Janet M. Lord

Fibroblasts regulate the switch from acute resolving to chronic persistent inflammation

Fibroblasts are important sentinel cells in the immune system and, here, it is proposed that these cells play a critical role in the switch from acute inflammation to adaptive immunity and tissue repair. It is suggested that chronic inflammation occurs because of disordered fibroblast behaviour in which failure to switch off their inflammatory programme leads to the inappropriate survival and retention of leukocytes within inflamed tissue.

RGD peptides induce apoptosis by direct caspase-3 activation

Synthetic peptides containing the arginine–glycine–aspartate (RGD) motif have been used extensively as inhibitors of integrin–ligand interactions in studies of cell adhesion, migration, growth and differentiation,,, because the RGD motif is an integrin-recognition motif found in many ligands. Here we report that RGD-containing peptides are able to directly induce apoptosis without any requirement for integrin-mediated cell clustering or signals. We show that RGD-containing peptides enter cells and directly induce autoprocessing and enzymatic activity of pro-caspase-3, a pro-apoptotic protein. Using the breast carcinoma cell line MCF-7, which has a functional deletion of the caspase-3 gene, we confirm that caspase-3 is required for RGD-mediated cell death. In addition to an RGD motif, pro-caspase-3 also contains a potential RGD-binding motif, aspartate–aspartate–methionine (DDM), near the site of processing to produce the p12 and p17 subunits. On the basis of the ability of RGD–DDX interactions to trigger integrin activation, we suggest that RGD peptides induce apoptosis by triggering conformational changes that promote pro-caspase-3 autoprocessing and activation. These findings provide an alternative molecular explanation for the potent pro-apoptotic properties of RGD peptides in models of angiogenesis, inflammation and cancer metastasis,,.

Aging of the innate immune system

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.

Human innate immunosenescence: causes and consequences for immunity in old age

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 proinflammatory or antiviral cytokine production, 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, natural killer and natural killer T (NKT) cells and dendritic cells-with a focus on consequences for the response to infection or vaccination in old age.

Senescence in innate immune responses: reduced neutrophil phagocytic capacity and CD16 expression in elderly humans

Elderly humans are more susceptible to bacterial infections because of declining immune status. We have investigated the effect of aging on neutrophil bactericidal responses, comparing neutrophil function in healthy, young (23–35 years) and elderly (>65 years) volunteers. Superoxide generation in response to fMLP was slightly increased in neutrophils from elderly donors, and serum from the elderly was able to opsonize E. coli efficiently. In contrast, phagocytic index was significantly lower in neutrophils from the elderly, compared with young donors (P<0.005). CD11a and CD11b expression was not affected by age, but CD16 was significantly reduced in neutrophils from elderly donors (P<0.0001). CD16 expression and phagocytic index were measured in the same neutrophils using FITC‐labeled E. coli, PE‐conjugated anti‐CD16 antibody, and CD16 expression correlated with phagocytic index (r=0.83;P<0.05). In elderly patients with bacterial infection, CD16 expression remained low. We propose that reduced neutrophil CD16 expression and phagocytosis contribute to human immunesenescence.

PKC-delta is an apoptotic lamin kinase.

Protein kinase C-δ is activated during apoptosis, following proteolytic cleavage by caspase 3. Furthermore, overexpression of the catalytic kinase fragment of PKC-δ induces the nuclear phenotype associated with apoptosis, though the molecular basis of this effect has not been determined. In these studies we have examined the role of PKC-δ in the disassembly of the nuclear lamina at apoptosis. The nuclear lamina is disassembled during mitosis and apoptosis and mitotic disassembly involves hyperphosphorylation of lamin proteins by mitotic lamin kinases. During apoptosis, lamin proteins are degraded by caspase 6 and the contribution made by phosphorylation has not been proven. We show here that protein kinase C-δ co-localized with lamin B during apoptosis and activation of PKC-δ by caspase 3 was concomitant with lamin B phosphorylation and proteolysis. Inhibition of PKC-δ delayed lamin proteolysis, even in the presence of active caspase 6, whilst inhibitors of mitotic lamin kinases were without effect. In addition recombinant human PKC-δ was able to phosphorylate lamin B in vitro suggesting that its actions are direct and not via an intermediary kinase. We propose that PKC-δ is an apoptotic lamin kinase and that efficient lamina disassembly at apoptosis requires both lamin hyperphosphorylation and caspase mediated proteolysis.

Diacylglycerol and Phosphatidate Generated by Phospholipases C and D, Respectively, Have Distinct Fatty Acid Compositions and Functions PHOSPHOLIPASE D-DERIVED DIACYLGLYCEROL DOES NOT ACTIVATE PROTEIN KINASE C IN PORCINE AORTIC ENDOTHELIAL CELLS

Stimulation of cells with certain agonists often activates both phospholipases C and D. These generate diacylglycerol and phosphatidate, respectively, although the two lipids are also apparently interconvertable through the actions of phosphatidate phosphohydrolase and diacylglycerol kinase. Diacylglycerol activates protein kinase C while one role for phosphatidate is the activation of actin stress fiber formation. Therefore, if the two lipids are interconvertable, it is theoretically possible that an uncontrolled signaling loop could arise. To address this issue structural analysis of diacylglycerol, phosphatidate, and phosphatidylbutanol (formed in the presence of butan-1-ol) from both Swiss 3T3 and porcine aortic endothelial cells was performed. This demonstrated that phospholipase C activation generates primarily polyunsaturated species while phospholipase D activation generates saturated/monounsaturated species. In the endothelial cells, where phospholipase D was activated by lysophosphatidic acid independently of phospholipase C, there was no activation of protein kinase C. Thus we propose that only polyunsaturated diacylglycerols and saturated/monounsaturated phosphatidates function as intracellular messengers and that their interconversion products are inactive.

Neutrophil ageing and immunesenescence.

As humans age, their morbidity and mortality from infection increases, their response to vaccination declines and they have an increased incidence of inflammatory diseases and cancer. The reasons for these effects are clearly complex, but reduced efficiency of the innate and adaptive immune system is likely to be important in the pathology of old age. Age-related changes in the adaptive immune system are well-documented and include alterations in T cell phenotype and effector functions and a reduced ability of B cells to produce high affinity antibody. In contrast, the innate immune system has been less well researched and the perception amongst many immunogerontologists is that this branch of the immune system is only moderately affected by age. However, it is becoming increasingly clear that the adaptive and innate immune systems co-operate at several levels to ensure the optimal immune response and any decline in adaptive immunity will impact upon the function of the innate immune system and vice-versa. Here, we review the literature concerning intrinsic age-related changes in neutrophil responses and consider how changes in lymphocyte function with age might further compromise efficiency of neutrophil function.

IFN-α and IFN-β: a link between immune memory and chronic inflammation

Abstract The majority of expanded T cells generated during an immune response are cleared by apoptosis. Prevention of death in some activated T cells enables the persistence of a memory T-cell pool. Here, observations that IFN-α and IFN-β inhibit activated T-cell apoptosis are described. Although this enables memory T cells to persist without antigen, excessive IFN-α or IFN-γ secretion might lead to chronic inflammation.

The death-inducing signalling complex is recruited to lipid rafts in Fas-induced apoptosis

Membrane microdomains known as lipid rafts have been shown recently to be involved in Fas signalling and apoptosis in T and B cell lines. Here, we have investigated further the role of lipid rafts in Fas-induced apoptosis in non-transformed human CD4 T cells. We show that Fas-induced apoptosis in CD4 T cells was inhibited by the lipid raft disrupter methyl-beta-cyclodextrin. When lipid rafts were isolated from control and Fas ligand treated cells, we found that a small proportion of Fas was present in the raft fraction in untreated cells and that this was greatly increased upon Fas ligation. The other components of the Death Inducing Signalling Complex (DISC), FADD, and procaspase 8, were also present at higher levels in the raft fraction isolated from Fas ligand treated cells. We conclude that formation of the DISC occurs in lipid rafts and that these membrane microdomains are required for efficient Fas signalling and apoptosis.