Pain management research from the NIH HEAL Initiative.

This article presents an overview of the pain research programs within the National Institutes of Health (NIH) Helping to End Addiction Long-term® Initiative, or NIH HEAL Initiative®. Launched in 2018 to address the opioid crisis, the NIH HEAL Initiative supports research on addiction prevention and treatment. A key component of addiction prevention is the development of new, effective, non-addictive treatments for acute and chronic pain. HEAL's innovate research portfolio spans the spectrum from therapeutic discovery and development through clinical trials and into clinical practice.

The Partnership for Accelerating Cancer Therapies.

As a part of the Cancer Moonshot, the National Cancer Institute, part of the National Institutes of Health, the Foundation for National Institutes of Health, the US Food and Drug Administration, and 12 pharmaceutical companies have formed a 5-year, $220 million precompetitive public-private research collaboration called the Partnership for Accelerating Cancer Therapies. A systematic cross-sector effort to identify and develop robust, standardized biomarkers and related clinical data, Partnership for Accelerating Cancer Therapies will support the selection and testing of promising immunotherapies for the treatment of cancer, with the goal of bringing effective therapy to more patients.

Induction of innate immune memory via microRNA targeting of chromatin remodelling factors.

Prolonged exposure to microbial products such as lipopolysaccharide can induce a form of innate immune memory that blunts subsequent responses to unrelated pathogens, known as lipopolysaccharide tolerance. Sepsis is a dysregulated systemic immune response to disseminated infection that has a high mortality rate. In some patients, sepsis results in a period of immunosuppression (known as 'immunoparalysis')1 characterized by reduced inflammatory cytokine output2, increased secondary infection3 and an increased risk of organ failure and mortality4. Lipopolysaccharide tolerance recapitulates several key features of sepsis-associated immunosuppression5. Although various epigenetic changes have previously been observed in tolerized macrophages6-8, the molecular basis of tolerance, immunoparalysis and other forms of innate immune memory has remained unclear. Here we perform a screen for tolerance-associated microRNAs and identify miR-221 and miR-222 as regulators of the functional reprogramming of macrophages during lipopolysaccharide tolerization. Prolonged stimulation with lipopolysaccharide in mice leads to increased expression of miR-221 and mir-222, both of which regulate brahma-related gene 1 (Brg1, also known as Smarca4). This increased expression causes the transcriptional silencing of a subset of inflammatory genes that depend on chromatin remodelling mediated by SWI/SNF (switch/sucrose non-fermentable) and STAT (signal transducer and activator of transcription), which in turn promotes tolerance. In patients with sepsis, increased expression of miR-221 and miR-222 correlates with immunoparalysis and increased organ damage. Our results show that specific microRNAs can regulate macrophage tolerization and may serve as biomarkers of immunoparalysis and poor prognosis in patients with sepsis.

SLP-76 is required for optimal CXCR4-stimulated T lymphocyte firm arrest to ICAM-1 under shear flow.

Rapid arrest of T cells at target sites upon engagement of chemokine receptors is crucial to the proper functioning of the immune system. Although T-cell arrest always occurs under hydrodynamic forces in vivo, most studies investigating the molecular mechanisms of arrest have been performed under static conditions. While the requirement of the adapter protein SLP-76 (Src homology 2-domain containing leukocyte-specific phosphoprotein of 76 kDa) in TCR-induced integrin activation has been demonstrated, its role in chemokine-triggered T-cell adhesion is unknown. Using a flow chamber system, we show that SLP-76 plays an important role in regulating the transition from tethering and rolling to firm adhesion of T cells under physiological shear flow in response to CXCL12α (stromal cell-derived factor-1α); SLP-76-deficient primary T cells exhibited defective adhesion with a significant decrease in the number of firmly arrested cells. We further demonstrate the N-terminal phosphotyrosines of SLP-76 play a critical role in T-cell adhesion under flow. These findings reveal a novel role for SLP-76 in CXCR4-mediated T lymphocyte trafficking.

NF-κB, inflammation, and metabolic disease.

Metabolic disorders including obesity, type 2 diabetes, and atherosclerosis have been viewed historically as lipid storage disorders brought about by overnutrition. It is now widely appreciated that chronic low-grade inflammation plays a key role in the initiation, propagation, and development of metabolic diseases. Consistent with its central role in coordinating inflammatory responses, numerous recent studies have implicated the transcription factor NF-κB in the development of such diseases, thereby further establishing inflammation as a critical factor in their etiology and offering hope for the development of new therapeutic approaches for their treatment.

Independent and cooperative roles of adaptor molecules in proximal signaling during FcepsilonRI-mediated mast cell activation.

Activation through FcepsilonRI, a high-affinity IgE-binding receptor, is critical for mast cell function during allergy. The formation of a multimolecular proximal signaling complex nucleated by the adaptor molecules SLP-76 and LAT1 is required for activation through this receptor. Based on previous T-cell studies, current dogma dictates that LAT1 is required for plasma membrane recruitment and function of SLP-76. Unexpectedly, we found that the recruitment and phosphorylation of SLP-76 were preserved in LAT1(-/-) mast cells and that SLP-76(-/-) and LAT1(-/-) mast cells harbored distinct functional and biochemical defects. The LAT1-like molecule LAT2 was responsible for the preserved membrane localization and phosphorylation of SLP-76 in LAT1(-/-) mast cells. Although LAT2 supported SLP-76 phosphorylation and recruitment to the plasma membrane, LAT2 only partially compensated for LAT1-mediated cell signaling due to its decreased ability to stabilize interactions with phospholipase Cgamma (PLCgamma). Comparison of SLP-76(-/-) LAT1(-/-) and SLP-76(-/-) mast cells revealed that some functions of LAT1 could occur independently of SLP-76. We propose that while SLP-76 and LAT1 depend on each other for many of their functions, LAT2/SLP-76 interactions and SLP-76-independent LAT1 functions also mediate a positive signaling pathway downstream of FcepsilonRI in mast cells.

Cutting edge: rescue of pre-TCR but not mature TCR signaling in mice expressing membrane-targeted SLP-76.

SLP-76 (Src homology 2 domain-containing leukocyte phosphoprotein of 76 kDa) organizes signaling from immunoreceptors, including the platelet collagen receptor, the pre-TCR, and the TCR, and is required for T cell development. In this study we examine a mouse in which wild-type SLP-76 is replaced with a mutant constitutively targeted to the cell membrane. Membrane-targeted SLP-76 (MTS) supports ITAM signaling in platelets and from the pre-TCR. Signaling from the mature TCR, however, is defective in MTS thymocytes, resulting in failed T cell differentiation. Defective thymic selection by MTS is not rescued by a SLP-76 mutant whose localization is restricted to the cytosol. Thus, fixed localization of SLP-76 reveals differential requirements for the subcellular localization of signaling complexes downstream of the pre-TCR vs mature TCR.

Regulation of T cell integrin function by adapter proteins.

Integrins are cell surface heterodimers that bind adhesion molecules expressed on other cells or in the extracellular matrix. Integrin-mediated interactions are critical for T cell development in the thymus, migration of T cells in the periphery, and induction of T cell effector functions. In resting T cells, integrins are maintained in a low affinity state. Engagement of the T cell receptor or chemokine receptors increases integrin affinity, enabling integrins to bind their ligands and initiate a signaling cascade resulting in altered cell morphology and motility. Our laboratory is interested how adapter proteins, mediators of intracellular signal transduction, regulate both signals from the T cell receptor to integrins (inside-out signaling) and (outside-in) signals from integrins into the cell.

Indirect involvement of allergen-captured mast cells in antigen presentation.

It is generally thought that mast cells influence T-cell activation nonspecifically through the release of inflammatory mediators. In this report, we provide evidence that mast cells may also affect antigen-specific T-cell responses by internalizing immunoglobulin E-bound antigens for presentation to antigen-specific T cells. Surprisingly, T-cell activation did not require that mast cells express major histocompatibility complex class II, indicating that mast cells were not involved in the direct presentation of the internalized antigens. Rather, the antigen captured by mast cells is presented by other major histocompatibility complex class II(+) antigen-presenting cells. To explore how this may occur, we investigated the fate of mast cells stimulated by antigen and found that FcepsilonRI crosslinking enhances mast cell apoptosis. Cell death by antigen-captured mast cells was required for efficient presentation because protection of mast cell death significantly decreased T-cell activation. These results suggest that mast cells may be involved in antigen presentation by acting as an antigen reservoir after antigen capture through specific immunoglobulin E molecules bound to their FcepsilonRI. This mechanism may contribute to how mast cells impact the development of T-cell responses.