuclear cells (PBMC), which then produced virus in the absence of extraneous interleukin 2. However, when inoculated into macaques, the virus failed to replicate productively or cause disease. Thus, while these results confirmed that the nef/LTR region of SIVsmmPBj14 played a major role in the activation of resting PBMC, duplication of the cellular activation process in macaques may require a further interaction between nef and the envelope glycoprotein of simian immunodeficiency virus because SHIV, containing the envelope of human immunodeficiency virus type 1, failed to cause activation in vivo.