The social, health, and economic challenges of a steadily increasing aging population demand the use of appropriate translational animal models to address questions like healthy aging, vaccination ...strategies, or potential interventions during the aging process. Due to their genetic proximity to humans, especially nonhuman primates (NHPs) with a relatively short generation period compared to humans, qualify as excellent animal models for these purposes. The use of common marmosets (Callithrix jacchus) in gerontology research steadily increased over the last decades, yet important information about their aging parameters are still missing. We therefore aimed to characterize their aging immune system by comprehensive flow cytometric phenotyping of blood immune cells from juvenile, adult, aging, and geriatric animals. Aged and geriatric animals displayed clear signs of immunosenescence. A decline in CD4/CD8 ratio, increased expression of HLA‐DR and PD‐1, higher frequencies of CD95+ memory cells, alterations in cytokine secretion, and a decline in the proliferative capacity proved T cell senescence in aging marmosets. Also, the B cell compartment was affected by age‐related changes: while overall B cell numbers remained stable with advancing age, expression of the activation marker CD80 increased and immunoglobulin M expression decreased. Interestingly, marmoset B cell memory subset distribution rather mirrored the human situation than that of other NHP. CD21+ CD27− naïve B cell frequencies decreased while those of CD21− CD27− tissue memory B cells increased with age. Furthermore, frequencies and numbers of NK cells as part of the innate immune system declined with advancing age.
Thus, the observed immunological changes in common marmosets over their life span revealed several similarities to age‐related changes in humans and encourages further studies to strengthen the common marmoset as a potential aging model.
Correlation between immune cell frequencies and age in juvenile, adult, aging, and geriatric common marmosets.
Research Highlights
Immunological investigations of the aging process in common marmosets revealed changes in different immune cell subsets.
Due to similar immunological changes during aging, this species is proposed as a potential translational animal model in gerontology research.
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.
Natural killer (NK) cells play a critical understudied role during HIV infection in tissues. In a natural host of SIV, the African green monkey (AGM), NK cells mediate a strong control of SIVagm ...infection in secondary lymphoid tissues. We demonstrate that SIVagm infection induces the expansion of terminally differentiated NKG2a
NK cells in secondary lymphoid organs displaying an adaptive transcriptional profile and increased MHC-E-restricted cytotoxicity in response to SIV Env peptides while expressing little IFN-γ. Such NK cell differentiation was lacking in SIVmac-infected macaques. Adaptive NK cells displayed no increased NKG2C expression. This study reveals a previously unknown profile of NK cell adaptation to a viral infection, thus accelerating strategies toward NK-cell directed therapies and viral control in tissues.
The MHC class I chain‐related molecule A (MICA) is a highly polymorphic ligand for the activating natural killer (NK)‐cell receptor NKG2D. A single nucleotide polymorphism causes a valine to ...methionine exchange at position 129. Presence of a MICA‐129Met allele in patients (n = 452) undergoing hematopoietic stem cell transplantation (HSCT) increased the chance of overall survival (hazard ratio HR = 0.77, P = 0.0445) and reduced the risk to die due to acute graft‐versus‐host disease (aGVHD) (odds ratio OR = 0.57, P = 0.0400) although homozygous carriers had an increased risk to experience this complication (OR = 1.92, P = 0.0371). Overall survival of MICA‐129Val/Val genotype carriers was improved when treated with anti‐thymocyte globulin (HR = 0.54, P = 0.0166). Functionally, the MICA‐129Met isoform was characterized by stronger NKG2D signaling, triggering more NK‐cell cytotoxicity and interferon‐γ release, and faster co‐stimulation of CD8+ T cells. The MICA‐129Met variant also induced a faster and stronger down‐regulation of NKG2D on NK and CD8+ T cells than the MICA‐129Val isoform. The reduced cell surface expression of NKG2D in response to engagement by MICA‐129Met variants appeared to reduce the severity of aGVHD.
Synopsis
Allogeneic hematopoietic stem cell transplantation still has a high risk of post‐transplant complications including graft versus host disease and relapse of malignancy. Detection of single nucleotide polymorphisms may help to provide a risk‐adapted treatment.
The MICA‐129Met/Val dimorphism was associated with an increased survival and a reduced risk to die due to acute graft versus host disease in a cohort of 452 patients.
The survival of MICA‐129Val/Val genotype carriers was improved when treated with antithymocyte globulin (ATG).
The MICA‐129Met isoform triggered more NK‐cell cytotoxicity and interferon‐γ release and it co‐stimulated cytotoxic T cells faster.
The MICA‐129Met isoform also induced a faster and stronger down‐regulation of NKG2D on NK and cytotoxic T cells limiting the initially stronger functional effects.
MICA‐129Val/Val carriers might profit from a T‐cell depleting treatment since this MICA variant has a lower ability to down‐regulate NKG2D and to limit the activation of alloreactive T cells.
Allogeneic hematopoietic stem cell transplantation still has a high risk of post‐transplant complications including graft versus host disease and relapse of malignancy. Detection of single nucleotide polymorphisms may help to provide a risk‐adapted treatment.
Measurement of the health and disease status of free‐ranging primates is often limited by a lack of available biomarkers of immune activation and inflammation that can be applied noninvasively via ...the measurement of urine or fecal samples. Here, we evaluate the potential usefulness of noninvasive urinary measurements of a number of cytokines, chemokines, and other markers of inflammation and infection. We took advantage of surgery‐associated inflammation in seven captive rhesus macaques, collecting urine samples before and after the medical interventions. We measured these urine samples for 33 different markers of inflammation and immune activation that are known to be responsive to inflammation and infection in rhesus macaque blood samples, via the Luminex platform. We also measured all samples for concentrations of the soluble urokinase plasminogen activator receptor (suPAR), which we had validated in a prior study as an effective biomarker of inflammation. Despite urine samples being collected in captivity under ideal conditions (clean, no contamination with feces or soil, frozen quickly), 13/33 biomarkers measured via Luminex were found at concentrations below detection limits in >50% of samples. Of the remaining 20 markers, only 2 showed significant increases in response to surgery—IL18 and MPO (myeloperoxidase). However, suPAR measurements of the same samples show a consistent marked increase in response to surgery that is absent from the patterns of IL18 and MPO measurement. Given that our samples were collected under conditions that are greatly preferable to those usually encountered in the field, urinary cytokine measurements via the Luminex platform seem overall unpromising for primate field studies.
Urinary cytokine measurements do not reflect surgery‐induced inflammation in rhesus macaques.
Research Highlights
13/33 cytokines validated for measurement in rhesus macaque plasma were measured below minimum assay sensitivity in urine in more than 50% of samples.
Only two cytokines measured in urine increased in response to surgically‐induced tissue trauma, and neither responded to the same extent as measurements of suPAR, a previously validated urinary biomarker of inflammation.
Urinary measurements of cytokines using the Luminex platform are overall unpromising for studies of macaques, and potentially other primates.
The SARS-CoV-2 pandemic has so far claimed over three and a half million lives worldwide. Though the SARS-CoV-2 mediated disease COVID-19 has first been characterized by an infection of the upper ...airways and the lung, recent evidence suggests a complex disease including gastrointestinal symptoms. Even if a direct viral tropism of intestinal cells has recently been demonstrated, it remains unclear, whether gastrointestinal symptoms are caused by direct infection of the gastrointestinal tract by SARS-CoV-2 or whether they are a consequence of a systemic immune activation and subsequent modulation of the mucosal immune system. To better understand the cause of intestinal symptoms we analyzed biopsies of the small intestine from SARS-CoV-2 infected individuals. Applying qRT-PCR and immunohistochemistry, we detected SARS-CoV-2 RNA and nucleocapsid protein in duodenal mucosa. In addition, applying imaging mass cytometry and immunohistochemistry, we identified histomorphological changes of the epithelium, which were characterized by an accumulation of activated intraepithelial CD8
T cells as well as epithelial apoptosis and subsequent regenerative proliferation in the small intestine of COVID-19 patients. In summary, our findings indicate that intraepithelial CD8
T cells are activated upon infection of intestinal epithelial cells with SARS-CoV-2, providing one possible explanation for gastrointestinal symptoms associated with COVID-19.
Comprehensive knowledge of immunoglobulin genetics is required to advance our understanding of B cell biology. Validated immunoglobulin variable (V) gene databases are close to completion only for ...human and mouse. We present a novel computational approach, IgDiscover, that identifies germline V genes from expressed repertoires to a specificity of 100%. IgDiscover uses a cluster identification process to produce candidate sequences that, once filtered, results in individualized germline V gene databases. IgDiscover was tested in multiple species, validated by genomic cloning and cross library comparisons and produces comprehensive gene databases even where limited genomic sequence is available. IgDiscover analysis of the allelic content of the Indian and Chinese-origin rhesus macaques reveals high levels of immunoglobulin gene diversity in this species. Further, we describe a novel human IGHV3-21 allele and confirm significant gene differences between Balb/c and C57BL6 mouse strains, demonstrating the power of IgDiscover as a germline V gene discovery tool.
The intesinal microbiome is considered important in human immunodeficiency virus (HIV) pathogenesis and therefore represents a potential therapeutic target to improve the patients' health status. ...Longitudinal alterations in the colonic mucosa-associated microbiome during simian immunodeficiency virus (SIV) infection were investigated using a 16S rRNA amplicon approach on the Illumina sequencing platform and bioinformatics analyses. Following SIV infection of six animals, no alterations in microbial composition were observed before the viral load peaked in the colon. At the time of acute mucosal SIV replication, the phylum Bacteroidetes including the Bacteroidia class as well as the phylum Firmicutes and its families Ruminococcaceae and Eubacteriaceae became more abundant. Enrichment of Bacteroidetes was maintained until the chronic phase of SIV infection. The shift towards Bacteroidetes in the mucosa-associated microbiome was associated with the extent of SIV infection-induced mucosal CD4
T cell depletion and correlated with increasing rates of enterocyte damage. These observations suggest that Bacteroidetes strains increase during virus-induced mucosal immune destruction. As Bacteroidetes belong to the lipopolysaccharide- and short chain fatty acids-producing bacteria, their rapid enrichment may contribute to inflammatory tissue damage and metabolic alterations in SIV/HIV infection. These aspects should be considered in future studies on therapeutic interventions.
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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