RESUMO
Gαs-coupled receptors signaling through cAMP provide a key mechanism for the sensitization of nociceptive sensory neurons, and the cAMP effector Epac has been implicated in the transition from acute to chronic pain. Epac exerts its effects through Rap1 and protein kinase C (PKC). To identify targets of Epac-PKC signaling in sensory neurons of the mouse dorsal root ganglion (DRG), we profiled PKC substrate proteins phosphorylated in response to the activation of Epac with the proinflammatory prostaglandin E2 (PGE2). A prominent Epac-dependent phospho-protein band induced by PGE2 was identified by mass spectrometry as the mitochondrial enzyme pyruvate dehydrogenase (Pdha1). In dissociated DRG from both males and females, the recruitment of Pdha1 to phospho-protein fractions was rapidly induced by PGE2 and prevented by selective inhibition of Epac2. Epac activation increased mitochondrial respiration, consistent with an increase in Pdha1 function mediated by Epac2. Hindpaw injection of PGE2 induced heat hyperalgesia in males and females, but Pdha1 phosphorylation occurred only in males. Hyperalgesia was attenuated in males but not in females by systemic inhibition of Epac2, and also by a mitochondrial membrane potential uncoupler, dinitrophenol, supporting a role for mitochondrial regulation in acute hyperalgesia. These findings identify a mechanism for the regulation of mitochondrial function by Epac2 that contributes to acute inflammatory hyperalgesia in male mice. Systemic administration of the cyclooxygenase 2 inhibitor celecoxib suppressed both PGE2-induced heat hyperalgesia and Pdha1 phosphorylation in DRG of males but not females, suggesting that prostaglandin synthesis within the DRG mediates the phosphorylation of Pdha1 in response to hindpaw insult.SIGNIFICANCE STATEMENT There has been extensive investigation of mitochondrial dysfunction as a causative factor in neuropathic pain disorders. In contrast, results reported here implicate enhanced mitochondrial function as a contributing factor in the development of acute inflammatory hyperalgesia. We describe a mechanism in which Epac2 activation by prostaglandin receptors leads to phosphorylation of pyruvate dehydrogenase and an increase in mitochondrial respiration in peripheral sensory neurons. Although Epac2 activation leads to Pdha1 (pyruvate dehydrogenase) phosphorylation in dissociated neurons from mice of both sexes, induction of this pathway in vivo by hindpaw insult is restricted to males and appears to require intraganglionic prostaglandin synthesis. These findings support a model in which Gs-coupled receptor modulation of mitochondrial function promotes acute nociceptive signaling and inflammatory hyperalgesia.
Assuntos
Gânglios Espinais/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Hiperalgesia/metabolismo , Mitocôndrias/metabolismo , Medição da Dor/métodos , Animais , Inibidores Enzimáticos/farmacologia , Feminino , Gânglios Espinais/efeitos dos fármacos , Inflamação/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/efeitos dos fármacos , Nociceptores/metabolismo , Medição da Dor/efeitos dos fármacos , Piruvato Desidrogenase (Lipoamida)/antagonistas & inibidores , Piruvato Desidrogenase (Lipoamida)/metabolismo , Células Receptoras Sensoriais/efeitos dos fármacos , Células Receptoras Sensoriais/metabolismoRESUMO
Infection with human immunodeficiency virus 1 (HIV-1) results in the dissemination of virus to gut-associated lymphoid tissue. Subsequently, HIV-1 mediates massive depletion of gut CD4+ T cells, which contributes to HIV-1-induced immune dysfunction. The migration of lymphocytes to gut-associated lymphoid tissue is mediated by integrin alpha4beta7. We demonstrate here that the HIV-1 envelope protein gp120 bound to an activated form of alpha4beta7. This interaction was mediated by a tripeptide in the V2 loop of gp120, a peptide motif that mimics structures presented by the natural ligands of alpha4beta7. On CD4+ T cells, engagement of alpha4beta7 by gp120 resulted in rapid activation of LFA-1, the central integrin involved in the establishment of virological synapses, which facilitate efficient cell-to-cell spreading of HIV-1.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Proteína gp120 do Envelope de HIV/metabolismo , Infecções por HIV/imunologia , HIV-1/imunologia , Integrinas/metabolismo , Mucosa Intestinal/imunologia , Linfócitos T CD4-Positivos/virologia , Movimento Celular/imunologia , Células Cultivadas , Fibroblastos/metabolismo , Fibroblastos/virologia , Citometria de Fluxo , Humanos , Mucosa Intestinal/virologia , Células Matadoras Naturais/imunologia , Ligantes , Antígeno-1 Associado à Função Linfocitária/metabolismo , Ligação Proteica/imunologia , Transdução de Sinais/imunologiaRESUMO
CD28 and CTLA-4 are cell surface cosignaling molecules essential for the control of T cell activation upon the engagement of their ligands B7-1 and B7-2 from antigen-presenting cells. By employing a receptor array assay, we have demonstrated that B7-H2, best known as the ligand of inducible costimulator, was a ligand for CD28 and CTLA-4 in human, whereas these interactions were not conserved in mouse. B7-H2 and B7-1 or B7-2 interacted with CD28 through distinctive domains. B7-H2-CD28 interaction was essential for the costimulation of human T cells' primary responses to allogeneic antigens and memory recall responses. Similar to B7-1 and B7-2, B7-H2 costimulation via CD28 induced survival factor Bcl-xL, downregulated cell cycle inhibitor p27(kip1), and triggered signaling cascade of ERK and AKT kinase-dependent pathways. Our findings warrant re-evaluation of CD28 and CTLA-4's functions previously attributed exclusively to B7-1 and B7-2 and have important implications in therapeutic interventions against human diseases.
Assuntos
Antígenos CD/imunologia , Antígenos CD28/imunologia , Animais , Antígenos CD/química , Antígenos de Diferenciação de Linfócitos T/imunologia , Sítios de Ligação , Antígenos CD28/química , Antígeno CTLA-4 , Linhagem Celular , Proliferação de Células , Humanos , Ligante Coestimulador de Linfócitos T Induzíveis , Proteína Coestimuladora de Linfócitos T Induzíveis , Ligantes , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Modelos Moleculares , Estrutura Quaternária de Proteína , Linfócitos T/citologia , Linfócitos T/imunologiaRESUMO
Exchange protein directly activated by cAMP (Epac) and protein kinase A are effectors for cAMP with distinct actions and regulatory mechanisms. Epac is a Rap guanine nucleotide exchange factor that activates Rap1; protein kinase C (PKC) is a major downstream target of Epac-Rap1 signaling that has been implicated in a variety of pathophysiological processes, including cardiac hypertrophy, cancer, and nociceptor sensitization leading to chronic pain. Despite the implication of both Epac and PKC in these processes, few downstream targets of Epac-PKC signaling have been identified. This study characterized the regulation of PKC activity downstream of Epac activation. Using an antibody that recognizes phospho-serine residues within the consensus sequence phosphorylated by PKC, we analyzed the 1-dimensional banding profile of PKC substrate protein phosphorylation from the Neuro2A mouse neuroblastoma cell line. Activation of Epac either indirectly by prostaglandin PGE2, or directly by 8-pCPT-2-O-Me-cAMP-AM (8pCpt), produced distinct PKC phospho-substrate protein bands that were suppressed by co-administration of the Epac inhibitor ESI09. Different PKC isoforms contributed to the induction of individual phospho-substrate bands, as determined using isoform-selective PKC inhibitors. Moreover, the banding profile after Epac activation was altered by disruption of the cytoskeleton, suggesting that the orchestration of Epac-dependent PKC signaling is regulated in part by interactions with the cytoskeleton. The approach described here provides an effective means to characterize Epac-dependent PKC activity.
Assuntos
Citoesqueleto/enzimologia , Dinoprostona/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteína Quinase C/metabolismo , Transdução de Sinais , Proteínas rap1 de Ligação ao GTP/metabolismo , Animais , Linhagem Celular Tumoral , Citoesqueleto/genética , Fatores de Troca do Nucleotídeo Guanina/genética , Camundongos , Fosforilação , Proteína Quinase C/genética , Proteínas rap1 de Ligação ao GTP/genéticaRESUMO
Mucosal HIV-1 transmission is inefficient. However, certain viral and host characteristics may play a role in facilitating HIV acquisition and systemic expansion. Cells expressing high levels of integrin α4ß7 have been implicated in favoring the transmission process and the infusion of an anti-α4ß7 mAb (RM-Act-1) prior to, and during a repeated low-dose vaginal challenge (RLDC) regimen with SIVmac251 reduced SIV acquisition and protected the gut-associated lymphoid tissues (GALT) in the macaques that acquired SIV. α4ß7 expression is required for lymphocyte trafficking to the gut lamina propria and gut inductive sites. Several therapeutic strategies that target α4ß7 have been shown to be effective in treating inflammatory conditions of the intestine, such as inflammatory bowel disease (IBD). To determine if blocking α4ß7 with ELN, an orally available anti-α4 small molecule, would inhibit SHIV-SF162P3 acquisition, we tested its ability to block MAdCAM-1 (α4ß7 natural ligand) and HIV-gp120 binding in vitro. We studied the pharmacokinetic profile of ELN after oral and vaginal delivery in macaques. Twenty-six macaques were divided into 3 groups: 9 animals were treated with ELN orally, 9 orally and vaginally and 8 were used as controls. All animals were challenged intra-vaginally with SHIV-SF162P3 using the RLDC regimen. We found that ELN did not protect macaques from SHIV acquisition although it reduced the SHIV-induced inflammatory status during the acute phase of infection. Notably, integrins can exist in different activation states and, comparing the effect of ELN and the anti-α4ß7 mAb RM-Act-1 that reduced susceptibility to SIV infection, we determined that ELN induces the active conformation of α4ß7, while RM-Act-1 inhibits its activation through an allosteric mechanism. These results suggest that inhibition of α4ß7 activation may be necessary to reduce susceptibility to SIV/SHIV infection and highlight the complexity of anti-integrins therapeutic approach in HIV as well as in IBD and other autoimmune diseases.
Assuntos
Antivirais/farmacologia , Imunoglobulinas/metabolismo , Integrinas/antagonistas & inibidores , Mucoproteínas/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/transmissão , Animais , Anticorpos Monoclonais Humanizados/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Moléculas de Adesão Celular , Células Cultivadas , Feminino , Citometria de Fluxo , Humanos , Macaca mulatta , Mucosa/virologia , Vírus da Imunodeficiência Símia , Vagina/virologia , Carga ViralRESUMO
[This corrects the article DOI: 10.1371/journal.ppat.1005720.].
RESUMO
The tissue microenvironment shapes the characteristics and functions of dendritic cells (DCs), which are important players in HIV infection and dissemination. Notably, DCs in the gut have the daunting task of orchestrating the balance between immune response and tolerance. They produce retinoic acid (RA), which imprints a gut-homing phenotype and influences surrounding DCs. To investigate how the gut microenvironment impacts the ability of DCs to drive HIV infection, we conditioned human immature monocyte-derived DCs (moDCs) with RA (RA-DCs), before pulsing them with HIV and mixing them with autologous T cells. RA-DCs showed a semimature, mucosal-like phenotype and released higher amounts of TGF-ß1 and CCL2. Using flow cytometry, Western blot, and microscopy, we determined that moDCs express the cell adhesion molecule mucosal vascular addressin cell adhesion molecule-1 (MAdCAM-1) and that RA increases its expression. MAdCAM-1 was also detected on a small population of DCs in rhesus macaque (Macaca mulata) mesenteric lymph node. RA-DCs formed more DC-T cell conjugates and promoted significantly higher HIV replication in DC-T cell mixtures compared with moDCs. This correlated with the increase in MAdCAM-1 expression. Blocking MAdCAM-1 partially inhibited the enhanced HIV replication. In summary, RA influences DC phenotype, increasing their ability to exacerbate HIV infection. We describe a previously unknown mechanism that may contribute to rapid HIV spread in the gut, a major site of HIV replication after mucosal exposure.
Assuntos
Células Dendríticas/efeitos dos fármacos , Infecções por HIV/imunologia , HIV-1/imunologia , Mucosa Intestinal/efeitos dos fármacos , Linfócitos T/efeitos dos fármacos , Tretinoína/farmacologia , Animais , Moléculas de Adesão Celular , Quimiocina CCL2/biossíntese , Quimiocina CCL2/metabolismo , Técnicas de Cocultura , Células Dendríticas/imunologia , Células Dendríticas/patologia , Células Dendríticas/virologia , Feminino , Expressão Gênica , Infecções por HIV/genética , Infecções por HIV/patologia , Infecções por HIV/virologia , Humanos , Imunoglobulinas/genética , Imunoglobulinas/imunologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/patologia , Mucosa Intestinal/virologia , Linfonodos/imunologia , Linfonodos/patologia , Linfonodos/virologia , Macaca mulatta , Mucoproteínas/genética , Mucoproteínas/imunologia , Fenótipo , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Linfócitos T/imunologia , Linfócitos T/patologia , Linfócitos T/virologia , Fator de Crescimento Transformador beta1/biossíntese , Fator de Crescimento Transformador beta1/metabolismo , Tretinoína/imunologia , Replicação ViralRESUMO
The availability of highly susceptible HIV target cells that can rapidly reach the mucosal lymphoid tissues may increase the chances of an otherwise rare transmission event to occur. Expression of α4ß7 is required for trafficking of immune cells to gut inductive sites where HIV can expand and it is expressed at high level on cells particularly susceptible to HIV infection. We hypothesized that HSV-2 modulates the expression of α4ß7 and other homing receptors in the vaginal tissue and that this correlates with the increased risk of HIV acquisition in HSV-2 positive individuals. To test this hypothesis we used an in vivo rhesus macaque (RM) model of HSV-2 vaginal infection and a new ex vivo model of macaque vaginal explants. In vivo we found that HSV-2 latently infected RMs appeared to be more susceptible to vaginal SHIVSF162P3 infection, had higher frequency of α4ß7high CD4+ T cells in the vaginal tissue and higher expression of α4ß7 and CD11c on vaginal DCs. Similarly, ex vivo HSV-2 infection increased the susceptibility of the vaginal tissue to SHIVSF162P3. HSV-2 infection increased the frequencies of α4ß7high CD4+ T cells and this directly correlated with HSV-2 replication. A higher amount of inflammatory cytokines in vaginal fluids of the HSV-2 infected animals was similar to those found in the supernatants of the infected explants. Remarkably, the HSV-2-driven increase in the frequency of α4ß7high CD4+ T cells directly correlated with SHIV replication in the HSV-2 infected tissues. Our results suggest that the HSV-2-driven increase in availability of CD4+ T cells and DCs that express high levels of α4ß7 is associated with the increase in susceptibility to SHIV due to HSV-2. This may persists in absence of HSV-2 shedding. Hence, higher availability of α4ß7 positive HIV target cells in the vaginal tissue may constitute a risk factor for HIV transmission.
Assuntos
Coinfecção/virologia , Suscetibilidade a Doenças/fisiopatologia , Infecções por HIV/fisiopatologia , Herpes Genital/complicações , Herpesvirus Humano 2/fisiologia , Integrinas/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/fisiopatologia , Animais , Antígeno CD11c/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Coinfecção/patologia , Coinfecção/fisiopatologia , Células Dendríticas/metabolismo , Células Dendríticas/patologia , Modelos Animais de Doenças , Suscetibilidade a Doenças/metabolismo , Feminino , HIV/isolamento & purificação , HIV/fisiologia , Infecções por HIV/metabolismo , Infecções por HIV/patologia , Herpes Genital/metabolismo , Herpes Genital/fisiopatologia , Herpesvirus Humano 2/isolamento & purificação , Macaca mulatta , Síndrome de Imunodeficiência Adquirida dos Símios/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/patologia , Vírus da Imunodeficiência Símia/isolamento & purificação , Vírus da Imunodeficiência Símia/fisiologia , Regulação para Cima , Vagina/metabolismo , Vagina/patologia , Vagina/virologiaRESUMO
Chemotherapy is often a life-saving treatment, but the development of intractable pain caused by chemotherapy-induced peripheral neuropathy (CIPN) is a major dose-limiting toxicity that restricts cancer survival rates. Recent reports demonstrate that paclitaxel (PTX) robustly increases anti-inflammatory CD4+ T cells in the dorsal root ganglion (DRG), and that T cells and anti-inflammatory cytokines are protective against CIPN. However, the mechanism by which CD4+ T cells are activated, and the extent cytokines released by CD4+ T cells target DRG neurons are unknown. Here, we are the first to detect major histocompatibility complex II (MHCII) protein in mouse DRG neurons and to find CD4+ T cells breaching the satellite glial cell barrier to be in close proximity to neurons, together suggesting CD4+ T cell activation and targeted cytokine release. MHCII protein is primarily expressed in small nociceptive neurons in male and female mouse DRG but increased after PTX in small nociceptive neurons in only female DRG. Reducing one copy of MHCII in small nociceptive neurons decreased anti-inflammatory IL-10 and IL-4 producing CD4+ T cells in naïve male DRG and increased their hypersensitivity to cold. Administration of PTX to male and female mice that lacked one copy of MHCII in nociceptive neurons decreased anti-inflammatory CD4+ T cells in the DRG and increased the severity of PTX-induced cold hypersensitivity. Collectively, our results demonstrate expression of MHCII protein in mouse DRG neurons, which modulates cytokine producing CD4+ T cells in the DRG and attenuates cold hypersensitivity during homeostasis and after PTX treatment.
Assuntos
Síndromes Periódicas Associadas à Criopirina , Paclitaxel , Doenças do Sistema Nervoso Periférico , Ratos , Camundongos , Masculino , Feminino , Animais , Paclitaxel/toxicidade , Paclitaxel/metabolismo , Gânglios Espinais/metabolismo , Hiperalgesia/etiologia , Ratos Sprague-Dawley , Doenças do Sistema Nervoso Periférico/induzido quimicamente , Citocinas/metabolismo , Neurônios/metabolismo , Anti-Inflamatórios/uso terapêuticoRESUMO
Chemotherapy is often a life-saving treatment, but the development of intractable pain caused by chemotherapy-induced peripheral neuropathy (CIPN) is a major dose-limiting toxicity that restricts survival rates. Recent reports demonstrate that paclitaxel (PTX) robustly increases anti-inflammatory CD4+ T cells in the dorsal root ganglion (DRG), and that T cells and anti-inflammatory cytokines are protective against CIPN. However, the mechanism by which CD4+ T cells are activated, and the extent cytokines released by CD4+ T cells target DRG neurons are unknown. Here, we found novel expression of functional major histocompatibility complex II (MHCII) protein in DRG neurons, and CD4+ T cells in close proximity to DRG neurons, together suggesting CD4+ T cell activation and targeted cytokine release. MHCII protein is primarily expressed in small nociceptive neurons in male mouse DRG regardless of PTX, while MHCII is induced in small nociceptive neurons in female DRG after PTX. Accordingly, reducing MHCII in small nociceptive neurons increased hypersensitivity to cold only in naive male mice, but increased severity of PTX-induced cold hypersensitivity in both sexes. Collectively, our results demonstrate expression of MHCII on DRG neurons and a functional role during homeostasis and inflammation.
RESUMO
Both activated and resting CD4(+) T cells in mucosal tissues play important roles in the earliest phases of infection after sexual transmission of HIV-1, a process that is inefficient. HIV-1 gp120 binds to integrin alpha(4)beta(7) (alpha(4)beta(7)), the gut mucosal homing receptor. We find that alpha(4)beta(7)(high) CD4(+) T cells are more susceptible to productive infection than are alpha(4)beta(7)(low-neg) CD4(+) T cells in part because this cellular subset is enriched with metabolically active CD4(+) T cells. alpha(4)beta(7)(high) CD4(+) T cells are CCR5(high) and CXCR4(low); on these cells, alpha(4)beta(7) appears in a complex with CD4. The specific affinity of gp120 for alpha(4)beta(7) provides a mechanism for HIV-1 to target activated cells that are critical for efficient virus propagation and dissemination following sexual transmission.
Assuntos
Antígenos CD4/imunologia , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Integrinas/imunologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/virologia , Anticorpos Monoclonais/farmacologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Membrana Celular/efeitos dos fármacos , Membrana Celular/imunologia , Membrana Celular/virologia , Feminino , Genitália Feminina/efeitos dos fármacos , Genitália Feminina/imunologia , HIV-1/efeitos dos fármacos , HIV-1/fisiologia , Humanos , Imunoprecipitação , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/imunologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Receptores CCR5/metabolismo , Subpopulações de Linfócitos T/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
Chemotherapy is often dose limiting due to the emergence of a debilitating neuropathy. IL-10 and IL-4 are protective against peripheral neuropathy, yet the contribution by CD4+ T cells is unknown. Using flow cytometry, we found that naïve females had a greater frequency of anti-inflammatory CD4+ T cells in the dorsal root ganglion (DRG) compared to males. In response to paclitaxel, females had reduced mechanical hypersensitivity and a greater frequency of anti-inflammatory CD4+ T cells (FoxP3, IL-10, IL-4) in the DRG than male and ovariectomized female mice. These findings support a model in which estrogen promotes anti-inflammatory CD4+ T cells in female DRG to suppress peripheral neuropathy.
Assuntos
Gânglios Espinais , Doenças do Sistema Nervoso Periférico , Animais , Anti-Inflamatórios/uso terapêutico , Linfócitos T CD4-Positivos , Feminino , Humanos , Interleucina-10/uso terapêutico , Interleucina-4 , Masculino , Camundongos , Ovariectomia , Paclitaxel/toxicidade , Doenças do Sistema Nervoso Periférico/induzido quimicamente , RatosRESUMO
Zika virus (ZIKV) recently emerged in the Western Hemisphere with previously unrecognized or unreported clinical presentations. Here, we identify two putative binding mechanisms of ancestral and emergent ZIKV strains featuring the envelope (E) protein residue asparagine 154 (ASN154) and viral phosphatidylserine (PS). Synthetic peptides representing the region containing ASN154 from strains PRVABC59 (Puerto Rico 2015) and MR_766 (Uganda 1947) were exposed to neuronal cells and fibroblasts to model ZIKV E protein/cell interactions and bound MDCK or Vero cells and primary neurons significantly. Peptides significantly inhibited Vero cell infectivity by ZIKV strains MR_766 and PRVABC59, indicating that this region represents a putative binding mechanism of ancestral African ZIKV strains and emergent Western Hemisphere strains. Pretreatment of ZIKV strains MR_766 and PRVABC59 with the PS-binding protein annexin V significantly inhibited replication of PRVABC59 but not MR_766, suggesting that Western hemisphere strains may additionally be capable of utilizing PS-mediated entry to infect host cells. These data indicate that the region surrounding E protein ASN154 is capable of binding fibroblasts and primary neuronal cells and that PS-mediated entry may be a secondary mechanism for infectivity utilized by Western Hemisphere strains.
Assuntos
Proteínas do Envelope Viral/metabolismo , Infecção por Zika virus/virologia , Zika virus/fisiologia , Motivos de Aminoácidos , Animais , Asparagina/metabolismo , Chlorocebus aethiops , Cães , Fibroblastos/virologia , Humanos , Células Madin Darby de Rim Canino , Neurônios/virologia , Fosfatidilserinas/metabolismo , Células Vero , Proteínas do Envelope Viral/genética , Ligação ViralRESUMO
Prevalent HSV-2 infection increases the risk of HIV acquisition both in men and women even in asymptomatic subjects. Understanding the impact of HSV-2 on the mucosal microenvironment may help to identify determinants of susceptibility to HIV. Vaginal HSV-2 infection increases the frequency of cells highly susceptible to HIV in the vaginal tissue of women and macaques and this correlates with increased susceptibility to vaginal SHIV infection in macaques. However, the effect of rectal HSV-2 infection on HIV acquisition remains understudied. We developed a model of rectal HSV-2 infection in macaques in combination with rectal SIVmac239Δnef (SIVΔnef) vaccination and our results suggest that rectal HSV-2 infection may increase the susceptibility of macaques to rectal SIVmac239 wild-type (wt) infection even in SIVΔnef-infected animals. Rectal SIVΔnef infection/vaccination protected 7 out of 7 SIVΔnef-infected macaques from SIVmac239wt rectal infection (vs 12 out of 16 SIVΔnef-negative macaques), while 1 out of 3 animals co-infected with SIVΔnef and HSV-2 acquired SIVmac239wt infection. HSV-2/SIVmac239wt co-infected animals had increased concentrations of inflammatory factors in their plasma and rectal fluids and a tendency toward higher acute SIVmac239wt plasma viral load. However, they had higher blood CD4 counts and reduced depletion of CCR5+ CD4+ T cells compared to SIVmac239wt-only infected animals. Thus, rectal HSV-2 infection generates a pro-inflammatory environment that may increase susceptibility to rectal SIV infection and may impact immunological and virological parameters during acute SIV infection. Studies with larger number of animals are needed to confirm these findings.
Assuntos
Produtos do Gene nef/metabolismo , Herpesvirus Humano 2/fisiologia , Reto/virologia , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/fisiologia , Vacinação , Animais , Contagem de Linfócito CD4 , Coinfecção/sangue , Coinfecção/imunologia , Coinfecção/virologia , Citocinas/metabolismo , Humanos , Inflamação/patologia , Linfonodos/patologia , Macaca mulatta , Masculino , Fenótipo , Síndrome de Imunodeficiência Adquirida dos Símios/sangue , Carga Viral/imunologiaRESUMO
Several studies suggest that progesterone and estrogens may affect HIV transmission in different, possibly opposing ways. Nonetheless, a direct comparison of their effects on the mucosal immune system has never been done. We hypothesize that sex hormones might impact the availability of cells and immune factors important in early stages of mucosal transmission, and, in doing so influence the risk of HIV acquisition. To test this hypothesis, we employed 15 ovarectomized rhesus macaques: 5 were treated with Depot Medroxy Progesterone Acetate (DMPA), 6 with 17-ß estradiol (E2) and 4 were left untreated. All animals were euthanized 5 weeks after the initiation of hormone treatment, a time post-DMPA injection associated with high susceptibility to SIV infection. We found that DMPA-treated macaques exhibited higher expression of integrin α4ß7 (α4ß7) on CD4+ T cells, the gut homing receptor and a marker of cells highly susceptible to HIV, in the endocervix than did the E2-treated animals. In contrast, the frequency of CCR5+ CD4+ T cells in DMPA-treated macaques was higher than in the E2-treated group in vaginal tissue, but lower in endocervix. α4ß7 expression on dendritic cells (DCs) was higher in the DMPA-treated group in the endocervical tissue, but lower in vaginal tissue and on blood DCs compared with the E2-treated animals. Soluble MAdCAM-1, the α4ß7 ligand, was present in the vaginal fluids of the control and E2-treated groups, but absent in the fluids from DMPA-treated animals. Both hormones modulated the expression and release of inflammatory factors and modified the distribution of sialomucins in the endocervix. In summary, we found that sex hormones profoundly impact mucosal immune factors that are directly implicated in HIV transmission. The effect is particularly significant in the endocervix. This may increase our understanding of the potential hormone-driven modulation of HIV susceptibility and potentially guide contraceptive policies in high-risk settings.
Assuntos
Colo do Útero/metabolismo , Endométrio/metabolismo , Infecções por HIV/transmissão , Acetato de Medroxiprogesterona/farmacologia , Mucosa/metabolismo , Animais , Linfócitos T CD4-Positivos/metabolismo , Colo do Útero/efeitos dos fármacos , Quimiocina CCL21/genética , Quimiocina CCL21/metabolismo , Quimiocina CCL4/genética , Quimiocina CCL4/metabolismo , Endométrio/efeitos dos fármacos , Estradiol/farmacologia , Estrogênios/farmacologia , Estrogênios/fisiologia , Feminino , Expressão Gênica , Integrina alfa4/genética , Integrina alfa4/metabolismo , Cadeias beta de Integrinas/genética , Cadeias beta de Integrinas/metabolismo , Macaca mulatta , Mucinas/genética , Mucinas/metabolismo , Mucosa/efeitos dos fármacos , Receptores CCR5/genética , Receptores CCR5/metabolismoRESUMO
BACKGROUND: Integrin α4ß7(high) (α4ß7(high)) mediates the homing of CD4⺠T cells to gut-associated lymphoid tissues, which constitute a highly favorable environment for HIV expansion and dissemination. HIV and simian immunodeficiency virus (SIV) envelope proteins bind to and signal through α4ß7(high) and during acute infection SIV preferentially infects α4ß7(high) CD4⺠T cells. We postulated that the availability of these cells at the time of challenge could influence mucosal SIV transmission and acute viral load (VL). METHODS: We challenged 17 rhesus macaques with 3000 TCID50 of SIVmac239 rectally and followed the subsets of α4ß7(high) T cells and dendritic cells (DCs) by flow cytometry in blood and tissues, before and after challenge. RESULTS: We found that the frequency of memory CD4⺠T cells that expressed high levels of α4ß7(high) (α4ß7(high) memory CD4⺠T cells) in blood before challenge correlated strongly with susceptibility to infection and acute VL. Notably, not only at the time of challenge but also their frequency 3 weeks before challenge correlated with infection. This association extended to the rectal tissue as we observed a strong direct correlation between the frequency of α4ß7(high) memory CD4⺠T cells in blood and rectum before and after challenge. The frequency of α4ß7 myeloid DCs and α4ß7(high) CD80⺠DCs also correlated with infection and acute VL, whereas blood CCR5⺠and CD69⺠CD4⺠T cells could not be associated with infection. CONCLUSIONS: Our results suggest that animals with higher frequency of α4ß7(high) CD4⺠T cells in circulation and in rectal tissue could be more susceptible to SIV rectal transmission.
Assuntos
Linfócitos T CD4-Positivos/classificação , Linfócitos T CD4-Positivos/fisiologia , Integrinas/metabolismo , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Vírus da Imunodeficiência Símia , Animais , Suscetibilidade a Doenças , Citometria de Fluxo , Memória Imunológica/fisiologia , Macaca mulatta , Masculino , RetoRESUMO
Plasmacytoid dendritic cells (pDCs) play a central role in innate and adaptive immune responses against viral infections. pDCs secrete type I IFNs and proinflammatory cytokines upon stimulation by either TLR7 or TLR9. Throughout the course of HIV infection, the production of type-I IFNs is profoundly impaired, and total pDC cell counts in peripheral blood correlates inversely with viral load and positively with CD4(+) T cell count. The origin of these defects is unclear. pDCs express CD4, CCR5, and CXCR4, the primary receptor and coreceptors, respectively, for the HIV envelope; yet little is known concerning the effects of the viral envelope on these cells. Here, we show that exposure of pDCs to gp120 results in the suppression of activation of these cells. This suppression is specific for TLR9-mediated responses, because TLR7-mediated responses are unaffected by gp120. gp120 also suppressed TLR9-mediated induction of proinflammatory cytokines and expression of CD83, a marker of DC activation. Finally, gp120 suppressed pDC-induced cytolytic activity of natural killer cells. Taken together, these data demonstrate that the direct interaction of HIV-1 gp120 with pDCs interferes with TLR9 activation resulting in a decreased ability of pDCs to secrete antiviral and inflammatory factors that play a central role in initiating host immune responses against invading pathogens.