We evaluated Toll-like receptor (TLR) function in primary human dendritic cells from 104 young (age 21–30) and older (≥ 65 years) individuals. We used multicolor flow cytometry and intracellular cytokine staining of myeloid (mDC) and plasmacytoid (pDC) DCs and found substantial decreases in older, compared to young individuals in TNF-α, IL-6 and/or IL-12 (p40) production in mDCs and in TNF-α and IFN-α production in pDCs in response to TLR1/2, TLR2/6, TLR3, TLR5, and TLR8 engagement in mDCs and TLR7 and TLR9 in pDCs. These differences were highly significant after adjustment for heterogeneity between young and older groups (e.g. gender, race, body mass index [BMI], number of comorbid medical conditions) using mixed effect statistical modeling. Studies of surface and intracellular expression of TLR proteins, and of TLR gene expression in purified mDCs and pDCs revealed potential contributions for both transcriptional and post-transcriptional mechanisms in these age-associated effects. Moreover, intracellular cytokine production in the absence of TLR ligand stimulation was elevated in cells from older, compared to young individuals, suggesting a dysregulation of cytokine production that may limit further activation by TLR engagement. Our results provide evidence for immunosenescence in dendritic cells; notably, defects in cytokine production were strongly associated with poor antibody response to influenza immunization, a functional consequence of impaired TLR function in the aging innate immune response.

Summary The innate immune system is composed of a network of cells including neutrophils, NK and NKT cells, monocytes/macrophages, and dendritic cells that mediate the earliest interactions with pathogens. Age-associated defects are observed in the activation of all of these cell types, linked to compromised signal transduction pathways including the Toll-like Receptors. However, aging is also characterized by a constitutive pro-inflammatory environment (Inflamm-aging) with persistent low-grade innate immune activation that may augment tissue damage caused by infections in elderly individuals. Thus, immunosenescence in the innate immune system appears to reflect dysregulation, rather than exclusively impaired function.

The past decade has seen an explosion in research focusing on innate immunity. Through a wide range of mechanisms including phagocytosis, intracellular killing, and activation of pro-inflammatory or antiviral cytokine production via pattern recognition receptors, the cells of the innate immune system initiate and support adaptive immunity. The effects of aging on innate immune responses remain incompletely understood, particularly in humans. Here, we review advances in the study of human immunosenescence in the diverse cells of the innate immune system, including neutrophils, monocytes, macrophages, NK and NKT cells, and dendritic cells—with a focus on consequences for the response to infection or vaccination in old age.

Studies addressing immunosenescence in the immune system have expanded to focus on the innate as well as the adaptive responses. In particular, aging results in alterations in the function of Toll-like receptors (TLRs), the first described pattern recognition receptor family of the innate immune system. Recent studies have begun to elucidate the consequences of aging on TLR function in human cohorts and add to existing findings performed in animal models. In general, these studies show that human TLR function is impaired in the context of aging, and in addition there is evidence for inappropriate persistence of TLR activation in specific systems. These findings are consistent with an overarching theme of age-associated dysregulation of TLR signaling that likely contributes to the increased morbidity and mortality from infectious diseases found in geriatric patients.

Endothelial nitric-oxide synthase (eNOS) is an important regulator of endothelial function and vascular tone in biological tissues. While endothelial dysfunction occurs following ischemia and has been attributed to altered NO ⅐ formation, the biochemical basis for this dysfunction is unknown. Therefore, studies were performed to determine the effects of myocardial ischemia and reperfusion on eNOS in isolated rat hearts subjected to periods of global ischemia or ischemia followed by reperfusion. eNOS activity was assayed by L-[ C]arginine to L-[14 C]citrulline conversion and alterations in the amount and distribution of eNOS determined by Western blotting and immunohistochemistry. While activity was preserved after 30 min of ischemia with a value of 1.1 ؎ 0.1 pmol ؋ min ؊1 ؋ mg of protein ؊1 , it decreased by 77% after 60 min and became nearly undetectable after 120 min. Reperfusion resulted in only a partial restoration of activity. The decline in activity with ischemia was due, in part, to a loss of eNOS protein. Hemodynamic studies showed that the onset of impaired vascular reactivity paralleled the loss of functional eNOS. Subjecting isolated eNOS to conditions of acidosis, which occur during ischemia, followed by restoration of pH as occurs on reperfusion, caused a combination of reversible and irreversible loss of activity similar to that seen in ischemic and reperfused hearts. Thus, loss of endothelial function following ischemia is paralleled by a loss of eNOS activity due to a combination of pH-dependent denaturation and proteolysis.Over the last decade it has been shown that the endothelium plays a critical role in the control of vascular tone (1). A labile vasodilating substance termed endothelium-derived relaxing factor was identified as nitric oxide (NO ⅐ ), 1 which is synthesized by a calcium dependent nitric-oxide synthase (NOS) in endothelial cells (2-4). Subsequently, it was observed that ischemia causes impaired endothelial reactivity (5). In the heart it was observed that coronary artery occlusion, as occurs in heart attack, results in endothelial dysfunction. In both isolated vascular ring and in vivo models, endothelium-dependent vasodilation is markedly decreased after myocardial ischemia and reperfusion (6 -8). Studies in humans have identified a similar decline in acetylcholine-induced vasodilation or even paradoxical vasoconstriction in areas adjacent to atherosclerotic plaques present in coronary arteries (9). Likewise, a diffuse vasomotor impairment in hypercholesterolemic and diabetic patients was seen showing that vascular dysfunction might occur in different pathological conditions (10, 11).Because of the major pathophysiological significance of impaired endothelial reactivity following ischemia, there has been great interest in determining its underlying mechanisms. Earlier studies verified that while receptor mediated endothelialdependent responses to acetylcholine as well as receptor-independent responses to calcium ionophore A23187 were lost, endothelium-independent agents such a...