The ability of long term non progressors to maintain very low levels of HIV/SIV and a healthy state, involves various host genetic and immunological factors. CD8+ non-cytolytic antiviral response ...(CNAR) most likely plays an important role in this regard. In order to gain a deeper insight into this unique phenomenon, the ability of CD8+ T cells to suppress viral replication in vitro was investigated in 16 uninfected, longitudinally in 23 SIV-infected long-term non-progressing (LTNPs), and 10 SIV-infected rhesus macaques with progressing disease. An acute infection assay utilizing CD4+ cells from MHC-mismatched monkeys to avoid cytolytic responses was employed. The study has identified CNAR as a long-term stable activity that inversely correlated with plasma viral load. The activity was also detected in CD8+ cells of uninfected macaques, which indicates that CNAR is not necessarily a virus specific response but increases after SIV-infection. Physical contact between CD4+ and CD8+ cells was mainly involved in mediating viral inhibition. Loss of this activity appeared to be due to a loss of CNAR-expressing CD8+ cells as well as a reduction of CNAR-responsive CD4+ cells. In contrast, in vitro viral replication did not differ in CD4+ cells from un-infected macaques, CNAR(+) and CNAR(-) LTNPs. A role for transitional memory cells in supporting CNAR in the macaque model of AIDS was questionable. CNAR appears to represent an important part of the immune response displayed by CD8+ T cells which might be underestimated up to now.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
HIV-1 causes chronic inflammation and AIDS in humans, whereas related simian immunodeficiency viruses (SIVs) replicate efficiently in their natural hosts without causing disease. It is currently ...unknown to what extent virus-specific properties are responsible for these different clinical outcomes. Here, we incorporate two putative HIV-1 virulence determinants, i.e., a Vpu protein that antagonizes tetherin and blocks NF-κB activation and a Nef protein that fails to suppress T cell activation via downmodulation of CD3, into a non-pathogenic SIVagm strain and test their impact on viral replication and pathogenicity in African green monkeys. Despite sustained high-level viremia over more than 4 years, moderately increased immune activation and transcriptional signatures of inflammation, the HIV-1-like SIVagm does not cause immunodeficiency or any other disease. These data indicate that species-specific host factors rather than intrinsic viral virulence factors determine the pathogenicity of primate lentiviruses.
IFN-γ promotes tumoral immune surveillance, but its involvement in controlling metastases is less clear. Using a mouse model of pulmonary metastases, we show that local IFN-γ treatment inhibits ...formation of metastases through its regulation of IRF-1 in tumor cells. IRF-1 is an IFN-γ-induced transcription factor pivotal in the regulation of infection and inflammation. IRF-1 blockade abolished the inhibitory effect of IFN-γ on tumor metastases, whereas ectopic expression of IRF-1 phenocopied the inhibitory effects of IFN-γ. IRF-1 did not affect the survival of tumor cells in the circulation or their infiltration into lungs, but it was essential to support the pulmonary attraction and activation of natural killer (NK) cells. Depleting NK cells from mice abolished the protective effect of IFN-γ or IRF-1 on metastases. In addition, cytotoxicity assays revealed that tumor cells expressing IRF-1 were targeted more effectively by NK cells than IRF-1 nonexpressing tumor cells. Moreover, NK cells isolated from lungs inoculated with IRF-1-expressing tumor cells exhibit a greater cytotoxic activity. Mechanistic investigations revealed that IRF-1-induced NK cell cytotoxicity was independent of perforin and granzyme B but dependent on the NK cell activating receptor DNAM-1. Taken together, our findings establish IRF-1 as an essential mediator of the cross-talk between tumor cells and NK cells that mediate immune surveillance in the metastatic niche.
Ligament‐to‐bone and tendon‐to‐bone interfaces (entheses, osteotendinous junctions OTJs) serve to dissipate stress between soft tissue and bone. Surgical reconstruction of these interfaces is an ...issue of considerable importance as they are prone to injury and the integration of bone and tendon/ligament is in general not satisfactory. We report here the stem cell‐dependent spontaneous formation of fibrocartilaginous and fibrous entheses in heterotopic locations of the mouse if progenitors possess a tenogenic and osteo‐/chondrogenic capacity. This study followed the hypothesis that enhanced Bone Morphogenetic Protein (BMP)‐signaling in adult mesenchymal stem cells that are induced for tendon formation may overcome the tendon‐inherent interference with bone formation and may thus allow the stem cell‐dependent formation of tendon‐bone interfaces. The tenogenic and osteo‐/chondrogenic competence was mediated by the adeno‐ and/or lentiviral expression of the biologically active Smad8 signaling mediator (Smad8ca) and of Bone Morphogenetic Protein 2 (BMP2). Modified mesenchymal progenitors were implanted in subcutaneous or intramuscular sites of the mouse. The stem cell‐dependent enthesis formation was characterized histologically by immunohistological approaches and by in situ hybridization. Transplantation of modified murine stem cells resulted in the formation of tendinous and osseous structures exhibiting fibrocartilage‐type OTJs, while, in contrast, the viral modification of primary human bone marrow‐derived mesenchymal stromal/stem cells showed evidence of fibrous tendon‐bone interface formation. Moreover, it could be demonstrated that Smad8ca expression alone was sufficient for the formation of tendon/ligament‐like structures. These findings may contribute to the establishment of stem cell‐dependent regenerative therapies involving tendon/ligaments and to the improvement of the insertion of tendon grafts at bony attachment sites, eventually. STEM CELLS 2010; 28:1590–1601.
We recently described that IRF-1 is important for IFNγ mediated immune surveillance in metastasis. Despite the upregulation of MHC Class I in tumor cells, IRF-1 leads to a NK cell-dependent ...elimination of tumor cells. This mechanism is independent on inhibitory receptors and cytotoxic granules but dependent on DNAM-1.
Cryopreservation of peripheral blood mononuclear cells (PBMCs) is common for large clinical trials for which phenotypical characterization of lymphocytes is retrospectively performed in specialized ...core laboratories. It is therefore essential to assess the comparability between fresh and frozen samples. No side-by-side comparison of B and plasma cells of rhesus macaques (RM), which serve as useful models for several human diseases has been conducted until now. Hence, we performed an extensive comparative analysis between fresh and thawed mononuclear cells (MNCs) from blood and various tissues of healthy RM to analyze for the possible effects of cryopreservation on phenotype and functionality. Our data demonstrate that −80°C cryopreservation induces profound changes compared to fresh ex vivo-derived material. Percentages of B cells were stable in PBMCs, but were increased in all organs analyzed. The expression of CD27, a marker for differentiation between naïve and memory B cells, was massively reduced in PBMCs and MNC from organs with the most severe changes observed in cells from bone marrow (BM). Additionally, similar low percentages of CD27+ memory B cells were detected in PBMCs and BM samples stored in liquid nitrogen. Therefore, cryopreservation is not suitable for the phenotypical and functional characterization of B cells. Further optimization of cryoconservation protocols monitoring the surface expression of CD27, which was identified as a marker for the quality of cryopreserved material of RM, will be essential.
A comprehensive analysis of rhesus B cell subsets and plasma cells as a basis for studies on disease‐related B and plasma cell disorders.
B cells, as an important part of the humoral immune response, ...are generated in the BM, migrate to secondary lymphoid organs, and upon activation, differentiate into antibody‐producing memory B cells or plasma cells. Despite the pivotal roles that they play in different diseases, a comprehensive characterization in healthy rhesus macaques, which serve as valuable models for a variety of human diseases, is still missing. With the use of multiparameter flow cytometry, we analyzed B cells in BM collected from two locations, i.e., the iliac crest (BMca) and the femur (BMfem), PB, as well as secondary lymphoid organs of healthy rhesus macaques. We assessed the frequencies of immature and mature B cells, as well as CD19+ CD20– CD38+/++ CD138+/++ plasmablasts/plasma cells. Furthermore, we found site‐specific differences in the expression of markers for B cell activation and proliferation, chemokine receptors and Igs, as well as the distribution of memory B cell subpopulations. As secondary lymphoid organs harbor the highest frequencies of naive B cells, expression of CD80, CD95, and Ki67 was lower compared with B cells in the periphery and BM, whereas expression of IgD, CXCR4 (CD184), and CCR7 (CD197) was higher. Interestingly, BMca differed from BMfem regarding frequencies of B cells, their expression of CD80 and CXCR4, T cells, and plasma cells. In summary, these data identify baseline values for the above‐mentioned parameters and provide the foundation for future studies on B and plasma cells in different diseases.
The ability of long term non progressors to maintain very low levels of HIV/SIV and a healthy state, involves various host genetic and immunological factors. CD8+ non-cytolytic antiviral response ...(CNAR) most likely plays an important role in this regard. In order to gain a deeper insight into this unique phenomenon, the ability of CD8+ T cells to suppress viral replication in vitro was investigated in 16 uninfected, longitudinally in 23 SIV-infected long-term non-progressing (LTNPs), and 10 SIV-infected rhesus macaques with progressing disease. An acute infection assay utilizing CD4.sup.+ cells from MHC-mismatched monkeys to avoid cytolytic responses was employed. The study has identified CNAR as a long-term stable activity that inversely correlated with plasma viral load. The activity was also detected in CD8.sup.+ cells of uninfected macaques, which indicates that CNAR is not necessarily a virus specific response but increases after SIV-infection. Physical contact between CD4.sup.+ and CD8.sup.+ cells was mainly involved in mediating viral inhibition. Loss of this activity appeared to be due to a loss of CNAR-expressing CD8+ cells as well as a reduction of CNAR-responsive CD4+ cells. In contrast, in vitro viral replication did not differ in CD4+ cells from un-infected macaques, CNAR(+) and CNAR(-) LTNPs. A role for transitional memory cells in supporting CNAR in the macaque model of AIDS was questionable. CNAR appears to represent an important part of the immune response displayed by CD8+ T cells which might be underestimated up to now.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Granzyme B‐expressing (GrB+) B cells are thought to contribute to immune dysfunctions in HIV patients, but so far their exact role is unknown. This report demonstrates for the first time the ...existence of GrB+ B cells in SIV‐infected rhesus macaques, which represent the most commonly used nonhuman primate model for HIV research. Similar to HIV patients, we found significantly higher frequencies of these cells in the blood of chronically SIV‐infected rhesus monkeys compared with uninfected healthy ones. These frequencies correlated with plasma viral load and inversely with absolute CD4 T‐cell counts. When investigating GrB+ B cells in different compartments, levels were highest in blood, spleen and bone marrow, but considerably lower in lymph nodes and tonsils. Analysis of expression of various surface markers on this particular B‐cell subset in SIV‐infected macaques revealed differences between the phenotype in macaques and in humans. GrB+ B cells in SIV‐infected rhesus macaques exhibit an elevated expression of CD5, CD10, CD25 and CD27, while expression of CD19, CD185 and HLA‐DR is reduced. In contrast to human GrB+ B cells, we did not observe a significantly increased expression of CD43 and CD86. B‐cell receptor stimulation in combination with IL‐21 of purified B cells from healthy animals led to the induction of GrB expression. Furthermore, initial functional analyses indicated a regulatory role on T‐cell proliferation. Overall, our data pave the way for longitudinal analyses including studies on the functionality of GrB+ B cells in the nonhuman primate model for AIDS.
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