Miguel Galán

Resident memory CD8+ T cells (Trms) are essential for protecting barrier nonlymphoid tissues (NLTs) against reinfection, yet the involvement of dendritic cells (DCs) in this process and the nature of Trm–DC interactions within these tissues remain poorly understood. Our study demonstrates that upon reactivation, memory CD8+ T cells located in the skin—independently of circulating memory counterparts—initiate the infiltration and maturation of plasmacytoid DCs (pDCs) in the tissue. This, in turn, promotes the maturation of conventional type 1 DCs (cDC1s) through type I IFN (IFN-I) signaling in a pDC-dependent manner. Depletion of pDCs or blocking IFN-I signaling disrupts this axis, severely impairing Trm-driven protection against secondary infections with vaccinia virus (VACV) in the skin. Notably, this pDC-dependent, IFN-I-mediated pathway is also essential for Trm-mediated protection against secondary respiratory infections with influenza A virus (IAV). Our findings uncover a crucial collaboration between Trm, pDCs, and cDC1s, offering new insights for enhancing vaccines.

Atherosclerosis is the main underlying cause of cardiovascular diseases. Its prevention is based on the detection and treatment of traditional cardiovascular risk factors¹. However, individuals at risk for early vascular disease often remain unidentified². Recent research has identified new molecules in the pathophysiology of atherosclerosis³, highlighting the need for alternative disease biomarkers and therapeutic targets to improve early diagnosis and therapy efficacy. Here, we observed that imidazole propionate (ImP), produced by microorganisms, is associated with the extent of atherosclerosis in mice and in two independent human cohorts. Furthermore, ImP administration to atherosclerosis-prone mice fed with chow diet was sufficient to induce atherosclerosis without altering the lipid profile, and was linked to activation of both systemic and local innate and adaptive immunity and inflammation. Specifically, we found that ImP caused atherosclerosis through the imidazoline-1 receptor (I1R, also known as nischarin) in myeloid cells. Blocking this ImP–I1R axis inhibited the development of atherosclerosis induced by ImP or high-cholesterol diet in mice. Identification of the strong association of ImP with active atherosclerosis and the contribution of the ImP–I1R axis to disease progression opens new avenues for improving the early diagnosis and personalized therapy of atherosclerosis.

BACKGROUND: Atherosclerosis is characterized by immune cell accumulation in the arterial wall and adaptive CD4 ⁺ Th1 immunity contributes to atherosclerosis development. However, how conventional dendritic cells (cDCs) orchestrate this adaptive response remains controversial. This study unveils strategies for the gain and loss of function of cDCs to decipher their role in atherosclerosis induction in relation to adaptive T-cell immunity. METHODS: We tested atherosclerosis in Ldlr ^−/− mice fed a high-cholesterol diet (HCD). Expansion of cDCs in vivo was achieved by overexpression of FLT3L (Fms-like tyrosine kinase 3 ligand), while the effect of ablation of conventional type 1 dendritic cells (cDC1s) in atherosclerosis was analyzed by grafting bone marrow from different mouse models of cDC1 depletion, including Xcr1 ^Cre−DTA and Irf8 Δ32 mice, into lethally irradiated Ldlr ^−/− recipients before HCD. CD3 ⁺ T-cell subsets were analyzed using flow cytometry or scRNA-seq. Nanoparticles loaded with dexamethasone and decorated with anti-CLEC9A antibody to target cDC1s were tested for immunotherapy. RESULTS: Expansion of dendritic cells in Ldlr ^{− / −} mice fed HCD for 8 weeks led to increased atherosclerotic lesion, which was prevented when Ldlr ^{− / −} mice were grafted before dendritic cell expansion with Xcr1 ^Cre−DTA cDC1-depleted bone marrow compared with controls. Consistently, even in the absence of dendritic cell expansion, cDC1 deficiency prevented HCD-induced atherosclerosis. The scRNA-seq analysis of aortic CD3 ⁺ T cells in this experimental approach showed a local reduction in CD4 ⁺ Th1 and CD8 ⁺ IFN (interferon)-γ ⁺ T cells in the absence of cDC1s compared with control mice. Mechanistically, stimulator of IFN genes in cDC1s was required for the proatherogenic function of cDC1s. As a potential cDC1-targeted immunotherapy for atherosclerosis, we generated lipid nanoparticles decorated with an anti-CLEC9A antibody to specifically target cDC1s. When loaded with the immunosuppressive drug dexamethasone, these nanoparticles promoted a reduction of the atherosclerotic lesion in Ldlr ^{− / −} mice fed HCD, correlating with decreased CD4 ⁺ Th1 and CD8 ⁺ IFN-γ ⁺ T cells in the spleen. These immunosuppressive nanoparticles, however, did not impair antiviral response. CONCLUSIONS: Using state-of-the-art strategies, our results establish that cDC1s have a proatherogenic role in atherosclerosis by boosting CD4 ⁺ and CD8 ⁺ T-cell immunity and propose that cDC1s can be targeted with an immunosuppressive drug to decrease atherosclerosis progression.

The clinical presentations of COVID-19 may range from an asymptomatic or mild infection to a critical or fatal disease. Several host factors such as elderly age, male gender, and previous comorbidities seem to be involved in the most severe outcomes, but also an impaired immune response that causes a hyperinflammatory state but is unable to clear the infection. In order to get further understanding about this impaired immune response, we aimed to determine the association of specific HLA alleles with different clinical presentations of COVID-19. Therefore, we analyzed HLA Class I and II, as well as KIR gene sequences, in 72 individuals with Spanish Mediterranean Caucasian ethnicity who presented mild, severe, or critical COVID-19, according to their clinical characteristics and management. This cohort was recruited in Madrid (Spain) during the first and second pandemic waves between April and October 2020. There were no significant differences in HLA-A or HLA-B alleles among groups. However, despite the small sample size, we found that HLA-C alleles from group C1 HLA-C*08:02, -C*12:03, or -C*16:01 were more frequently associated in individuals with mild COVID-19 (43.8%) than in individuals with severe (8.3%; p = 0.0030; pc = 0.033) and critical (16.1%; p = 0.0014; pc = 0.0154) disease. C1 alleles are supposed to be highly efficient to present peptides to T cells, and HLA-C*12:03 may present a high number of verified epitopes from abundant SARS-CoV-2 proteins M, N, and S, thereby being allegedly able to trigger an efficient antiviral response. On the contrary, C2 alleles are usually poorly expressed on the cell surface due to low association with β2-microglobulin (β2M) and peptides, which may impede the adequate formation of stable HLA-C/β2M/peptide heterotrimers. Consequently, this pilot study described significant differences in the presence of specific HLA-C1 alleles in individuals with different clinical presentations of COVID-19, thereby suggesting that HLA haplotyping could be valuable to get further understanding in the underlying mechanisms of the impaired immune response during critical COVID-19.

Purpose: Bevacizumab is a monoclonal antibody that binds to vascular endothelial growth factor A. It is currently used in combination with chemotherapy to treat metastatic colorectal cancer. This therapy is not equally effective in every patient; in some, mechanisms of resistance arise that remain poorly understood. The aim of the present work was to determine whether the expression of 26 miRNAs could be associated with the effectiveness of bevacizumab plus chemotherapy, with progression-free survival (PFS), and with overall survival (OS) in metastatic colorectal cancer. Patients and Methods: Paraffin-embedded biopsies from 76 patients with metastatic colorectal cancer were collected to isolate miRNAs. The expression of 26 miRNAs was analyzed by quantitative RT-PCR. For the purpose of analysis, patients were classified as either “responders” (PFS ≥ 6 months since beginning treatment) or “non-responders” (PFS < 6 months). For the analysis of PFS and OS, patients were classified into two groups using the median gene expression value as the cut-off point (“high” [≥ 50% percentile] or “low” [< 50% percentile]). Time-to-event data were analyzed using the Kaplan–Meier method and compared by the log rank test. Cox regression was used to estimate hazard ratios (HR) and their 95% confidence intervals. Results: miR-7-5p and miR-10a-5p were more strongly expressed in non-responders than responders (p=0.049 and p=0.043, respectively), and OS was poorer in patients showing these higher expression levels (HR=2.54, 95% CI 1.42– 4.55, p=0. 001, and HR=1.81, 95% CI 1.02– 3.20, p=0.039, respectively). The overexpression of miR-143-3p, however, was associated with a better prognosis and significantly better PFS (HR=0.57; 95% CI: 0.33– 0.96; p=0.033). Conclusion: High expression values for miR-7-5p and miR-10a-5p might be considered markers of a poorer prognosis in patients with metastatic colorectal cancer treated with bevacizumab plus chemotherapy, while the same for miR-143-3p might be a marker of better outcomes.

Angiogenesis pathway genes show substantial genetic variability causing inter-individual differences in responses to anti-angiogenic drugs. We examined 20 single nucleotide polymorphisms (SNPs) in 13 of these genes to predict tumour response and clinical outcome measured as progression free survival (PFS) and overall survival (OS) in 57 patients with metastatic colorectal cancer (mCRC) given bevacizumab plus chemotherapy. SNPs were detected (iPLEX® Assay) in genomic DNA extracted from formalin-fixed paraffin-embedded tumour specimens. The variant allele CD39 rs11188513 was associated with a good tumour response (p = 0.024). Patients homozygous for the wild-type allele FGF2 rs1960669 showed a median PFS of 10.95 months versus 5.44 months for those with at least one variant allele-A (HR 3.30; 95% CI: 1.52–7.14; p = 0.001). Patients homozygous for wild-type MMP9 rs2236416 and rs2274755 showed a median PFS of 9.48 months versus 6 and 6.62 months, respectively, for those with at least one variant allele (p = 0.022, p = 0.043, respectively). OS was also lengthened to 30.92 months (p = 0.034) in carriers of wild-type ANGPT1 rs2445365 versus 22.07 months for those carrying at least one variant allele-A. These gene variants were able to predict clinical outcome and tumour response in mCRC patients given bevacizumab-based therapy.

BCR-ABL is an aberrant tyrosine kinase responsible for chronic myeloid leukemia (CML). Tyrosine kinase inhibitors (TKIs) induce a potent antileukemic response mostly based on the inhibition of BCR-ABL, but they also increase the activity of Natural Killer (NK) and CD8+ T cells. After several years, patients may interrupt treatment due to sustained, deep molecular response. By unknown reasons, half of the patients relapse during treatment interruption, whereas others maintain a potent control of the residual leukemic cells for several years. In this study, several immunological parameters related to sustained antileukemic control were analyzed. According to our results, the features more related to poor antileukemic control were as follows: low levels of cytotoxic cells such as NK, (Natural Killer T) NKT and CD8±TCRγβ+ T cells; low expression of activating receptors on the surface of NK and NKT cells; impaired synthesis of proinflammatory cytokines or proteases from NK cells; and HLA-E*0103 homozygosis and KIR haplotype BX. A Random Forest algorithm predicted 90% of the accuracy for the classification of CML patients in groups of relapse or non-relapse according to these parameters. Consequently, these features may be useful as biomarkers predictive of CML relapse in patients that are candidates to initiate treatment discontinuation.

Rigid adsorbents used as matrix skeleton have advantages over soft gel media for downstream processing of proteins. The adsorption of bovine serum albumin (BSA) has been investigated on a rigid ceramic monolith coated with cross-linked microporous agarose (D5). The physical properties of the adsorbent and the adsorption equilibria, adsorption kinetics, and mass transfer behavior have been studied for five different flow rates, with a pH value ranging from pH 4.5 to 7.0. The optimal working flow rate was 14.0 cm³/min, and using this flow rate, increasing the pH does not generate a significant improvement in the adsorption capacity. The rates of BSA adsorption have been measured and it was possible to describe a theoretical model, in which the mass transfer involves a dispersion coefficient (k_disp), which describes the mass transfer in the adsorbent surface, from the volume of the protein solution to the agarose surface. This parameter presents an exponential tendency by increasing the flow rate from 2.37 × 10⁻⁶ to 87.40 × 10⁻⁶ cm/s for n = 1. Values obtained for the adsorption kinetic constant (k_ads) followed the trend of the mass transfer parameter, increasing with the flow rate from 1.94 × 10⁴ to 117.39 × 10⁴ cm²/mol s. The theoretical model predicts the protein concentration in equilibrium for successive column reuses and it can be readily used to determine the optimal reuse of column. Likewise, for a maximum flow rate of 14 cm³/min, pressure drop was 0.04 MPa, being an advantage in front of packed columns that have higher pressure drop.