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BACKGROUND: SARS-CoV-2 binding to ACE2 is potentially associated with severe pneumonia due to COVID-19. The aim of the study was to test whether Mas-receptor activation by 20-hydroxyecdysone (BIO101) could restore the Renin-Angiotensin System equilibrium and limit the frequency of respiratory failure and mortality in adults hospitalized with severe COVID-19. METHODS: Double-blind, randomized, placebo-controlled phase 2/3 trial. Randomization: 1:1 oral BIO101 (350 mg BID) or placebo, up to 28 days or until an endpoint was reached. Primary endpoint: mortality or respiratory failure requiring high-flow oxygen, mechanical ventilation, or extra-corporeal membrane oxygenation. Key secondary endpoint: hospital discharge following recovery (ClinicalTrials.gov Number, NCT04472728). FINDINGS: Due to low recruitment the planned sample size of 310 was not reached and 238 patients were randomized between August 26, 2020 and March 8, 2022. In the modified ITT population (233 patients; 126 BIO101 and 107 placebo), respiratory failure or early death by day 28 was 11.4% lower in the BIO101 (13.5%) than in the placebo (24.3%) group, (p = 0.0426). At day 28, proportions of patients discharged following recovery were 80.1%, and 70.9% in the BIO101 and placebo group respectively, (adjusted difference 11.0%, 95% CI [-0.4%, 22.4%], p = 0.0586). Hazard Ratio for time to death over 90 days: 0.554 (95% CI [0.285, 1.077]), a 44.6% mortality reduction in the BIO101 group (not statistically significant). Treatment emergent adverse events of respiratory failure were more frequent in the placebo group. INTERPRETATION: BIO101 significantly reduced the risk of death or respiratory failure supporting its use in adults hospitalized with severe respiratory symptoms due to COVID-19. FUNDING: Biophytis.

Developing drugs for brain infection by Naegleria fowleri is an unmet medical need. We used a combination of cheminformatics, target-, and phenotypic-based drug discovery methods to identify inhibitors that target an essential N. fowleri enzyme, sterol 14-demethylase (NfCYP51). A total of 124 compounds preselected in silico were tested against N. fowleri. Nine primary hits with EC50 ≤ 10 μM were phenotypically identified. Cocrystallization with NfCYP51 focused attention on one primary hit, miconazole-like compound 2a. The S-enantiomer of 2a produced a 1.74 Å cocrystal structure. A set of analogues was then synthesized and evaluated to confirm the superiority of the S-configuration over the R-configuration and the advantage of an ether linkage over an ester linkage. The two compounds, S-8b and S-9b, had an improved EC50 and KD compared to 2a. Importantly, both were readily taken up into the brain. The brain-to-plasma distribution coefficient of S-9b was 1.02 ± 0.12, suggesting further evaluation as a lead for primary amoebic meningoencephalitis.

O objetivo deste estudo foi descrever a morbidade e a mortalidade hospitalar das neoplasias malignas de lábio, cavidade oral e faringe no Brasil (período de 2016 a 2021). Trata-se de um estudo epidemiológico, observacional, analítico e transversal de notificações de neoplasias malignas, utilizando informações do DATASUS (Base de Dados do Serviço Público de Saúde Brasileiro) e analisadas por meio de estatística descritiva. Foram analisadas 150.025 notificações de neoplasias malignas de lábio, cavidade oral e faringe no Brasil, de 2016 a 2021. Em 2016, a taxa nacional de neoplasias malignas de lábio, cavidade oral e faringe foi de 12,43 por 100 mil habitantes, diminuindo para 10,25 em 2021. Entre as regiões do país, as maiores taxas foram encontradas na região Sul, variando de 19,35 em 2016 a 16,40 em 2021. Quanto às características das internações, a maior incidência foi notada para o sexo masculino (73,76 %), a faixa etária estava entre 50 e 64 anos (45,56%) e a raça/cor branca (43%). Considerando os pacientes internados no período, faleceram 18.185, representando letalidade de 12,12%. No Brasil, a incidência de câncer bucal no período analisado diferiu nas diferentes regiões. Embora o número de internações por neoplasias malignas de lábio, cavidade oral e faringe continue elevado, houve diminuição no período analisado, com tendência de redução das internações por câncer bucal, enfatizando cada vez mais a necessidade de atuação precoce, tais como prevenção, diagnóstico e tratamento por profissionais da Odontologia, além de políticas públicas para prevenção dessa doença.

Abstract Background SARS-CoV-2 belongs to the coronaviridae family and infects human cells by directly interacting with the angiotensin-converting enzyme-2 (ACE-2) through the viral Spike Protein (SP). While vaccines are crucial, much attention has been directed towards managing the symptoms of acute respiratory distress syndrome. Our present study highlights the potential in counteracting lung inflammation triggered by SARS-CoV-2 SP of the intranasal administration of the engineered probiotic Lactobacillus paracasei F19 expressing the enzyme NAPE-PLD (pNAPE-LP) able to in situ release palmitoylethanolamide (PEA) under a super-low boost of palmitate. Methods C57BL/6J mice undergo prophylactic treatment with intranasal pNAPE-LP/palmitate for 7 days before a 7 days challenge with intranasal SARS-CoV-2 SP. Then the capability of pNAPE-LP of colonizing the lungs and actively release PEA in situ have been determined by immunofluorescence, western blot and HPLC-MS. Moreover, the innate immune system downregulation and the histological damage rescue exerted by pNAPE-LP have been tested by immunofluorescence, hematoxylin and eosin staining, western blot analysis and ELISA test for the release of the pro-inflammatory mediators. Results pNAPE-LP effectively colonizes mice lungs and releases the anti-inflammatory compound PEA. Moreover, pNAPE-LP exhibits a protective effect on alveolar morphology, innate immune cells infiltration and in the reduction of neutrophil count, effectively reducing lung injury induced by SARS-CoV-2 SP. This is achieved by mitigating TLR4-mediated NLRP3 activation and the downstream pro-inflammatory products such as ILs, TNFα, C-reactive protein and the myeloperoxidase activity. Interestingly we observed a global reduction ACE2 expression in the lungs. Conclusion pNAPE-LP actively protect from severe inflammatory-related symptoms in SP-challenged mice. Also, it can downregulate the expression of ACE-2 receptors at the lung site potentially preventing the spreading of the infection.

Nonlinear extensions of classical Maxwell's electromagnetism are among the prominent candidates for theories admitting regular black hole solutions. A quest for such examples has been fruitful, but mostly unsystematic and littered by the introduction of physically unrealistic Lagrangians. We provide a procedure which admits the reconstruction of a nonlinear electromagnetic Lagrangian, consistent with the Euler–Heisenberg Lagrangian in the weak-field limit, from a given metric representing a regular, magnetically charged black hole.

While current anti-Spike protein (SP) vaccines have been pivotal in managing the pandemic, their limitations in delivery, storage, and the inability to provide mucosal immunization (preventing infections) highlight the ongoing necessity for research and innovation. To tackle these constraints, our research group developed a bacterial-based vaccine using a non-pathogenic E. coli Nissle 1917 (EcN) strain genetically modified to express the SARS-CoV-2 spike protein on its surface (EcN-pAIDA1-SP). We intranasally delivered the EcN-pAIDA1-SP in two doses and checked specific IgG/IgA production as well as the key immune mediators involved in the process. Moreover, following the initial and booster vaccine doses, we exposed both immunized and non-immunized mice to intranasal delivery of SARS-CoV-2 SP to assess the effectiveness of EcN-pAIDA1-SP in protecting lung tissue from the inflammation damage. We observed detectable levels of anti-SARS-CoV-2 spike IgG in serum samples and IgA in bronchoalveolar lavage fluid two weeks after the initial treatment, with peak concentrations in the respective samples on the 35th day. Moreover, immunoglobulins displayed a progressively enhanced avidity index, suggesting a selective binding to the spike protein. Finally, the pre-immunized group displayed a decrease in proinflammatory markers (TLR4, NLRP3, ILs) following SP challenge, compared to the non-immunized groups, along with better preservation of tissue morphology. Our probiotic-based technology provides an effective immunobiotic tool to protect individuals against disease and control infection spread.

Abstract Background Sepsis is characterized by a dysregulated immune response and metabolic alterations, including decreased high-density lipoprotein cholesterol (HDL-C) levels. HDL exhibits beneficial properties, such as lipopolysaccharides (LPS) scavenging, exerting anti-inflammatory effects and providing endothelial protection. We investigated the effects of CER-001, an engineered HDL-mimetic, in a swine model of LPS-induced acute kidney injury (AKI) and a Phase 2a clinical trial, aiming to better understand its molecular basis in systemic inflammation and renal function. Methods We carried out a translational approach to study the effects of HDL administration on sepsis. Sterile systemic inflammation was induced in pigs by LPS infusion. Animals were randomized into LPS (n = 6), CER20 (single dose of CER-001 20 mg/kg; n = 6), and CER20 × 2 (two doses of CER-001 20 mg/kg; n = 6) groups. Survival rate, endothelial dysfunction biomarkers, pro-inflammatory mediators, LPS, and apolipoprotein A-I (ApoA-I) levels were assessed. Renal and liver histology and biochemistry were analyzed. Subsequently, we performed an open-label, randomized, dose-ranging (Phase 2a) study included 20 patients with sepsis due to intra-abdominal infection or urosepsis, randomized into Group A (conventional treatment, n = 5), Group B (CER-001 5 mg/kg BID, n = 5), Group C (CER-001 10 mg/kg BID, n = 5), and Group D (CER-001 20 mg/kg BID, n = 5). Primary outcomes were safety and efficacy in preventing AKI onset and severity; secondary outcomes include changes in inflammatory and endothelial dysfunction markers. Results CER-001 increased median survival, reduced inflammatory mediators, complement activation, and endothelial dysfunction in endotoxemic pigs. It enhanced LPS elimination through the bile and preserved liver and renal parenchyma. In the clinical study, CER-001 was well-tolerated with no serious adverse events related to study treatment. Rapid ApoA-I normalization was associated with enhanced LPS removal and immunomodulation with improvement of clinical outcomes, independently of the type and gravity of the sepsis. CER-001-treated patients had reduced risk for the onset and progression to severe AKI (stage 2 or 3) and, in a subset of critically ill patients, a reduced need for organ support and shorter ICU length of stay. Conclusions CER-001 shows promise as a therapeutic strategy for sepsis management, improving outcomes and mitigating inflammation and organ damage. Trial registration The study was approved by the Agenzia Italiana del Farmaco (AIFA) and by the Local Ethic Committee (N° EUDRACT 2020–004202-60, Protocol CER-001- SEP_AKI_01) and was added to the EU Clinical Trials Register on January 13, 2021.

Metarhizium (Hypocreales: Clavicipitaceae) has a multifunctional life cycle, establishing as a plant endophyte and acting as entomopathogenic fungi. Metarhizium robertsii and Metarhizium brunneum can be associated with coffee plants and provide enhanced protection against a major pest of coffee, the coffee leaf miner (CLM) (Leucoptera coffeella). This association would be an easily deployable biological control option. Here we tested the potential of inoculating coffee seeds with M. robertsii and M. brunneum collected from the soil of coffee crops in the Cerrado (Brazil) for control of the CLM and the enhancement of plant growth with a commonly used fungicide. We conducted the experiment in a greenhouse and after the seedlings grew, we placed them in a cage with two couples of CLMs. We evaluated the CLM development time, reproduction, and plant growth traits. We observed a longer development time of CLMs when fed on plants inoculated with both isolates. In addition, the CLMs laid fewer eggs compared to those fed on plants without fungal inoculation. Plant growth was promoted when seeds were inoculated with fungi, and the fungicide did not affect any evaluated parameter. Coffee seed inoculation with M. robertsii and M. brunneum appears to provide protection against CLMs and promote growth improvement.

O presente trabalho apresenta o recorte de uma pesquisa de dissertação aplicada em uma turma do 9º ano do Ensino Fundamental. Por meio de um questionário com 10 questões, buscou-se averiguar quais habilidades relacionadas ao letramento estatístico os alunos desenvolveram durante os anos que compõem o Ensino Fundamental. As habilidades avaliadas foram selecionadas conforme proposto pela Base Nacional Comum Curricular (BNCC), na disciplina de matemática. A pesquisa adota uma abordagem mista e justifica-se pela necessidade de conhecer quais fragilidades devem ser superadas no processo de ensino-aprendizagem de conceitos estatísticos na Educação Básica. Dessa forma, se favorece a busca contínua pela melhoria por meio da aplicação e adaptação de novas metodologias, atividades e estratégias. Os resultados obtidos indicaram que a maior parte da turma pesquisada desenvolveu habilidades relacionadas à interpretação de gráficos de colunas, gráficos de setores e interpretação de tabelas simples. No entanto, apresentaram dificuldades ao utilizar medidas de tendência central (média e moda), interpretar gráficos de linhas, tabelas de dupla entrada e interpretar enunciados. Como possibilidade para auxiliar na superação das fragilidades identificadas, sugere-se a utilização de uma Unidade de Ensino Potencialmente Significativa (UEPS), que inclua a elaboração e execução de uma pesquisa estatística com os alunos, bem como a utilização de planilhas digitais, adotando uma perspectiva interdisciplinar.

Glycopyrrolate is a competitive muscarinic receptor antagonist used in the treatment of sialorrhea, especially in pediatrics. Degradation research was conducted to better understand the stability of the active pharmaceutical ingredient (API). Using an HPLC-UV method, we evaluated the chemical stability of the oral solution of the galenic compound glycopyrrolate 0.5 mg/mL under different storage conditions. Method validation was performed according to the International Council for Harmonization (ICH) Q2(R2) guidelines. The results of the stability study of the galenic compound in different storage conditions, with the exception of those stored in glass containers at 45 °C for more than 3 months, were stable (100 ± 10% of the nominal concentration). The aim of this work was to study the stability of the galenic compound glycopyrrolate in two different types of containers and at three different storage temperatures. Glycopyrrolate showed degradation beyond the limits only in glass at 45 °C and after 2 months of storage. The results indicate that oral liquid dosage forms of glycopyrrolate are stable for at least 210 days when stored at room temperature or at 4 °C, in glass or PET, for at least 7 months, maintaining product quality according to the standards established by the European Pharmacopoeia, ensuring long-term coverage for pediatric patient therapies.

In forensic investigation, determining the time and cause of death becomes challenging, especially in cases where the remains are found in advanced decomposition, rendering traditional toxicological samples unavailable or unreliable. Entomotoxicology, an emerging methodology within forensic science, leverages insect specimens collected from cadavers as alternative toxicological samples. Several laboratory and field research studies have highlighted the efficacy in detecting various drugs, toxins, and elements absorbed by insects feeding on cadaveric tissues, even at low concentrations. However, correlation studies between drug concentrations in conventional matrices and insects remain controversial due to unknown factors influencing drug metabolism and larval feeding activity. This paper presents four real cases in which human cadavers were discovered in advanced stages of decomposition, and toxicological analyses were performed on both insect samples and available matrices. The results presented complement the scant literature currently available on the application of entomotoxicology in real cases, providing insights into the correlation between larvae and human specimen results. Furthermore, guidelines to collect and preserve entomological evidence at the crime scene and during the autopsy for use in entomotoxicological analyses are provided. This advancement holds promise in aiding forensic investigations, particularly in cases where traditional methods cannot be applied or require supporting data for further validation.

Bile has emerged as an alternative matrix for toxicological investigation of drugs in suspected forensic cases of overdose in adults and intoxications in children. Toxicological investigation consists in screening and, subsequently, confirming the result with specific techniques, such as liquid chromatography with tandem mass spectrometry (LC-MS/MS). As there is no screening test on the market to test postmortem bile specimens, the novelty of this study was in investigating the applicability of a chemiluminescence immunoassay, designed for other matrices and available on the market, on bile and validate its use, testing the agreement with LC-MS/MS analysis. Bile specimens were obtained from 25 forensic cases of suspected death from overdose and intoxication. Sample preparation for bile screening consists simply in centrifugation and dilution. Confirmation analysis allows simultaneous identification of 108 drugs and was validated on bile. Kappa analysis assessed a perfect agreement (0.81–1) between the assays for benzodiazepines, methadone, opiates, cocaine, oxycodone, cannabinoids, buprenorphine and pregabalin; a substantial agreement (0.41–0.6) was reported for barbiturates. No agreement was assessed for amphetamines, due to an abundance of putrefactive amines in postmortem specimens. In conclusion, this fast and easy immunoassay could be used for initial screening of bile specimens, identifying presence of drugs, except amphetamines, with reliability.

Sorghum plants naturally produce dhurrin, a cyanogenic glycoside that may be hydrolysed to cyanide, resulting in often-lethal toxicoses. Ruminants are particularly sensitive to cyanogenic glycosides due to the active role of rumen microbiota in dhurrin hydrolysis. This work provides an overview of a poisoning outbreak that occurred in 5 farms in Northwest Italy in August 2022; a total of 66 cows died, and many others developed acute toxicosis after being fed on either cultivated (Sorghum bicolor) or wild Sorghum (Sorghum halepense). Clinical signs were recorded, and all cows received antidotal/supportive therapy. Dead animals were subjected to necropsy, and dhurrin content was determined in Sorghum specimens using an LC–MS/MS method. Rapid onset, severe respiratory distress, recumbency and convulsions were the main clinical features; bright red blood, a bitter almond smell and lung emphysema were consistently observed on necropsy. The combined i.v. and oral administration of sodium thiosulphate resulted in a rapid improvement of clinical signs. Dhurrin concentrations corresponding to cyanide levels higher than the tolerated threshold of 200 mg/kg were detected in sorghum specimens from 4 out of 5 involved farms; thereafter, such levels declined, reaching tolerable concentrations in September–October. Feeding cattle with wild or cultivated Sorghum as green fodder is a common practice in Northern Italy, especially in summer. However, care should be taken in case of adverse climatic conditions, such as severe drought and tropical temperatures (characterising summer 2022), which are reported to increase dhurrin synthesis and storage.

Acyclovir and ganciclovir comprise the prophylaxis and treatment of herpesvirus and cytomegalovirus infections occurring in immunocompromised patients. Their therapeutic drug monitoring is fundamental because of interindividual variability leading to side effects and drug resistance and is performed through several techniques, such as liquid chromatography coupled with UV spectrophotometry (HPLC-UV) or mass spectrometry (LC-MS/MS). Therefore, we developed and validated a low-cost, non-time-consuming, and low-sample-consuming HPLC-UV method. Briefly, 100 µL of sample was used for sample preparation, mainly consisting of precipitation through organic solvent. In total, 20 µL was injected into the instrument. Chromatographic separation was obtained eluting mobile phases A (10 mM ammonium formiate 0.01% formic acid) and B (acetonitrile) on a Poroshell 120 SB-C8 2.1 × 150 mm, 2.7 µm for 12 min isocratically (97:3; A:B) at a flow rate of 0.2 mL/min. The linearity range (0.5–40 mg/L) of the method allowed us to quantify both the Cmin and Cmax of acyclovir and ganciclovir. Plasma concentrations measured on a small cohort of patients undergoing acyclovir (31) and ganciclovir (9) treatment by the proposed method and the LC-MS/MS methods, already in use, were significantly correlated. The proposed HPLC-UV method may be implemented in diagnostics as an alternative method in case of the unavailability of the LC-MS/MS system.

Abstract Background To date, measurement of intracellular cystine is used for the therapeutic monitoring of patients affected by cystinosis in treatment with cysteamine. Since this method is time and sample consuming, development of a faster method to quantify cysteamine would be extremely useful in order to help clinicians to adjust dosages of cysteamine and to define better the pharmacokinetic profile of this drug. The aim of the study was to develop a liquid chromatography tandem mass spectrometry method for the quantification of cysteamine in plasma samples and to test its applicability on plasma samples derived from patients with nephropathic infantile cystinosis in treatment with cysteamine. Results The percentage of accuracy of the developed method varied between 97.80 and 106.00% and CV% between 0.90 and 6.93%. There was no carry over. The calibration curves were built from 2.5 to 50 µM. The limit of detection and the lower limit of quantification occurred at 0.25 and 1.25 µM respectively. Cysteamine was stable up to 2 months at -20 °C. Concentrations of cysteamine and intracellular cystine of 4 patients were in line with data previously reported. Conclusion The proposed method showed an appropriate selectivity, specificity, linearity, sensibility, accuracy, precision and good applicability to samples.

Introduction: Melanomas are malignant neoplasms that occur in various anatomical sites, including the eye. Ocular melanomas account for 5% of all melanomas and are mainly described in Caucasian and older individuals. This study describes the clinical and pathological characteristics of uveal (choroid) melanoma in a Caucasian patient. Case report: A 41-year-old Caucasian female patient, brown eyes, without history of ophthalmological diseases or family history of cancer experienced pain and loss of visual acuity in the left eye. On clinical examination, an increase of ocular pressure was detected. Ultrasound showed a mushroom-like neoformation. Moreover, magnetic resonance imaging showed a mass with spontaneous hypersignal on T1-weighted images, intense gadolinium enhancement, and marked hyposignal on T2-weighted images. The patient was referred to the Oncology Ophthalmology department for enucleation due to suspected uveal melanoma. Anatomopathological analysis revealed a blackened mass in the eyeball. Histologically, the mass comprised spindle cells (50%) and epithelioid cells (50%). A diagnosis of choroidal melanoma was established based on the identification of ophthalmoscopic, imaging, and histological characteristics of the tumor. Conclusion: Choroidal melanomas usually occur in males, clear-eyed, and older individuals. A wide variety of ocular lesions may mimic choroidal melanoma, which should be included in the differential diagnosis of choroidal nevus and peripheral hemorrhages.