Ehrlich tumor induces TRPV1-dependent evoked and non-evoked pain-like behavior in mice

We standardized a model by injecting Ehrlich tumor cells into the paw to evaluate cancer pain mechanisms and pharmacological treatments. Opioid treatment, but not cyclooxygenase inhibitor or tricyclic antidepressant treatments reduces Ehrlich tumor pain. To best use this model for drug screening it is essential to understand its pathophysiological mechanisms. Herein, we investigated the contribution of the transient receptor potential cation channel subfamily V member 1 (TRPV1) in the Ehrlich tumor-induced pain model. Dorsal root ganglia (DRG) neurons from the Ehrlich tumor mice presented higher activity (calcium levels using fluo-4 fluorescent probe) and an increased response to capsaicin (TRPV1 agonist) than the saline-injected animals

Renal function evaluation in patients with American Cutaneous Leishmaniasis after specific treatment with pentavalent antimonial

Background\ud Renal evaluation studies are rare in American Cutaneous Leishmaniasis (ACL). The aim of this study is to investigate whether specific treatment reverts ACL-associated renal dysfunction.\ud \ud Methods\ud A prospective study was conducted with 37 patients with ACL. Urinary concentrating and acidification ability was assessed before and after treatment with pentavalent antimonial.\ud \ud Results\ud The patients mean age was 35.6 ± 12 years and 19 were male. Before treatment, urinary concentrating defect (U/Posm <2.8) was identified in 27 patients (77%) and urinary acidification defect in 17 patients (46%). No significant glomerular dysfunction was observed before and after specific ACL treatment. There was no reversion of urinary concentrating defects, being observed in 77% of the patients before and in 88% after treatment (p = 0.344). Urinary acidification defect was corrected in 9 patients after treatment, reducing its prevalence from 40% before to only 16% after treament, (p = 0.012). Microalbuminuria higher than 30 mg/g was found in 35% of patients before treatment and in only 8% after treatment. Regarding fractional excretion of sodium, potassium, calcium, phosphorus and magnesium, there was no significant difference between pre and post-treatment period.\ud \ud Conclusion\ud As previously described, urinary concentrating and acidification defects were found in an important number of patients with ACL. Present results demonstrate that only some patients recover urinary acidification capacity, while no one returned to normal urinary concentration capacity.This research was supported by the Brazilian National Council for Scientific and Technological Development (CNPq). The desmopressin acetate was provided by Ferring of Brazil

A“Dirty” Footprint: Macroinvertebrate diversity in Amazonian Anthropic Soils

International audienceAmazonian rainforests, once thought to be pristine wilderness, are increasingly known to have been widely inhabited, modified, and managed prior to European arrival, by human populations with diverse cultural backgrounds. Amazonian Dark Earths (ADEs) are fertile soils found throughout the Amazon Basin, created by pre-Columbian societies with sedentary habits. Much is known about the chemistry of these soils, yet their zoology has been neglected. Hence, we characterized soil fertility, macroinvertebrate communities, and their activity at nine archeological sites in three Amazonian regions in ADEs and adjacent reference soils under native forest (young and old) and agricultural systems. We found 673 morphospecies and, despite similar richness in ADEs (385 spp.) and reference soils (399 spp.), we identified a tenacious pre-Columbian footprint, with 49% of morphospecies found exclusively in ADEs. Termite and total macroinvertebrate abundance were higher in reference soils, while soil fertility and macroinvertebrate activity were higher in the ADEs, and associated with larger earthworm quantities and biomass. We show that ADE habitats have a unique pool of species, but that modern land use of ADEs decreases their populations, diversity, and contributions to soil functioning. These findings support the idea that humans created and sustained high-fertility ecosystems that persist today, altering biodiversity patterns in Amazonia

Safety and efficacy of the ChAdOx1 nCoV-19 vaccine (AZD1222) against SARS-CoV-2: an interim analysis of four randomised controlled trials in Brazil, South Africa, and the UK.

BACKGROUND: A safe and efficacious vaccine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), if deployed with high coverage, could contribute to the control of the COVID-19 pandemic. We evaluated the safety and efficacy of the ChAdOx1 nCoV-19 vaccine in a pooled interim analysis of four trials. METHODS: This analysis includes data from four ongoing blinded, randomised, controlled trials done across the UK, Brazil, and South Africa. Participants aged 18 years and older were randomly assigned (1:1) to ChAdOx1 nCoV-19 vaccine or control (meningococcal group A, C, W, and Y conjugate vaccine or saline). Participants in the ChAdOx1 nCoV-19 group received two doses containing 5 × 1010 viral particles (standard dose; SD/SD cohort); a subset in the UK trial received a half dose as their first dose (low dose) and a standard dose as their second dose (LD/SD cohort). The primary efficacy analysis included symptomatic COVID-19 in seronegative participants with a nucleic acid amplification test-positive swab more than 14 days after a second dose of vaccine. Participants were analysed according to treatment received, with data cutoff on Nov 4, 2020. Vaccine efficacy was calculated as 1 - relative risk derived from a robust Poisson regression model adjusted for age. Studies are registered at ISRCTN89951424 and ClinicalTrials.gov, NCT04324606, NCT04400838, and NCT04444674. FINDINGS: Between April 23 and Nov 4, 2020, 23 848 participants were enrolled and 11 636 participants (7548 in the UK, 4088 in Brazil) were included in the interim primary efficacy analysis. In participants who received two standard doses, vaccine efficacy was 62·1% (95% CI 41·0-75·7; 27 [0·6%] of 4440 in the ChAdOx1 nCoV-19 group vs71 [1·6%] of 4455 in the control group) and in participants who received a low dose followed by a standard dose, efficacy was 90·0% (67·4-97·0; three [0·2%] of 1367 vs 30 [2·2%] of 1374; pinteraction=0·010). Overall vaccine efficacy across both groups was 70·4% (95·8% CI 54·8-80·6; 30 [0·5%] of 5807 vs 101 [1·7%] of 5829). From 21 days after the first dose, there were ten cases hospitalised for COVID-19, all in the control arm; two were classified as severe COVID-19, including one death. There were 74 341 person-months of safety follow-up (median 3·4 months, IQR 1·3-4·8): 175 severe adverse events occurred in 168 participants, 84 events in the ChAdOx1 nCoV-19 group and 91 in the control group. Three events were classified as possibly related to a vaccine: one in the ChAdOx1 nCoV-19 group, one in the control group, and one in a participant who remains masked to group allocation. INTERPRETATION: ChAdOx1 nCoV-19 has an acceptable safety profile and has been found to be efficacious against symptomatic COVID-19 in this interim analysis of ongoing clinical trials. FUNDING: UK Research and Innovation, National Institutes for Health Research (NIHR), Coalition for Epidemic Preparedness Innovations, Bill & Melinda Gates Foundation, Lemann Foundation, Rede D'Or, Brava and Telles Foundation, NIHR Oxford Biomedical Research Centre, Thames Valley and South Midland's NIHR Clinical Research Network, and AstraZeneca

Safety and efficacy of the ChAdOx1 nCoV-19 vaccine (AZD1222) against SARS-CoV-2: an interim analysis of four randomised controlled trials in Brazil, South Africa, and the UK

Background A safe and efficacious vaccine against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), if deployed with high coverage, could contribute to the control of the COVID-19 pandemic. We evaluated the safety and efficacy of the ChAdOx1 nCoV-19 vaccine in a pooled interim analysis of four trials. Methods This analysis includes data from four ongoing blinded, randomised, controlled trials done across the UK, Brazil, and South Africa. Participants aged 18 years and older were randomly assigned (1:1) to ChAdOx1 nCoV-19 vaccine or control (meningococcal group A, C, W, and Y conjugate vaccine or saline). Participants in the ChAdOx1 nCoV-19 group received two doses containing 5 × 1010 viral particles (standard dose; SD/SD cohort); a subset in the UK trial received a half dose as their first dose (low dose) and a standard dose as their second dose (LD/SD cohort). The primary efficacy analysis included symptomatic COVID-19 in seronegative participants with a nucleic acid amplification test-positive swab more than 14 days after a second dose of vaccine. Participants were analysed according to treatment received, with data cutoff on Nov 4, 2020. Vaccine efficacy was calculated as 1 - relative risk derived from a robust Poisson regression model adjusted for age. Studies are registered at ISRCTN89951424 and ClinicalTrials.gov, NCT04324606, NCT04400838, and NCT04444674. Findings Between April 23 and Nov 4, 2020, 23 848 participants were enrolled and 11 636 participants (7548 in the UK, 4088 in Brazil) were included in the interim primary efficacy analysis. In participants who received two standard doses, vaccine efficacy was 62·1% (95% CI 41·0–75·7; 27 [0·6%] of 4440 in the ChAdOx1 nCoV-19 group vs71 [1·6%] of 4455 in the control group) and in participants who received a low dose followed by a standard dose, efficacy was 90·0% (67·4–97·0; three [0·2%] of 1367 vs 30 [2·2%] of 1374; pinteraction=0·010). Overall vaccine efficacy across both groups was 70·4% (95·8% CI 54·8–80·6; 30 [0·5%] of 5807 vs 101 [1·7%] of 5829). From 21 days after the first dose, there were ten cases hospitalised for COVID-19, all in the control arm; two were classified as severe COVID-19, including one death. There were 74 341 person-months of safety follow-up (median 3·4 months, IQR 1·3–4·8): 175 severe adverse events occurred in 168 participants, 84 events in the ChAdOx1 nCoV-19 group and 91 in the control group. Three events were classified as possibly related to a vaccine: one in the ChAdOx1 nCoV-19 group, one in the control group, and one in a participant who remains masked to group allocation. Interpretation ChAdOx1 nCoV-19 has an acceptable safety profile and has been found to be efficacious against symptomatic COVID-19 in this interim analysis of ongoing clinical trials

Repurposing of the Nootropic Drug Vinpocetine as an Analgesic and Anti-Inflammatory Agent: Evidence in a Mouse Model of Superoxide Anion-Triggered Inflammation

Clinically active drugs for the treatment of acute pain have their prescription limited due to the significant side effects they induce. An increase in reactive oxygen species (ROS) has been linked to several conditions, including inflammation and pain processing. Therefore, new or repurposed drugs with the ability of reducing ROS-triggered responses are promising candidates for analgesic drugs. Vinpocetine is a clinically used nootropic drug with antioxidant, anti-inflammatory, and analgesic properties. However, the effects of vinpocetine have not been investigated in a model with a direct relationship between ROS, inflammation, and pain. Based on that, we aimed to investigate the effects of vinpocetine in a model of superoxide anion-induced pain and inflammation using potassium superoxide (KO2) as a superoxide anion donor to trigger inflammation and pain. In the KO2 model, vinpocetine dose-dependently reduced pain-like behaviors (spontaneous pain and hyperalgesia), paw edema, and neutrophil and mononuclear cell recruitment to the paw skin (assessed by H&E staining, fluorescence, and enzymatic assays) and to the peritoneal cavity. Vinpocetine also restored tissue endogenous antioxidant ability and Nrf2 and Ho-1 mRNA expression and reduced superoxide anion production and gp91phox mRNA expression. We also observed the inhibition of IκBα degradation by vinpocetine, which demonstrates a reduction in the activation of NF-κB explaining the diminished production of IL-33, IL-1β, and TNF-α. Collectively, our data show that vinpocetine alleviates pain and inflammation induced by KO2, which is a mouse model with a direct role of ROS in triggering pain and other inflammatory phenomena. Thus, the results suggest the repurposing of vinpocetine as an anti-inflammatory and analgesic drug

Inhibition of caspase-1 or gasdermin-D enable caspase-8 activation in the Naip5/NLRC4/ASC inflammasome

<div><p><i>Legionella pneumophila</i> is a Gram-negative, flagellated bacterium that survives in phagocytes and causes Legionnaires’ disease. Upon infection of mammalian macrophages, cytosolic flagellin triggers the activation of Naip/NLRC4 inflammasome, which culminates in pyroptosis and restriction of bacterial replication. Although NLRC4 and caspase-1 participate in the same inflammasome, <i>Nlrc4</i>^<i>-/-</i> mice and their macrophages are more permissive to <i>L</i>. <i>pneumophila</i> replication compared with <i>Casp1/11</i>^<i>-/-</i>. This feature supports the existence of a pathway that is NLRC4-dependent and caspase-1/11-independent. Here, we demonstrate that caspase-8 is recruited to the Naip5/NLRC4/ASC inflammasome in response to flagellin-positive bacteria. Accordingly, caspase-8 is activated in <i>Casp1/11</i>^<i>-/-</i> macrophages in a process dependent on flagellin, Naip5, NLRC4 and ASC. Silencing caspase-8 in <i>Casp1/11</i>^<i>-/-</i> cells culminated in macrophages that were as susceptible as <i>Nlrc4</i>^<i>-/-</i> for the restriction of <i>L</i>. <i>pneumophila</i> replication. Accordingly, macrophages and mice deficient in <i>Asc/Casp1/11</i>^<i>-/-</i> were more susceptible than <i>Casp1/11</i>^<i>-/-</i> and as susceptible as <i>Nlrc4</i>^<i>-/-</i> for the restriction of infection. Mechanistically, we found that caspase-8 activation triggers gasdermin-D-independent pore formation and cell death. Interestingly, caspase-8 is recruited to the Naip5/NLRC4/ASC inflammasome in wild-type macrophages, but it is only activated when caspase-1 or gasdermin-D is inhibited. Our data suggest that caspase-8 activation in the Naip5/NLRC4/ASC inflammasome enable induction of cell death when caspase-1 or gasdermin-D is suppressed.</p></div

<i>Legionella</i> triggers the formation of NLRC4/caspase-8 puncta in a process that is dependent on ASC and flagellin and independent of caspase-1/11.

<p>Bone marrow-derived macrophages (BMDMs) obtained from <i>Casp1/11</i>^<i>-/-</i> and <i>Asc/Casp1/11</i>^<i>-/-</i> mice were transduced with retrovirus encoding NLRC4-GFP. Cells were infected with wild-type <i>L</i>. <i>pneumophila</i> (WT Lp), motility-deficient mutants expressing flagellin (<i>fliI</i>^-) or with flagellin-deficient mutants (<i>flaA</i>^<i>-</i>) at a MOI of 1, 3 or 10. (A) After 2, 4 and 8 hours of infection, the cells were fixed, and the percentage of transduced cells containing NLRC4-GFP puncta were determined using an epifluorescence microscope. <i>*</i>, <i>P</i><0.05 compared with BMDMs infected with <i>flaA</i>^<i>-</i>, Student’s <i>t</i> test. (B-D) Transduced cells were infected with <i>fliI</i>^<i>-</i> (MOI 10) and fixed after 8 hours of infection. (B) The percentages of transduced <i>Casp1/11</i>^<i>-/-</i> and <i>Asc/Casp1/11</i>^<i>-/-</i> cells containing NLRC4-GFP puncta were determined. (C) The percentages of NLRC4-GFP puncta that colocalized with ASC and caspase-8 were determined. (D) Representative images of a transduced BMDM infected with <i>fliI</i>^- at a MOI of 10. The cultures were stained with anti-caspase-8 (red) and anti-ASC (purple). The cell nuclei were stained with DAPI (cyan); NLRC4-GFP is shown in green. The images show the colocalization of NLRC4-GFP, ASC and caspase-8 in <i>Casp1/11</i>^<i>-/-</i> BMDMs. The images were acquired by multiphoton microscopy using a 63x oil immersion objective and analyzed using ImageJ Software. Scale bar, 10μm. Data show the average ± SD of triplicate wells. NI, uninfected. Data are presented for one representative experiment of five (A) and two (B-D) experiments with similar results.</p

ASC is important for NLRC4/caspase-8-mediated restriction <i>of L</i>. <i>pneumophila</i> replication independently of caspase-1/11.

<p>(A-D) Bone marrow-derived macrophages (BMDMs) from C57BL/6 (open circles), <i>Nlrc4</i>^<i>-/-</i> (closed squares), <i>Casp1/11</i>^<i>-/-</i> (closed triangles) and <i>Asc/Casp1/11</i>^<i>-/-</i> (open triangles) mice were infected with motility-deficient mutants expressing flagellin (<i>fliI</i>^-, A), with flagellin-deficient mutants (<i>flaA</i>^<i>-</i>, B), <i>L</i>. <i>gratiana</i> (C) or with <i>L</i>. <i>micdadei</i> (D) at a MOI of 10. The cells were incubated for 24, 48 and 72 hours for CFU determination. Data show the average ± SD of triplicate wells. <i>*</i>, <i>P</i><0.05 compared with <i>Casp-1/11</i>^<i>-/-</i> BMDMs. ^#, <i>P</i><0.05 compared with C57BL/6 BMDMs, ANOVA. (E-I) BMDMs from <i>Casp1/11</i>^<i>-/-</i> and <i>Asc/Casp1/11</i>^<i>-/-</i> mice were transduced with a retrovirus encoding shRNA sequences to target caspase-8 (Seq1, Seq2) and a non-target shRNA sequence (NT). (E) Caspase-8 silencing was confirmed by western blot analysis. Cell lysates were separated by SDS-PAGE, blotted and probed with anti-caspase-8 (pro-caspase-8 p55) and anti-α-actin. (F-I) Transduced <i>Casp1/11</i>^<i>-/-</i> (F, H) and <i>Asc/Casp1/11</i>^<i>-/-</i> (G, I) BMDMs were infected with <i>fliI</i>^<i>-</i> (F, G) or <i>flaA</i>^<i>-</i> (H, I) at a MOI of 10 and incubated for 24, 48 and 72 hours for CFU determination. Data show the average ± SD of triplicate wells. <i>*</i>, <i>P</i><0.05 compared with NT shRNA, ANOVA. NT, non-target shRNA. Data are presented for one representative experiment of four (A), two (B-D) and one (F-I) experiments performed with similar results.</p