Publications by authors named "Ruy M Ribeiro"

145 Publications

Bamlanivimab reduces nasopharyngeal SARS-CoV-2 RNA levels but not symptom duration in non-hospitalized adults with COVID-19.

medRxiv 2021 Dec 21. Epub 2021 Dec 21.

Importance: The antiviral activity and efficacy of anti-SARS-CoV-2 monoclonal antibody (mAb) therapies to accelerate recovery from COVID-19 is important to define.

Objective: To determine safety and efficacy of the mAb bamlanivimab to reduce nasopharyngeal (NP) SARS-CoV-2 RNA levels and symptom duration.

Design: ACTIV-2/A5401 is a randomized, blinded, placebo-controlled platform trial. Two dose cohorts were enrolled between August 19 and November 17, 2020 for phase 2 evaluation: in the first, participants were randomized 1:1 to bamlanivimab 7000 mg versus placebo, and in the second to bamlanivimab 700 mg versus placebo. Randomization was stratified by time from symptom onset (≤ or >5 days) and risk of progression to severe COVID-19 ("higher" vs "lower").

Setting: Multicenter trial conducted at U.S. sites.

Participants: Non-hospitalized adults ≥18 years of age with positive SARS-CoV-2 antigen or nucleic acid test within 7 days, ≤10 days of COVID-19 symptoms, and with oxygen saturation ≥92% within 48 hours prior to study entry.

Intervention: Single infusion of bamlanivimab (7000 or 700 mg) or placebo.

Main Outcomes And Measures: Detection of NP SARS-CoV-2 RNA at days 3, 7, 14, 21, and 28, time to improvement of all of 13 targeted COVID-19 symptoms by daily self-assessment through day 28, and grade 3 or higher treatment emergent adverse events (TEAEs) through day 28. Secondary measures included quantitative NP SARS-CoV-2 RNA, all-cause hospitalizations and deaths (composite), area under the curve of symptom scores from day 0 through day 28, plasma bamlanivimab concentrations, plasma and serum inflammatory biomarkers, and safety through week 24.

Results: Ninety-four participants were enrolled to the 7000 mg cohort and 223 to the 700 mg cohort and initiated study intervention. The proportion meeting protocol criteria for "higher" risk for COVID-19 progression was 42% and 51% for the 7000 and 700 mg cohort, respectively. Median time from symptom onset at study entry for both cohorts was 6 days. There was no difference in the proportion with undetectable NP SARS-CoV-2 RNA at any post-treatment timepoints (risk ratio compared to placebo, 0.82-1.05 for 7000 mg dose [overall p=0.88] and 0.81-1.21 for 700 mg dose [overall p=0.49]), time to symptom improvement (median of 21 vs 18.5 days, p=0.97, for 7000 mg bamlanivimab vs placebo and 24 vs 20.5 days, p=0.08, for 700 mg bamlanivimab vs placebo), or grade 3+ TEAEs with either dose compared to placebo. Median NP SARS-CoV-2 RNA levels were lower at day 3 and C-reactive protein, ferritin, and fibrinogen levels significantly reduced at days 7 and 14 for bamlanivimab 700 mg compared to placebo, with similar trends observed for bamlanivimab 7000 mg. Viral decay modeling supported more rapid decay with bamlanivimab compared to placebo.

Conclusions And Relevance: Treatment with bamlanivimab 7000 mg and 700 mg was safe and compared to placebo led to more rapid reductions in NP SARS-CoV-2 RNA and inflammatory biomarkers, but did not decrease time to symptom improvement. The clinical utility of mAbs for outcomes other than hospitalizations and deaths is uncertain.

Trial Registration: ClinicalTrials.gov Identifier: NCT04518410.

Key Points: What is the safety and efficacy of bamlanivimab monoclonal antibody (mAb) treatment for mild to moderate COVID-19? In this randomized, placebo-controlled phase 2 trial of 317 non-hospitalized adults with COVID-19, there was no relationship between symptoms or disease progression risk and nasopharyngeal (NP) virus shedding. Bamlanivimab was safe and reduced NP SARS-CoV-2 RNA levels and inflammatory biomarker levels more than placebo, but did not shorten symptom duration. Nasal virus shedding was not associated with symptoms or baseline risk factors for severe COVID-19. Bamlanivimab, which has been associated with reduced hospitalizations in high-risk individuals, demonstrated antiviral activity with early post-treatment NP sampling but did not accelerate symptom improvement. The clinical utility of bamlanivimab for outcomes other than hospitalizations and deaths, including longer-term outcomes, is uncertain.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2021.12.17.21268009DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8722620PMC
December 2021

In vivo kinetics of SARS-CoV-2 infection and its relationship with a person's infectiousness.

Proc Natl Acad Sci U S A 2021 12;118(49)

Theoretical Biology and Biophysics Group, Theoretical Division, Los Alamos National Laboratory, Los Alamos, NM 87545;

The within-host viral kinetics of SARS-CoV-2 infection and how they relate to a person's infectiousness are not well understood. This limits our ability to quantify the impact of interventions on viral transmission. Here, we develop viral dynamic models of SARS-CoV-2 infection and fit them to data to estimate key within-host parameters such as the infected cell half-life and the within-host reproductive number. We then develop a model linking viral load (VL) to infectiousness and show a person's infectiousness increases sublinearly with VL and that the logarithm of the VL in the upper respiratory tract is a better surrogate of infectiousness than the VL itself. Using data on VL and the predicted infectiousness, we further incorporated data on antigen and RT-PCR tests and compared their usefulness in detecting infection and preventing transmission. We found that RT-PCR tests perform better than antigen tests assuming equal testing frequency; however, more frequent antigen testing may perform equally well with RT-PCR tests at a lower cost but with many more false-negative tests. Overall, our models provide a quantitative framework for inferring the impact of therapeutics and vaccines that lower VL on the infectiousness of individuals and for evaluating rapid testing strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1073/pnas.2111477118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8670484PMC
December 2021

Comparison of Three Assays for Total and Free PSA Using Hybritech and WHO Calibrations.

In Vivo 2021 Nov-Dec;35(6):3431-3439

Department of Clinical Pathology, Centro Hospitalar Universitário de São João, Porto, Portugal.

Background/aim: Lack of interchangeability between prostate-specific antigen (PSA) assays could have a clinical impact. We compared PSA assays from different manufacturers and calibrations.

Patients And Methods: A total of 233 men who underwent prostate biopsy (PSA: 2-10 ng/ml; Beckman Coulter Access Hybritech as reference) were enrolled. Total (tPSA) and free PSA (fPSA) were also measured using the Roche cobas and the Abbott Architect methods.

Results: Roche tPSA values were ≈1% higher than Beckman, while Abbott values were ≈5% lower. Roche had the highest diagnostic sensitivity (92%) compared to Beckman Coulter (87%) and Abbott (85%). Roche fPSA was ≈3% lower and Abbott ≈17% higher than that of Beckman. For the percentage of fPSA, Roche had the highest sensitivity (98%).

Conclusion: Roche cobas and Beckman Coulter Access Hybritech tPSA were almost interchangeable. While the agreement was acceptable for tPSA, this did not happen with fPSA and greater efforts for harmonization are required.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.21873/invivo.12643DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8627779PMC
October 2021

Longitudinal SARS-CoV-2 seroprevalence in Portugal and antibody maintenance 12 months after infection.

Eur J Immunol 2022 01 10;52(1):149-160. Epub 2021 Nov 10.

Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.

During the COVID-19 pandemic, Portugal has experienced three distinct SARS-CoV-2 infection waves. We previously documented the prevalence of SARS-CoV-2 immunity, measured by specific antibodies, in September 2020, 6 months after the initial moderate wave. Here, we show the seroprevalence changes 6 months later, up to the second week of March 2021, shortly following the third wave, which was one of the most severe in the world, and 2 months following the start of the vaccination campaign. A longitudinal epidemiological study was conducted, with a stratified quota sample of the Portuguese population. Serological testing was performed, including ELISA determination of antibody class and titers. The proportion of seropositives, which was 2.2% in September 2020, rose sharply to 17.3% (95% CI: 15.8-18.8%) in March 2021. Importantly, circulating IgG and IgA antibody levels were very stable 6 months after the initial determination and up to a year after initial infection, indicating long-lasting infection immunity against SARS-CoV-2. Moreover, vaccinated people had higher IgG levels from 3 weeks post-vaccination when compared with previously infected people at the same time post-infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/eji.202149619DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8646574PMC
January 2022

Antiretroviral therapy for HIV and intrahepatic hepatitis C virus replication.

AIDS 2022 Mar;36(3):337-346

The Johns Hopkins Medical Institutions, Baltimore, Maryland.

Objective: HIV alters host responses to hepatitis C virus (HCV). However, the impact of antiretroviral therapy (ART) on HCV is rarely understood in relevant tissues and never before within individual hepatocytes.

Design: HIV and HCV kinetics were studied before and after ART initiation among 19 HIV/HCV co-infected persons. From five persons with the largest decline in plasma HCV RNA, liver tissues collected before and during ART, when plasma HIV RNA was undetectable, were studied.

Methods: We used single-cell laser capture microdissection and quantitative PCR to assess intrahepatic HCV. Immunohistochemistry was performed to characterize intrahepatic immune cell populations.

Results: Plasma HCV RNA declined by 0.81 (0.52-1.60) log10 IU/ml from a median (range) 7.26 (6.05-7.29) log10 IU/ml and correlated with proportions of HCV-infected hepatocytes (r = 0.89, P = 2 × 10-5), which declined from median (range) of 37% (6-49%) to 23% (0.5-52%) after plasma HIV clearance. Median (range) HCV RNA abundance within cells was unchanged in four of five participants. Liver T-cell abundance unexpectedly decreased, whereas natural killer (NK) and NK T-cell infiltration increased, correlating with changes in proportions of HCV-infected hepatocytes (r = -0.82 and r = -0.73, respectively). Hepatocyte expression of HLA-E, an NK cell restriction marker, correlated with proportions of HCV-infected hepatocytes (r = 0.79).

Conclusion: These are the first data to show that ART control of HIV reduces the intrahepatic burden of HCV. Furthermore, our data suggest that HIV affects the pathogenesis of HCV infection by an NK/NK T-cell-mediated mechanism that may involve HLA-E and can be rescued, at least in part, by ART.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1097/QAD.0000000000003116DOI Listing
March 2022

Emergence of SARS-CoV-2 Resistance with Monoclonal Antibody Therapy.

medRxiv 2021 Sep 15. Epub 2021 Sep 15.

Resistance mutations to monoclonal antibody (mAb) therapy has been reported, but in the non-immunosuppressed population, it is unclear if emergence of SARS-CoV-2 resistance mutations alters either viral replication dynamics or therapeutic efficacy. In ACTIV-2/A5401, non-hospitalized participants with symptomatic SARS-CoV-2 infection were randomized to bamlanivimab (700mg or 7000mg) or placebo. Treatment-emergent resistance mutations were significantly more likely detected after bamlanivimab 700mg treatment than placebo (7% of 111 vs 0% of 112 participants, P=0.003). There were no treatment-emergent resistance mutations among the 48 participants who received bamlanivimab 7000mg. Participants with emerging mAb resistant virus had significantly higher pre-treatment nasopharyngeal and anterior nasal viral load. Intensive respiratory tract viral sampling revealed the dynamic nature of SARS-CoV-2 evolution, with evidence of rapid and sustained viral rebound after emergence of resistance mutations, and worsened symptom severity. Participants with emerging bamlanivimab resistance often accumulated additional polymorphisms found in current variants of concern/interest and associated with immune escape. These results highlight the potential for rapid emergence of resistance during mAb monotherapy treatment, resulting in prolonged high level respiratory tract viral loads and clinical worsening. Careful virologic assessment should be prioritized during the development and clinical implementation of antiviral treatments for COVID-19.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2021.09.03.21263105DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8452115PMC
September 2021

Prevalence of SARS-CoV-2 Antibodies after First 6 Months of COVID-19 Pandemic, Portugal.

Emerg Infect Dis 2021 11 26;27(11):2878-2881. Epub 2021 Aug 26.

In September 2020, we tested 13,398 persons in Portugal for antibodies against severe acute respiratory syndrome coronavirus 2 by using a quota sample stratified by age and population density. We found a seroprevalence of 2.2%, 3-4 times larger than the official number of cases at the end of the first wave of the pandemic.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3201/eid2711.210636DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8544986PMC
November 2021

Viral Load Kinetics of Severe Acute Respiratory Syndrome Coronavirus 2 in Hospitalized Individuals With Coronavirus Disease 2019.

Open Forum Infect Dis 2021 Aug 17;8(8):ofab153. Epub 2021 Apr 17.

Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts, USA.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) kinetics remain understudied, including the impact of remdesivir. In hospitalized individuals, peak sputum viral load occurred in week 2 of symptoms, whereas viremia peaked within 1 week of symptom-onset, suggesting early systemic seeding of SARS-CoV-2. Remdesivir treatment was associated with faster viral decay.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/ofid/ofab153DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8083268PMC
August 2021

Telemonitoring of daily activities compared to the six-minute walk test further completes the puzzle of oximetry-guided interventions.

Sci Rep 2021 08 16;11(1):16600. Epub 2021 Aug 16.

Instituto de Saúde Ambiental, Faculdade de Medicina, Universidade de Lisboa, Lisbon, Portugal.

Pulmonary rehabilitation is based on a thorough patient assessment, including peripheral oxygen saturation (SpO) and heart rate (HR) at rest and on exertion. To understand whether exercise-field tests identify patients who desaturate (SpO < 90%) during physical activities, this study compared the six-minute walk test (6MWT) and daily-life telemonitoring. Cross-sectional study including 100 patients referred for pulmonary rehabilitation. The 6MWT was performed in hospital with continuous assessment of SpO, HR, walked distance and calculated metabolic equivalent of tasks (METs). Patients were also evaluated in real-life by SMARTREAB telemonitoring, a combined oximetry-accelerometery with remote continuous assessment of SpO, HR and METs. SMARTREAB telemonitoring identified 24% more desaturators compared with the 6MWT. Moreover, there were significant mean differences between 6MWT and SMARTREAB in lowest SpO of 7.2 ± 8.4% (P < 0.0005), in peak HR of - 9.3 ± 15.5% (P < 0.0005) and also in activity intensity of - 0.3 ± 0.8 METs (P < 0.0005). The 6MWT underestimates the proportion of patients with exercise-induced oxygen desaturation compared to real-life telemonitoring. These results help defining oximetry-guided interventions, such as telemedicine algorithms, oxygen therapy titration and regular physical activity assessment in pulmonary rehabilitation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41598-021-96060-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8367992PMC
August 2021

In vivo kinetics of SARS-CoV-2 infection and its relationship with a person's infectiousness.

medRxiv 2021 Jun 30. Epub 2021 Jun 30.

Theoretical Biology and Biophysics Group, Theoretical Division, Los Alamos National Laboratory, Los Alamos, NM 87545, USA.

The within-host viral kinetics of SARS-CoV-2 infection and how they relate to a person's infectiousness are not well understood. This limits our ability to quantify the impact of interventions on viral transmission. Here, we develop data-driven viral dynamic models of SARS-CoV-2 infection and estimate key within-host parameters such as the infected cell half-life and the within-host reproductive number. We then develop a model linking VL to infectiousness, showing that a person's infectiousness increases sub-linearly with VL. We show that the logarithm of the VL in the upper respiratory tract (URT) is a better surrogate of infectiousness than the VL itself. Using data on VL and the predicted infectiousness, we further incorporated data on antigen and reverse transcription polymerase chain reaction (RT-PCR) tests and compared their usefulness in detecting infection and preventing transmission. We found that RT-PCR tests perform better than antigen tests assuming equal testing frequency; however, more frequent antigen testing may perform equally well with RT-PCR tests at a lower cost, but with many more false-negative tests. Overall, our models provide a quantitative framework for inferring the impact of therapeutics and vaccines that lower VL on the infectiousness of individuals and for evaluating rapid testing strategies.

Significance: Quantifying the kinetics of SARS-CoV-2 infection and individual infectiousness is key to quantitatively understanding SARS-CoV-2 transmission and evaluating intervention strategies. Here we developed data-driven within-host models of SARS-CoV-2 infection and by fitting them to clinical data we estimated key within-host viral dynamic parameters. We also developed a mechanistic model for viral transmission and show that the logarithm of the viral load in the upper respiratory tract serves an appropriate surrogate for a person's infectiousness. Using data on how viral load changes during infection, we further evaluated the effectiveness of PCR and antigen-based testing strategies for averting transmission and identifying infected individuals.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1101/2021.06.26.21259581DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8259912PMC
June 2021

Relevance of Circulating Nucleosomes, HMGB1 and sRAGE for Prostate Cancer Diagnosis.

In Vivo 2021 Jul-Aug;35(4):2207-2212

Institute of Laboratory Medicine, Munich Biomarker Research Center, Deutsches Herzzentrum München, Technische Universität München, Munich, Germany.

Background/aim: Evasion from cell death occurs in prostate cancer (PCa). We verified whether serum levels of cell death markers can have diagnostic value in PCa.

Patients And Methods: A total of 233 men scheduled for prostate biopsy [prostate specific antigen (PSA) level: 2-10 ng/ml] were enrolled. Serum nucleosomes, nucleosomes containing the H3 histone (H3), high mobility group box 1 (HMGB1), and soluble receptor for advanced glycation end products (sRAGE) were analyzed by enzyme immunoassays.

Results: There were no differences (p>0.05) in nucleosomes, H3, and sRAGE levels between patients with and without PCa or clinically significant PCa (csPCa). HMGB1 had lower levels in PCa patients (p=0.023) and was a predictor of PCa (p=0.047), but not of csPCa (p=0.180).

Conclusion: In patients with critical PSA levels between 2-10 ng/ml, HMGB1 had some diagnostic value for overall PCa detection, but it was not predictive of csPCa. Nucleosomes, H3 and sRAGE did not discriminate between PCa or csPCa and controls.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.21873/invivo.12492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8286487PMC
June 2021

The prostate health index (PHI) density: Are there advantages over PHI or over the prostate-specific antigen density?

Clin Chim Acta 2021 Sep 5;520:133-138. Epub 2021 Jun 5.

Department of Clinical Pathology, Centro Hospitalar Universitário de São João, Porto, Portugal; Department of Biomedicine, Faculdade de Medicina da Universidade do Porto, Porto, Portugal; EPIUnit, Instituto de Saúde Pública, Universidade do Porto, Porto, Portugal.

Background And Aims: Overdiagnosis of prostate cancer (PCa) should be minimized. We wanted to evaluate the diagnostic performance of the prostate health index density (PHID) and compare it with that of the prostate health index (PHI) alone and of the prostate-specific antigen density (PSAD).

Materials And Methods: 232 men scheduled for a prostate biopsy (prostate-specific antigen level: 2-10 µg/L), were enrolled. PHI, PHID and PSAD were evaluated considering PCa and clinically significant PCa (csPCa) as the outcomes.

Results: For PCa, the area under the curve (AUC) was higher for PHID (0.823) than for PHI (0.779) and PSAD (0.776). For csPCa, the AUC was also higher for PHID (0.851) but closer to that of PSAD (0.819) and PHI (0.813). For equal sensitivities (90%) for PCa, PHID and PSAD offered the highest specificities (37%), missing the same number of cancers (n = 11). Considering csPCa, PHI and PHID had similar specificities. PSAD reached the highest specificity (50.0%), sparing 32.8% of biopsies, while missing 9 cases of csPCa.

Conclusions: PHID has a better diagnostic performance than PHI for overall PCa detection, but very close to the PSAD performance. Considering csPCa, PHI and PHID perform almost equally, but PSAD has a better diagnostic performance.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cca.2021.06.006DOI Listing
September 2021

Are Proinflammatory Cytokines Relevant for the Diagnosis of Prostate Cancer?

Anticancer Res 2021 Jun;41(6):3067-3073

Institute of Laboratory Medicine, Munich Biomarker Research Center, Deutsches Herzzentrum München, Technische Universität München, Munich, Germany.

Background/aim: Inflammation plays an important role in prostate cancer (PCa). We evaluated proinflammatory cytokines regarding differential diagnosis of PCa in men with PSA levels between 2-10 ng/ml.

Patients And Methods: Serum samples of 79 men (PSA 2-10 ng/ml) were analyzed for 10 proinflammatory cytokines (IL-6, IL-8, TNF-α, IFN-γ, IL-10, IL-1β, IL-2, IL-4, IL-12p70, IL-13) and results were evaluated regarding presence of PCa and disease severity.

Results: Significant differences between PCa patients and controls were found for IL-6 (p=0.002), IL-8 (p=0.030), and TNF-α (p=0.009), although they were not predictors of PCa in a logistic regression analysis. In addition, IL-6 and TNF-α levels were significantly higher in patients with high-risk PCa (p<0.05). No significant differences were observed regarding the other cytokines.

Conclusion: In patients with PSA levels between 2-10 ng/ml, IL-6, IL-8, and TNF-α are associated with PCa, and IL-6 and TNF-α are associated with high-risk PCa.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.21873/anticanres.15090DOI Listing
June 2021

Serum lipids and prostate cancer.

J Clin Lab Anal 2021 Apr 16;35(4):e23705. Epub 2021 Mar 16.

Department of Clinical Pathology, Sao Joao University Hospital Center & Department of Biomedicine, School of Medicine & Institute of Public Health, University of Porto, Porto, Portugal.

Background: Conflicting results are found in the literature relating serum lipids levels and prostate cancer. Some results imply a relationship between them; others contradict this association. The purpose of this study was to investigate a possible association between serum lipids levels and prostate cancer, at time of diagnosis.

Methods: We measured serum levels of total cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides in 237 patients submitted to a prostate biopsy, with PSA between 2 and 10 ng/ml. Patients without cancer at biopsy were used as controls, and the others were considered as cases. No information about lipid-lowering therapy, including statins, was available neither in cases nor in controls. Cases were divided into risk groups, according to the disease severity, based on staging. Lipids levels were compared between groups, using parametric and nonparametric tests. Logistic regression analysis and odds ratios were calculated.

Results: LDL and total cholesterol levels were lower in patients with cancer, with the difference being statistically significant for LDL cholesterol (p = 0.010) and borderline for total cholesterol (p = 0.050). No significant differences were found between the several risk groups. Odds ratios for low LDL cholesterol (<130 mg/dl) and low total cholesterol (<200 mg/dl), with prostate cancer as the outcome, were 1.983 and 1.703, respectively. There were no significant differences between cases and controls for the other lipids.

Conclusion: Lower LDL cholesterol (<130 mg/dl) and lower total cholesterol (<200 mg/dl) serum levels seem to associate with prostate cancer, at time of diagnosis.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/jcla.23705DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8059719PMC
April 2021

Infective Endocarditis as the Cause of Death: A Populationbased Study in Portugal, from 2002 to 2018.

Acta Med Port 2021 Dec 8;34(12):833-841. Epub 2021 Mar 8.

Centro Cardiovascular da Universidade de Lisboa (CCUL). Faculdade de Medicina. Universidade de Lisboa. Lisboa. Departamento Coração e Vasos. Centro Hospitalar e Universitário Lisboa Norte (CHULN). Lisboa. Portugal.

Introduction: Infective endocarditis presents a high rate of morbidity and mortality. Population-based studies addressing mortality caused by infective endocarditis in Portugal are scarce. We aimed to study deaths caused by Infective endocarditis, as well as corresponding demographics and temporal trends.

Material And Methods: Retrospective cohort study of all patients whose main cause of death was Infective endocarditis in Portugal from 2002 to 2018. The data was obtained from the national death certificate information system.

Results: In Portugal, 3634 people died from infective endocarditis throughout the 17-year study period - infective endocarditis specific mortality rate of 2.1 per 100 000 habitants. Of all deceased, 89% were at least 60 years old, and most were women (55%). Overall, 72% died in a healthcare institution. An annual 9% increase in the incidence death rate from Infective endocarditis was observed, with a significant upward trend during the colder months.

Discussion: In Portugal, mortality by infective endocarditis increased, mainly affecting older patients and women, and which can partially be explained by factors such as ageing of the population. The management of older patients with infective endocarditis is challenging as they present a higher number of comorbidities, more valvular heart disease and valve implants, invasive medical procedures and are less likely to undergo cardiac surgery.

Conclusion: In addition to data on the evolution of demographics in Portugal, it is crucial to study the incidence of infective endocarditis over time to help explain these findings. The identification of factors that can be used to better model national health policies to improve clinical outcomes of infective endocarditis in Portugal is also required.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.20344/amp.14609DOI Listing
December 2021

The burden of infective endocarditis in Portugal in the last 30 years - a systematic review of observational studies.

Rev Port Cardiol (Engl Ed) 2021 Mar 27;40(3):205-217. Epub 2021 Feb 27.

Centro Cardiovascular da Universidade de Lisboa (CCUL), Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal; Departamento Coração e Vasos, Centro Hospitalar e Universitário Lisboa Norte (CHULN), EPE, Lisboa, Portugal.

Introduction: Infective endocarditis affects cardiac valves or devices and has a potentially uncertain prognosis. Little information is available on the epidemiology of this disease in Portugal.

Objective: A systematic review of all evidence published in the last 30 years to assess epidemiological data in patients hospitalized with infective endocarditis in Portuguese hospital centers.

Methods: Extensive search of all published evidence using Medline, Scopus, general search databases and in addition Portuguese medical journals was performed. All relevant studies in Portuguese or English that reported short- or long-term mortality were included.

Results: Eighteen retrospective cohort studies (15 medical and three surgical series) were included with a total of 1872 patients assessed. The medical series included 1279 patients. Older males with predominant native left heart valve involvement were identified. Staphylococcus and streptococcus were the most frequent reported pathogens. Surgical intervention was performed on average in 29.8% of cases. The short-term mortality rate ranged from three to 37.2% (average 21.9%). Surgical cases involved older males with affected native left heart valves, emergent/urgent indication was dominant and short-term mortality ranged from 13.6 to 16%.

Conclusions: The current study provides a descriptive analysis of the published series of infective endocarditis in Portugal over the last 30 years. Therefore, it may serve as a starting point for the development and implementation of a multicentric prospective registry on infective endocarditis patients in Portugal that will allow a better and more accurate characterization of this special patient population.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.repc.2020.07.014DOI Listing
March 2021

Zika virus dynamics: Effects of inoculum dose, the innate immune response and viral interference.

PLoS Comput Biol 2021 01 20;17(1):e1008564. Epub 2021 Jan 20.

Theoretical Biology and Biophysics, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America.

Experimental Zika virus infection in non-human primates results in acute viral load dynamics that can be well-described by mathematical models. The inoculum dose that would be received in a natural infection setting is likely lower than the experimental infections and how this difference affects the viral dynamics and immune response is unclear. Here we study a dataset of experimental infection of non-human primates with a range of doses of Zika virus. We develop new models of infection incorporating both an innate immune response and viral interference with that response. We find that such a model explains the data better than models with no interaction between virus and the immune response. We also find that larger inoculum doses lead to faster dynamics of infection, but approximately the same total amount of viral production.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1371/journal.pcbi.1008564DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7817008PMC
January 2021

HIV influences clustering and intracellular replication of hepatitis C virus.

J Viral Hepat 2021 02 24;28(2):334-344. Epub 2020 Nov 24.

Theoretical Biology and Biophysics, Los Alamos National Laboratory, Los Alamos, NM, USA.

HCV and HIV coinfection is common and HIV leads to increased HCV viraemia and accelerated disease progression. However, the biological basis of this interaction remains poorly understood and little is known about the impact of HIV on HCV replication at the cellular level. We analysed HCV RNA, based on single-cell laser-capture microdissection, in liver biopsies from monoinfected (n = 4) and HCV/HIV-coinfected (n = 5) participants. HCV RNA was assayed in 3200 hepatocytes with information of spatial position. We compared HCV RNA levels and clustering properties of infection between mono- and coinfected participants, and developed a mathematical model of infection. Although the median plasma HCV RNA level and the fraction of infected cells were comparable in monoinfected (7.0 log IU/mL and ~ 30%) and coinfected (7.3 log IU/mL and ~ 40%) participants, the median HCV RNA per infected hepatocyte in monoinfected (2.8IU) was significantly lower than in coinfected (8.2IU) participants (p = .03). Clustering of infected cells was more prominent in monoinfected participants (91% of samples) than in coinfected participants (~48%), p = .0045, suggesting that spatial spread may be influenced by HIV coinfection. Interestingly, when clustering does occur, the size of clusters is similar in both types of infection. A mathematical model of infection suggested that HIV allows higher intracellular accumulation of HCV RNA by impeding the export of HCV RNA. Our observations show that HIV coinfection impacts intracellular accumulation of HCV RNA and the clustering of HCV-infected cells, but to a less extent the fraction of HCV-infected cells.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/jvh.13429DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7855861PMC
February 2021

Single hepatocytes show persistence and transcriptional inactivity of hepatitis B.

JCI Insight 2020 10 2;5(19). Epub 2020 Oct 2.

Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.

There is no cure for the more than 270 million people chronically infected with HBV. Nucleos(t)ide analogs (NUCs), the mainstay of anti-HBV treatment, block HBV reverse transcription. NUCs do not eliminate the intranuclear covalently closed circular DNA (cccDNA), from which viral RNAs, including pregenomic RNA (pgRNA), are transcribed. A key gap in designing a cure is understanding how NUCs affect HBV replication and transcription because serum markers yield an incomplete view of intrahepatic HBV. We applied single-cell laser capture microdissection and droplet digital PCR to paired liver biopsies collected from 5 HBV/HIV-coinfected persons who took NUCs over 2-4 years. From biopsy 1 to 2, proportions of HBV-infected hepatocytes declined with adherence to NUC treatment (P < 0.05); we extrapolated that eradication of HBV will take over 10 decades with NUCs in these participants. In individual hepatocytes, pgRNA levels diminished 28- to 73-fold during NUC treatment, corresponding with decreased tissue HBV core antigen staining (P < 0.01). In 4 out of 5 participants, hepatocytes with cccDNA but undetectable pgRNA (transcriptionally inactive) were present, and these were enriched in 3 participants during NUC treatment. Further work to unravel mechanisms of cccDNA transcriptional inactivation may lead to therapies that can achieve this in all hepatocytes, resulting in a functional cure.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/jci.insight.140584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7566712PMC
October 2020

Novel Input for Designing Patient-Tailored Pulmonary Rehabilitation: Telemonitoring Physical Activity as a Vital Sign-SMARTREAB Study.

J Clin Med 2020 Jul 31;9(8). Epub 2020 Jul 31.

Instituto de Saúde Ambiental (ISAMB), Faculdade de Medicina, Universidade de Lisboa, 1649 028 Lisbon, Portugal.

Physical inactivity may be a consequence of chronic diseases but also a potential modifiable risk factor. Therefore, it should be clinically assessed as a vital sign of patients' general physical condition prior to any exercise-based intervention. This cross-sectional study describes physical activity in the daily life of 100 chronic respiratory patients before pulmonary rehabilitation, comparing subjective and objective measures. The assessment combined the International Physical Activity Questionnaire (IPAQ) and 4-day accelerometer and oximeter telemonitoring with SMARTREAB technology, assessing heart rate, transcutaneous oxygen saturation and activity-related energy expenditure by metabolic equivalent of task (MET). According to IPAQ, 49% of patients had a moderate level of physical activity in daily life (PADL), a weekly mean level of 2844 ± 2925 MET.min/week, and a mean sedentary time of 5.8 ± 2.7 h/day. Alongside this, SMARTREAB telemonitoring assessed maximum activity ranging from 1.51 to 4.64 METs, with 99.6% daytime spent on PADL below 3 METs and 93% of patients with daily desaturation episodes. Regardless of the self-reported IPAQ, patients spend at least 70% of daytime on PADL below 2 METs. SMARTREAB was demonstrated to be an innovative methodology to measure PADL as a vital sign, combining oximetry with accelerometry, crossmatched with qualitative patient data, providing important input for designing patient-tailored pulmonary rehabilitation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jcm9082450DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464888PMC
July 2020

The Role of Health Preconditions on COVID-19 Deaths in Portugal: Evidence from Surveillance Data of the First 20293 Infection Cases.

J Clin Med 2020 Jul 24;9(8). Epub 2020 Jul 24.

IMPSP-Instituto de Medicina Preventiva e Saúde Pública, Faculdade de Medicina, Universidade de Lisboa, Avenida Professor Egas Moniz, 1649-028 Lisboa, Portugal.

Background: It is essential to study the effect of potential co-factors on the risk of death in patients infected by COVID-19. The identification of risk factors is important to allow more efficient public health and health services strategic interventions with a significant impact on deaths by COVID-19. This study aimed to identify factors associated with COVID-19 deaths in Portugal.

Methods: A national dataset with the first 20,293 patients infected with COVID-19 between 1 January and 21 April 2020 was analyzed. The primary outcome measure was mortality by COVID-19, measured (registered and confirmed) by Medical Doctors serving as health delegates on the daily death registry. A logistic regression model using a generalized linear model was used for estimating Odds Ratio (OR) with 95% confidence intervals (95% CI) for each potential risk indicator.

Results: A total of 502 infected patients died of COVID-19. The risk factors for increased odds of death by COVID-19 were: sex (male: OR = 1.47, ref = female), age ((56-60) years, OR = 6.01; (61-65) years, OR = 10.5; (66-70) years, OR = 20.4; (71-75) years, OR = 34; (76-80) years, OR = 50.9; (81-85) years, OR = 70.7; (86-90) years, OR = 83.2; (91-95) years, OR = 91.8; (96-104) years, OR = 140.2, ref = (0-55)), Cardiac disease (OR = 2.86), Kidney disorder (OR = 2.95), and Neuromuscular disorder (OR = 1.58), while condition (None (absence of precondition); OR = 0.49) was associated with a reduced chance of dying after adjusting for other variables of interest.

Conclusions: Besides age and sex, preconditions justify the risk difference in mortality by COVID-19.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jcm9082368DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7464004PMC
July 2020

Untangling the immune basis of disease susceptibility.

Elife 2020 05 14;9. Epub 2020 May 14.

Instituto de Medicina Molecular, Faculdade de Medicina, Universidade de Lisboa, Lisboa, Portugal.

Interactions between immune cell receptors and proteins that determine disease susceptibility shed light on how different arms of the immune system are involved in three viral infections and Crohn's disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.7554/eLife.56886DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7224693PMC
May 2020

The impact of negative lymph nodes in the survival outcomes of pN+ patients following radical gastrectomy: the inverse lymph node ratio as a better score to study negative lymph nodes.

Updates Surg 2020 Dec 9;72(4):1031-1040. Epub 2020 May 9.

Laboratório de Biomatemática, Lisboa, Portugal.

The impact of negative lymph nodes (LNs) on survival of pN+ patients has been recognized. The weight of negative LNs in an inverse lymph node ratio (nR) should be related to its prognostic impact. Five hundred and two consecutive gastric cancer patients, who underwent radical gastrectomy, were included. Patients were split into groups according to the number of harvested nodes and a cross-tabulation with pTNM stages was performed to test differences in the tumor burden. pN+ patients (n = 296) were split into groups of negative LNs harvested. We tested an alternative formula for computing a lymph node ratio: nR = total number of harvested nodes/total number of positive nodes. The median number of negative LNs was significantly different (p < 0.01) between dissection groups, but not the median of positive nodes (p > 0.05). No difference in pTNM percentage distribution was found between these groups (p > 0.05). When tested, the overall survival improved significantly for groups with larger numbers of negative LNs (p < 0.001). A cutoff of nR ≥ 6 was an independent prognostic factor for survival (p = 0.001), and the survival of pN+ patients with nR ≥ 6 was not different from pN0 patients. The impact of the number of negative LNs on the survival of the pN+ patients was demonstrated. The higher numbers in the numerator of the nR was due to the disproportion between harvested negative LNs and metastatic LNs. Larger ratios imply more negative lymph nodes in relation to positive lymph nodes, which was significantly associated with survival. We believe that the proposed nR is a friendlier to use format because of its intuitive interpretation.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1007/s13304-020-00757-yDOI Listing
December 2020

GO-DACT: a phase 3b randomised, double-blind, placebo-controlled trial of GOlimumab plus methotrexate (MTX) versus placebo plus MTX in improving DACTylitis in MTX-naive patients with psoriatic arthritis.

Ann Rheum Dis 2020 04;79(4):490-498

Rheumatology Research Unit, Instituto de Medicina Molecular, Faculdade de Medicina da Universidade de Lisboa, Lisboa, Portugal.

Objectives: To assess the efficacy of golimumab in combination with methotrexate (MTX) versus MTX monotherapy in psoriatic arthritis (PsA) dactylitis.

Methods: Multicentre, investigator-initiated, randomised, double-blind, placebo-controlled, parallel-design phase 3b trial in 11 Portuguese rheumatology centres. Patients with PsA along with active dactylitis and naive to MTX and biologic disease-modifying antirheumatic drugs (bDMARDs) were randomly assigned to golimumab or placebo, both in combination with MTX. The primary endpoint was Dactylitis Severity Score (DSS) change from baseline to week 24. Key secondary endpoints included DSS and Leeds Dactylitis Index (LDI) response, and changes from baseline in the LDI and MRI dactylitis score. Analysis was by intention-to-treat for the primary endpoint.

Results: Twenty-one patients received golimumab plus MTX and 23 MTX monotherapy for 24 weeks. One patient from each arm discontinued. Patient inclusion was halted at 50% planned recruitment due to a favourable interim analysis. Median baseline DSS was 6 in both arms. By week 24, patients treated with golimumab plus MTX exhibited significantly greater improvements in DSS relative to MTX monotherapy (median change of 5 vs 2 points, respectively; p=0.026). In the golimumab plus MTX arm, significantly higher proportions of patients achieved at least 50% or 70% improvement in DSS and 20%, 50% or 70% improvement in LDI in comparison to MTX monotherapy.

Conclusions: The combination of golimumab and MTX as first-line bDMARD therapy is superior to MTX monotherapy for the treatment of PsA dactylitis.

Trial Registration Number: NCT02065713.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/annrheumdis-2019-216500DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7147178PMC
April 2020

High-fat diet exacerbates SIV pathogenesis and accelerates disease progression.

J Clin Invest 2019 12;129(12):5474-5488

Center for Vaccine Research.

Consuming a high-fat diet (HFD) is a risk factor for obesity and diabetes; both of these diseases are also associated with systemic inflammation, similar to HIV infection. A HFD induces intestinal dysbiosis and impairs liver function and coagulation, with a potential negative impact on HIV/SIV pathogenesis. We administered a HFD rich in saturated fats and cholesterol to nonpathogenic (African green monkeys) and pathogenic (pigtailed macaques) SIV hosts. The HFD had a negative impact on SIV disease progression in both species. Thus, increased cell-associated SIV DNA and RNA occurred in the HFD-receiving nonhuman primates, indicating a potential reservoir expansion. The HFD induced prominent immune cell infiltration in the adipose tissue, an important SIV reservoir, and heightened systemic immune activation and inflammation, altering the intestinal immune environment and triggering gut damage and microbial translocation. Furthermore, HFD altered lipid metabolism and HDL oxidation and also induced liver steatosis and fibrosis. These metabolic disturbances triggered incipient atherosclerosis and heightened cardiovascular risk in the SIV-infected HFD-receiving nonhuman primates. Our study demonstrates that dietary intake has a discernable impact on the natural history of HIV/SIV infections and suggests that dietary changes can be used as adjuvant approaches for HIV-infected subjects, to reduce inflammation and the risk of non-AIDS comorbidities and possibly other infectious diseases.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1172/JCI121208DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6877342PMC
December 2019

Disentangling the lifespans of hepatitis C virus-infected cells and intracellular vRNA replication-complexes during direct-acting anti-viral therapy.

J Viral Hepat 2020 03 22;27(3):261-269. Epub 2019 Nov 22.

Theoretical Biology and Biophysics Group, Los Alamos National Laboratory, Los Alamos, NM, USA.

The decay rate of hepatitis C virus (HCV)-infected cells during therapy has been used to determine the duration of treatment needed to attain a sustained virologic response, but with direct-acting anti-virals (DAA), this rate has been difficult to estimate. Here, we show that it is possible to estimate it, by simultaneously analysing the viral load and alanine aminotransferase (ALT) kinetics during combination DAA therapy. We modelled the HCV RNA and ALT serum kinetics in 26 patients with chronic HCV genotype 1b infection, under four different sofosbuvir-based combination treatments. In all patients, ALT decayed exponentially to a set point in the normal range by 1-3 weeks after initiation of therapy. The model indicates that the ALT decay rate during the first few weeks after initiation of therapy reflects the death rate of infected cells, with an estimated median half-life of 2.5 days in this patient population. This information allows independent estimation of the rate of loss of intracellular replication complexes during therapy. Our model also predicts that the final ALT set point is not related to the release of ALT by dying HCV-infected cells. Using ALT data, one can separately obtain information about the rate of 'cure' of HCV-infected cells versus their rate of death, something not possible when analysing only HCV RNA data. This information can be used to compare the effects of different DAA combinations and to rationally evaluate their anti-viral effects.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/jvh.13229DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7031045PMC
March 2020

Modeling the immune response to HIV infection.

Curr Opin Syst Biol 2018 Dec 8;12:61-69. Epub 2018 Nov 8.

Laboratorio de Biomatematica, Faculdade de Medicina da Universidade de Lisboa, Portugal and Theoretical Biology and Biophysics, Los Alamos National Laboratory, Los Alamos, NM 87545, USA.

The interplay between immune response and HIV is intensely studied via mathematical modeling, with significant insights but few direct answers. In this short review, we highlight advances and knowledge gaps across different aspects of immunity. In particular, we identify the innate immune response and its role in priming the adaptive response as ripe for modeling. The latter have been the focus of most modeling studies, but we also synthesize key outstanding questions regarding effector mechanisms of cellular immunity and development of broadly neutralizing antibodies. Thus far, most modeling studies aimed to infer general immune mechanisms; we foresee that significant progress will be made next by detailed quantitative fitting of models to data, and prediction of immune responses.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.coisb.2018.10.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6713454PMC
December 2018

Naive and Stem Cell Memory T Cell Subset Recovery Reveals Opposing Reconstitution Patterns in CD4 and CD8 T Cells in Chronic Graft vs. Host Disease.

Front Immunol 2019 6;10:334. Epub 2019 Mar 6.

JLacerda Lab, Hematology and Transplantation Immunology, Instituto de Medicina Molecular, Faculdade de Medicina da Universidade de Lisboa, Lisbon, Portugal.

The success of allogeneic hematopoietic stem cell transplantation (allo-HSCT) in the treatment of hematological malignancies remains hampered by life-threatening chronic graft vs. host disease (cGVHD). Although multifactorial in nature, cGVHD has been associated with imbalances between effector and regulatory T cells (Treg). To further elucidate this issue, we performed a prospective analysis of patients undergoing unrelated donor allo-HSCT after a reduced intensity conditioning (RIC) regimen containing anti-thymocyte globulin (ATG) and the same GVHD prophylaxis, at a single institution. We studied T cell subset homeostasis over a 24-month follow-up after HSCT in a comparative analysis of patients with and without cGVHD. We also quantified naive and memory T cell subsets, proliferation and expression of the apoptosis-related proteins Bcl-2 and CD95. Finally, we assessed thymic function by T cell receptor excision circle (TREC) quantification and T cell receptor (TCR) diversity by TCRVβ spectratyping. While the total number of conventional CD4 (Tcon) and CD8 T cells was similar between patient groups, Treg were decreased in cGVHD patients. Interestingly, we also observed divergent patterns of Naive and Stem Cell Memory (SCM) subset recovery in Treg and Tcon compared to CD8. Patients with cGVHD showed impaired recovery of Naive and SCM Tcon and Treg, but significantly increased frequencies and absolute numbers of Naive and SCM were observed in the CD8 pool. Markedly increased EMRA CD8 T cells were also noted in cGVHD. Taken together, these results suggest that Naive, SCM and EMRA CD8 play a role in the emergence of cGHVD. Reduced Naive and recent thymic emigrant Tcon and Treg in cGVHD was likely due to impaired thymic output, as it was accompanied by decreased CD4 TREC and TCR diversity. On the other hand, CD8 TCR diversity was similar between patient groups. Furthermore, no correlation was observed between CD8 TREC content and Naive CD8 numbers, suggesting limited thymic production of Naive CD8 T cells in patients after transplant, especially in those developing cGVHD. The mechanisms behind the opposing patterns of CD4 and CD8 subset cell recovery in cGVHD remain elusive, but may be linked to thymic damage associated with the conditioning regimen and/or acute GVHD.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fimmu.2019.00334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6414429PMC
May 2020
-->