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Gene therapy is being investigated for a range of serious lung diseases, such as cystic fibrosis and emphysema. Recombinant adeno-associated virus (rAAV) is a well-established, safe, viral vector for gene delivery with multiple naturally occurring and artificial serotypes available displaying alternate cell, tissue, and species-specific tropisms. Efficient AAV serotypes for the transduction of the conducting airways have been identified for several species; however, efficient serotypes for human lung parenchyma have not yet been identified. Here, we screened the ability of multiple AAV serotypes to transduce lung bud organoids (LBOs)—a model of human lung parenchyma generated from human embryonic stem cells. Microinjection of LBOs allowed us to model transduction from the luminal surface, similar to dosing via vector inhalation. We identified the naturally occurring rAAV2 and rAAV6 serotypes, along with synthetic rAAV6 variants, as having tropism for the human lung parenchyma. Positive staining of LBOs for surfactant proteins B and C confirmed distal lung identity and suggested the suitability of these vectors for the transduction of alveolar type II cells. Our findings establish LBOs as a new model for pulmonary gene therapy and stress the relevance of LBOs as a viral infection model of the lung parenchyma as relevant in SARS-CoV-2 research .

The incidence of opportunistic fungal infections has increased in recent decades due to the growing proportion of immunocompromised patients in our society. Candida krusei has been described as a causative agent of disseminated fungal infections in susceptible patients. Although its prevalence remains low among yeast infections (2-5%), its intrinsic resistance to fluconazole makes this yeast important from epidemiologic aspects. Non mammalian organisms are feasible models to study fungal virulence and drug efficacy. In this work we have used the lepidopteran Galleria mellonella and the nematode Caenorhabditis elegans as models to assess antifungal efficacy during infection by C. krusei. This yeast killed G. mellonella at 25, 30 and 37°C and reduced haemocytic density. Infected larvae melanized in a dose-dependent manner. Fluconazole did not protect against C. krusei infection, in contrast to amphotericin B, voriconazole or caspofungin. However, the doses of these antifungals required to obtain larvae protection were always higher during C. krusei infection than during C. albicans infection. Similar results were found in the model host C. elegans. Our work demonstrates that non mammalian models are useful tools to investigate in vivo antifungal efficacy and virulence of C. krusei.

The Spanish guideline for COPD (GesEPOC) recommends COPD treatment according to four clinical phenotypes: non-exacerbator phenotype with either chronic bronchitis or emphysema (NE), asthma-COPD overlap syndrome (ACOS), frequent exacerbator phenotype with emphysema (FEE) or frequent exacerbator phenotype with chronic bronchitis (FECB). However, little is known on the distribution and outcomes of the four suggested phenotypes. We aimed to determine the distribution of these COPD phenotypes, and their relation with one-year clinical outcomes. We followed a cohort of well-characterized patients with COPD up to one-year. Baseline characteristics, health status (CAT), BODE index, rate of exacerbations and mortality up to one year of follow-up were compared between the four phenotypes. Overall, 831 stable COPD patients were evaluated. They were distributed as NE, 550 (66.2%); ACOS, 125 (15.0%); FEE, 38 (4.6%); and FECB, 99 (11.9%); additionally 19 (2.3%) COPD patients with frequent exacerbations did not fulfill the criteria for neither FEE nor FECB. At baseline, there were significant differences in symptoms, FEV1 and BODE index (all p<0.05). The FECB phenotype had the highest CAT score (17.1±8.2, p<0.05 compared to the other phenotypes). Frequent exacerbator groups (FEE and FECB) were receiving more pharmacological treatment at baseline, and also experienced more exacerbations the year after (all p<0.05) with no differences in one-year mortality. Most of NE (93%) and half of exacerbators were stable after one year. There is an uneven distribution of COPD phenotypes in stable COPD patients, with significant differences in demographics, patient-centered outcomes and health care resources use.

Background Mammographic density (MD) is an established biomarker of breast cancer (BC) risk. However, its relationship to BC pathological subtypes remains unclear. This study aimed to investigate this association and assess whether it differs by body mass index (BMI) and menopausal status. Methods MD percentage was assessed in the diagnostic mammograms of the contralateral breast of 714 BC patients recruited from eight Spanish hospitals. Participants completed an epidemiological questionnaire, and hospital researchers collected clinical and pathological data. Standardized prevalences (SPs) and standardized prevalence ratios (SPRs) for each BC pathological subtype across MD categories were estimated based on multinomial logistic regression models, both overall and stratified by BMI and menopausal status. Results Mean MD was 26.1% (SD = 17.3). Although no statistically significant differences were detected, women with MD ≥ 50% had a 13% lower SP of hormone receptor positive tumors (SPR = 0.87; 95% CI 0.67–1.13), a 36% higher SP of human epidermal growth factor receptor 2 positive (HER2+) tumors (SPR = 1.36; 95% CI 0.72–2.58), and a 23% higher SP of triple negative (TN) tumors (SPR = 1.23; 95% CI 0.47–3.22), compared to those with MD < 10%. These patterns were mainly observed in pre/perimenopausal women and in those with BMI ≥ 25 kg/m 2 . Conclusions High MD might be mainly associated with the development of more aggressive and non-hormone-dependent cancers, such as HER2+ and TN BC, especially among pre/perimenopausal an overweight women.

This study investigates the still uncertain association between serum phospholipid fatty acids (PL-FA), and anthropometric and adiposity variables. A cross-sectional study was conducted with 1443 Spanish premenopausal women. Participants answered an epidemiological and a food frequency questionnaire. Anthropometric variables were measured using a bioimpedance scale. Serum PL-FAs levels were determined by gas chromatography–mass spectrometry. The association between body mass index (BMI), weight gain, body fat percentage, visceral fat index, and waist circumference with serum PL-FAs and desaturation indices was evaluated using multivariable linear regression models. BMI was positively associated with the relative concentration of saturated fatty acids (SFAs) (β = 0.94, q-val = 0.001), and with palmitoleic, dihomo-γ-linolenic (DGLA), arachidonic (AA) and α-linolenic acids, and was inversely associated with oleic, gondoic, trans-vaccenic, linoleic and γ-linolenic acids. Total fat percentage was positively associated with DGLA and AA, and inversely with linoleic and γ-linolenic acids. Low relative concentrations of some SFAs and high levels of n-6 PUFAs were associated with greater waist circumference. While the oleic/stearic and AA/DGLA acid ratios were inversely associated with BMI, DGLA/linoleic acid ratio was positively related to almost all variables. In addition to BMI, total fat percentage and waist circumference were also associated with certain individual fatty acids.

Sphingolipids, ceramides and cholesterol are integral components of cellular membranes, and they also play important roles in signal transduction by regulating the dynamics of membrane receptors through their effects on membrane fluidity. Here, we combined biochemical and functional assays with single-particle tracking analysis of diffusion in the plasma membrane to demonstrate that the local lipid environment regulates CXCR4 organization and function and modulates chemokine-triggered directed cell migration. Prolonged treatment of T cells with bacterial sphingomyelinase promoted the complete and sustained breakdown of sphingomyelins and the accumulation of the corresponding ceramides, which altered both membrane fluidity and CXCR4 nanoclustering and dynamics. Under these conditions CXCR4 retained some CXCL12-mediated signaling activity but failed to promote efficient directed cell migration. Our data underscore a critical role for the local lipid composition at the cell membrane in regulating the lateral mobility of chemokine receptors, and their ability to dynamically increase receptor density at the leading edge to promote efficient cell migration.

Macrophages are a heterogeneous population of innate immune cells that support tissue homeostasis through their involvement in tissue development and repair, and pathogen defense. Emerging data reveal that metabolism may control macrophage polarization and function and, conversely, phenotypic polarization may drive metabolic reprogramming. Here we use biochemical analysis, correlative cryogenic fluorescence microscopy and cryo-focused ion-beam scanning electron microscopy. We demonstrate that growth hormone (GH) reprograms inflammatory GM-CSF-primed monocyte-derived macrophages (GM-MØ) by functioning as a metabolic modulator. We found that exogenous treatment of GM-MØ with recombinant human GH reduced glycolysis and lactate production to levels similar to those found in anti-inflammatory M-MØ. Moreover, GH treatment of GM-MØ augmented mitochondrial volume and altered mitochondrial dynamics, including the remodeling of the inner membrane to increase the density of cristae. Our data demonstrate that GH likely serves a modulatory role in the metabolism of inflammatory macrophages and suggest that metabolic reprogramming of macrophages should be considered as a new target to intervene in inflammatory diseases.

CCR5 and CXCR4, the respective cell surface coreceptors of R5 and X4 HIV-1 strains, both form heterodimers with CD4, the principal HIV-1 receptor. Using several resonance energy transfer techniques, we determined that CD4, CXCR4, and CCR5 formed heterotrimers, and that CCR5 coexpression altered the conformation of both CXCR4/CXCR4 homodimers and CD4/CXCR4 heterodimers. As a result, binding of the HIV-1 envelope protein gp120 IIIB to the CD4/CXCR4/CCR5 heterooligomer was negligible, and the gp120-induced cytoskeletal rearrangements necessary for HIV-1 entry were prevented. CCR5 reduced HIV-1 envelope-induced CD4/CXCR4-mediated cell-cell fusion. In nucleofected Jurkat CD4 cells and primary human CD4 ⁺ T cells, CCR5 expression led to a reduction in X4 HIV-1 infectivity. These findings can help to understand why X4 HIV-1 strains infection affect T-cell types differently during AIDS development and indicate that receptor oligomerization might be a target for previously unidentified therapeutic approaches for AIDS intervention.

Rheumatoid arthritis (RA) is a chronic inflammatory disease characterized by an autoimmune response in the joints and an exacerbation of cytokine responses. A minority of patients with RA experience spontaneous remission, but most will show moderate/high disease activity, with aggressive joint damage and multiple systemic manifestations. There is thus is a great need to identify prognostic biomarkers for disease risk to improve diagnosis and prognosis, and to inform on the most appropriate therapy. Here we focused on suppressor of cytokine signaling 1 (SOCS1), a physiological negative regulator of cytokines that modulates cell activation. Using four independent cohorts of patients with arthritis, we characterized the correlation between mRNA levels and clinical outcome. We found a significant inverse correlation between mRNA expression and disease activity throughout the follow-up of patients with RA. Lower baseline levels were associated with poorer disease control in response to methotrexate and other conventional synthetic disease-modifying anti-rheumatic drugs in early arthritis, and to rituximab in established (active) RA. Moreover, we identified several single nucleotide polymorphisms in the gene that correlated with mRNA expression, and that might identify those patients with early arthritis that fulfill RA classification criteria. One of them, rs4780355, is in linkage disequilibrium with a microsatellite (TTTTC) , mapped 0.9 kb downstream of the SNP, and correlated with reduced expression . Overall, our data support the association between expression and disease progression, disease severity and response to treatment in RA. These observations underlie the relevance of mRNA levels for stratifying patients prognostically and guiding therapeutic decisions.