HMGB1, a non-histone nuclear factor, acts extracellularly as a mediator of delayed endotoxin lethality, which raises the question of how a nuclear protein can reach the extracellular space. We show that activation of monocytes results in the redistribution of HMGB1 from the nucleus to cytoplasmic organelles, which display ultrastructural features of endolysosomes. HMGB1 secretion is induced by stimuli triggering lysosome exocytosis. The early mediator of inflammation interleukin (IL)-1β is also secreted by monocytes through a non-classical pathway involving exocytosis of secretory lysosomes. However, in keeping with their respective role of early and late inflammatory factors, IL-1β and HMGB1 respond at different times to different stimuli: IL-1β secretion is induced earlier by ATP, autocrinally released by monocytes soon after activation; HMGB1 secretion is triggered by lysophosphatidylcholine, generated later in the inflammation site. Thus, in monocytes, non-classical secretion can occur through vescicle compartments that are at least partially distinct.

T lymphocytes are defective in cystine uptake and thus require exogenous thiols for activation and function. Here we show that monocyte-derived human dendritic cells (DCs) release cysteine in the extracellular space. Cysteine generation is increased by lipopolysaccharide and tumor necrosis factor alpha, and by contact with T cells specifically recognizing soluble or alloantigens. These stimuli also induce thioredoxin (TRX) accumulation in DCs. However, only the contact with antigen-specific T cells triggers TRX secretion by the antigen-presenting cells. Fewer extracellular thiols are recovered after DC-T cell interactions when cystine uptake or TRX activity are inhibited. In addition, glutamate (Glu) and anti-TRX-inactivating antibodies inhibit antigen-dependent T lymphocyte proliferation. These findings indicate that, during antigen presentation, DCs uptake cystine and release cysteine and TRX, thus providing a reducing microenvironment that facilitates immune response.

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Here we show that dendritic cells accumulate the precursor form of the leaderless secretory protein interleukin-18 (pro-interleukin-18) in the cell cytosol and in organelles cofractionating with endolysosomes. Upon antigen specific contact with T lymphocytes, particulated pro-interleukin-18 decreases rapidly, and the cytokine appears extracellularly, suggesting that exocytosis of pro-interleukin-18-containing organelles is induced. Exocytosis of secretory lysosomes is modulated by calcium: in agreement with this, calcium influx results in secretion of prointerleukin-18. In turn, pro-interleukin-18 secretion induced by T cells is prevented by the calcium channel blocker nifedipine. Our results demonstrate a novel, calcium-mediated mechanism of post-translational regulation of secretion for interleukin-18, that allows a fast release of the cytokine. ß

We recently reported that human dendritic cells release the leaderless secretory protein interleukin-1␤ (IL-1␤) following specific interaction with alloreactive T lymphocytes. To clarify the molecular mechanism underlying this secretion, this study investigated the intracellular trafficking of IL-1␤ in dendritic cells and the signal ( IntroductionRegulated secretion is traditionally considered as a specialized process occurring in only a few polarized cell types, namely, exocrine, endocrine, and neuronal cells. 1 However, former observations that, on binding to antigen-presenting B cells, T-helper cells release lymphokines preferentially over the membrane area where T-cell receptor cross-linking is occurring, 2,3 led to the hypothesis that a regulated polarized secretion may exist also in nonpolarized cells.It is now clear that many hemopoietic cells use regulated secretion; examples are mast cells and granulocytes, which degranulate in response to Fc receptor cross-linking, 4 or platelets, which respond to vascular lesions by releasing small molecules and proteins from intracellular granules. 5 Unlike conventional secretory cells, endowed with specific structures for storage and release, hemopoietic cells use secretory lysosomes, a mixture organelle between lysosomes and secretory granules. 6 Interestingly, other cell types, including fibroblasts, are able to transform conventional lysosomes into a secretory organelle underlying inducible exocytosis 7 ; thus, regulated secretion seems to be a more widespread phenomenon than previously thought. However, even if secretory lysosomes are quite ubiquitous, the ability of directing their content toward a given target, resulting in polarized secretion, has been described so far only for a few immune cells, namely, T lymphocytes or natural killer cells. 8,9 This may depend on how exocytosis is induced. Indeed, in most cell types exocytosis is driven by stimuli triggering structures distributed uniformly on the plasma membrane (such as IgE receptors on mast cells); in contrast, in the case of T and natural killer cells, binding to target cells with engagement of a single or few receptor complexes results in local activation leading to polarized degranulation.Although it is known that increases in intracellular free calcium concentration ([Ca ϩϩ ] i ) and cytoskeleton rearrangement occur during the process, 7 the molecular mechanisms underlying regulated lysosome exocytosis are still unclear.We have recently demonstrated in monocytes an adenosine triphosphate (ATP)-dependent exocytosis of secretory lysosomes, resulting in release of lysosomal enzymes and of the mature form of interleukin-1␤ (IL-1␤). 10 The latter belongs to the family of leaderless secretory proteins, which despite their extracellular localization and function, lack a secretory signal peptide and are externalized through nonclassical pathways, avoiding the endoplasmic reticulum-Golgi exocytotic route. 11,12 IL-1␤ is synthesized on free cytosolic ribosomes as a 35-kd precursor (pro-IL-1␤), which undergoe...

The role of interleukin-1β (IL-1β) as a regulator of the immune response, although extensively investigated, is still debated. We then studied the expression of IL-1β by human dendritic cells (DCs), the professional antigen presenting cells, and its modulation during immune reactions in vitro. Our results show that, on maturation or tetanus toxoid presentation to specific CD4+ CD40L+T lymphocytes, DCs begin to accumulate IL-1β precursor (pro–IL-1β) but do not secrete bioactive IL-1β. In contrast, interaction with alloreactive T cells results in both stimulation of pro–IL-1β synthesis and secretion of processed isoforms of the cytokine, that display biologic activity. Both CD4+ and CD8+ subsets of allospecific T lymphocytes are required: CD4+ T cells drive the synthesis of pro–IL-1β through CD40 engagement but have no effects on pro–IL-1β processing; CD8+ T cells, unable to induce synthesis of pro–IL-1β per se, are responsible for the generation of mature IL-1β by pro–IL-1β–producing DCs. Interleukin-1β–converting enzyme (ICE) inhibitors do not prevent the recovery of IL-1β bioactivity after allorecognition, indicating that allospecific CD8+ T cells may induce the release of bioactive IL-1β via mechanism(s) other than ICE activation. Altogether, these findings suggest that CD4+ and CD8+ T-lymphocyte subsets have distinct roles in the induction of IL-1β secretion by DCs and support the hypothesis that IL-1β plays a role in cell-mediated immune responses.

The role of interleukin-1β (IL-1β) as a regulator of the immune response, although extensively investigated, is still debated. We then studied the expression of IL-1β by human dendritic cells (DCs), the professional antigen presenting cells, and its modulation during immune reactions in vitro. Our results show that, on maturation or tetanus toxoid presentation to specific CD4+ CD40L+T lymphocytes, DCs begin to accumulate IL-1β precursor (pro–IL-1β) but do not secrete bioactive IL-1β. In contrast, interaction with alloreactive T cells results in both stimulation of pro–IL-1β synthesis and secretion of processed isoforms of the cytokine, that display biologic activity. Both CD4+ and CD8+ subsets of allospecific T lymphocytes are required: CD4+ T cells drive the synthesis of pro–IL-1β through CD40 engagement but have no effects on pro–IL-1β processing; CD8+ T cells, unable to induce synthesis of pro–IL-1β per se, are responsible for the generation of mature IL-1β by pro–IL-1β–producing DCs. Interleukin-1β–converting enzyme (ICE) inhibitors do not prevent the recovery of IL-1β bioactivity after allorecognition, indicating that allospecific CD8+ T cells may induce the release of bioactive IL-1β via mechanism(s) other than ICE activation. Altogether, these findings suggest that CD4+ and CD8+ T-lymphocyte subsets have distinct roles in the induction of IL-1β secretion by DCs and support the hypothesis that IL-1β plays a role in cell-mediated immune responses.