Transforming growth factor-beta (TGF-β) induces HIV-1 restriction in Langerhans cells

Transforming growth factor-beta (TGF-β) drives the development of immature LC from hematopoietic progenitor cells and shapes the cells functions. Here I showed that two LC model cells, MuLC and MDLC, used exchangeably in the research, differ significantly in their phenotype and immune responses. Discrepancies between these models were specifically visible during stimulation with type-I IFN, where MuLC failed to up-regulate ISG levels. Yet both MuLC and MDLC demonstrated low susceptibility to HIV-1 infection, even in the absence of SAMHD-1. This post-entry restriction was conferred by the action of TGF-β on differentiation cells as indicated by our study. Indeed, in the absence of TGF-β supplementation, derived cells showed MDDC phenotype related to high susceptibility of the cells to HIV-1 infection during co-infection with SIV-Vpx. Additionally blocking of the TGF-β signalling, reversed the restrictive phenotype of LC. Importantly this pattern was also confirmed in skin extracted real epidermal LC versus dermal DC, suggesting that SAMHD-1-independent restriction activity operates in TGF-β derived cells. Accordingly to PCR analysis virus replication in LC is interrupted prior to integration, suggesting the role of additional restriction factors at early stages of virus infection or lack of essential viral dependency factors such as dNTPs. Interestingly maturation of MDLC with a synthetic bacterial triacylated lipopeptide or TNF-alpha significantly increased their susceptibility to HIV-1 infection, which may explain why HIV-1 acquisition is increased during co-infection with other STIs. In summary, our study strongly supports the action of SAMHD-1-independent HIV-1 restriction mechanisms in LC. A better understanding of the balance between HIV-1 restriction and propagation from LC to CD4+ T cells may help in the development of new microbicides or vaccines to curb HIV-1 infection at its earliest stages

Effects of creatine supplementation on brain in the healthy population

Introduction and purpose:  Creatine supplementation is playing an increasingly important role not only in sports, but is beneficial in the context of brain health (e.g., cognitive processing, brain function and recovery from injury). This is a growing field of research, and the purpose of this brief review is to provide an update on the effects of creatine supplementation on brain health in healthy people. Description of the state of knowledge: Creatine, is an organic compound produced mainly by the liver and kidneys. From the liver, kidneys and gastrointestinal tract, creatine enters through the bloodstream into skeletal muscle, where a total of about 95% of the body's total creatine is stored. It is stored in the muscles, heart and brain. Summary: There is potential for creatine supplementation to improve cognitive processing, especially in conditions characterized by creatine deficits in the brain, which can be triggered by acute stress factors (e.g., exercise, lack of sleep) or chronic, pathological conditions (e.g., creatine synthesis enzyme deficiencies, mild traumatic brain injury, aging, Alzheimer's disease). People with depression, the elderly, people with diseases associated with certain genetic defects (where creatine production and storage in the brain is impaired) are just some of the groups of people in whom creatine supplementation may be helpful. More research is needed to determine the clinical impact of long-term creatine supplementation dosing strategies on brain function and health

Langerin (CD207) represents a novel interferon-stimulated gene in Langerhans cells

Interferons (IFNs) are “warning signal” cytokines released upon pathogen sensing. IFNs control the expression of interferon-stimulated genes (ISGs), which are often crucial to restrict viral infections and establish a cellular antiviral state.1,2 Langerin (CD207), a well-known surface receptor on Langerhans cells (LC), belongs to the C-type lectin receptor (CLR) family and constitutes a major pathogen binding receptor able to regulate both innate and adaptive immune responses.3,4 Importantly, this CLR was reported as an antiviral receptor, notably able to bind and internalize incoming human immunodeficiency virus (HIV) virions in Birbeck granules for degradation.5,6 However, langerin was never viewed as a contributor to the interferon-mediated antiviral immune response. We now provide evidence that langerin is an ISG showing upregulated expression upon IFN treatment in monocyte-derived and ex vivo human skin-isolated LCs

Effective in vivo gene modification in mouse tissue-resident peritoneal macrophages by intraperitoneal delivery of lentiviral vectors

Tissue-resident macrophages exhibit specialized phenotypes dependent on their in vivo physiological niche. Investigation of their function often relies upon complex whole mouse transgenic studies. While some appropriate lineage-associated promoters exist, there are no options for tissue-specific targeting of macrophages. We have developed full protocols for in vivo productive infection (defined by stable transgene expression) of tissue-resident macrophages with lentiviral vectors, enabling RNA and protein overexpression, including expression of small RNA species such as shRNA, to knock down and modulate gene expression. These approaches allow robust infection of peritoneal tissue-resident macrophages without significant infection of other cell populations. They permit rapid functional study of macrophages in homeostatic and inflammatory settings, such as thioglycolate-induced peritonitis, while maintaining the cells in their physiological context. Here we provide detailed protocols for the whole workflow: viral production, purification, and quality control; safety considerations for administration of the virus to mice; and assessment of in vivo transduction efficiency and the low background levels of inflammation induced by the virus. In summary, we present a quick and accessible protocol for the rapid assessment of gene function in peritoneal tissue-resident macrophages in vivo

TGFβ induces a SAMHD1-independent post-entry restriction to HIV-1 infection of human epithelial Langerhans cells

Sterile alpha motif (SAM) and histidine-aspartic (HD) domains protein 1 (SAMHD1) was previously identified as a critical post-entry restriction factor to HIV-1 infection in myeloid dendritic cells. Here we show that SAMHD1 is also expressed in epidermis-isolated Langerhans cells (LC), but degradation of SAMHD1 does not rescue HIV-1 or vesicular stomatitis virus G-pseudotyped lentivectors infection in LC. Strikingly, using Langerhans cells model systems (mutz-3-derived LC, monocyte-derived LC [MDLC], and freshly isolated epidermal LC), we characterize previously unreported post-entry restriction activity to HIV-1 in these cells, which acts at HIV-1 reverse transcription, but remains independent of restriction factors SAMHD1 and myxovirus resistance 2 (MX2). We demonstrate that transforming growth factor-β signaling confers this potent HIV-1 restriction in MDLC during their differentiation and blocking of mothers against decapentaplegic homolog 2 (SMAD2) signaling in MDLC restores cells’ infectivity. Interestingly, maturation of MDLC with a toll-like receptor 2 agonist or transforming growth factor-α significantly increases cells’ susceptibility to HIV-1 infection, which may explain why HIV-1 acquisition is increased during coinfection with sexually transmitted infections. In conclusion, we report a SAMHD1-independent post-entry restriction in MDLC and LC isolated from epidermis, which inhibits HIV-1 replication. A better understanding of HIV-1 restriction and propagation from LC to CD4+ T cells may help in the development of new microbicides or vaccines to curb HIV-1 infection at its earliest stages during mucosal transmission

Tissue‐resident macrophages actively suppress IL‐1beta release via a reactive prostanoid/IL‐10 pathway

The alarm cytokine interleukin‐1β (IL‐1β) is a potent activator of the inflammatory cascade following pathogen recognition. IL‐1β production typically requires two signals: first, priming by recognition of pathogen‐associated molecular patterns leads to the production of immature pro‐IL‐1β; subsequently, inflammasome activation by a secondary signal allows cleavage and maturation of IL‐1β from its pro‐form. However, despite the important role of IL‐1β in controlling local and systemic inflammation, its overall regulation is still not fully understood. Here we demonstrate that peritoneal tissue‐resident macrophages use an active inhibitory pathway, to suppress IL‐1β processing, which can otherwise occur in the absence of a second signal. Programming by the transcription factor Gata6 controls the expression of prostacyclin synthase, which is required for prostacyclin production after lipopolysaccharide stimulation and optimal induction of IL‐10. In the absence of secondary signal, IL‐10 potently inhibits IL‐1β processing, providing a previously unrecognized control of IL‐1β in tissue‐resident macrophages

Lentiviral vector preparation for efficient gene and microRNA modulation of peritoneal cavity tissue-resident macrophages in vivo in mice

Peritoneal tissue-resident macrophages have broad functions in the maintenance of homeostasis and are involved in pathologies within local and neighboring tissues. Their functions are dictated by microenvironmental cues; thus, it is essential to investigate their behavior in an in vivo physiological niche. Currently, specific peritoneal macrophage-targeting methodologies employ whole-mouse transgenic models. Here, a protocol for effective in vivo modulation of mRNA and small RNA species (e.g., microRNA) expression in peritoneal macrophages using lentivirus particles is described. Lentivirus preparations were made in HEK293T cells and purified on a single sucrose layer. In vivo validation of lentivirus effectivity following intraperitoneal injection revealed predominant infection of macrophages restricted to local tissue. Targeting of peritoneal macrophages was successful during homeostasis and thioglycolate-induced peritonitis. The limitations of the protocol, including low-level inflammation induced by intraperitoneal delivery of lentivirus and time restrictions for potential experiments, are discussed. Overall, this study presents a quick and accessible protocol for the rapid assessment of gene function in peritoneal macrophages in vivo

Tissue-specific transcriptional programming of macrophages controls the microRNA transcriptome targeting multiple functional pathways

Recent interest in the biology and function of peritoneal tissue resident macrophages (pMΦ) has led to a better understanding of their cellular origin, programming and renewal. The programming of pMΦ is dependent on microenvironmental cues and tissue specific transcription factors, including GATA6. However, the contribution of microRNAs remains poorly defined. We conducted a detailed analysis of the impact of GATA6-deficiency on microRNA expression in mouse pMΦ. Our data suggest that for many of the pMΦ, microRNA composition may be established during tissue specialization, and that the effect of GATA6 knockout is largely unable to be rescued in the adult by exogenous GATA6. The data are consistent with GATA6 modulating the expression pattern of specific microRNAs, directly or indirectly, and including miR-146a, -223, and -203 established by the lineage-determining transcription factor PU.1, to achieve a differentiated pMΦ phenotype. Lastly, we showed a significant dysregulation of miR-708 in pMΦ in the absence of GATA6 during homeostasis and in response to LPS/IFN-γ stimulation. Overexpression of miR-708 in mouse pMΦ in vivo altered 167 mRNA species demonstrating functional downregulation of predicted targets, including cell immune responses and cell cycle regulation. In conclusion, we demonstrate dependence of the microRNA transcriptome on tissue-specific programming of tissue macrophages as exemplified by the role of GATA6 in pMΦ specialization