Autoantibodies against chemokines post-SARS-CoV-2 infection correlate with disease course.

Infection with severe acute respiratory syndrome coronavirus 2 associates with diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse coronavirus disease 2019 (COVID-19) outcomes. Here we discovered that antibodies against specific chemokines were omnipresent post-COVID-19, were associated with favorable disease outcome and negatively correlated with the development of long COVID at 1 yr post-infection. Chemokine antibodies were also present in HIV-1 infection and autoimmune disorders, but they targeted different chemokines compared with COVID-19. Monoclonal antibodies derived from COVID-19 convalescents that bound to the chemokine N-loop impaired cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising chemokine antibodies may modulate the inflammatory response and thus bear therapeutic potential.

Recognition and inhibition of SARS-CoV-2 by humoral innate immunity pattern recognition molecules.

The humoral arm of innate immunity includes diverse molecules with antibody-like functions, some of which serve as disease severity biomarkers in coronavirus disease 2019 (COVID-19). The present study was designed to conduct a systematic investigation of the interaction of human humoral fluid-phase pattern recognition molecules (PRMs) with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Of 12 PRMs tested, the long pentraxin 3 (PTX3) and mannose-binding lectin (MBL) bound the viral nucleocapsid and spike proteins, respectively. MBL bound trimeric spike protein, including that of variants of concern (VoC), in a glycan-dependent manner and inhibited SARS-CoV-2 in three in vitro models. Moreover, after binding to spike protein, MBL activated the lectin pathway of complement activation. Based on retention of glycosylation sites and modeling, MBL was predicted to recognize the Omicron VoC. Genetic polymorphisms at the MBL2 locus were associated with disease severity. These results suggest that selected humoral fluid-phase PRMs can play an important role in resistance to, and pathogenesis of, COVID-19, a finding with translational implications.

HMGB1 promotes CXCL12-dependent egress of murine B cells from Peyer's patches in homeostasis.

High mobility group box-1 protein (HMGB1) is an alarmin that, once released, promotes inflammatory responses, alone and as a complex with the chemokine CXCL12. Here, we report that the HMGB1-CXCL12 complex plays an essential role also in homeostasis by controlling the migration of B lymphocytes. We show that extracellular HMGB1 is critical for the CXCL12-dependent egress of B cells from the Peyer's patches (PP). This promigratory function of the complex was restricted to the PPs, since HMGB1 was not required for B-cell migratory processes in other locations. Accordingly, we detected higher constitutive levels of the HMGB1-CXCL12 complex in PPs than in other lymphoid organs. HMGB1-CXCL12 in vivo inhibition was associated with a reduced basal IgA production in the gut. Collectively, our results demonstrate a role for the HMGB1-CXCL12 complex in orchestrating B-cell trafficking in homeostasis, and provide a novel target to control lymphocyte migration in mucosal immunity.

Impairment of CCR6+ and CXCR3+ Th Cell Migration in HIV-1 Infection Is Rescued by Modulating Actin Polymerization.

CD4+ T cell repopulation of the gut is rarely achieved in HIV-1-infected individuals who are receiving clinically effective antiretroviral therapy. Alterations in the integrity of the mucosal barrier have been indicated as a cause for chronic immune activation and disease progression. In this study, we present evidence that persistent immune activation causes impairment of lymphocytes to respond to chemotactic stimuli, thus preventing their trafficking from the blood stream to peripheral organs. CCR6+ and CXCR3+ Th cells accumulate in the blood of aviremic HIV-1-infected patients on long-term antiretroviral therapy, and their frequency in the circulation positively correlates to levels of soluble CD14 in plasma, a marker of chronic immune activation. Th cells show an impaired response to chemotactic stimuli both in humans and in the pathogenic model of SIV infection, and this defect is due to hyperactivation of cofilin and inefficient actin polymerization. Taking advantage of a murine model of chronic immune activation, we demonstrate that cytoskeleton remodeling, induced by okadaic acid, restores lymphocyte migration in response to chemokines, both in vitro and in vivo. This study calls for novel pharmacological approaches in those pathological conditions characterized by persistent immune activation and loss of trafficking of T cell subsets to niches that sustain their maturation and activities.

Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4.

The chemokine receptor, CXC chemokine receptor 4 (CXCR4), is selective for CXC chemokine ligand 12 (CXCL12), is broadly expressed in blood and tissue cells, and is essential during embryogenesis and hematopoiesis. CXCL14 is a homeostatic chemokine with unknown receptor selectivity and preferential expression in peripheral tissues. Here, we demonstrate that CXCL14 synergized with CXCL12 in the induction of chemokine responses in primary human lymphoid cells and cell lines that express CXCR4. Combining subactive concentrations of CXCL12 with 100-300 nM CXCL14 resulted in chemotaxis responses that exceeded maximal responses that were obtained with CXCL12 alone. CXCL14 did not activate CXCR4-expressing cells (i.e., failed to trigger chemotaxis and Ca2+ mobilization, as well as signaling via ERK1/2 and the small GTPase Rac1); however, CXCL14 bound to CXCR4 with high affinity, induced redistribution of cell-surface CXCR4, and enhanced HIV-1 infection by >3-fold. We postulate that CXCL14 is a positive allosteric modulator of CXCR4 that enhances the potency of CXCR4 ligands. Our findings provide new insights that will inform the development of novel therapeutics that target CXCR4 in a range of diseases, including cancer, autoimmunity, and HIV.-Collins, P. J., McCully, M. L., Martínez-Muñoz, L., Santiago, C., Wheeldon, J., Caucheteux, S., Thelen, S., Cecchinato, V., Laufer, J. M., Purvanov, V., Monneau, Y. R., Lortat-Jacob, H., Legler, D. F., Uguccioni, M., Thelen, M., Piguet, V., Mellado, M., Moser, B. Epithelial chemokine CXCL14 synergizes with CXCL12 via allosteric modulation of CXCR4.

Tumor cells express and maintain HMGB1 in the reduced isoform to enhance CXCR4-mediated migration.

During inflammation and tissue regeneration, the alarmin High Mobility Group Box 1 (HMGB1), in its reduced isoform, enhances the activity of the chemokine CXCL12, forming a heterocomplex that acts via the chemokine receptor CXCR4. Despite the established roles of both HMGB1 and CXCL12 in tumor progression and metastatic spread to distal sites, the role of the CXCL12/HMGB1 heterocomplex in cancer has never been investigated. By employing a newly established mass spectrometry protocol that allows an unambiguous distinction between reduced (red-HMGB1) and oxidized (ox-HMGB1) HMGB1 isoforms in cell lysates, we demonstrate that human epithelial cells derived from breast (MCF-7 and MDA-MB-231) and prostate (PC-3) cancer predominantly express red-HMGB1, while primary CD3+ T lymphocytes from peripheral blood express both HMGB1 isoforms. All these cancer cells release HMGB1 in the extracellular microenvironment together with varying concentrations of thioredoxin and thioredoxin reductase. The CXCL12/HMGB1 heterocomplex enhances, via CXCR4, the directional migration and invasiveness of cancer cells characterized by high metastatic potential that possess a fully active thioredoxin system, contributing to maintain red-HMGB1. On the contrary, cancer cells with low metastatic potential, lack thioredoxin reductase, promptly uptake CXCL12 and fail to respond to the heterocomplex. Our study demonstrates that the responsiveness of cancer cells to the CXCL12/HMGB1 heterocomplex, resulting in enhanced cell migration and invasiveness, depends on the maintenance of HMGB1 in its reduced isoform, and suggests disruption of the heterocomplex as a potential therapeutic target to inhibit invasion and metastatic spread in cancer therapies.

Patients with ankylosing spondylitis present a distinct CD8 T cell subset with osteogenic and cytotoxic potential.

OBJECTIVES: Ankylosing spondylitis (AS) is a chronic inflammatory rheumatic disease affecting mainly the axial skeleton. Peripheral involvement (arthritis, enthesitis and dactylitis) and extra-musculoskeletal manifestations, including uveitis, psoriasis and bowel inflammation, occur in a relevant proportion of patients. AS is responsible for chronic and severe back pain caused by local inflammation that can lead to osteoproliferation and ultimately spinal fusion. The association of AS with the human leucocyte antigen-B27 gene, together with elevated levels of chemokines, CCL17 and CCL22, in the sera of patients with AS, led us to study the role of CCR4+ T cells in the disease pathogenesis. METHODS: CD8+CCR4+ T cells isolated from the blood of patients with AS (n=76) or healthy donors were analysed by multiparameter flow cytometry, and gene expression was evaluated by RNA sequencing. Patients with AS were stratified according to the therapeutic regimen and current disease score. RESULTS: CD8+CCR4+ T cells display a distinct effector phenotype and upregulate the inflammatory chemokine receptors CCR1, CCR5, CX3CR1 and L-selectin CD62L, indicating an altered migration ability. CD8+CCR4+ T cells expressing CX3CR1 present an enhanced cytotoxic profile, expressing both perforin and granzyme B. RNA-sequencing pathway analysis revealed that CD8+CCR4+ T cells from patients with active disease significantly upregulate genes promoting osteogenesis, a core process in AS pathogenesis. CONCLUSIONS: Our results shed light on a new molecular mechanism by which T cells may selectively migrate to inflammatory loci, promote new bone formation and contribute to the pathological ossification process observed in AS.

The chemokine landscape: one system multiple shades.

Leukocyte trafficking is mainly governed by chemokines, chemotactic cytokines, which can be concomitantly produced in tissues during homeostatic conditions or inflammation. After the discovery and characterization of the individual chemokines, we and others have shown that they present additional properties. The first discoveries demonstrated that some chemokines act as natural antagonists on chemokine receptors, and prevent infiltration of leukocyte subsets in tissues. Later on it was shown that they can exert a repulsive effect on selective cell types, or synergize with other chemokines and inflammatory mediators to enhance chemokine receptors activities. The relevance of the fine-tuning modulation has been demonstrated in vivo in a multitude of processes, spanning from chronic inflammation to tissue regeneration, while its role in the tumor microenvironment needs further investigation. Moreover, naturally occurring autoantibodies targeting chemokines were found in tumors and autoimmune diseases. More recently in SARS-CoV-2 infection, the presence of several autoantibodies neutralizing chemokine activities distinguished disease severity, and they were shown to be beneficial, protecting from long-term sequelae. Here, we review the additional properties of chemokines that influence cell recruitment and activities. We believe these features need to be taken into account when designing novel therapeutic strategies targeting immunological disorders.

Tracking unlabeled cancer cells imaged with low resolution in wide migration chambers via U-NET class-1 probability (pseudofluorescence).

Cell migration is a pivotal biological process, whose dysregulation is found in many diseases including inflammation and cancer. Advances in microscopy technologies allow now to study cell migration in vitro, within engineered microenvironments that resemble in vivo conditions. However, to capture an entire 3D migration chamber for extended periods of time and with high temporal resolution, images are generally acquired with low resolution, which poses a challenge for data analysis. Indeed, cell detection and tracking are hampered due to the large pixel size (i.e., cell diameter down to 2 pixels), the possible low signal-to-noise ratio, and distortions in the cell shape due to changes in the z-axis position. Although fluorescent staining can be used to facilitate cell detection, it may alter cell behavior and it may suffer from fluorescence loss over time (photobleaching).Here we describe a protocol that employs an established deep learning method (U-NET), to specifically convert transmitted light (TL) signal from unlabeled cells imaged with low resolution to a fluorescent-like signal (class 1 probability). We demonstrate its application to study cancer cell migration, obtaining a significant improvement in tracking accuracy, while not suffering from photobleaching. This is reflected in the possibility of tracking cells for three-fold longer periods of time. To facilitate the application of the protocol we provide WID-U, an open-source plugin for FIJI and Imaris imaging software, the training dataset used in this paper, and the code to train the network for custom experimental settings.

The interplay between previous infection and mental health condition on antibody response to COVID-19 mRNA vaccination.

Increasing evidence has been pointing towards the existence of a bi-directional interplay between mental health condition and immunity. Data collected during the COVID-19 outbreak suggest that depressive symptoms may impact the production of antibodies against SARS-CoV-2, while a previous infection could affect the immune response and cause neuropsychological disturbances. A prospective observational study was designed to investigate the association between mental health conditions and immune response over time. We analyzed the mental health at baseline and the antibodies before and after immunization with the COVID-19 mRNA vaccine in a cohort of healthcare professionals in southern Switzerland. One-hundred and six subjects were enrolled. Anxiety, distress and depression correlated to each other. There were no correlations between the mentioned variables and the vaccine induced IgG antibodies against the receptor binding domain (RBD) of the spike protein. For those who had a previous COVID-19 infection, the antibodies increased according to the grade of depression. For those who did not, the anti-RBD IgG levels remained similar when comparing presence or absence of depression symptoms. Our results show that previous SARS-CoV-2 natural infection in subjects with mental health conditions enhances the immune response to COVID-19 mRNA vaccination. The correlation between immune response to COVID-19 vaccination, a previous exposure to the virus, and symptoms of mood disorders, makes it necessary to explore the direction of the causality between immune response and depressive symptoms.

Human neutralizing antibodies to cold linear epitopes and subdomain 1 of the SARS-CoV-2 spike glycoprotein.

Emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants diminishes the efficacy of vaccines and antiviral monoclonal antibodies. Continued development of immunotherapies and vaccine immunogens resilient to viral evolution is therefore necessary. Using coldspot-guided antibody discovery, a screening approach that focuses on portions of the virus spike glycoprotein that are both functionally relevant and averse to change, we identified human neutralizing antibodies to highly conserved viral epitopes. Antibody fp.006 binds the fusion peptide and cross-reacts against coronaviruses of the four genera, including the nine human coronaviruses, through recognition of a conserved motif that includes the S2' site of proteolytic cleavage. Antibody hr2.016 targets the stem helix and neutralizes SARS-CoV-2 variants. Antibody sd1.040 binds to subdomain 1, synergizes with antibody rbd.042 for neutralization, and, similar to fp.006 and hr2.016, protects mice expressing human angiotensin-converting enzyme 2 against infection when present as a bispecific antibody. Thus, coldspot-guided antibody discovery reveals donor-derived neutralizing antibodies that are cross-reactive with Orthocoronavirinae, including SARS-CoV-2 variants.

Anti-chemokine antibodies after SARS-CoV-2 infection correlate with favorable disease course.

Infection by SARS-CoV-2 leads to diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse COVID-19 outcomes. Instead, we discovered that antibodies against specific chemokines are omnipresent after COVID-19, associated with favorable disease, and predictive of lack of long COVID symptoms at one year post infection. Anti-chemokine antibodies are present also in HIV-1 infection and autoimmune disorders, but they target different chemokines than those in COVID-19. Monoclonal antibodies derived from COVID- 19 convalescents that bind to the chemokine N-loop impair cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising anti-chemokine antibodies associated with favorable COVID-19 may be beneficial by modulating the inflammatory response and thus bear therapeutic potential. One-Sentence Summary: Naturally arising anti-chemokine antibodies associate with favorable COVID-19 and predict lack of long COVID.

Human neutralizing antibodies to cold linear epitopes and to subdomain 1 of SARS-CoV-2.

Emergence of SARS-CoV-2 variants diminishes the efficacy of vaccines and antiviral monoclonal antibodies. Continued development of immunotherapies and vaccine immunogens resilient to viral evolution is therefore necessary. Using coldspot-guided antibody discovery, a screening approach that focuses on portions of the virus spike that are both functionally relevant and averse to change, we identified human neutralizing antibodies to highly conserved viral epitopes. Antibody fp.006 binds the fusion peptide and cross-reacts against coronaviruses of the four genera , including the nine human coronaviruses, through recognition of a conserved motif that includes the S2' site of proteolytic cleavage. Antibody hr2.016 targets the stem helix and neutralizes SARS-CoV-2 variants. Antibody sd1.040 binds to subdomain 1, synergizes with antibody rbd.042 for neutralization and, like fp.006 and hr2.016, protects mice when present as bispecific antibody. Thus, coldspot-guided antibody discovery reveals donor-derived neutralizing antibodies that are cross-reactive with Orthocoronavirinae , including SARS-CoV-2 variants. One sentence summary: Broadly cross-reactive antibodies that protect from SARS-CoV-2 variants are revealed by virus coldspot-driven discovery.

Synthetic double-stranded RNAs are adjuvants for the induction of T helper 1 and humoral immune responses to human papillomavirus in rhesus macaques.

Toll-like receptor (TLR) ligands are being considered as adjuvants for the induction of antigen-specific immune responses, as in the design of vaccines. Polyriboinosinic-polyribocytoidylic acid (poly I:C), a synthetic double-stranded RNA (dsRNA), is recognized by TLR3 and other intracellular receptors. Poly ICLC is a poly I:C analogue, which has been stabilized against the serum nucleases that are present in the plasma of primates. Poly I:C(12)U, another analogue, is less toxic but also less stable in vivo than poly I:C, and TLR3 is essential for its recognition. To study the effects of these compounds on the induction of protein-specific immune responses in an animal model relevant to humans, rhesus macaques were immunized subcutaneously (s.c.) with keyhole limpet hemocyanin (KLH) or human papillomavirus (HPV)16 capsomeres with or without dsRNA or a control adjuvant, the TLR9 ligand CpG-C. All dsRNA compounds served as adjuvants for KLH-specific cellular immune responses, with the highest proliferative responses being observed with 2 mg/animal poly ICLC (p = 0.002) or 6 mg/animal poly I:C(12)U (p = 0.001) when compared with immunization with KLH alone. Notably, poly ICLC -- but not CpG-C given at the same dose -- also helped to induce HPV16-specific Th1 immune responses while both adjuvants supported the induction of strong anti-HPV16 L1 antibody responses as determined by ELISA and neutralization assay. In contrast, control animals injected with HPV16 capsomeres alone did not develop substantial HPV16-specific immune responses. Injection of dsRNA led to increased numbers of cells producing the T cell-activating chemokines CXCL9 and CXCL10 as detected by in situ hybridization in draining lymph nodes 18 hours after injections, and to increased serum levels of CXCL10 (p = 0.01). This was paralleled by the reduced production of the homeostatic T cell-attracting chemokine CCL21. Thus, synthetic dsRNAs induce an innate chemokine response and act as adjuvants for virus-specific Th1 and humoral immune responses in nonhuman primates.

Basal and IL-1ß enhanced chondrocyte chemotactic activity on monocytes are co-dependent on both IKKα and IKKß NF-κB activating kinases.

IKKα and IKKß are essential kinases for activating NF-κB transcription factors that regulate cellular differentiation and inflammation. By virtue of their small size, chemokines support the crosstalk between cartilage and other joint compartments and contribute to immune cell chemotaxis in osteoarthritis (OA). Here we employed shRNA retroviruses to stably and efficiently ablate the expression of each IKK in primary OA chondrocytes to determine their individual contributions for monocyte chemotaxis in response to chondrocyte conditioned media. Both IKKα and IKKß KDs blunted both the monocyte chemotactic potential and the protein levels of CCL2/MCP-1, the chemokine with the highest concentration and the strongest association with monocyte chemotaxis. These findings were mirrored by gene expression analysis indicating that the lowest levels of CCL2/MCP-1 and other monocyte-active chemokines were in IKKαKD cells under both basal and IL-1ß stimulated conditions. We find that in their response to IL-1ß stimulation IKKαKD primary OA chondrocytes have reduced levels of phosphorylated NFkappaB p65pSer536 and H3pSer10. Confocal microscopy analysis revealed co-localized p65 and H3pSer10 nuclear signals in agreement with our findings that IKKαKD effectively blunts their basal level and IL-1ß dependent increases. Our results suggest that IKKα could be a novel OA disease target.

Systematic Development of Peptide Inhibitors Targeting the CXCL12/HMGB1 Interaction.

During inflammatory reactions, the production and release of chemotactic factors guide the recruitment of selective leukocyte subpopulations. The alarmin HMGB1 and the chemokine CXCL12, both released in the microenvironment, can form a heterocomplex, which exclusively acts on the chemokine receptor CXCR4, enhancing cell migration, and in some pathological conditions such as rheumatoid arthritis exacerbates the immune response. An excessive cell influx at the inflammatory site can be diminished by disrupting the heterocomplex. Here, we report the computationally driven identification of the first peptide (HBP08) binding HMGB1 and selectively inhibiting the activity of the CXCL12/HMGB1 heterocomplex. Furthermore, HBP08 binds HMGB1 with the highest affinity reported so far (Kd of 0.8 ± 0.4 µM). The identification of this peptide represents an important step toward the development of innovative pharmacological tools for the treatment of severe chronic inflammatory conditions characterized by an uncontrolled immune response.

Risk assessment and seroprevalence of SARS-CoV-2 infection in healthcare workers of COVID-19 and non-COVID-19 hospitals in Southern Switzerland.

BACKGROUND: Hospital healthcare workers (HCW), in particular those involved in the clinical care of COVID-19 cases, are presumably exposed to a higher risk of acquiring the disease than the general population. METHODS: Between April 16 and 30, 2020 we conducted a prospective, SARS-CoV-2 seroprevalence study in HCWs in Southern Switzerland. Participants were hospital personnel with varying COVID-19 exposure risk depending on job function and working site. They provided personal information (including age, sex, occupation, and medical history) and self-reported COVID-19 symptoms. Odds ratio (OR) of seropositivity to IgG antibodies was estimated by univariate and multivariate logistic regressions. FINDINGS: Among 4726 participants, IgG antibodies to SARS-CoV-2 were detected in 9.6% of the HCWs. Seropositivity was higher among HCWs working on COVID-19 wards (14.1% (11.9-16.5)) compared to other hospital areas at medium (10.7% (7.6-14.6)) or low risk exposure (7.3% (6.4-8.3)). OR for high vs. medium wards risk exposure was 1.42 (0.91-2.22), P = 0.119, and 1.98 (1.55-2.53), P<0.001 for high vs. low wards risk exposure. The same was for true for doctors and nurses (10.1% (9.0-11.3)) compared to other employees at medium (7.1% (4.8-10.0)) or low risk exposure (6.6% (5.0-8.4)). OR for high vs. medium profession risk exposure was 1.37 (0.89-2.11), P = 0.149, and 1.75 (1.28-2.40), P = 0.001 for high vs. low profession risk exposure. Moreover, seropositivity was higher among HCWs who had household exposure to COVID-19 cases compared to those without (18.7% (15.3-22.5) vs. 7.7% (6.9-8.6), OR 2.80 (2.14-3.67), P<0.001). INTERPRETATION: SARS-CoV-2 antibodies are detectable in up to 10% of HCWs from acute care hospitals in a region with high incidence of COVID-19 in the weeks preceding the study. HCWs with exposure to COVID-19 patients have only a slightly higher absolute risk of seropositivity compared to those without, suggesting that the use of PPE and other measures aiming at reducing nosocomial viral transmission are effective. Household contact with known COVID-19 cases represents the highest risk of seropositivity. FUNDING: Henry Krenter Foundation, Ente Ospedaliero Cantonale and Vir Biotechnology.

Perivascular expression of CXCL9 and CXCL12 in primary central nervous system lymphoma: T-cell infiltration and positioning of malignant B cells.

Primary central nervous system lymphomas (PCNSL) are aggressive malignancies confined to the CNS, mostly of diffuse large B-cell histotype. Despite improved understanding of the malignant B cells, little is known on the tumor microenvironment and on the response of the adaptive immunity against PCNSL. We investigated the phenotype of tumor infiltrating lymphocytes (TILs), and the expression of chemokines that could affect malignant B cells and trafficking of TILs. TILs and chemokine expression were evaluated by immunohistochemistry and in situ hybridization. Furthermore, we performed in vitro migration assays to analyze the migratory capacity of lymphocytes and malignant B cells toward chemokines and chemokine heterocomplexes. We show in 22 cases of PCNSL from immunocompetent patients that CD8(+) T cells represent the majority of TILs in the tumor mass. They tend to accumulate in perivascular areas, show Granzyme B expression and proliferate in situ. Their localization and density correlates with the expression of the inflammatory chemokine CXCL9, which is transcribed and translated by perivascular macrophages and pericytes in the perivascular microenvironment. Moreover, CXCL9 and CXCL12 are coexpressed on the tumor vasculature and form heterocomplexes. In the presence of CXCL9, CXCL12-induced migration is enhanced not only on CXCR4(+)/CXCR3(+)/CD8(+) T cells but also on CXCR4(+)/CXCR3(-) malignant B cells. These findings indicate the presence of a strong chemoattractant stimulus in the perivascular microenvironment, which might serve as regulator for the recruitment of TILs and for the angiocentric positioning of malignant B cells in the perivascular cuff.

Regulation of dendritic cell migration to the draining lymph node: impact on T lymphocyte traffic and priming.

Antigen-pulsed dendritic cells (DCs) are used as natural adjuvants for vaccination, but the factors that influence the efficacy of this treatment are poorly understood. We investigated the parameters that affect the migration of subcutaneously injected mouse-mature DCs to the draining lymph node. We found that the efficiency of DC migration varied with the number of injected DCs and that CCR7+/+ DCs migrating to the draining lymph node, but not CCR7-/- DCs that failed to do so, efficiently induced a rapid increase in lymph node cellularity, which was observed before the onset of T cell proliferation. We also report that DC migration could be increased up to 10-fold by preinjection of inflammatory cytokines that increased the expression of the CCR7 ligand CCL21 in lymphatic endothelial cells. The magnitude and quality of CD4+ T cell response was proportional to the number of antigen-carrying DCs that reached the lymph node and could be boosted up to 40-fold by preinjection of tumor necrosis factor that conditioned the tissue for increased DC migration. These results indicate that DC number and tissue inflammation are critical parameters for DC-based vaccination.