Development and characterization of a non-human primate model of disseminated synucleinopathy.

Introduction: The presence of a widespread cortical synucleinopathy is the main neuropathological hallmark underlying clinical entities such as Parkinson's disease with dementia (PDD) and dementia with Lewy bodies (DLB). There currently is a pressing need for the development of non-human primate (NHPs) models of PDD and DLB to further overcome existing limitations in drug discovery. Methods: Here we took advantage of a retrogradely-spreading adeno-associated viral vector serotype 9 coding for the alpha-synuclein A53T mutated gene (AAV9-SynA53T) to induce a widespread synucleinopathy of cortical and subcortical territories innervating the putamen. Four weeks post-AAV deliveries animals were sacrificed and a comprehensive biodistribution study was conducted, comprising the quantification of neurons expressing alpha-synuclein, rostrocaudal distribution and their specific location. Results: Intraputaminal deliveries of AAV9-SynA53T lead to a disseminated synucleinopathy throughout ipsi- and contralateral cerebral cortices, together with transduced neurons located in the ipsilateral caudal intralaminar nuclei and in the substantia nigra pars compacta (leading to thalamostriatal and nigrostriatal projections, respectively). Cortical afferent systems were found to be the main contributors to putaminal afferents (superior frontal and precentral gyri in particular). Discussion: Obtained data extends current models of synucleinopathies in NHPs, providing a reproducible platform enabling the adequate implementation of end-stage preclinical screening of new drugs targeting alpha-synuclein.

ß-Variational autoencoders and transformers for reduced-order modelling of fluid flows.

Variational autoencoder architectures have the potential to develop reduced-order models for chaotic fluid flows. We propose a method for learning compact and near-orthogonal reduced-order models using a combination of a ß-variational autoencoder and a transformer, tested on numerical data from a two-dimensional viscous flow in both periodic and chaotic regimes. The ß-variational autoencoder is trained to learn a compact latent representation of the flow velocity, and the transformer is trained to predict the temporal dynamics in latent-space. Using the ß-variational autoencoder to learn disentangled representations in latent-space, we obtain a more interpretable flow model with features that resemble those observed in the proper orthogonal decomposition, but with a more efficient representation. Using Poincaré maps, the results show that our method can capture the underlying dynamics of the flow outperforming other prediction models. The proposed method has potential applications in other fields such as weather forecasting, structural dynamics or biomedical engineering.

Inhibition of RIP1 improves immune reconstitution and reduces GVHD mortality while preserving graft-versus-leukemia effects.

Graft-versus-host disease (GVHD) remains the major cause of morbidity and nonrelapse mortality (NRM) after hematopoietic cell transplantation (HCT). Inflammatory cytokines mediate damage to key GVHD targets such as intestinal stem cells (ISCs) and also activate receptor interacting protein kinase 1 (RIP1; RIPK1), a critical regulator of apoptosis and necroptosis. We therefore investigated the role of RIP1 in acute GVHD using samples from HCT patients, modeling GVHD damage in vitro with both human and mouse gastrointestinal (GI) organoids, and blocking RIP1 activation in vivo using several well-characterized mouse HCT models. Increased phospho-RIP1 expression in GI biopsies from patients with acute GVHD correlated with tissue damage and predicted NRM. Both the genetic inactivation of RIP1 and the RIP1 inhibitor GNE684 prevented GVHD-induced apoptosis of ISCs in vivo and in vitro. Daily administration of GNE684 for 14 days reduced inflammatory infiltrates in three GVHD target organs (intestine, liver, and spleen) in mice. Unexpectedly, GNE684 administration also reversed the marked loss of regulatory T cells in the intestines and liver during GVHD and reduced splenic T cell exhaustion, thus improving immune reconstitution. Pharmacological and genetic inhibition of RIP1 improved long-term survival without compromising the graft-versus-leukemia (GVL) effect in lymphocytic and myeloid leukemia mouse models. Thus, RIP1inhibition may represent a nonimmunosuppressive treatment for GVHD.

Boolean analysis shows a high proportion of dopamine D2 receptors interacting with adenosine A2A receptors in striatal medium spiny neurons of mouse and non-human primate models of Parkinson's disease.

The antagonistic effect of adenosine on dopaminergic transmission in the basal ganglia indirect motor control pathway is mediated by dopamine D2 (D2R) and adenosine A2A (A2AR) receptors co-expressed on medium spiny striatal neurons. The pathway is unbalanced in Parkinson's disease (PD) and an A2AR blocker has been approved for use with levodopa in the therapy of the disease. However, it is not known whether the therapy is acting on individually expressed receptors or in receptors forming A2A-D2 receptor heteromers, whose functionality is unique. For two proteins prone to interact, a very recently developed technique, MolBoolean, allows to determine the number of proteins that are either non-interacting or interacting. After checking the feasibility of the technique and reliability of data in transfected cells and in striatal primary neurons, the Boolean analysis of receptors in the striatum of rats and monkeys showed a high percentage of D2 receptors interacting with the adenosine receptor, while, on the contrary, a significant proportion of A2A receptors do not interact with dopamine receptors. The number of interacting receptors increased when rats and monkeys were lesioned to become a PD model. The use of a tracer of the indirect pathway in monkeys confirmed that the data was restricted to the population of striatal neurons projecting to the GPe. The results are not only relevant for being the first study quantifying individual versus interacting G protein-coupled receptors, but also for showing that the D2R in these specific neurons, in both control and PD animals, is under the control of the A2AR. The tight adenosine/dopamine receptor coupling suggest benefits of early antiparkinsonian treatment with adenosine receptor blockers.

Lactate dehydrogenase contribution to symptom persistence in long COVID: A pooled analysis.

There's critical need for risk predictors in long COVID. This meta-analysis evaluates the evidence for an association between plasma lactate dehydrogenase (LDH) and long COVID and explores the contribution of LDH to symptoms persistent across the distinct post-acute sequelae of COVID-19 (PASC) domains. PubMed, EMBASE, Web of Science, and Google Scholar were searched for articles published up to 20 March 2023 for studies that reported data on LDH levels in COVID-19 survivors with and without PASC. Random-effect meta-analysis was employed to estimate the standardized mean difference (SMD) with corresponding 95% confidence interval of each outcome. There were a total of 8289 study participants (3338 PASC vs. 4951 controls) from 46 studies. Our meta-analysis compared to the controls showed a significant association between LDH elevation and Resp-PASC [SMD = 1.07, 95%CI = 0.72, 1.41, p = 0.01] but not Cardio-PASC [SMD = 1.79, 95%CI = -0.02, 3.61, p = 0.05], Neuro-PASC [SMD = 0.19, 95%CI = -0.24, 0.61, p = 0.40], and Gastrointestinal-PASC [SMD = 0.45, 95%CI = -1.08, 1.98, p = 0.56]. This meta-analysis suggests elevated LDH can be used for predicting Resp-PASC, but not Cardio-PASC, Neuro-PASC or gastrointestinal-PASC. Thus, elevated plasma LDH following COVID infection may be considered as a disease biomarker.

Neuromelanin accumulation drives endogenous synucleinopathy in non-human primates.

Although neuromelanin is a dark pigment characteristic of dopaminergic neurons in the human substantia nigra pars compacta, its potential role in the pathogenesis of Parkinson's disease (PD) has often been neglected since most commonly used laboratory animals lack neuromelanin. Here we took advantage of adeno-associated viral vectors encoding the human tyrosinase gene for triggering a time-dependent neuromelanin accumulation within substantia nigra pars compacta dopaminergic neurons in macaques up to similar levels of pigmentation as observed in elderly humans. Furthermore, neuromelanin accumulation induced an endogenous synucleinopathy mimicking intracellular inclusions typically observed in PD together with a progressive degeneration of neuromelanin-expressing dopaminergic neurons. Moreover, Lewy body-like intracellular inclusions were observed in cortical areas of the frontal lobe receiving dopaminergic innervation, supporting a circuit-specific anterograde spread of endogenous synucleinopathy by permissive trans-synaptic templating. In summary, the conducted strategy resulted in the development and characterization of a new macaque model of PD matching the known neuropathology of this disorder with unprecedented accuracy. Most importantly, evidence is provided showing that intracellular aggregation of endogenous α-synuclein is triggered by neuromelanin accumulation, therefore any therapeutic approach intended to decrease neuromelanin levels may provide appealing choices for the successful implementation of novel PD therapeutics.

Determinants of poor clinical outcome in patients with influenza pneumonia: A systematic review and meta-analysis.

BACKGROUND: The clinical burden of influenza is increasing worldwide. Aging, immunosuppression, and underlying respiratory illness are determinants of poor clinical outcomes, including greater mortality. Bacterial infections seem to be the main reason. Updated information on the role of bacterial infection as the cause of complications would be of value in improving the prognosis of patients with influenza. METHODS: A systematic review and meta-analysis were performed by using the PubMed repository using keywords like: Influenza, H1N1, Streptococcus pneumoniae, bacterial coinfection, secondary coinfection, bacterial complications in pneumonia, and seasonal influenza. Only articles written in English were included in publications from 2010 to 2020. The analyses were conducted following the preferred reporting items for systematic review and meta-analyses guidelines. The results were independently validated using a TrinetX database cohort of roughly 4 million patients. RESULTS: We included 135 studies that contained data from 48,259 patients hospitalized with influenza of any age. Bacterial infections were diagnosed in 5391 (11.2%). Streptococcus pneumoniae (30.7%) and Staphylococcus aureus (30.4%) were the most frequent microorganisms, followed by Haemophilus influenzae (7.1%) and Pseudomonas aeruginosa (5.9%). The random-effects model of the meta-analysis indicated that bacterial infections posed a 3.4-fold increased risk of death compared with influenza infection alone. Unexpectedly, asthma was protective (odds ratio 0.8). CONCLUSION: Bacterial infections diagnosed in 11.2% of patients with influenza increase 3.4-fold the mortality risk. S. pneumoniae, S. aureus, H. influenzae, and P. aeruginosa account for nearly 75% of the cases. Earlier diagnosis and use of antibiotics should improve outcomes in this population.

Hemodialysis-Associated Immune Dysregulation in SARS-CoV-2-Infected End-Stage Renal Disease Patients.

Patients on hemodialysis show dysregulated immunity, basal hyperinflammation and a marked vulnerability to COVID-19. We evaluated the immune profile in COVID-19 hemodialysis patients and the changes associated with clinical deterioration after the hemodialysis session. Recruited patients included eight hemodialysis subjects with active, PCR-confirmed SARS-CoV-2 infection, five uninfected hemodialysis patients and five healthy controls. In SARS-CoV-2-infected hemodialysis patients TNF-α, IL-6 and IL-8 were particularly increased. Lymphopenia was mostly due to reduction in CD4+ T, B and central memory CD8+ T cells. There was a predominance of classical and intermediate monocytes with reduced HLA-DR expression and enhanced production of pro-inflammatory molecules. Immune parameters were analysed pre- and post-hemodialysis in three patients with COVID-19 symptoms worsening after the hemodialysis session. There was a higher than 2.5-fold increase in GM-CSF, IFN-γ, IL-1ß, IL-2, IL-6, IL-17A and IL-21 in serum, and augmentation of monocytes-derived TNF-α, IL-1ß and IL-8 and CXCL10 (p < 0.05). In conclusion, COVID-19 in hemodialysis patients associates with alteration of lymphocyte subsets, increasing of pro-inflammatory cytokines and monocyte activation. The observed worsening during the hemodialysis session in some patients was accompanied by augmentation of particular inflammatory cytokines, which might suggest biomarkers and therapeutic targets to prevent or mitigate the hemodialysis-related deterioration during SARS-CoV-2 infection.

Cellular and humoral immune responses and breakthrough infections after three SARS-CoV-2 mRNA vaccine doses.

Background: SARS-CoV-2 vaccination has proven the most effective measure to control the COVID-19 pandemic. Booster doses are being administered with limited knowledge on their need and effect on immunity. Objective: To determine the duration of specific T cells, antibodies and neutralization after 2-dose vaccination, to assess the effect of a third dose on adaptive immunity and to explore correlates of protection against breakthrough infection. Methods: 12-month longitudinal assessment of SARS-CoV-2-specific T cells, IgG and neutralizing antibodies triggered by 2 BNT162b2 doses followed by a third mRNA-1273 dose in a cohort of 77 healthcare workers: 17 with SARS-CoV-2 infection prior to vaccination (recovered) and 60 naïve. Results: Peak levels of cellular and humoral response were achieved 2 weeks after the second dose. Antibodies declined thereafter while T cells reached a plateau 3 months after vaccination. The decline in neutralization was specially marked in naïve individuals and it was this group who benefited most from the third dose, which resulted in a 20.9-fold increase in neutralization. Overall, recovered individuals maintained higher levels of T cells, antibodies and neutralization 1 to 6 months post-vaccination than naïve. Seventeen asymptomatic or mild SARS-CoV-2 breakthrough infections were reported during follow-up, only in naïve individuals. This viral exposure boosted adaptive immunity. High peak levels of T cells and neutralizing antibodies 15 days post-vaccination associated with protection from breakthrough infections. Conclusion: Booster vaccination in naïve individuals and the inclusion of viral antigens other than spike in future vaccine formulations could be useful strategies to prevent SARS-CoV-2 breakthrough infections.

Adeno-Associated Viral Vectors as Versatile Tools for Neurological Disorders: Focus on Delivery Routes and Therapeutic Perspectives.

It is without doubt that the gene therapy field is currently in the spotlight for the development of new therapeutics targeting unmet medical needs. Thus, considering the gene therapy scenario, neurological diseases in general and neurodegenerative disorders in particular are emerging as the most appealing choices for new therapeutic arrivals intended to slow down, stop, or even revert the natural progressive course that characterizes most of these devastating neurodegenerative processes. Since an extensive coverage of all available literature is not feasible in practical terms, here emphasis was made in providing some advice to beginners in the field with a narrow focus on elucidating the best delivery route available for fulfilling any given AAV-based therapeutic approach. Furthermore, it is worth nothing that the number of ongoing clinical trials is increasing at a breath-taking speed. Accordingly, a landscape view of preclinical and clinical initiatives is also provided here in an attempt to best illustrate what is ongoing in this quickly expanding field.

Development of Potent Cellular and Humoral Immune Responses in Long-Term Hemodialysis Patients After 1273-mRNA SARS-CoV-2 Vaccination.

Long-term hemodialysis (HD) patients are considered vulnerable and at high-risk of developing severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection due to their immunocompromised condition. Since COVID-19 associated mortality rates are higher in HD patients, vaccination is critical to protect them. The response towards vaccination against COVID-19 in HD patients is still uncertain and, in particular the cellular immune response is not fully understood. We monitored the humoral and cellular immune responses by analysis of the serological responses and Spike-specific cellular immunity in COVID-19-recovered and naïve HD patients in a longitudinal study shortly after vaccination to determine the protective effects of 1273-mRNA vaccination against SARS-CoV-2 in these high-risk patients. In naïve HD patients, the cellular immune response measured by IL-2 and IFN-É£ secretion needed a second vaccine dose to significantly increase, with a similar pattern for the humoral response. In contrast, COVID-19 recovered HD patients developed a potent and rapid cellular and humoral immune response after the first vaccine dose. Interestingly, when comparing COVID-19 recovered healthy volunteers (HV), previously vaccinated with BNT162b2 vaccine to HD patients vaccinated with 1273-mRNA, these exhibited a more robust immune response that is maintained longitudinally. Our results indicate that HD patients develop strong cellular and humoral immune responses to 1273-mRNA vaccination and argue in favor of personalized immune monitoring studies in HD patients, especially if COVID-19 pre-exposed, to adapt COVID-19 vaccination protocols for this immunocompromised population.

An Early Th1 Response Is a Key Factor for a Favorable COVID-19 Evolution.

The Th1/Th2 balance plays a crucial role in the progression of different pathologies and is a determining factor in the evolution of infectious diseases. This work has aimed to evaluate the early, or on diagnosis, T-cell compartment response, T-helper subsets and anti-SARS-CoV-2 antibody specificity in COVID-19 patients and to classify them according to evolution based on infection severity. A unicenter, randomized group of 146 COVID-19 patients was divided into four groups in accordance with the most critical events during the course of disease. The immunophenotype and T-helper subsets were analyzed by flow cytometry. Asymptomatic SARS-CoV-2 infected individuals showed a potent and robust Th1 immunity, with a lower Th17 and less activated T-cells at the time of sample acquisition compared not only with symptomatic patients, but also with healthy controls. Conversely, severe COVID-19 patients presented with Th17-skewed immunity, fewer Th1 responses and more activated T-cells. The multivariate analysis of the immunological and inflammatory parameters, together with the comorbidities, showed that the Th1 response was an independent protective factor for the prevention of hospitalization (OR 0.17, 95% CI 0.03-0.81), with an AUC of 0.844. Likewise, the Th1 response was found to be an independent protective factor for severe forms of the disease (OR 0.09, 95% CI: 0.01-0.63, p = 0.015, AUC: 0.873). In conclusion, a predominant Th1 immune response in the acute phase of the SARS-CoV-2 infection could be used as a tool to identify patients who might have a good disease evolution.

Longitudinal dynamics of SARS-CoV-2-specific cellular and humoral immunity after natural infection or BNT162b2 vaccination.

The timing of the development of specific adaptive immunity after natural SARS-CoV-2 infection, and its relevance in clinical outcome, has not been characterized in depth. Description of the long-term maintenance of both cellular and humoral responses elicited by real-world anti-SARS-CoV-2 vaccination is still scarce. Here we aimed to understand the development of optimal protective responses after SARS-CoV-2 infection and vaccination. We performed an early, longitudinal study of S1-, M- and N-specific IFN-γ and IL-2 T cell immunity and anti-S total and neutralizing antibodies in 88 mild, moderate or severe acute COVID-19 patients. Moreover, SARS-CoV-2-specific adaptive immunity was also analysed in 234 COVID-19 recovered subjects, 28 uninfected BNT162b2-vaccinees and 30 uninfected healthy controls. Upon natural infection, cellular and humoral responses were early and coordinated in mild patients, while weak and inconsistent in severe patients. The S1-specific cellular response measured at hospital arrival was an independent predictive factor against severity. In COVID-19 recovered patients, four to seven months post-infection, cellular immunity was maintained but antibodies and neutralization capacity declined. Finally, a robust Th1-driven immune response was developed in uninfected BNT162b2-vaccinees. Three months post-vaccination, the cellular response was comparable, while the humoral response was consistently stronger, to that measured in COVID-19 recovered patients. Thus, measurement of both humoral and cellular responses provides information on prognosis and protection from infection, which may add value for individual and public health recommendations.

A Short Corticosteroid Course Reduces Symptoms and Immunological Alterations Underlying Long-COVID.

Despite the growing number of patients with persistent symptoms after acute SARS-CoV-2 infection, the pathophysiology underlying long-COVID is not yet well characterized, and there is no established therapy. We performed a deep immune profiling in nine patients with persistent symptoms (PSP), before and after a 4-day prednisone course, and five post-COVID-19 patients without persistent symptoms (NSP). PSP showed a perturbed distribution of circulating mononuclear cell populations. Symptoms in PSP were accompanied by a pro-inflammatory phenotype characterized by increased conventional dendritic cells and augmented expression of antigen presentation, co-stimulation, migration, and activation markers in monocytes. The adaptive immunity compartment in PSP showed a Th1-predominance, decreased naïve and regulatory T cells, and augmentation of the PD-1 exhaustion marker. These immune alterations reverted after the corticosteroid treatment and were maintained during the 4-month follow-up, and their normalization correlated with clinical amelioration. The current work highlights an immunopathogenic basis together with a possible role for steroids in the treatment for long-COVID.

Alterations in Circulating Monocytes Predict COVID-19 Severity and Include Chromatin Modifications Still Detectable Six Months after Recovery.

An early analysis of circulating monocytes may be critical for predicting COVID-19 course and its sequelae. In 131 untreated, acute COVID-19 patients at emergency room arrival, monocytes showed decreased surface molecule expression, including low HLA-DR, in association with an inflammatory cytokine status and limited anti-SARS-CoV-2-specific T cell response. Most of these alterations had normalized in post-COVID-19 patients 6 months after discharge. Acute COVID-19 monocytes transcriptome showed upregulation of anti-inflammatory tissue repair genes such as BCL6, AREG and IL-10 and increased accessibility of chromatin. Some of these transcriptomic and epigenetic features still remained in post-COVID-19 monocytes. Importantly, a poorer expression of surface molecules and low IRF1 gene transcription in circulating monocytes at admission defined a COVID-19 patient group with impaired SARS-CoV-2-specific T cell response and increased risk of requiring intensive care or dying. An early analysis of monocytes may be useful for COVID-19 patient stratification and for designing innate immunity-focused therapies.

Adeno-Associated Viral Vectors as Versatile Tools for Parkinson's Research, Both for Disease Modeling Purposes and for Therapeutic Uses.

It is without any doubt that precision medicine therapeutic strategies targeting neurodegenerative disorders are currently witnessing the spectacular rise of newly designed approaches based on the use of viral vectors as Trojan horses for the controlled release of a given genetic payload. Among the different types of viral vectors, adeno-associated viruses (AAVs) rank as the ones most commonly used for the purposes of either disease modeling or for therapeutic strategies. Here, we reviewed the current literature dealing with the use of AAVs within the field of Parkinson's disease with the aim to provide neuroscientists with the advice and background required when facing a choice on which AAV might be best suited for addressing a given experimental challenge. Accordingly, here we will be summarizing some insights on different AAV serotypes, and which would be the most appropriate AAV delivery route. Next, the use of AAVs for modeling synucleinopathies is highlighted, providing potential readers with a landscape view of ongoing pre-clinical and clinical initiatives pushing forward AAV-based therapeutic approaches for Parkinson's disease and related synucleinopathies.

Circulatory follicular helper T lymphocytes associate with lower incidence of CMV infection in kidney transplant recipients.

Primary infection and/or reactivation of cytomegalovirus (CMV) in kidney transplant recipients (KTR) favor rejection and mortality. T follicular helper cells (TFH) could contribute to protection against CMV. Circulatory TFH (cTFH) were studied pretransplant and early posttransplant in 90 CMV seropositive KTR not receiving antithymocyte globulin or antiviral prophylaxis, followed-up for 1 year. Patients who presented CMV infection had significantly lower cTFH and activated cTFH pretransplant and early posttransplant. Pretransplant activated cTFH were also lower within patients who developed CMV disease. Pre- and 14 days posttransplant activated cTFH were an independent protective factor for CMV infection (HR 0.41, p = .01; and 0.52, p = .02, respectively). KTR with low cTFH 7 days posttransplant (<11.9%) had lower CMV infection-free survival than patients with high cTFH (28.2% vs. 67.6%, p = .002). cTFH were associated with CMV-specific neutralizing antibodies (Nabs). In addition, IL-21 increased interferon-γ secretion by CMV-specific CD8+ T cells in healthy controls. Thus, we show an association between cTFH and lower incidence of CMV infection, probably through their cooperation in CMV-specific Nab production and IL-21-mediated enhancement of CD8+ T cell activity. Moreover, monitoring cTFH pre- and early posttransplant could improve CMV risk stratification and help select KTR catalogued at low/intermediate risk who could benefit from prophylaxis.

Glucocerebrosidase Gene Therapy Induces Alpha-Synuclein Clearance and Neuroprotection of Midbrain Dopaminergic Neurons in Mice and Macaques.

Mutations in the GBA1 gene coding for glucocerebrosidase (GCase) are the main genetic risk factor for Parkinson's disease (PD). Indeed, identifying reduced GCase activity as a common feature underlying the typical neuropathological signatures of PD-even when considering idiopathic forms of PD-has recently paved the way for designing novel strategies focused on enhancing GCase activity to reduce alpha-synuclein burden and preventing dopaminergic cell death. Here we have performed bilateral injections of a viral vector coding for the mutated form of alpha-synuclein (rAAV9-SynA53T) for disease modeling purposes, both in mice as well as in nonhuman primates (NHPs), further inducing a progressive neuronal death in the substantia nigra pars compacta (SNpc). Next, another vector coding for the GBA1 gene (rAAV9-GBA1) was unilaterally delivered in the SNpc of mice and NHPs one month after the initial insult, together with the contralateral delivery of an empty/null rAAV9 for control purposes. Obtained results showed that GCase enhancement reduced alpha-synuclein burden, leading to improved survival of dopaminergic neurons. Data reported here support using GCase gene therapy as a disease-modifying treatment for PD and related synucleinopathies, including idiopathic forms of these disorders.

IL-6-based mortality prediction model for COVID-19: Validation and update in multicenter and second wave cohorts.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is a highly variable condition. Validated tools to assist in the early detection of patients at high risk of mortality can help guide medical decisions. OBJECTIVE: We sought to validate externally, as well as in patients from the second pandemic wave in Europe, our previously developed mortality prediction model for hospitalized COVID-19 patients. METHODS: Three validation cohorts were generated: 2 external with 185 and 730 patients from the first wave and 1 internal with 119 patients from the second wave. The probability of death was calculated for all subjects using our prediction model, which includes peripheral blood oxygen saturation/fraction of inspired oxygen ratio, neutrophil-to-lymphocyte ratio, lactate dehydrogenase, IL-6, and age. Discrimination and calibration were evaluated in the validation cohorts. The prediction model was updated by reestimating individual risk factor effects in the overall cohort (N = 1477). RESULTS: The mortality prediction model showed good performance in the external validation cohorts 1 and 2, and in the second wave validation cohort 3 (area under the receiver-operating characteristic curve, 0.94, 0.86, and 0.86, respectively), with excellent calibration (calibration slope, 0.86, 0.94, and 0.79; intercept, 0.05, 0.03, and 0.10, respectively). The updated model accurately predicted mortality in the overall cohort (area under the receiver-operating characteristic curve, 0.91), which included patients from both the first and second COVID-19 waves. The updated model was also useful to predict fatal outcome in patients without respiratory distress at the time of evaluation. CONCLUSIONS: This is the first COVID-19 mortality prediction model validated in patients from the first and second pandemic waves. The COR+12 online calculator is freely available to facilitate its implementation (https://utrero-rico.shinyapps.io/COR12_Score/).

Expression of GPR55 and either cannabinoid CB1 or CB2 heteroreceptor complexes in the caudate, putamen, and accumbens nuclei of control, parkinsonian, and dyskinetic non-human primates.

Endocannabinoids are neuromodulators acting on specific cannabinoid CB1 and CB2 G-protein-coupled receptors (GPCRs), representing potential therapeutic targets for neurodegenerative diseases. Cannabinoids also regulate the activity of GPR55, a recently "deorphanized" GPCR that directly interacts with CB1 and with CB2 receptors. Our hypothesis is that these heteromers may be taken as potential targets for Parkinson's disease (PD). This work aims at assessing the expression of heteromers made of GPR55 and CB1/CB2 receptors in the striatum of control and parkinsonian macaques (with and without levodopa-induced dyskinesia). For this purpose, double blind in situ proximity ligation assays, enabling the detection of GPCR heteromers in tissue samples, were performed in striatal sections of control, MPTP-treated and MPTP-treated animals rendered dyskinetic by chronic treatment with levodopa. Image analysis and statistical assessment were performed using dedicated software. We have previously demonstrated the formation of heteromers between GPR55 and CB1 receptor (CB1-GPR55_Hets), which is highly expressed in the central nervous system (CNS), but also with the CB2 receptor (CB2-GPR55_Hets). Compared to the baseline expression of CB1-GPR55_Hets in control animals, our results showed increased expression levels in basal ganglia input nuclei of MPTP-treated animals. These observed increases in CB1-GPR55_Hets returned back to baseline levels upon chronic treatment with levodopa in dyskinetic animals. Obtained data regarding CB2-GPR55_Hets were quite similar, with somehow equivalent amounts in control and dyskinetic animals, and with increased expression levels in MPTP animals. Taken together, the detected increased expression of GPR55-endocannabinoid heteromers appoints these GPCR complexes as potential non-dopaminergic targets for PD therapy.