Grandparent-grandchild relationships, generativity, subjective well-being and self-rated health of older people in Chile.

With increasing life expectancy, grandparents and grandchildren have more years available to share. Furthermore, with lower fertility rates and fewer grandchildren, relationships can be more frequent and profound. Intergenerational relationships are expected to be associated with older people's quality of life, especially in Latin American countries such as Chile, with high intergenerational co-residence and contact between generations. This research aims to analyze the associations between the characteristics of intergenerational relationships and grandparents' subjective well-being (Diener Scale and Satisfaction) and self-rated health. The novelty stems from including the structural characteristics of relationships with grandchildren (frequency of contact, closeness, and care), the activities they share (generativity), and the quality of relationships (ambivalence). This study is based on data from a specific face-to-face grandparenting survey conducted on a sample of 464 grandparents in January 2020. It is representative of older Chilean grandparents living in private dwellings. Multiple logistic and ordinary regression models were estimated using the Diener Scale, unique satisfaction question, and health self-perception. The results demonstrated that subjective well-being, but not self-rated health, was highly associated with the characteristics of intergenerational relationships, especially with the quality of relationships and with generative activities such as recreational activities and family identity. In conclusion, intergenerational relationships' quality and content are strongly associated with subjective well-being in old age, but not with health self-perception. Even in a Latin American country like Chile, with high co-residence and intergenerational contact, the variations in quality and generativity activities significantly explain the variations in subjective well-being. For this reason, policies for the promotion of well-being in older people must consider the family environment in which older people live, encompassing wider family networks, including grandchildren.

A longitudinal study monitoring the quality of life in a national cohort of older adults in Chile before and during the COVID-19 outbreak.

BACKGROUND: Confinement during the COVID-19 pandemic has placed great stress on older adults, which may be affecting their quality of life. Thus, this study aims to describe the changes in mental and physical health, isolation and loneliness, residence and socioeconomic resources in a national cohort of Chilean older adults before and during the COVID-19 outbreak. It also analyzes the changes in depressive symptoms by changes in the other quality of life indicators before and during the COVID-19 outbreak. Possible methodological biases of telephone surveys in older adults living in non-developed countries are also discussed. METHODS: Between June and September 2020, a random subsample of 720 people who had participated in the face-to-face V National Survey on Quality of Life in Older Adults in Chile conducted at the end of 2019 was followed up by telephone. Descriptive bivariate analyses were performed using t-test and non-parametric tests for independent variables, comparing the baseline sample with the current 2020 follow-up sample during the peak of the pandemic outbreak in Latin America. Furthermore, descriptive bivariate analysis through t-test and non-parametric test for paired samples compared the follow-up subsample at baseline with the not-included sample, examining possible biases of the telephone interview compared with the face-to-face interview. RESULTS: In the panel, there was no variation in self-rated health. The health symptoms that worsened were memory, stomach, and mood problems. Depressive symptoms and anxiety increased; similarly, smartphone users, social contacts, intergenerational co-residence and resilience increased. The telephone follow-up sample had a higher educational level and greater smartphone use than those not included in the subsample. CONCLUSIONS: Although some physical and mental health indicators have worsened during the pandemic, older adults mobilized resources that could allow them to maintain their quality of life, such as improved resilience. Thus, these findings can guide future research and the development of efficient strategies to improve these resources among older adults to ensure wellbeing.

Pluripotent Stem Cell-Derived Hepatocytes Inhibit T Cell Proliferation In Vitro through Tryptophan Starvation.

Regenerative medicine aims to replace damaged tissues by stimulating endogenous tissue repair or by transplanting autologous or allogeneic cells. Due to their capacity to produce unlimited numbers of cells of a given cell type, pluripotent stem cells, whether of embryonic origin or induced via the reprogramming of somatic cells, are of considerable therapeutic interest in the regenerative medicine field. However, regardless of the cell type, host immune responses present a barrier to success. The aim of this study was to investigate in vitro the immunological properties of human pluripotent stem cell (PSC)-derived hepatocyte-like cells (HLCs). These cells expressed MHC class I molecules while they lacked MHC class II and co-stimulatory molecules, such as CD80 and CD86. Following stimulation with IFN-γ, HLCs upregulated CD40, PD-L1 and MHC class I molecules. When co-cultured with allogeneic T cells, HLCs did not induce T cell proliferation; furthermore, when T cells were stimulated via αCD3/CD28 beads, HLCs inhibited their proliferation via IDO1 and tryptophan deprivation. These results demonstrate that PSC-derived HLCs possess immunoregulatory functions, at least in vitro.

Does the selective attrition of a panel survey of older people affect the multivariate estimations of subjective well-being?

PURPOSE: The increased population aging has resulted in a growing need for longitudinal studies about the quality of life among older people. Nevertheless, the results of these investigations could be biased because more disadvantaged people leave the original sample. The purpose of this study is to examine how the selective attrition observed in a panel survey affect multivariate models of subjective well-being (SWB). The question is if we could do reliable longitudinal investigations concerning the predictors of SWB in old age. METHODS: This paper examines attrition in a panel of older people in Chile. Attrition was evaluated in the variables that affect elderly SWB. Probit models were fitted to compare dropouts with nondropouts. Then, multivariate probit models were estimated on satisfaction and depressive symptoms, comparing dropouts and nondropouts. Finally, we compared weighted and unweighted multivariate probit models on SWB. RESULTS: The attrition rate in 2 years was 38.8%, including deaths and 32.9%, excluding them. Survey dropouts had lower satisfaction but not higher depressive symptoms. Among SWB predictors, people without a partner and with lower self-efficacy abandoned more the study. When applying the Becketti, Gould, Lillard, and Welch test, the probit coefficients of the predictor variables on SWB outcome variables were similar for dropouts and nondropouts. Finally, the comparison of multivariate models on SWB with weighting methods did not find substantial differences in the explanatory coefficients. CONCLUSION: Although some predictors of attrition were associated with SWB, attrition did not produce biased estimates in multivariate models of life satisfaction life or depressive symptoms in old age.

Characterizing the B-Cell and Humoral Response in Tertiary Lymphoid Organs in Kidney Allografts.

OBJECTIVES: Tertiary lymphoid organs are formed at sites of chronic inflammation and are thought to contribute to the immune response. Here, we aimed to characterize the structure and function of tertiary lymphoid organs in a model of murine kidney allotransplant to understand their role in alloimmunity. MATERIALS AND METHODS: We transplanted 4 C57BL/6 mouse kidneys (isograft group) and 17 DBA/2 mouse kidneys into C57BL/6 mouse recipients. Three DBA/2-to-C57BL/6 transplant mice that rejected their grafts acutely (before 10 days posttransplant) were excluded from the study. The 14 surviving DAB2 grafts were retrieved at day 45 posttransplant and evaluated histologically. The presence of antibody-secreting cells and circulating levels of donor-specific antibodies were also evaluated. RESULTS: We found that tertiary lymphoid organs can be associated with a beneficial response in a kidney allotransplant model. Characterization of B-cell subsets within tertiary lymphoid organs in mouse kidney allografts revealed naive, plasma, and memory B cells, which were mostly grouped within or in close proximity of tertiary lymphoid organs. Staining for intracellular immunoglobulin G showed that many of the B cells within tertiary lymphoid organs were capable of producing antibodies. Although allospecific antibodies were found in the serum of recipient mice and were deposited in the transplanted kidneys, graft function was not affected in this model. CONCLUSIONS: B cells within tertiary lymphoid organs are functional and contribute to the humoral arm of the alloresponse. However, tertiary lymphoid organs are not necessarily associated with graft rejection, suggesting that protective mechanisms are at play.

Nox2 in regulatory T cells promotes angiotensin II-induced cardiovascular remodeling.

The superoxide-generating enzyme Nox2 contributes to hypertension and cardiovascular remodeling triggered by activation of the renin-angiotensin system. Multiple Nox2-expressing cells are implicated in angiotensin II-induced (Ang II-induced) pathophysiology, but the importance of Nox2 in leukocyte subsets is poorly understood. Here, we investigated the role of Nox2 in T cells, particularly Tregs. Mice globally deficient in Nox2 displayed increased numbers of Tregs in the heart at baseline, whereas Ang II-induced effector T cell (Teff) infiltration was inhibited. To investigate the role of Treg Nox2, we generated a mouse line with CD4-targeted Nox2 deficiency (Nox2fl/flCD4Cre+). These animals showed inhibition of Ang II-induced hypertension and cardiac remodeling related to increased tissue-resident Tregs and reduction in infiltrating Teffs, including Th17 cells. The protection in Nox2fl/flCD4Cre+ mice was reversed by anti-CD25 antibody depletion of Tregs. Mechanistically, Nox2-/y Tregs showed higher in vitro suppression of Teff proliferation than WT Tregs, increased nuclear levels of FoxP3 and NF-κB, and enhanced transcription of CD25, CD39, and CD73. Adoptive transfer of Tregs confirmed that Nox2-deficient cells had greater inhibitory effects on Ang II-induced heart remodeling than WT cells. These results identify a previously unrecognized role of Nox2 in modulating suppression of Tregs, which acts to enhance hypertension and cardiac remodeling.

Spatial and Single-Cell Transcriptional Profiling Identifies Functionally Distinct Human Dermal Fibroblast Subpopulations.

Previous studies have shown that mouse dermis is composed of functionally distinct fibroblast lineages. To explore the extent of fibroblast heterogeneity in human skin, we used a combination of comparative spatial transcriptional profiling of human and mouse dermis and single-cell transcriptional profiling of human dermal fibroblasts. We show that there are at least four distinct fibroblast populations in adult human skin, not all of which are spatially segregated. We define markers permitting their isolation and show that although marker expression is lost in culture, different fibroblast subpopulations retain distinct functionality in terms of Wnt signaling, responsiveness to IFN-γ, and ability to support human epidermal reconstitution when introduced into decellularized dermis. These findings suggest that ex vivo expansion or in vivo ablation of specific fibroblast subpopulations may have therapeutic applications in wound healing and diseases characterized by excessive fibrosis.

Endothelial Plasmalemma Vesicle-Associated Protein Regulates the Homeostasis of Splenic Immature B Cells and B-1 B Cells.

Plasmalemma vesicle-associated protein (Plvap) is an endothelial protein with roles in endothelial diaphragm formation and maintenance of basal vascular permeability. At the same time, Plvap has roles in immunity by facilitating leukocyte diapedesis at inflammatory sites and controlling peripheral lymph node morphogenesis and the entry of soluble Ags into lymph node conduits. Based on its postulated role in diapedesis, we have investigated the role of Plvap in hematopoiesis and show that deletion of Plvap results in a dramatic decrease of IgM+IgDlo B cells in both the spleen and the peritoneal cavity. Tissue-specific deletion of Plvap demonstrates that the defect is B cell extrinsic, because B cell and pan-hematopoietic Plvap deletion has no effect on IgM+IgDlo B cell numbers. Endothelial-specific deletion of Plvap in the embryo or at adult stage recapitulates the full Plvap knockout phenotype, whereas endothelial-specific reconstitution of Plvap under the Chd5 promoter rescues the IgM+IgDlo B cell phenotype. Taken together, these results show that Plvap expression in endothelial cells is important in the maintenance of IgM+ B cells in the spleen and peritoneal cavity.

IL-10-produced by human transitional B-cells down-regulates CD86 expression on B-cells leading to inhibition of CD4+T-cell responses.

A novel subset of human regulatory B-cells has recently been described. They arise from within the transitional B-cell subpopulation and are characterised by the production of IL-10. They appear to be of significant importance in regulating T-cell immunity in vivo. Despite this important function, the molecular mechanisms by which they control T-cell activation are incompletely defined. Here we show that transitional B-cells produced more IL-10 and expressed higher levels of IL-10 receptor after CD40 engagement compared to other B-cell subsets. Furthermore, under this stimulatory condition, CD86 expressed by transitional B-cells was down regulated and T-cell proliferation was reduced. We provide evidence to demonstrate that the down-regulation of CD86 expression by transitional B-cells was due to the autocrine effect of IL-10, which in turn leads to decreased T-cell proliferation and TNF-α production. This analysis was further extended to peripheral B-cells in kidney transplant recipients. We observed that B-cells from patients tolerant to the graft maintained higher IL-10 production after CD40 ligation, which correlates with lower CD86 expression compared to patients with chronic rejection. Hence, the results obtained in this study shed light on a new alternative mechanism by which transitional B-cells inhibit T-cell proliferation and cytokine production.

Retinoic Acid Signaling in B Cells Is Required for the Generation of an Effective T-Independent Immune Response.

Retinoic acid (RA) plays an important role in the balance of inflammation and tolerance in T cells. Furthermore, it has been demonstrated that RA facilitates IgA isotype switching in B cells in vivo. However, it is unclear whether RA has a direct effect on T-independent B cell responses in vivo. To address this question, we generated a mouse model where RA signaling is specifically silenced in the B cell lineage. This was achieved through the overexpression of a dominant negative receptor α for RA (dnRARα) in the B cell lineage. In this model, we found a dramatic reduction in marginal zone (MZ) B cells and accumulation of transitional 2 B cells in the spleen. We also observed a reduction in B1 B cells in the peritoneum with a defect in the T-independent B cell response against 2,4,6-trinitrophenyl. This was not a result of inhibited development of B cells in the bone marrow, but likely the result of both defective expression of S1P1 in MZ B cells and a defect in the development of MZ and B1 B cells. This suggests that RARα expression in B cells is important for B cell frequency in the MZ and peritoneum, which is crucial for the generation of T-independent humoral responses.

Cutting Edge: Retinoic Acid Signaling in B Cells Is Essential for Oral Immunization and Microflora Composition.

Retinoic acid (RA) is a critical regulator of the intestinal adaptive immune response. However, the intrinsic impact of RA on B cell differentiation in the regulation of gut humoral immunity in vivo has never been directly shown. To address this issue, we have been able to generate a mouse model where B cells specifically express a dominant-negative receptor α for RA. In this study, we show that the silencing of RA signaling in B cells reduces the numbers of IgA(+) Ab-secreting cells both in vitro and in vivo, suggesting that RA has a direct effect on IgA plasma cell differentiation. Moreover, the lack of RA signaling in B cells abrogates Ag-specific IgA responses after oral immunization and affects the microbiota composition. In conclusion, these results suggest that RA signaling in B cells through the RA receptor α is important to generate an effective gut humoral response and to maintain a normal microbiota composition.

Retinoic acid is essential for Th1 cell lineage stability and prevents transition to a Th17 cell program.

CD4(+) T cells differentiate into phenotypically distinct T helper cells upon antigenic stimulation. Regulation of plasticity between these CD4(+) T-cell lineages is critical for immune homeostasis and prevention of autoimmune disease. However, the factors that regulate lineage stability are largely unknown. Here we investigate a role for retinoic acid (RA) in the regulation of lineage stability using T helper 1 (Th1) cells, traditionally considered the most phenotypically stable Th subset. We found that RA, through its receptor RARα, sustains stable expression of Th1 lineage specifying genes, as well as repressing genes that instruct Th17-cell fate. RA signaling is essential for limiting Th1-cell conversion into Th17 effectors and for preventing pathogenic Th17 responses in vivo. Our study identifies RA-RARα as a key component of the regulatory network governing maintenance and plasticity of Th1-cell fate and defines an additional pathway for the development of Th17 cells.

Transitional-2 B cells acquire regulatory function during tolerance induction and contribute to allograft survival.

In humans, tolerance to renal transplants has been associated with alterations in B-cell gene transcription and maintenance of the numbers of circulating transitional B cells. Here, we use a mouse model of transplantation tolerance to investigate the contribution of B cells to allograft survival. We demonstrate that transfer of B cells from mice rendered tolerant to MHC class I mismatched skin grafts can prolong graft survival in a dose-dependent and antigen-specific manner to a degree similar to that afforded by graft-specific regulatory T (Treg) cells. Tolerance in this model was associated with an increase in transitional-2 (T2) B cells. Only T2 B cells from tolerized mice, not naïve T2 nor alloantigen experienced T2, were capable of prolonging skin allograft survival, and suppressing T-cell activation. Tolerized T2 B cells expressed lower levels of CD86, increased TIM-1, and demonstrated a preferential survival in vivo. Furthermore, we demonstrate a synergistic effect between tolerized B cells and graft-specific Treg cells. IL-10 production by T2 B cells did not contribute to tolerance, as shown by transfer of B cells from IL-10(-/-) mice. These results suggest that T2 B cells in tolerant patients may include a population of regulatory B cells that directly inhibit graft rejection.

CCR6-dependent positioning of memory B cells is essential for their ability to mount a recall response to antigen.

Chemokine-dependent localization of specific B cell subsets within the immune microarchitecture is essential to ensure successful cognate interactions. Although cognate interactions between T cells and memory B cells (B(mem)) are essential for the secondary humoral immune responses, the chemokine response patterns of B(mem) cells are largely unknown. In contrast to naive B cells, this study shows that Ag-specific B(mem) cells have heightened expression of CCR6 and a selective chemotactic response to the CCR6 ligand, CCL20. Although CCR6 appears be nonessential for the initial clonal expansion and maintenance of B(mem), CCR6 is essential for the ability of B(mem) to respond to a recall response to their cognate Ag. This dependency was deemed intrinsic by studies in CCR6-deficient mice and in bone marrow chimeric mice where CCR6 deficiency was limited to the B cell lineage. Finally, the mis-positioning of CCR6-deficient B(mem) was revealed by immunohistological analysis with an altered distribution of CCR6-deficient B(mem) from the marginal and perifollicular to the follicular/germinal center area.

The diaphragms of fenestrated endothelia: gatekeepers of vascular permeability and blood composition.

Fenestral and stomatal diaphragms are endothelial subcellular structures of unknown function that form on organelles implicated in vascular permeability: fenestrae, transendothelial channels, and caveolae. PV1 protein is required for diaphragm formation in vitro. Here, we report that deletion of the PV1-encoding Plvap gene in mice results in the absence of diaphragms and decreased survival. Loss of diaphragms did not affect the fenestrae and transendothelial channels formation but disrupted the barrier function of fenestrated capillaries, causing a major leak of plasma proteins. This disruption results in early death of animals due to severe noninflammatory protein-losing enteropathy. Deletion of PV1 in endothelium, but not in the hematopoietic compartment, recapitulates the phenotype of global PV1 deletion, whereas endothelial reconstitution of PV1 rescues the phenotype. Taken together, these data provide genetic evidence for the critical role of the diaphragms in fenestrated capillaries in the maintenance of blood composition.

A retinoic acid-dependent checkpoint in the development of CD4+ T cell-mediated immunity.

It is known that vitamin A and its metabolite, retinoic acid (RA), are essential for host defense. However, the mechanisms for how RA controls inflammation are incompletely understood. The findings presented in this study show that RA signaling occurs concurrent with the development of inflammation. In models of vaccination and allogeneic graft rejection, whole body imaging reveals that RA signaling is temporally and spatially restricted to the site of inflammation. Conditional ablation of RA signaling in T cells significantly interferes with CD4(+) T cell effector function, migration, and polarity. These findings provide a new perspective of the role of RA as a mediator directly controlling CD4(+) T cell differentiation and immunity.

The immortality of humoral immunity.

Decades of high-titered antibody are sustained due to the persistence of memory B cells and long-lived plasma cells (PCs). The differentiation of each of these subsets is antigen- and T-cell driven and is dependent on signals acquired and integrated during the germinal center response. Inherent in the primary immune response must be the delivery of signals to B cells to create these populations, which have virtual immortality. Differences in biology and chemotactic behavior disperse memory B cells and long-lived PCs to a spectrum of anatomic sites. Each subset must rely on survival factors that can support their longevity. This review focuses on the generation of each of these subsets, their survival, and renewal, which must occur to sustain serological memory. In this context, we discuss the role of antigen, bystander inflammation, and cellular niches. The contribution of BAFF (B-cell activating factor belonging to the tumor necrosis factor family) and APRIL (a proliferation-inducing ligand) to the persistence of memory B cells and PCs are also detailed. Insights that have been provided over the past few years in the regulation of long-lived B-cell responses will have profound impact on vaccine development, the treatment of pre-sensitized patients for organ transplantation, and therapeutic interventions in both antibody- and T-cell-mediated autoimmunity.

Distinction of the memory B cell response to cognate antigen versus bystander inflammatory signals.

The hypothesis that bystander inflammatory signals promote memory B cell (B(MEM)) self-renewal and differentiation in an antigen-independent manner is critically evaluated herein. To comprehensively address this hypothesis, a detailed analysis is presented examining the response profiles of B-2 lineage B220(+)IgG(+) B(MEM) toward cognate protein antigen in comparison to bystander inflammatory signals. After in vivo antigen encounter, quiescent B(MEM) clonally expand. Surprisingly, proliferating B(MEM) do not acquire germinal center (GC) B cell markers before generating daughter B(MEM) and differentiating into plasma cells or form structurally identifiable GCs. In striking contrast to cognate antigen, inflammatory stimuli, including Toll-like receptor agonists or bystander T cell activation, fail to induce even low levels of B(MEM) proliferation or differentiation in vivo. Under the extreme conditions of adjuvanted protein vaccination or acute viral infection, no detectable bystander proliferation or differentiation of B(MEM) occurred. The absence of a B(MEM) response to nonspecific inflammatory signals clearly shows that B(MEM) proliferation and differentiation is a process tightly controlled by the availability of cognate antigen.

The enigmatic role of mast cells in dominant tolerance.

PURPOSE OF REVIEW: The role of regulatory T cells (Treg) in peripheral tolerance has been studied extensively in transplantation research. Recently, mast cells have been shown to play an indispensable role in allograft tolerance. The purpose of this review is to inform the reader on the current standings of the role of mast cells in dominant tolerance with an emphasis on the interaction of mast cells with Treg. RECENT FINDINGS: Mast cells are required to sustain peripheral tolerance via Treg. Treg can stabilize mast cells degranulation by contact-dependent mechanisms through the interaction of OX40 and its ligand OX40L, and by production of soluble factors, such as interleukin-10 and transforming growth factor-beta. Conversely, the activation and subsequent degranulation of mast cells break peripheral tolerance. SUMMARY: Both mast cells and Treg are needed to create a local immunosuppressive environment in the transplant. Treg are not only necessary to suppress effector T-cell responses but also to stabilize mast cells. Mast cells in return could contribute to the immunosuppressive state by release of transforming growth factor-beta, interleukin-10 and specific proteases. However, the molecular basis for mast cells control of Treg suppression in organ transplantation is still unresolved.

Polyethylenimine-based siRNA nanocomplexes reprogram tumor-associated dendritic cells via TLR5 to elicit therapeutic antitumor immunity.

The success of clinically relevant immunotherapies requires reversing tumor-induced immunosuppression. Here we demonstrated that linear polyethylenimine-based (PEI-based) nanoparticles encapsulating siRNA were preferentially and avidly engulfed by regulatory DCs expressing CD11c and programmed cell death 1-ligand 1 (PD-L1) at ovarian cancer locations in mice. PEI-siRNA uptake transformed these DCs from immunosuppressive cells to efficient antigen-presenting cells that activated tumor-reactive lymphocytes and exerted direct tumoricidal activity, both in vivo and in situ. PEI triggered robust and selective TLR5 activation in vitro and elicited the production of hallmark TLR5-inducible cytokines in WT mice, but not in Tlr5-/- littermates. Thus, PEI is a TLR5 agonist that, to our knowledge, was not previously recognized. In addition, PEI-complexed nontargeting siRNA oligonucleotides stimulated TLR3 and TLR7. The nonspecific activation of multiple TLRs (specifically, TLR5 and TLR7) reversed the tolerogenic phenotype of human and mouse ovarian tumor-associated DCs. In ovarian carcinoma-bearing mice, this induced T cell-mediated tumor regression and prolonged survival in a manner dependent upon myeloid differentiation primary response gene 88 (MyD88; i.e., independent of TLR3). Furthermore, gene-specific siRNA-PEI nanocomplexes that silenced immunosuppressive molecules on mouse tumor-associated DCs elicited discernibly superior antitumor immunity and enhanced therapeutic effects compared with nontargeting siRNA-PEI nanocomplexes. Our results demonstrate that the intrinsic TLR5 and TLR7 stimulation of siRNA-PEI nanoparticles synergizes with the gene-specific silencing activity of siRNA to transform tumor-infiltrating regulatory DCs into DCs capable of promoting therapeutic antitumor immunity.