RESUMO
Patients infected with SARS-CoV-2 experience variable disease susceptibility, and patients with comorbidities such as sepsis are often hospitalized for COVID-19 complications. However, the extent to which initial infectious inoculum dose determines disease outcomes and whether this can be used for immunological priming in a genetically susceptible host has not been completely defined. We used an established SARS-like murine model in which responses to primary and/or secondary challenges with murine hepatitis virus type 1 (MHV-1) were analyzed. We compared the response to infection in genetically susceptible C3H/HeJ mice, genetically resistant C57BL/6J mice, and genetically diverse, variably susceptible outbred Swiss Webster mice. Although defined as genetically susceptible to MHV-1, C3H/HeJ mice displayed decreasing dose-dependent pathological changes in disease severity and lung infiltrate/edema, as well as lymphopenia. Importantly, an asymptomatic dose (500 PFU) was identified that yielded no measurable morbidity/mortality postinfection in C3H/HeJ mice. Polymicrobial sepsis induced via cecal ligation and puncture converted asymptomatic infections in C3H/HeJ and C57BL/6J mice to more pronounced disease, modeling the impact of sepsis as a comorbidity to ß-coronavirus infection. We then used low-dose infection as an immunological priming event in C3H/HeJ mice, which provided neutralizing Ab-dependent, but not circulating CD4/CD8 T cell-dependent, protection against a high-dose MHV-1 early rechallenge. Together, these data define how infection dose, immunological status, and comorbidities modulate outcomes of primary and secondary ß-coronavirus infections in hosts with variable susceptibility.
Assuntos
Vírus da Hepatite Murina , Sepse , Humanos , Camundongos , Animais , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos C3H , Camundongos Endogâmicos , Predisposição Genética para DoençaRESUMO
Long-lasting sepsis-induced immunoparalysis has been principally studied in primary (1°) memory CD8 T cells; however, the impact of sepsis on memory CD8 T cells with a history of repeated cognate Ag encounters is largely unknown but important in understanding the role of sepsis in shaping the pre-existing memory CD8 T cell compartment. Higher-order memory CD8 T cells are crucial in providing immunity against common pathogens that reinfect the host or are generated by repeated vaccination. In this study, we analyzed peripheral blood from septic patients and show that memory CD8 T cells with defined Ag specificity for recurring CMV infection proliferate less than bulk populations of central memory CD8 T cells. Using TCR-transgenic T cells to generate 1° and higher-order (quaternary [4°]) memory T cells within the same host, we demonstrate that the susceptibility and loss of both memory subsets are similar after sepsis induction, and sepsis diminished Ag-dependent and -independent (bystander) functions of these memory subsets equally. Both the 1° and 4° memory T cell populations proliferated in a sepsis-induced lymphopenic environment; however, due to the intrinsic differences in baseline proliferative capacity, expression of receptors (e.g., CD127/CD122), and responsiveness to homeostatic cytokines, 1° memory T cells become overrepresented over time in sepsis survivors. Finally, IL-7/anti-IL-7 mAb complex treatment early after sepsis induction preferentially rescued the proliferation and accumulation of 1° memory T cells, whereas recovery of 4° memory T cells was less pronounced. Thus, inefficient recovery of repeatedly stimulated memory cells after polymicrobial sepsis induction leads to changes in memory T cell pool composition, a notion with important implications in devising strategies to recover the number and function of pre-existing memory CD8 T cells in sepsis survivors.
Assuntos
Linfopenia , Sepse , Humanos , Animais , Camundongos , Células T de Memória , Linfócitos T CD8-Positivos , Citocinas/metabolismo , Linfopenia/metabolismo , Memória Imunológica , Camundongos Endogâmicos C57BLRESUMO
Sepsis reduces the number and function of memory CD8 T cells within the host, contributing to the long-lasting state of immunoparalysis. Interestingly, the relative susceptibility of memory CD8 T cell subsets to quantitative/qualitative changes differ after cecal ligation and puncture (CLP)-induced sepsis. Compared with circulatory memory CD8 T cells (TCIRCM), moderate sepsis (0-10% mortality) does not result in numerical decline of CD8 tissue-resident memory T cells (TRM), which retain their "sensing and alarm" IFN-γ-mediated effector function. To interrogate this biologically important dichotomy, vaccinia virus-immune C57BL/6 (B6) mice containing CD8 TCIRCM and skin TRM underwent moderate or severe (â¼50% mortality) sepsis. Severe sepsis led to increased morbidity and mortality characterized by increased inflammation compared with moderate CLP or sham controls. Severe CLP mice also displayed increased vascular permeability in the ears. Interestingly, skin CD103+ CD8 TRM, detected by i.v. exclusion or two-photon microscopy, underwent apoptosis and subsequent numerical loss following severe sepsis, which was not observed in mice that experienced moderate CLP or sham surgeries. Consequently, severe septic mice showed diminished CD8 T cell-mediated protection to localized skin reinfection. Finally, the relationship between severity of sepsis and demise in circulatory versus tissue-embedded memory CD8 T cell populations was confirmed by examining tumor-infiltrating and nonspecific CD8 T cells in B16 melanoma tumors. Thus, sepsis can differentially affect the presence and function of Ag-specific CD8 T cells that reside inside tissues/tumors depending on the severity of the insult, a notion with direct relevance to sepsis survivors and their ability to mount protective memory CD8 T cell-dependent responses to localized Ag re-encounter.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Coriomeningite Linfocítica/imunologia , Vírus da Coriomeningite Linfocítica/fisiologia , Sepse/imunologia , Subpopulações de Linfócitos T/imunologia , Animais , Circulação Sanguínea , Células Cultivadas , Progressão da Doença , Humanos , Memória Imunológica , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Especificidade de ÓrgãosRESUMO
CRISPR/Cas9 technology has revolutionized rapid and reliable gene editing in cells. Although many cell types have been subjected to CRISPR/Cas9-mediated gene editing, there is no evidence of success in genetic alteration of Ag-experienced memory CD8 T cells. In this study, we show that CRISPR/Cas9-mediated gene editing in memory CD8 T cells precludes their proliferation after Ag re-encounter in vivo. This defect is mediated by the proapoptotic transcription factor p53, a sensor of DNA damage. Temporarily inhibiting p53 function offers a window of opportunity for the memory CD8 T cells to repair the DNA damage, facilitating robust recall responses on Ag re-encounter. We demonstrate this by functionally altering memory CD8 T cells using CRISPR/Cas9-mediated targeted gene disruption under the aegis of p53siRNA in the mouse model. Our approach thus adapts the CRISPR/Cas9 technology for memory CD8 T cells to undertake gene editing in vivo, for the first time, to our knowledge.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Sistemas CRISPR-Cas , Proliferação de Células/genética , Memória Imunológica/genética , Proteína Supressora de Tumor p53 , Animais , Antígenos/imunologia , Dano ao DNA/genética , Dano ao DNA/imunologia , Camundongos , Camundongos Transgênicos , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/imunologiaRESUMO
Protective lung tissue-resident memory CD8+T cells (Trm) form after influenza A virus (IAV) infection. We show that IAV infection of mice generates CD69+CD103+and other memory CD8+T cell populations in lung-draining mediastinal lymph nodes (mLNs) from circulating naive or memory CD8+T cells. Repeated antigen exposure, mimicking seasonal IAV infections, generates quaternary memory (4M) CD8+T cells that protect mLN from viral infection better than 1M CD8+T cells. Better protection by 4M CD8+T cells associates with enhanced granzyme A/B expression and stable maintenance of mLN CD69+CD103+4M CD8+T cells, vs the steady decline of CD69+CD103+1M CD8+T cells, paralleling the durability of protective CD69+CD103+4M vs 1M in the lung after IAV infection. Coordinated upregulation in canonical Trm-associated genes occurs in circulating 4M vs 1M populations without the enrichment of canonical downregulated Trm genes. Thus, repeated antigen exposure arms circulating memory CD8+T cells with enhanced capacity to form long-lived populations of Trm that enhance control of viral infections of the mLN.
Assuntos
Linfócitos T CD8-Positivos , Linfonodos , Animais , Antígenos CD/genética , Antígenos CD/imunologia , Antígenos CD/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Células Cultivadas , Feminino , Vírus da Influenza A/imunologia , Pulmão/citologia , Pulmão/imunologia , Linfonodos/citologia , Linfonodos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Infecções por Orthomyxoviridae/imunologia , Transcriptoma/genéticaRESUMO
Plasmodium sporozoites inoculated by mosquitoes migrate to the liver and infect hepatocytes prior to release of merozoites that initiate symptomatic blood-stage malaria. Plasmodium parasites are thought to be restricted to hepatocytes throughout this obligate liver stage of development, and how liver-stage-expressed antigens prime productive CD8 T cell responses remains unknown. We found that a subset of liver-infiltrating monocyte-derived CD11c+ cells co-expressing F4/80, CD103, CD207, and CSF1R acquired parasites during the liver stage of malaria, but only after initial hepatocyte infection. These CD11c+ cells found in the infected liver and liver-draining lymph nodes exhibited transcriptionally and phenotypically enhanced antigen-presentation functions and primed protective CD8 T cell responses against Plasmodium liver-stage-restricted antigens. Our findings highlight a previously unrecognized aspect of Plasmodium biology and uncover the fundamental mechanism by which CD8 T cell responses are primed against liver-stage malaria antigens.