Hyperkinetic Movement Disorder Caused by the Recurrent c.892C>T NACC1 Variant.

BACKGROUND: Genetic syndromes of hyperkinetic movement disorders associated with epileptic encephalopathy and intellectual disability are becoming increasingly recognized. Recently, a de novo heterozygous NACC1 (nucleus accumbens-associated 1) missense variant was described in a patient cohort including one patient with a combined mitochondrial oxidative phosphorylation (OXPHOS) deficiency. OBJECTIVES: The objective is to characterize the movement disorder in affected patients with the recurrent c.892C>T NACC1 variant and study the NACC1 protein and mitochondrial function at the cellular level. METHODS: The movement disorder was analyzed on four patients with the NACC1 c.892C>T (p.Arg298Trp) variant. Studies on NACC1 protein and mitochondrial function were performed on patient-derived fibroblasts. RESULTS: All patients had a generalized hyperkinetic movement disorder with chorea and dystonia, which occurred cyclically and during sleep. Complex I was found altered, whereas the other OXPHOS enzymes and the mitochondria network seemed intact in one patient. CONCLUSIONS: The movement disorder is a prominent feature of NACC1-related disease.

Distinct metabolic responses to heme in inflammatory human and mouse macrophages - Role of nitric oxide.

Activation of inflammation is tightly associated with metabolic reprogramming in macrophages. The iron-containing tetrapyrrole heme can induce pro-oxidant and pro-inflammatory effects in murine macrophages, but has been associated with polarization towards an anti-inflammatory phenotype in human macrophages. In the current study, we compared the regulatory responses to heme and the prototypical Toll-like receptor (TLR)4 ligand lipopolysaccharide (LPS) in human and mouse macrophages with a particular focus on alterations of cellular bioenergetics. In human macrophages, bulk RNA-sequencing analysis indicated that heme led to an anti-inflammatory transcriptional profile, whereas LPS induced a classical pro-inflammatory gene response. Co-stimulation of heme with LPS caused opposing regulatory patterns of inflammatory activation and cellular bioenergetics in human and mouse macrophages. Specifically, in LPS-stimulated murine, but not human macrophages, heme led to a marked suppression of oxidative phosphorylation and an up-regulation of glycolysis. The species-specific alterations in cellular bioenergetics and inflammatory responses to heme were critically dependent on the availability of nitric oxide (NO) that is generated in inflammatory mouse, but not human macrophages. Accordingly, studies with an inducible nitric oxide synthase (iNOS) inhibitor in mouse, and a pharmacological NO donor in human macrophages, reveal that NO is responsible for the opposing effects of heme in these cells. Taken together, the current findings indicate that NO is critical for the immunomodulatory role of heme in macrophages.

Characterization of Chronic Graft-versus-host Disease After Haploidentical Stem Cell Transplantation With Posttransplant Cyclophosphamide: A Study on Behalf of GETH-TC.

BACKGROUND: Chronic graft-versus-host disease (cGVHD) is a cause of late morbidity and nonrelapse mortality (NRM) after allogenic hematopoietic stem cell transplantation (allo-HSCT). Although studies evaluating haploidentical allo-HSCT (haplo-HSCT) using posttransplant cyclophosphamide (PTCy) demonstrate lower cGVHD rates, comprehensive data describing the clinical profile, risk factors, or outcomes of cGVHD within this platform are scarce. METHODS: We conducted a retrospective multicenter analysis of 389 consecutive patients who underwent haplo-HSCT PTCy in 7 transplant centers of the Spanish Group Grupo Español de Trasplante Hematopoyético y Terapia Celular (GETH-TC) between 2008 and 2020 describing incidence, clinical profile, risk factors, and cGVHD outcomes. RESULTS: Ninety-five patients of 389 developed cGVHD. Our data revealed that the incidence and severity of cGVHD are lower than those reported for HLA-identical transplantation with conventional prophylaxis and that the strongest predictor for cGVHD was previous acute GVHD (P = 0.031). Also, recipient age ≥60 y (P = 0.044) was protective against cGVHD. Moreover, patients with moderate cGVHD had longer event-free survival at 3 y than other patients (P = 0.016) and a lower relapse rate at 3 y (P = 0.036). CONCLUSIONS: Our results support the fact that the incidence and severity of cGVHD are lower than those reported for HLA-identical transplantation with conventional prophylaxis. In this series, patients who develop moderate cGVHD after haplo-HSCT PTCy had a higher overall survival and event-free survival, and lower relapse, suggesting higher graft-versus-leukemia effect. Although this is the largest series focused on characterizing cGVHD in haplo-HSCT PTCy, further prospective studies are needed to confirm the findings.

SNARE chaperone Sly1 directly mediates close-range vesicle tethering.

The essential Golgi protein Sly1 is a member of the Sec1/mammalian Unc-18 (SM) family of SNARE chaperones. Sly1 was originally identified through remarkable gain-of-function alleles that bypass requirements for diverse vesicle tethering factors. Employing genetic analyses and chemically defined reconstitutions of ER-Golgi fusion, we discovered that a loop conserved among Sly1 family members is not only autoinhibitory but also acts as a positive effector. An amphipathic lipid packing sensor (ALPS)-like helix within the loop directly binds high-curvature membranes. Membrane binding is required for relief of Sly1 autoinhibition and also allows Sly1 to directly tether incoming vesicles to the Qa-SNARE on the target organelle. The SLY1-20 mutation bypasses requirements for diverse tethering factors but loses this ability if the tethering activity is impaired. We propose that long-range tethers, including Golgins and multisubunit tethering complexes, hand off vesicles to Sly1, which then tethers at close range to initiate trans-SNARE complex assembly and fusion in the early secretory pathway.

An association between plasma levels of α2-macroglobulin and α1-antitrypsin in PiMM and PiZZ individuals differing in COPD presentation.

INTRODUCTION: Compared to normal PiMM, individuals with severe α1-antitrypsin (AAT) PiZZ (Glu342Lys) genotype deficiency are at higher risk of developing early-onset chronic obstructive pulmonary disease (COPD)/emphysema associated with Z-AAT polymers and neutrophilic inflammation. We aimed to investigate putative differences in plasma levels of acute phase proteins (APP) between PiMM and PiZZ subjects and to determine plasma Z-AAT polymer levels in PiZZ subjects. MATERIALS AND METHODS: Nephelometric analysis of seven plasma APPs was performed in 67 PiMM and 44 PiZZ subjects, of whom 43 and 42, respectively, had stable COPD. Of the PiZZ-COPD patients, 21 received and 23 did not receive intravenous therapy with human AAT preparations (IV-AAT). Plasma levels of Z-AAT polymers were determined by Western blotting using specific mouse monoclonal antibodies (2C1 and LG96). RESULTS: In addition to lower plasma AAT, PiZZ patients had higher α2-macroglobulin (A2MG) levels than PiMM patients. In contrast, PiZZ who received IV-AAT had higher AAT values but lower A2MG values than PiZZ without IV-AAT. Regardless of the AAT genotype, AAT levels were inversely correlated with A2MG, and the AAT/A2MG ratio was correlated with lung diffusion capacity (DCLO%). All PiZZ patients had circulating Z-AAT polymer levels that correlated directly with A2MG. In PiZZ without IV-AAT therapy polymer levels correlated inversely with the ratio of forced expiratory volume in 1 s to forced vital capacity (FEV1/FVC). CONCLUSION: Combined measurement of plasma AAT and A2MG levels may be of clinical value in assessing the progression of COPD and requires further attention.

Human Bronchial Epithelial Cell Transcriptome Changes in Response to Serum from Patients with Different Status of Inflammation.

PURPOSE: To investigate the transcriptome of human bronchial epithelial cells (HBEC) in response to serum from patients with different degrees of inflammation. METHODS: Serum from 19 COVID-19 patients obtained from the Hannover Unified Biobank was used. At the time of sampling, 5 patients had a WHO Clinical Progression Scale (WHO-CPS) score of 9 (severe illness). The remaining 14 patients had a WHO-CPS of below 9 (range 1-7), and lower illness. Multiplex immunoassay was used to assess serum inflammatory markers. The culture medium of HBEC was supplemented with 2% of the patient's serum, and the cells were cultured at 37 °C, 5% CO2 for 18 h. Subsequently, cellular RNA was used for RNA-Seq. RESULTS: Patients with scores below 9 had significantly lower albumin and serum levels of E-selectin, IL-8, and MCP-1 than patients with scores of 9. Principal component analysis based on 500 "core genes" of RNA-seq segregated cells into two subsets: exposed to serum from 4 (I) and 15 (II) patients. Cells from a subset (I) treated with serum from 4 patients with a score of 9 showed 5566 differentially expressed genes of which 2793 were up- and 2773 downregulated in comparison with cells of subset II treated with serum from 14 patients with scores between 1 and 7 and one with score = 9. In subset I cells, a higher expression of TLR4 and CXCL8 but a lower CDH1, ACE2, and HMOX1, and greater effects on genes involved in metabolic regulation, cytoskeletal organization, and kinase activity pathways were observed. CONCLUSION: This simple model could be useful to characterize patient serum and epithelial cell properties.

Muscle Hypertrophy Is Linked to Changes in the Oxidative and Proteolytic Systems during Early Tenderization of the Spanish Breed "Asturiana de los Valles".

For fresh meat consumers, eating satisfaction is of utmost importance and tenderness is one of the most important characteristics in this regard. Our study examined beef of different animal biotypes of the autochthonous breed "Asturiana de los Valles" (AV) to determine if early postmortem oxidative and proteolytic processes may influence the final tenderness of the product. This meat-specialized breed shows different biotypes depending on the frequency of a myostatin mutation "mh" that induces double-muscling or muscular hypertrophy (mh/mh, mh/+, +/+). Samples from the longissimus dorsi muscles of yearling bulls were analyzed during the first 24 h postmortem. Changes in the redox balance of muscle cells were significant in the first hours after slaughter; total antioxidant activity was higher in the mh/mh biotype and it followed the shortening of the sarcomeres, a key parameter in understanding meat tenderness. The two proteolytic systems studied (proteasome and lysosome) followed distinct patterns. Proteasome activity was higher in the (mh/+) biotype, which correlated with higher protein damage. Lysosome proteolysis was increased in the more tender biotypes (mh genotypes). Autophagic activation showed significant differences between the biotypes, with (mh/mh) showing more intense basal autophagy at the beginning of the postmortem period that decreased gradually (p < 0.001), while in the normal biotype (+/+), it was slightly delayed and then increased progressively (p < 0.001). These results suggest that this type of catalytic process and antioxidant activity could contribute to the earlier disintegration of the myofibers, particularly in the mh/mh biotypes, and influence the conversion of muscle into meat.

Plasma levels of α1-antitrypsin-derived C-terminal peptides in PiMM and PiZZ COPD patients.

Plasma levels of α1-antitrypsin-derived C-terminal peptides might be valid as novel biomarkers to predict and/or characterise exacerbations in PiMM and PiZZ COPD patients, or to reflect the efficiency of augmentation therapy in PiZZ patients https://bit.ly/3rNJeLd.

Ultrafast 30-min infusion of a rituximab biosimilar (Truxima).

In this clinical trial, we demonstrate that ultrarapid fast infusion of rituximab (Truxima) in 30 min with oral premedication is feasible and secure for patients, and reduce the day-care hospital stays.

Melatonin Alleviates the Impairment of Muscle Bioenergetics and Protein Quality Control Systems in Leptin-Deficiency-Induced Obesity.

Leptin is critically compromised in the major common forms of obesity. Skeletal muscle is the main effector tissue for energy modification that occurs as a result of the effect of endocrine axes, such as leptin signaling. Our study was carried out using skeletal muscle from a leptin-deficient animal model, in order to ascertain the importance of this hormone and to identify the major skeletal muscle mechanisms affected. We also examined the therapeutic role of melatonin against leptin-induced muscle wasting. Here, we report that leptin deficiency stimulates fatty acid ß-oxidation, which results in mitochondrial uncoupling and the suppression of mitochondrial oxidative damage; however, it increases cytosolic oxidative damage. Thus, different nutrient-sensing pathways are disrupted, impairing proteostasis and promoting lipid anabolism, which induces myofiber degeneration and drives oxidative type I fiber conversion. Melatonin treatment plays a significant role in reducing cellular oxidative damage and regulating energy homeostasis and fuel utilization. Melatonin is able to improve both glucose and mitochondrial metabolism and partially restore proteostasis. Taken together, our study demonstrates melatonin to be a decisive mitochondrial function-fate regulator in skeletal muscle, with implications for resembling physiological energy requirements and targeting glycolytic type II fiber recovery.

Prevalence of asymptomatic Leishmania infection and knowledge, perceptions, and practices in blood donors in mainland Portugal.

BACKGROUND: Asymptomatic infection is the most common outcome of exposure to Leishmania parasites. In the Mediterranean region, where Leishmania infantum is endemic, studies on the prevalence of asymptomatic infection have often relied on serological testing in blood donors. In Spain, regional studies have shown seroprevalence in blood donors between 1 and 8%; in Portugal, values of 0 and 2% were suggested by two localized studies, in different populations. The purpose of this study was (i) to estimate the prevalence of asymptomatic Leishmania infection in blood donors in mainland Portugal, and (ii) to study the association between the detection of antibodies to Leishmania and sociodemographic factors, and also the knowledge, perceptions and practices (KPP) of the blood donors regarding leishmaniasis. METHODS: A cross-sectional study targeted the population of people who donated blood in mainland Portugal. Participants, distributed proportionally by municipality and aged between 18 and 65 years, were selected randomly in 347 blood collection points between February and June 2022, and completed a sociodemographic and a KPP questionnaire. Detection of anti-Leishmania antibodies in serum was performed using an ELISA commercial kit. Individual KPP scores were calculated by adding grades defined for each question. RESULTS: Globally, 201/3763 samples were positive. The estimated national true seroprevalence was 4.8% (95% CI 4.1-5.5%). The proportion of positive results was significantly different between NUTS (Nomenclature of Territorial Units for Statistics) regions. Models suggested that seropositivity was significantly higher in male sex, people older than 25 years, or residing in the Centro NUTS2 region, but not in dog owners nor people with lower KPP scores. Overall, 72.3% of participants had previously heard of leishmaniasis and, in multivariate analysis, a higher Knowledge score was associated with age 25-40 years, female sex, ownership of dogs, and higher education. CONCLUSIONS: Global estimated true seroprevalence (4.8%) was similar to previous regional studies in blood donors in neighboring Spain. Higher seroprevalence values in the NUTS2 Centro region were consistent with incidence data from humans and seroprevalence studies in dogs. On the other hand, the low values in the Alentejo and the high values in the northern subregions may be the result of geographical shifts in parasite circulation due to climate change and should prompt localized and integrated, vector, canine, and human research, following a One Health approach.

Acid Sphingomyelinase Deficiency Type B Patient-Derived Liver Organoids Reveals Altered Lysosomal Gene Expression and Lipid Homeostasis.

Acid sphingomyelinase deficiency (ASMD) or Niemann-Pick disease type A (NPA), type B (NPB) and type A/B (NPA/B), is a rare lysosomal storage disease characterized by progressive accumulation of sphingomyelin (SM) in the liver, lungs, bone marrow and, in severe cases, neurons. A disease model was established by generating liver organoids from a NPB patient carrying the p.Arg610del variant in the SMPD1 gene. Liver organoids were characterized by transcriptomic and lipidomic analysis. We observed altered lipid homeostasis in the patient-derived organoids showing the predictable increase in sphingomyelin (SM), together with cholesterol esters (CE) and triacylglycerides (TAG), and a reduction in phosphatidylcholine (PC) and cardiolipins (CL). Analysis of lysosomal gene expression pointed to 24 downregulated genes, including SMPD1, and 26 upregulated genes that reflect the lysosomal stress typical of the disease. Altered genes revealed reduced expression of enzymes that could be involved in the accumulation in the hepatocytes of sphyngoglycolipids and glycoproteins, as well as upregulated genes coding for different glycosidases and cathepsins. Lipidic and transcriptome changes support the use of hepatic organoids as ideal models for ASMD investigation.

Quantitative Lipid Profiling Reveals Major Differences between Liver Organoids with Normal Pi*M and Deficient Pi*Z Variants of Alpha-1-antitrypsin.

Different mutations in the SERPINA1 gene result in alpha-1 antitrypsin (AAT) deficiency and in an increased risk for the development of liver diseases. More than 90% of severe deficiency patients are homozygous for Z (Glu342Lys) mutation. This mutation causes Z-AAT polymerization and intrahepatic accumulation which can result in hepatic alterations leading to steatosis, fibrosis, cirrhosis, and/or hepatocarcinoma. We aimed to investigate lipid status in hepatocytes carrying Z and normal M alleles of the SERPINA1 gene. Hepatic organoids were developed to investigate lipid alterations. Lipid accumulation in HepG2 cells overexpressing Z-AAT, as well as in patient-derived hepatic organoids from Pi*MZ and Pi*ZZ individuals, was evaluated by Oil-Red staining in comparison to HepG2 cells expressing M-AAT and liver organoids from Pi*MM controls. Furthermore, mass spectrometry-based lipidomics analysis and transcriptomic profiling were assessed in Pi*MZ and Pi*ZZ organoids. HepG2 cells expressing Z-AAT and liver organoids from Pi*MZ and Pi*ZZ patients showed intracellular accumulation of AAT and high numbers of lipid droplets. These latter paralleled with augmented intrahepatic lipids, and in particular altered proportion of triglycerides, cholesterol esters, and cardiolipins. According to transcriptomic analysis, Pi*ZZ organoids possess many alterations in genes and cellular processes of lipid metabolism with a specific impact on the endoplasmic reticulum, mitochondria, and peroxisome dysfunction. Our data reveal a relationship between intrahepatic accumulation of Z-AAT and alterations in lipid homeostasis, which implies that liver organoids provide an excellent model to study liver diseases related to the mutation of the SERPINA1 gene.

NAFLD and AATD Are Two Diseases with Unbalanced Lipid Metabolism: Similarities and Differences.

Non-alcoholic fatty liver disease (NAFLD) is a type of steatosis commonly associated with obesity, dyslipidemia, hypertension, and diabetes. Other diseases such as inherited alpha-1 antitrypsin deficiency (AATD) have also been related to the development of liver steatosis. The primary reasons leading to hepatic lipid deposits can be genetic and epigenetic, and the outcomes range from benign steatosis to liver failure, as well as to extrahepatic diseases. Progressive hepatocellular damage and dysregulated systemic immune responses can affect extrahepatic organs, specifically the heart and lungs. In this review, we discuss the similarities and differences between the molecular pathways of NAFLD and AATD, and the putative value of hepatic organoids as novel models to investigate the physio pathological mechanisms of liver steatosis.

Results of haploidentical transplant in patients with donor-specific antibodies: a survey on behalf of the Spanish Group of Hematopoietic Transplant and Cell Therapy.

Background: Donor-specific antibodies (DSAs) are IgG allo-antibodies against mismatched donor HLA molecules and can cause graft failure (GF) in the setting of haploidentical hematopoietic stem cell transplantation (haplo-HSCT). Our aim was to report the experience of the Spanish Group of Hematopoietic Transplant (GETH-TC) in DSA-positive patients who had undergone haplo-HSCT. Methods: We conducted a survey of patients who underwent haplo-HSCT in GETH-TC centers between 2012 and 2021. Data were collected on the DSA assay used, monitoring strategy, complement fixation, criteria for desensitization, desensitization strategies and transplant outcomes. Results: Fifteen centers from the GETH-TC responded to the survey. During the study period, 1,454 patients underwent haplo-HSCT. Seventy of the transplants were performed in 69 DSA-positive patients, all of whom lacked a suitable alternative donor; 61 (88%) patients were female (90% with prior pregnancies). All patients received post-transplant cyclophosphamide-based graft-versus-host disease prophylaxis. Regarding baseline DSA intensity, 46 (67%) patients presented mean fluorescence intensity (MFI) >5,000, including 21 (30%) with MFI >10,000 and three (4%) with MFI >20,000. Six patients did not receive desensitization treatment, four of them with MFI <5,000. Of 63 patients receiving desensitization treatment, 48 (76%) were tested after desensitization therapy, and a reduction in intensity was confirmed in 45 (71%). Three patients (5%) experienced an increase in MFI after desensitization, two of whom experienced primary GF. Cumulative incidence of neutrophil engraftment at day 28 was 74% in a median of 18 days (IQR, 15─20); six patients died before engraftment due to toxicity or infection and eight patients had primary GF despite desensitization in seven of them. After a median follow-up of 30 months, two-year overall and event-free survival were 46.5% and 39%, respectively. The two-year cumulative incidence of relapse was 16% and non-relapse mortality (NRM) was 43%. Infection was the most frequent cause of NRM, followed by endothelial toxicity. Multivariate analysis identified baseline MFI >20,000 as an independent risk factor for survival and an increase in titers after infusion as an independent risk factor for GF. Conclusions: Haplo-HSCT is feasible in DSA-positive patients, with high rates of engraftment after desensitization guided by DSA intensity. Baseline MFI >20,000 and increased intensity after infusion are risk factors for survival and GF.

Dissecting the functions of cancer-associated fibroblasts to therapeutically target head and neck cancer microenvironment.

Head and neck cancers (HNC) are a diverse group of aggressive malignancies with high morbidity and mortality, leading to almost half-million deaths annually worldwide. A better understanding of the molecular processes governing tumor formation and progression is crucial to improve current diagnostic and prognostic tools as well as to develop more personalized treatment strategies. Tumors are highly complex and heterogeneous structures in which growth and dissemination is not only governed by the cancer cells intrinsic mechanisms, but also by the surrounding tumor microenvironment (TME). Cancer-associated fibroblasts (CAFs) emerge as predominant TME components and key players in the generation of permissive conditions that ultimately impact in tumor progression and metastatic dissemination. Although CAFs were initially considered a consequence of tumor development, it is now well established that they actively contribute to numerous cancer hallmarks i.e., tumor cell growth, migration and invasion, cancer cell stemness, angiogenesis, metabolic reprograming, inflammation, and immune system modulation. In this scenario, therapeutic strategies targeting CAF functions could potentially have a major impact in cancer therapeutics, providing avenues for new treatment options or for improving efficacy in established approaches. This review is focused on thoroughly dissecting existing evidences supporting the contribution of CAFs in HNC biology with an emphasis on current knowledge of the key molecules and pathways involved in CAF-tumor crosstalk, and their potential as novel biomarkers and/or therapeutic targets to effectively interfere the tumor-stroma crosstalk for HNC patients benefit. involved in CAF-tumor crosstalk, and their potential as novel biomarkers and/or therapeutic targets to effec- tively interfere the tumor-stroma crosstalk for HNC patients benefit.

CERT1 mutations perturb human development by disrupting sphingolipid homeostasis.

Neural differentiation, synaptic transmission, and action potential propagation depend on membrane sphingolipids, whose metabolism is tightly regulated. Mutations in the ceramide transporter CERT (CERT1), which is involved in sphingolipid biosynthesis, are associated with intellectual disability, but the pathogenic mechanism remains obscure. Here, we characterize 31 individuals with de novo missense variants in CERT1. Several variants fall into a previously uncharacterized dimeric helical domain that enables CERT homeostatic inactivation, without which sphingolipid production goes unchecked. The clinical severity reflects the degree to which CERT autoregulation is disrupted, and inhibiting CERT pharmacologically corrects morphological and motor abnormalities in a Drosophila model of the disease, which we call ceramide transporter (CerTra) syndrome. These findings uncover a central role for CERT autoregulation in the control of sphingolipid biosynthetic flux, provide unexpected insight into the structural organization of CERT, and suggest a possible therapeutic approach for patients with CerTra syndrome.

Mice inflammatory responses to inhaled aerosolized LPS: effects of various forms of human alpha1-antitrypsin.

Rodent models of lipopolysaccharide (LPS)-induced pulmonary inflammation are used for anti-inflammatory drug testing. We aimed to characterize mice responses to aerosolized LPS alone or with intraperitoneal (i.p.) delivery of alpha1-antitrypsin (AAT). Balb/c mice were exposed to clean air or aerosolized LPS (0.21 mg/mL) for 10 min per day, for 3 d. One hour after each challenge, animals were treated i.p. with saline or with (4 mg/kg body weight) one of the AAT preparations: native (AAT), oxidized (oxAAT), recombinant (recAAT), or peptide of AAT (C-36). Experiments were terminated 6 h after the last dose of AATs. Transcriptome data of mice lungs exposed to clean air versus LPS revealed 656 differentially expressed genes and 155 significant gene ontology terms, including neutrophil migration and toll-like receptor signaling pathways. Concordantly, mice inhaling LPS showed higher bronchoalveolar lavage fluid neutrophil counts and levels of myeloperoxidase, inducible nitric oxide synthase, IL-1ß, TNFα, KC, IL-6, and granulocyte-macrophage colony-stimulating factor (GM-CSF). Plasma inflammatory markers did not increase. After i.p. application of AATs, about 1% to 2% of proteins reached the lungs but, except for GM-CSF, none of the proteins significantly influenced inflammatory markers. All AATs and C-36 significantly inhibited LPS-induced GM-CSF release. Surprisingly, only oxAAT decreased the expression of several LPS-induced inflammatory genes, such as Cxcl3, Cd14, Il1b, Nfkb1, and Nfkb2, in lung tissues. According to lung transcriptome data, oxAAT mostly affected genes related to transcriptional regulation while native AAT or recAAT affected genes of inflammatory pathways. Hence, we present a feasible mice model of local lung inflammation induced via aerosolized LPS that can be useful for systemic drug testing.

Lung Adenocarcinoma Cell Sensitivity to Chemotherapies: A Spotlight on Lipid Droplets and SREBF1 Gene.

To explore the relationship between cancer cell SREBF1 expression, lipid droplets (LDs) formation, and the sensitivity to chemotherapies, we cultured lung adenocarcinoma cells H1299 (with LD) and H1563 (without LD) in a serum-free basal medium (BM) or neutrophil degranulation products containing medium (NDM), and tested cell responses to cisplatin and etoposide. By using the DESeq2 Bioconductor package, we detected 674 differentially expressed genes (DEGs) associated with NDM/BM differences between two cell lines, many of these genes were associated with the regulation of sterol and cholesterol biosynthesis processes. Specifically, SREBF1 markedly declined in both cell lines cultured in NDM or when treated with chemotherapeutics. Despite the latter, H1563 exhibited LD formation and resistance to etoposide, but not to cisplatin. Although H1299 cells preserved LDs, these cells were similarly sensitive to both drugs. In a cohort of 292 patients with non-small-cell lung cancer, a lower SREBF1 expression in tumors than in adjacent nontumor tissue correlated with overall better survival, specifically in patients with adenocarcinoma at stage I. Our findings imply that a direct correlation between SREBF1 and LD accumulation can be lost due to the changes in cancer cell environment and/or chemotherapy. The role of LDs in lung cancer development and response to therapies remains to be examined in more detail.

Loss of Serpina1 in Mice Leads to Altered Gene Expression in Inflammatory and Metabolic Pathways.

The SERPINA1 gene encodes alpha1-antitrypsin (AAT), an acute phase glycoprotein and serine protease inhibitor that is mainly (80-90%) produced in the liver. Point mutations in the SERPINA1 gene can lead to the misfolding, intracellular accumulation, and deficiency of circulating AAT protein, increasing the risk of developing chronic liver diseases or chronic obstructive pulmonary disease. Currently, siRNA technology can knock down the SERPINA1 gene and limit defective AAT production. How this latter affects other liver genes is unknown. Livers were taken from age- and sex-matched C57BL/6 wild-type (WT) and Serpina1 knockout mice (KO) aged from 8 to 14 weeks, all lacking the five serpin A1a-e paralogues. Total RNA was isolated and RNA sequencing, and transcriptome analysis was performed. The knockout of the Serpina1 gene in mice changed inflammatory, lipid metabolism, and cholesterol metabolism-related gene expression in the liver. Independent single-cell sequencing data of WT mice verified the involvement of Serpina1 in cholesterol metabolism. Our results from mice livers suggested that designing therapeutic strategies for the knockout of the SERPINA1 gene in humans must account for potential perturbations of key metabolic pathways and consequent mitigation of side effects.