Chemokines and lymphocyte homing in Sjögren's syndrome.

Sjögren's syndrome (SS) is a chronic systemic autoimmune disease that typically presents with lymphocyte, dendritic cell, and macrophage infiltration of exocrine gland ducts and the formation of ectopic germinal centers. The interactions of lymphocyte homing receptors and addressins and chemokines and their receptors, such as α4ß7/MAdCAM-1, LFA-1/ICAM-1, CXCL13/CXCR5, CCL25/CCR9, CX3CL1/CX3CR1, play important roles in the migration of inflammatory cells to the focal glands and the promotion of ectopic germinal center formation in SS. A variety of molecules have been shown to be involved in lymphocyte homing, including tumor necrosis factor-α, interferon (IFN)-α, IFN-ß, and B cell activating factor. This process mainly involves the Janus kinase-signal transducer and activator of transcription signaling pathway, lymphotoxin-ß receptor pathway, and nuclear factor-κB signaling pathway. These findings have led to the development of antibodies to cell adhesion molecules, antagonists of chemokines and their receptors, compounds interfering with chemokine receptor signaling, and gene therapies targeting chemokines and their receptors, providing new targets for the treatment of SS in humans. The aim of this study was to explore the relationship between lymphocyte homing and the pathogenesis of SS, and to provide a review of recent studies addressing lymphocyte homing in targeted therapy for SS.

Design, Pharmacological Characterization, and Molecular Docking of Minimalist Peptidomimetic Antagonists of α4ß1 Integrin.

Integrin receptors mediate cell-cell interactions via the recognition of cell-adhesion glycoproteins, as well as via the interactions of cells with proteins of the extracellular matrix, and upon activation they transduce signals bi-directionally across the cell membrane. In the case of injury, infection, or inflammation, integrins of ß2 and α4 families participate in the recruitment of leukocytes, a multi-step process initiated by the capturing of rolling leukocytes and terminated by their extravasation. In particular, α4ß1 integrin is deeply involved in leukocyte firm adhesion preceding extravasation. Besides its well-known role in inflammatory diseases, α4ß1 integrin is also involved in cancer, being expressed in various tumors and showing an important role in cancer formation and spreading. Hence, targeting this integrin represents an opportunity for the treatment of inflammatory disorders, some autoimmune diseases, and cancer. In this context, taking inspiration from the recognition motives of α4ß1 integrin with its natural ligands FN and VCAM-1, we designed minimalist α/ß hybrid peptide ligands, with our approach being associated with a retro strategy. These modifications are expected to improve the compounds' stability and bioavailability. As it turned out, some of the ligands were found to be antagonists, being able to inhibit the adhesion of integrin-expressing cells to plates coated with the natural ligands without inducing any conformational switch and any activation of intracellular signaling pathways. An original model structure of the receptor was generated using protein-protein docking to evaluate the bioactive conformations of the antagonists via molecular docking. Since the experimental structure of α4ß1 integrin is still unknown, the simulations might also shed light on the interactions between the receptor and its native protein ligands.

Preclinical Evaluation of 89Zr-Desferrioxamine-Bexmarilimab, a Humanized Antibody Against Common Lymphatic Endothelial and Vascular Endothelial Receptor-1, in a Rabbit Model of Renal Fibrosis.

Bexmarilimab is a new humanized monoclonal antibody against common lymphatic endothelial and vascular endothelial receptor-1 (CLEVER-1) and is in clinical trials for macrophage-guided cancer immunotherapy. In addition being associated with cancer, CLEVER-1 is also associated with fibrosis. To facilitate prospective human PET studies, we preclinically evaluated 89Zr-labeled bexmarilimab in rabbits. Methods: Bexmarilimab was conjugated with desferrioxamine (DFO) and radiolabeled with 89Zr. Retained immunoreactivity was confirmed by flow cytometry. The distribution kinetics of intravenously administered 89Zr-DFO-bexmarilimab (0.1 mg/kg) were determined for up to 7 d in a rabbit model of renal fibrosis mediated by unilateral ureteric obstruction. The in vivo stability of 89Zr-DFO-bexmarilimab was evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with autoradiography. Additionally, we estimated the human radiation dose from data obtained in healthy rabbits. Results: 89Zr-DFO-bexmarilimab cleared rapidly from the blood circulation and distributed to the liver and spleen. At 24 h after injection, PET/CT, ex vivo γ-counting, and autoradiography demonstrated that there was significantly higher 89Zr-DFO-bexmarilimab uptake in unilateral ureteric obstruction-operated fibrotic renal cortex, characterized by abundant CLEVER-1-positive cells, than in contralateral or healthy kidneys. The estimated effective dose for a 70-kg human was 0.70 mSv/MBq. Conclusion: The characteristics of 89Zr-DFO-bexmarilimab support future human PET studies to, for example, stratify patients for bexmarilimab treatment, evaluate the efficacy of treatment, or monitor disease progression.

The C-terminal peptide of CCL21 drastically augments CCL21 activity through the dendritic cell lymph node homing receptor CCR7 by interaction with the receptor N-terminus.

The endogenous chemokines CCL19 and CCL21 signal via their common receptor CCR7. CCL21 is the main lymph node homing chemokine, but a weak chemo-attractant compared to CCL19. Here we show that the 41-amino acid positively charged peptide, released through C-terminal cleavage of CCL21, C21TP, boosts the immune cell recruiting activity of CCL21 by up to 25-fold and the signaling activity via CCR7 by ~ 100-fold. Such boosting is unprecedented. Despite the presence of multiple basic glycosaminoglycan (GAG) binding motifs, C21TP boosting of CCL21 signaling does not involve interference with GAG mediated cell-surface retention. Instead, boosting is directly dependent on O-glycosylations in the CCR7 N-terminus. As dictated by the two-step binding model, the initial chemokine binding involves interaction of the chemokine fold with the receptor N-terminus, followed by insertion of the chemokine N-terminus deep into the receptor binding pocket. Our data suggest that apart from a role in initial chemokine binding, the receptor N-terminus also partakes in a gating mechanism, which could give rise to a reduced ligand activity, presumably through affecting the ligand positioning. Based on experiments that support a direct interaction of C21TP with the glycosylated CCR7 N-terminus, we propose that electrostatic interactions between the positively charged peptide and sialylated O-glycans in CCR7 N-terminus may create a more accessible version of the receptor and thus guide chemokine docking to generate a more favorable chemokine-receptor interaction, giving rise to the peptide boosting effect.

Layilin Anchors Regulatory T Cells in Skin.

Regulatory T cells (Tregs) reside in nonlymphoid tissues where they carry out unique functions. The molecular mechanisms responsible for Treg accumulation and maintenance in these tissues are relatively unknown. Using an unbiased discovery approach, we identified LAYN (layilin), a C-type lectin-like receptor, to be preferentially and highly expressed on a subset of activated Tregs in healthy and diseased human skin. Expression of layilin on Tregs was induced by TCR-mediated activation in the presence of IL-2 or TGF-ß. Mice with a conditional deletion of layilin in Tregs had reduced accumulation of these cells in tumors. However, these animals somewhat paradoxically had enhanced immune regulation in the tumor microenvironment, resulting in increased tumor growth. Mechanistically, layilin expression on Tregs had a minimal effect on their activation and suppressive capacity in vitro. However, expression of this molecule resulted in a cumulative anchoring effect on Treg dynamic motility in vivo. Taken together, our results suggest a model whereby layilin facilitates Treg adhesion in skin and, in doing so, limits their suppressive capacity. These findings uncover a unique mechanism whereby reduced Treg motility acts to limit immune regulation in nonlymphoid organs and may help guide strategies to exploit this phenomenon for therapeutic benefit.

Titanium Nanoparticles Enhance Production and Suppress Stabilin-1-Mediated Clearance of GDF-15 in Human Primary Macrophages.

Macrophages are key innate immune cells that mediate implant acceptance or rejection. Titanium implants degrade over time inside the body, which results in the release of implant wear-off particles. Titanium nanoparticles (TiNPs) favor pro-inflammatory macrophage polarization (M1) and lower tolerogenic activation (M2). GDF-15 regulates immune tolerance and fibrosis and is endocytosed by stabilin-1. How TiNPs affect the healing activities of macrophages and their release of circulating cytokines is an open question in regenerative medicine. In this study for the first time, we identified the transcriptional program induced and suppressed by TiNPs in human pro-inflammatory and healing macrophages. Microarray analysis revealed that TiNPs altered the expression of 5098 genes in M1 (IFN-γ-stimulated) and 4380 genes in M2 (IL-4-stimulated) macrophages. 1980 genes were differentially regulated in both M1 and M2. Affymetrix analysis, confirmed by RT-PCR, demonstrated that TiNPs upregulate expression of GDF-15 and suppress stabilin-1, scavenger receptor of GDF-15. TiNPs also significantly stimulated GDF-15 protein secretion in inflammatory and healing macrophages. Flow cytometry demonstrated, that scavenging activity of stabilin-1 was significantly suppressed by TiNPs. Confocal microscopy analysis showed that TiNPs impair internalization of stabilin-1 ligand acLDL and its transport to the endocytic pathway. Our data demonstrate that TiNPs have a dual effect on the GDF-15/stabilin-1 interaction in macrophage system, by increasing the production of GDF-15 and suppressing stabilin-1-mediated clearance function. In summary, this process can result in a significant increase of GDF-15 in the extracellular space and in circulation leading to unbalanced pro-fibrotic reactions and implant complications.

A molecular cell atlas of the human lung from single-cell RNA sequencing.

Although single-cell RNA sequencing studies have begun to provide compendia of cell expression profiles1-9, it has been difficult to systematically identify and localize all molecular cell types in individual organs to create a full molecular cell atlas. Here, using droplet- and plate-based single-cell RNA sequencing of approximately 75,000 human cells across all lung tissue compartments and circulating blood, combined with a multi-pronged cell annotation approach, we create an extensive cell atlas of the human lung. We define the gene expression profiles and anatomical locations of 58 cell populations in the human lung, including 41 out of 45 previously known cell types and 14 previously unknown ones. This comprehensive molecular atlas identifies the biochemical functions of lung cells and the transcription factors and markers for making and monitoring them; defines the cell targets of circulating hormones and predicts local signalling interactions and immune cell homing; and identifies cell types that are directly affected by lung disease genes and respiratory viruses. By comparing human and mouse data, we identified 17 molecular cell types that have been gained or lost during lung evolution and others with substantially altered expression profiles, revealing extensive plasticity of cell types and cell-type-specific gene expression during organ evolution including expression switches between cell types. This atlas provides the molecular foundation for investigating how lung cell identities, functions and interactions are achieved in development and tissue engineering and altered in disease and evolution.

Therapeutic Cytokine Inhibition Modulates Activation and Homing Receptors of Peripheral Memory B Cell Subsets in Rheumatoid Arthritis Patients.

Memory B cells have known to play an important role in the pathogenesis of rheumatoid arthritis (RA). With the emergence of B cell-targeted therapies, the modulation of memory B cells appears to be a key therapeutic target. Human peripheral memory B cells can be distinguished based on the phenotypic expression of CD27 and IgD, characterizing the three major B cell subpopulations: CD27+IgD+ pre-switch, CD27+IgD- post-switch, and CD27-IgD- double-negative memory B cells. We evaluated different memory cell populations for activation markers (CD95 and Ki-67) and chemokine receptors (CXCR3 and 4) expressing B cells in active RA, as well as under IL6-R blockade by tocilizumab (TCZ) and TNF-α blockade by adalimumab (ADA). Memory B cells were phenotypically analyzed from RA patients at baseline, week 12, and week 24 under TCZ or ADA treatment, respectively. Using flow cytometry, surface expression of CD95, intracellular Ki-67, and surface expressions of CXCR3 and CXCR4 were determined. Compared with healthy donors (n = 40), the phenotypic analysis of RA patients (n = 80) demonstrated that all three types of memory B cells were activated in RA patients. Surface and intracellular staining of B cells showed a significantly higher percentage of CD95+ (p < 0.0001) and Ki-67+ (p < 0.0001) cells, with numerically altered CXCR3+ and CXCR4+ cells in RA. CD95 and Ki-67 expressions were highest in post-switch memory B cells, whereas CD19+CXCR3+ and CD19+CXCR4+ expressing cells were substantially higher in the pre-switch compartment. In all subsets of the memory B cells, in vivo IL-6R, and TNF-α blockade significantly reduced the enhanced expressions of CD95 and Ki-67. Based on our findings, we conclude that the three major peripheral memory B cell populations, pre-, post-switch, and double-negative B cells, are activated in RA, demonstrating enhanced CD95 and Ki-67 expressions, and varied expression of CXCR3 and CXCR4 chemokine receptors when compared with healthy individuals. This activation can be efficaciously modulated under cytokine inhibition in vivo.

Single-cell sequencing reveals clonal expansions of pro-inflammatory synovial CD8 T cells expressing tissue-homing receptors in psoriatic arthritis.

Psoriatic arthritis (PsA) is a debilitating immune-mediated inflammatory arthritis of unknown pathogenesis commonly affecting patients with skin psoriasis. Here we use complementary single-cell approaches to study leukocytes from PsA joints. Mass cytometry demonstrates a 3-fold expansion of memory CD8 T cells in the joints of PsA patients compared to peripheral blood. Meanwhile, droplet-based and plate-based single-cell RNA sequencing of paired T cell receptor alpha and beta chain sequences show pronounced CD8 T cell clonal expansions within the joints. Transcriptome analyses find these expanded synovial CD8 T cells to express cycling, activation, tissue-homing and tissue residency markers. T cell receptor sequence comparison between patients identifies clonal convergence. Finally, chemokine receptor CXCR3 is upregulated in the expanded synovial CD8 T cells, while two CXCR3 ligands, CXCL9 and CXCL10, are elevated in PsA synovial fluid. Our data thus provide a quantitative molecular insight into the cellular immune landscape of psoriatic arthritis.

New tools to prevent cancer growth and spread: a 'Clever' approach.

Clever-1 (also known as Stabilin-1 and FEEL-1) is a scavenger receptor expressed on lymphatic endothelial cells, sinusoidal endothelial cells and immunosuppressive monocytes and macrophages. Its role in cancer growth and spread first became evident in Stab1-/- knockout mice, which have smaller primary tumours and metastases. Subsequent studies in mice and humans have shown that immunotherapeutic blockade of Clever-1 can activate T-cell responses, and that this response is mainly mediated by a phenotypic change in macrophages and monocytes from immunosuppressive to pro-inflammatory following Clever-1 inhibition. Analyses of human cancer cohorts have revealed marked associations between the number of Clever-1-positive macrophages and patient outcome. As hardly any reports to date have addressed the role of Clever-1 in immunotherapy resistance and T-cell dysfunction, we performed data mining using several published cancer cohorts, and observed a remarkable correlation between Clever-1 positivity and resistance to immune checkpoint therapies. This result provides impetus and potential for the ongoing clinical trial targeting Clever-1 in solid tumours, which has so far shown a shift towards immune activation when a particular epitope of Clever-1 is blocked.

Identifying the key genes and microRNAs in prostate cancer bone metastasis by bioinformatics analysis.

Prostate adenocarcinoma (PCa) is the most common cause of death due to malignancy among men, and bone metastasis is the leading cause of mortality in patients with PCa. Therefore, identifying the causes and molecular mechanism of bone metastasis is important for early detection, diagnosis and personalized therapy. In this study, we systematically analyzed molecular correlates of bone metastasis by bioinformatics analysis. A total of 12 differentially expressed microRNAs (miRNAs) and 102 differentially expressed genes were identified. Five miRNAs had prognostic significance in biochemical recurrence-free survival (miR-636, miR-491-5p, miR-199b-5p, miR-199b-3p, miR-28-3p). The differentially expressed genes were significantly enriched in extracellular matrix, cell-substrate adhesion, collagen and integrin. Seven hub genes (VCAN, COL3A1, COL1A1, APOE, COL1A2, SDC1, THY1) with worse biochemical recurrence-free survival and one hub gene (MMP9) with worse overall survival were detected. miR-636, a novel oncogene, was found to be up-regulated in bone metastatic PCa tissues and also predominately up-regulated in human PCa cell lines. miR-636 promoted cellular invasion and migration, and may promote bone metastasis via targeting MBNL2, TNS1 and STAB1. In conclusion, we have successfully defined molecular signatures of bone metastasis in PCa.

Microlocalization and clinical significance of stabilin-1+ macrophages in treatment-naïve patients with urothelial carcinoma of the bladder.

PURPOSE: Emerging evidence has shown that macrophages (Mφs) at different tumor sites have diverse clinical attributes. Stabilin-1 is a multi-functional scavenger marker for specialized tumor-associated Mφs. This study investigates the relationship between the density and microlocalization of stabilin-1+ Mφs within tumors and the clinical outcomes of patients with urothelial carcinoma of the bladder (UCB). METHODS: In this retrospective study, 283 UCB patients who received radical cystectomy or transurethral resection were examined. Immunohistochemistry and immunofluorescence analyses were used to colocalize the expression of stabilin-1 with other markers for Mφs (CD14, CD68, CD163, and CD206). Kaplan-Meier analysis and Cox proportional hazards regression models were applied to estimate overall survival (OS) and recurrence-free survival (RFS). RESULTS: In UCB tissues, stabilin-1 was primarily expressed on Mφs, as evident from triple immunofluorescence staining for stabilin-1 and Mφ markers. Stabilin-1+ Mφs were often more prominent in stromal regions rather than intratumoral regions in UCB tissues (P < 0.0001). After dichotomization at the median cell density for stabilin-1+ Mφs, only intratumoral stabilin-1+ Mφ density was a predictor of poor OS (P < 0.001) and RFS (P = 0.026). Moreover, intratumoral stabilin-1+ Mφ density was positively associated with tumor stage (P < 0.01) and histological grade (P < 0.01), and emerged as an independent prognostic factor for OS (HR 2.371; P < 0.0001), but not for RFS (HR 1.491; P = 0.061). CONCLUSIONS: Our findings indicate that intratumoral stabilin-1+ Mφs could potentially be used as a pro-tumoral prognostic marker for UCB patients.

The Role of Stabilin-1 in Lymphocyte Trafficking and Macrophage Scavenging in the Liver Microenvironment.

Chronic liver diseases are a major global health burden, and cases of these conditions continue to rise in many countries. A diverse range of insults can lead to chronic liver disease, but they are all characterised by the infiltration and accumulation of immune cells within liver tissue and, if progressive, can lead to tissue fibrosis and cirrhosis. In this review, we focus on the role of stabilin-1 in two key processes that contribute to liver disease, namely, the recruitment of lymphocytes into liver tissue and the response of macrophages to tissue injury. Stabilin-1 is constitutively expressed on the sinusoidal endothelium of the liver and contributes to the homeostatic scavenging function of these cells. Epithelial damage in the context of chronic liver disease leads to the upregulation of stabilin-1 at sites of tissue injury, specifically at sites of immune cell recruitment and on subpopulations of hepatic macrophages. Functionally, stabilin-1 has been shown to mediate transendothelial migration of lymphocyte subsets in the setting of pro-inflammatory-activated human liver endothelium. In experimental models of liver fibrosis, stabilin-1 promotes the uptake of products of chronic oxidative stress by a subset of hepatic macrophages and suppresses their release of pro-inflammatory mediators that regulate tissue remodelling. These studies highlight the active contribution that scavenger receptors such as stabilin-1 can make in regulating chronic inflammation and tissue fibrosis, and their potential as novel therapeutic targets for these conditions.

Not CD68 but stabilin-1 expression is associated with the risk of recurrence in patients with oral cavity squamous cell carcinoma.

BACKGROUND: This study's aim was to identify CD68 and stabilin-1 expressions and their prognostic implications in patients with oral cavity cancer (OCC). METHODS: Stabilin-1 and CD68 immunochemical staining was conducted in samples from patients with OCC who underwent curative surgery in our institute. We retrospectively analyzed patients' clinical information. We investigated the relationship between CD68 and stabilin-1 expression levels and their effect on prognostic factors and survival. RESULTS: Fifty-four patients' data were analyzed. CD68 and stabilin-1 expressions were positively correlated (P = .008). CD68 and stabilin-1 expressions were not correlated with other prognostic factors (eg, T classification or lymph node metastasis). Stabilin-1 expression and tumor recurrence risk were positively correlated (P = .015). CD68 expression was neither associated with recurrence-free nor OCC-specific survival. CONCLUSION: CD68 and stabilin-1 expressions in OCC were positively correlated. Stabilin-1 expression was significantly associated with OCC recurrence risk, but the prognostic value of CD68 was not confirmed.

Surface nanocavitation of titanium modulates macrophage activity.

Background: Nanoscale surface modifications are widely touted to improve the biocompatibility of medically relevant materials. Immune cells, such as macrophages, play a critical role in the initial healing events following implantation. Methods: To understand the response of macrophages to nanotopography better, we exposed U937-derived macrophages to a distinctive mesoporous titanium surface (TiNano) produced by a process of simple chemical nanocavitation, and to mechanically polished titanium (TiPolished) and glass coverslip (Glass) surfaces as controls. Cell numbers and morphology were evaluated. Osteopontin expression and that of the proinflammatory SPARC protein and its stabilin 1 receptor were analyzed. Release of inflammation-associated cytokines and chemokines was also measured. Results: Compared to the two control surfaces, there were fewer U937 cells on TiNano, and these exhibited a more rounded morphology with long filopodia. The cells showed areas of punctate actin distribution, indicating formation of podosomes. Of the three proteins examined, only osteopontin's immunofluorescence signal was clearly reduced. Irrespective of the substrate, the cytokine assay revealed important variations in expression levels of the multiple molecules analyzed and downregulation in a number of chemokines by the TiNano surface. Conclusion: These results indicate that macrophages sense and respond to the physicochemical cueing generated by the nanocavitated surface, triggering cellular and molecular changes consistent with lesser inflammatory propensity. Given the previously reported beneficial outcome of this mesoporous surface on osteogenic activity, it could be presumed that modulation of the macrophagic response it elicits may also contribute to initial bone-integration events.

The Rho regulator Myosin IXb enables nonlymphoid tissue seeding of protective CD8+ T cells.

T cells are actively scanning pMHC-presenting cells in lymphoid organs and nonlymphoid tissues (NLTs) with divergent topologies and confinement. How the T cell actomyosin cytoskeleton facilitates this task in distinct environments is incompletely understood. Here, we show that lack of Myosin IXb (Myo9b), a negative regulator of the small GTPase Rho, led to increased Rho-GTP levels and cell surface stiffness in primary T cells. Nonetheless, intravital imaging revealed robust motility of Myo9b-/- CD8+ T cells in lymphoid tissue and similar expansion and differentiation during immune responses. In contrast, accumulation of Myo9b-/- CD8+ T cells in NLTs was strongly impaired. Specifically, Myo9b was required for T cell crossing of basement membranes, such as those which are present between dermis and epidermis. As consequence, Myo9b-/- CD8+ T cells showed impaired control of skin infections. In sum, we show that Myo9b is critical for the CD8+ T cell adaptation from lymphoid to NLT surveillance and the establishment of protective tissue-resident T cell populations.

The effect of neoadjuvant chemotherapy on the correlation of tumor-associated macrophages with CD31 and LYVE-1.

Angiogenesis and lymphangiogenesis play a crucial role in tumor growth, invasion and metastasis. Tumor-associated macrophages (TAM) induce both angiogenesis and lymphangiogenesis in mouse breast cancer models and positively correlate with these processes in human breast cancer patients. Neoadjuvant chemotherapy (NAC) is a widely used therapeutic option for cancer treatment. However, the effect of NAC on the distribution of TAM within intratumoral compartments and their correlation with angiogenesis and lymphangiogenesis remained unknown. In the present study we analyzed the effect of NAC on the distribution of CD68+ and stabilin-1+ TAM in five functionally distinct areas of human breast cancer and their correlations with microvessel density (MVD) and lymphatic microvessel density (LMVD), identified by CD31 and LYVE1, respectively. We found that NAC enhances blood vessel density in soft fibrous stroma and in coarse fibrous stroma. Without NAC the amount of CD68+ TAM in gaps of ductal tumor structures positively correlate with CD31+ microvessel density in soft fibrous stroma. NAC had enhancing effect on the amount of CD68+ TAM but not stabilin-1+ TAM in soft fibrous stroma. However, no correlation between TAM and CD31+ microvessel density was identified after NAC. NAC did not enhance the lymphatic microvessel density. But after NAC stabilin-1 expressing subpopulation of TAM positively correlated with expression of LYVE-1. We hypothesized that CD68+ TAM can support tumor angiogenesis primarily before NAC, while stabilin-1+ TAM can contribute to the maintenance of lymphatic microvessel density after NAC.

Cardiac CD68+ and stabilin-1+ macrophages in wound healing following myocardial infarction: From experiment to clinic.

Myocardial infarction (MI) remains the leading cause of mortality and morbidity throughout the world. Macrophages are key innate immune cells that play a significant role in transition from the inflammatory to the regenerative phase during wound healing following MI. The scavenger receptor stabilin-1 is one of the most interesting macrophage biomarkers. This receptor contributes to wound healing, angiogenesis, and tissue remodeling. We suggested a research protocol using macrophage biomarkers to study the cellular basis of cardiac remodeling and healing in patients with acute MI. The purpose of the research was to translate experimental knowledge regarding macrophage subsets and their biomarkers in post-infarction myocardial regeneration into results observed in clinical settings. The study included 41 patients with fatal MI type 1. All patients were divided into four groups according to the timeline of MI histopathology. In addition to routine histopathological analysis, macrophage infiltration was assessed by immunohistochemistry. We used CD68 as a marker for the cells of the macrophage lineage and stabilin-1 as an M2-like macrophage biomarker. The number of CD68+ and stabilin-1+ macrophages in the infarct area increased and peaked in the regenerative phase and did not decrease in the late stage of MI. In the peri-infarct area, the number of CD68+ macrophages increased in the inflammatory phase, peaked during the reparative phase, and did not decrease in the late phase, while the number of stabilin-1+ macrophages increased in the regenerative phase and remained unchanged. Additionally, in the reparative phase, we observed increase in the number of CD68+ and stabilin-1+ macrophages in the non-infarct area. The research protocol suggested allowed us to translate experimental knowledge regarding macrophage subsets and their biomarkers in post-infarction myocardial regeneration into clinical data. Taken together, these results demonstrated biphasic cardiac macrophage response following acute MI somewhat similar to that in a murine model. The increase in stabilin-1+ macrophage infiltration noticed in the myocardium during the regenerative phase and the strong positive correlation between the number of these cells and timeline of MI histopathology enabled us to propose stabilin-1 as a diagnostic macrophage biomarker in myocardium wound healing in patients with acute MI.

Treatment-Naïve HIV-Infected Patients Have Fewer Gut-Homing ß7 Memory CD4 T Cells than Healthy Controls.

OBJECTIVES: The integrin α4ß7 is the gut-homing receptor for lymphocytes. It also is an important co-receptor for human immunodeficiency virus (HIV) via glycoprotein (gp)120 binding. Depletion of gut cluster of differentiation (CD)4 T cells is linked to chronic inflammation in patients with HIV; however, measuring CD4 cells in the gut is invasive and not routine. As such, establishing a peripheral marker for CD4 depletion of the gut is needed. We hypothesized that α4ß7 CD4 T cells are depleted in the peripheral blood of treatment-naïve patients with HIV compared with healthy controls. METHODS: The study groups were treatment-naïve patients with HIV and uninfected controls. Subjects were included if they were 18 years or older with no history of opportunistic infections, active tuberculosis, or cancer. We collected peripheral blood and examined on whole blood using flow cytometry for the following cell surface markers: CD4, CD45RO, chemokine receptor type 5, C-X-C chemokine receptor type 4 (CXCR4), and the integrin ß7. We collected demographic information, including age, sex, and ethnicity, as well as viral load (VL) and CD4 count. Two-sample t tests and Fisher exact tests were used to compare the differences between the two groups. Spearman correlation coefficients were calculated between CD4 count and log10- VL and percentage of CD4+/CD45RO+/ß7+ and log10- VL in patients. RESULTS: Twenty-two subjects were enrolled in the study (12 patients with HIV and 10 controls). There were no differences in age or sex between the two groups. There were more Hispanics and fewer Asians in the group comprising patients with HIV compared with the control group (7 vs 2 and 0 vs 4, P = 0.05, respectively). Patients infected with HIV had significantly lower frequencies of CD4+/CD45RO+/ß7+ cells (median 12%, range 5-18 compared with uninfected controls: median 20%, range 11-26, P = 0.0007). There was a statistically significant difference in the percentage of CD4+/CD45RO+/C-X-C chemokine receptor type 4+ cells between patients (72%, range 60%-91%) compared with controls (79%, range 72%-94%, P = 0.04). The percentage of CD4+/CD45RO+/chemokine receptor type 5+ did not differ between the group of patients with HIV and the control groups (22%, range 11%-57% vs 27%, range 14%-31%; P = 0.8, respectively). There was no correlation between percentage of CD4+/CD45RO+/ß+ cells and log10- VL as measured by the Spearman correlation coefficient (r = 0.05, P = 0.88) in patients infected with HIV. CONCLUSIONS: Memory CD4 ß7+ cells are reduced significantly in the peripheral blood of untreated patients infected with HIV, which could be used as a noninvasive indicator of intestinal CD4 T cell loss and recovery. Further studies are needed to examine whether depletion of these CD4+/CD45RO+/ß7+ cells in the peripheral blood parallels depletion in the gut of treatment-naïve patients with HIV and whether levels return to control levels after treatment.