Insights of potential trypanocidal effect of the synthetic derivative (2E)-1-(4-aminophenyl)-3-(2,4-dichlorophenyl)prop-2-en-1-one: in vitro assay, MEV analysis, quantum study, molecular docking, molecular dynamics, MPO analysis, and predictive ADMET.

This study aims to evaluate the antitrypanosomiasis activity of a synthetic dichloro-substituted aminochalcone via in vitro assays against infected cell cultures, as well as a theoretical characterization of pharmacokinetics and pharmacodynamics against the protein targets of the evolutionary cycle of T. cruzi. The in vitro evaluation of parasite proliferation inhibition was performed via cytotoxicity analysis on mammalian host cells, effect on epimastigote and trypomastigote forms, and cell death analysis, while computer simulations characterized the electronic structure of (2E)-1-(4-aminophenyl)-3-(2,4-dichlorophenyl)prop-2-en-1-one (DCl), the mechanism of action against the proteins of the evolutionary cycle of T. cruzi: Cruzain, Trypanothione reductase, TcGAPDH, and CYP51 by molecular docking and dynamics and predictive pharmacokinetics by MPO-based ADMET. The in vitro tests showed that the DCl LC50 in order of 178.9 ± 23.9 was similar to the BZN, evidencing the effectiveness of chalcone against Trypomastigotes. Molecular docking and dynamics simulations suggest that DCl acts on the active site of the CYP51 receptor, with hydrogen interactions that showed a high degree of occupation, establishing a stable complex with the target. MPO analysis and ADMET prediction tests suggest that the compound presents an alignment between permeability and hepatic clearance, although it presents low metabolic stability. Chalcone showed stable pharmacodynamics against the CYP51 target, but can form reactive metabolites from N-conjugation and C = C epoxidation, as an indication of controlled oral dose, although the estimated LD50 rate > 500 mg/kg is a indicative of low incidence of lethality by ingestion, constituting a promising therapeutic strategy.

Elongation on aliphatic chain improves selectivity of 2-hydroxy-3,4,6-trimethoxyphenyl chalcone on Trypanosoma cruzi.

Aim: Our objective was to investigate the trypanocidal effect of the chalcone (2E,4E)-1-(2-hydroxy-3,4,6-trimethoxyphenyl)-5-phenylpenta-2,4-dien-1-one (CPNC). Material & methods: Cytotoxicity toward LLC-MK2 host cells was assessed by MTT assay, and the effect on Trypanosoma cruzi life forms (epimastigotes, trypomastigotes and amastigotes) was evaluated by counting. Flow cytometry analysis was performed to evaluate the possible mechanisms of action. Finally, molecular docking simulations were performed to evaluate interactions between CPNC and T. cruzi enzymes. Results: CPNC showed activity against epimastigote, trypomastigote and amastigote life forms, induced membrane damage, increased cytoplasmic reactive oxygen species and mitochondrial dysfunction on T. cruzi. Regarding molecular docking, CPNC interacted with both trypanothione reductase and TcCr enzymes. Conclusion: CPNC presented a trypanocidal effect, and its effect is related to oxidative stress, mitochondrial impairment and necrosis.

Effect from dinoponeratoxin M-PONTXDq3a arginine and lysine substituted analogues against Staphylococcus aureus strains.

The growing incidence of methicillin-resistant Staphylococcus aureus (MRSA) infections is associated with increased mortality rates, which has generated interest in the development of antimicrobial peptides (AMP), such as those found in the giant ant Dinoponera quadríceps. In order to improve the net positive charge and the antibacterial activity of the AMP, amino acids with positive side chain single substituted analogues have been proposed, mainly arginine or lysine. The present work aims to study the antimicrobial activity of the analogues of M-PONTX-Dq3a, a 23 amino acid AMP identified in the D. quadriceps venom. M-PONTX-Dq3a[1-15], a fragment containing the 15 central amino acids, and eight derivatives of single arginine or lysine substituted analogues were proposed. The antimicrobial activity of peptides was evaluated against Staphylococcus aureus ATCC 6538 P (MSSA) and ATCC 33591 (MRSA) strains, followed by minimum inhibitory concentration (MIC), minimum lethal concentration (MLC), and minimum biofilm inhibitory concentration (MBIC) measurement. The membrane permeability was then assessed via crystal violet assay and flow cytometry analysis. The effect of exposure time on microbial viability (Time-Kill) was evaluated. Finally, ultrastructural alterations were evaluated through scanning electron microscopy (SEM). Both arginine-substituted peptides [Arg]3M-PONTX-Dq3a[1-15] and [Arg]4M-PONTX-Dq3a[1-15], showed lowest MIC and MLC values (each 0.78 µM). In the biofilm formation assays, the peptide [Arg]3M-PONTX-Dq3a [1-15] showed MBIC of 3.12 µM against the two tested strains. Both peptides were able to alter the membrane permeability approximately by 80%. The treatment with MIC was able to eliminate bacteria after 2 h of contact on the other hand, treatment with half of the MIC, the population of both bacterial strains remained constant for up to 12 h, indicating a possible bacteriostatic effect. The SEM results showed that the treatment with the lowest concentration (0.78 µM) of both peptides caused disruption of the cell membrane, destabilization of the intercellular interaction and the CLM of [Arg]4M-PONTX-Dq3a [1-15] resulted in the complete eradication of the bacteria. Thus, this study describes two AMPs active against MSSA and MRSA and also inhibits the biofilm formation of these stains. This study finds [Arg]3M-PONTX-Dq3a[1-15] and [Arg]4M-PONTX-Dq3a[1-15] as alternative substances to treat resistant and/or biofilm-forming strains.

GABAA and serotonergic receptors participation in anxiolytic effect of chalcones in adult zebrafish.

The prevalence of anxiety is a significant public health problem, being the 24th leading cause of disability in individuals affected by this disorder. In this context, chalcones, a flavonoid subclass obtained from natural or synthetic sources, interact with central nervous system (CNS) receptors at the same binding site as benzodiazepines, the primary drugs used in the treatment of anxiety. Thus, our study investigates the anxiolytic effect of synthetic chalcones derived from the natural product 2-hydroxy-3,4,6-trimethoxyacetophenone isolated from Croton anisodontus Müll.Arg. in modulating anxiolytic activity via GABAergic and serotoninergic neurotransmission in an adult zebrafish model. Chalcones 1 and 2 were non-toxic to adult zebrafish and showed anxiolytic activity via GABAA receptors. Chalcone 2 also had its anxiolytic action reversed by the antagonist granisetron, indicating the participation of serotonergic receptors 5HTR3A/3B in the anxiolytic effect. In addition, molecular docking results showed that chalcones have a higher affinity for the GABAA receptor than DZP and binding in the same region of the DZP binding site, indicating a similar effect to the drug. Furthermore, the interaction of chalcones with GABAA and 5-HT3A receptors demonstrates the anxiolytic effect potential of these molecules.Communicated by Ramaswamy H. Sarma.

Vitamin D3 actions on astrocyte cells: A target for therapeutic strategy in Parkinson's disease?

Parkinson's disease (PD) is a neurodegenerative disease characterized by the loss of dopaminergic cells in the substantia nigra pars compacta. PD patients' brains show neuroinflammation, oxidative stress, and mitochondrial dysfunction. The present study aims to evaluate the neuroprotective activity of VD3 on astrocytes after their exposure to rotenone (ROT) a natural pesticide known to exhibit neurotoxic potential via the inhibition of mitochondrial complex I. Cell viability parameters were evaluated by the MTT test and staining with 7-AAD in cultures of astrocytes treated and untreated with VD3 (0.1, 0.5, and 1.0 ng/mL) and/or ROT (10 µg/mL or 5 µg/mL), and the cytoplasmic production of ROS and the cell death profile were measured by flow cytometry. Glutathione accumulation and ultrastructural changes were evaluated and immunocytochemistry assays for NF-kB and Nrf2 were also carried out. The results showed that VD3 improved the viability of cells previously treated with VD3 and then exposed to ROT, reducing the occurrence of necrotic and apoptotic events. Furthermore, cells exposed to ROT showed increased production of ROS, which decreased significantly with previous treatment with VD3. Importantly, the decrease by ROT in the mitochondrial transmembrane potential was significantly prevented after treating cells with VD3, especially at a concentration of 1 ng/mL. Therefore, treatment with VD3 protected astrocytes from damage caused by ROT, decreasing oxidative stress, decreasing NF-kB and Nrf2 expressions, and improving mitochondrial function. However, further investigation is needed regarding the participation and mechanism of action of VD3 in this cellular model of PD focusing on the crosstalk between Nrf2 and NF-kB.

1α,25-Dihydroxyvitamin D3 (VD3) Shows a Neuroprotective Action Against Rotenone Toxicity on PC12 Cells: An In Vitro Model of Parkinson's Disease.

Parkinson's disease (PD) is characterized by dopaminergic cell loss in the substantia nigra, and PD brains show neuroinflammation, oxidative stress, and mitochondrial dysfunction. The study evaluated the neuroprotective activity of 1α,25-dihydroxy vitamin D3 (VD3), on the rotenone (ROT)-induced cytotoxicity in PC12 cells. The viability parameters were assessed by the MTT and flow cytometry, on cells treated or not with VD3 and/or ROT. Besides, ROS production, cell death, mitochondrial transmembrane potential, reduced GSH, superoxide accumulation, molecular docking (TH and Keap1-Nrf2), and TH, Nrf2, NF-kB, and VD3 receptor protein contents by western blot were evaluated. VD3 was shown to improve the viability of ROT-exposed cells. Cells exposed to ROT showed increased production of ROS and superoxide, which decreased after VD3. ROT decrease in the mitochondrial transmembrane potential was prevented, after VD3 treatment and, VD3 was shown to interact with tyrosine hydroxylase (TH) and Nrf2. While ROT decreased TH, Nrf2, and NF-kB expressions, these effects were reversed by VD3. In addition, VD3 also increased VD3 receptor protein contents and values went back to those of controls after ROT exposure. VD3 protects PC12 cells against ROT damage, by decreasing oxidative stress and improving mitochondrial function. One target seems to be the TH molecule and possibly an indirect Nrf2 activation could also justify its neuroprotective actions on this PC12 cell model of PD.

Rational design of a trypanocidal peptide derived from Dinoponera quadriceps venom.

Chagas disease is caused by the parasite Trypanosoma cruzi and affects millions of people worldwide, having no effective cure. The main sanitary emergency is related to patients with chronic infection, which accumulate comorbidities causing patient death. However, actual chemotherapeutic treatments do not effectively address the chronic forms of the disease. Invertebrates are a relevant source of antimicrobial peptides (AMPs) as part of the innate immune system for their protection. The AMP M-PONTX-Dq3a, isolated from the Dinoponera quadriceps ant venom, has shown very effective antimicrobial and trypanocidal activities. Although M-PONTX-Dq3a has better activity that the current therapies, the peptide length has limited its possibilities to reach clinical application. In this investigation, we aimed to dissect the trypanocidal effect of M-PONTX-Dq3a fragments and to study the activity of substituted analogs, to improve not only peptide trypanocidal activity and bioavailability, but also production costs. Our studies have led to the identification of two smaller peptides, M-PONTX-Dq3a [1-15] and [Lys]3-M-PONTX-Dq3a [3-153-15 with similar trypanocidal activities that the parent peptide has against the three forms of T. cruzi benznidazole-resistant Y strain. Both peptides represent promising candidates to develop novel and effective trypanocidal bio-therapeutic agents, opening new avenues for the treatment of chronic patients.

Antiparasitary and antiproliferative activities in vitro of a 1,2,4-oxadiazole derivative on Trypanosoma cruzi.

Chagas disease (CD) is a neglected disease, prevalent and endemic in Latin America, but also present in Europe and North America. The main treatment used for this disease is benznidazole, but its efficacy is variable in the chronic phase and presents high toxicity. So, there is a need for the development of new therapeutic agents. The five-membered heterocyclic 1,2,4-oxadiazole ring has received attention for its unique properties and a broad spectrum of biological activities and is therefore a potential candidate for the development of new drugs. Thus, the aim of this study was to evaluate the activity of the N-cyclohexyl-3-(3-methylphenyl)-1,2,4-oxadiazol-5-amine (2) on the evolutionary forms of Trypanosoma cruzi strain Y, as well as its mechanisms of action and in silico theoretical approach. The results by computational method showed an interaction of the 1,2,4-oxadiazole (2) with TcGAPDH, cruzain, and trypanothione reductase, showing good charge distribution and affinity in those three targets. Furthermore, cytotoxicity in LLC-MK2 cells was performed by the MTT method. In the assays with different parasite forms, the tested compound showed similar time-dependent concentration effect. The evaluation of the antiamastigote effect between the two concentrations tested showed a reduction in the number of infected cells and also in the number of amastigotes per infected cell. By flow cytometry, the compound (2) displayed alterations suggestive of necrotic events. Finally, in scanning electron microscopy structural alterations were present, characteristic of necrosisin the epimastigote forms. Overall, the 1,2,4-oxadiazole derivative (2) here evaluated opens perspectives to the development of new antichagasic agents.

Chloride substitution on 2-hydroxy-3,4,6-trimethoxyphenylchalcones improves in vitro selectivity on Trypanosoma cruzi strain Y.

Chagas disease is a disease that is emerging in North America and Europe countries. Benznidazole is the main drug available, but it has high toxicity and low efficacy in the chronic phase. In this way, researching new antichagasic agents is necessary. Thus, the aim of this study is to evaluate the effect of novel chalcones and the influence of chlorine substitutions on Trypanosoma cruzi and host cells. Unsubstituted (1), 4-chlorine substituted (2) and 2,4-chlorine substituted (3) chalcones were synthesized by Claisen-Schmidt condensation, characterized, and electrical distribution was assessed by Density Fuctional Theory (DFT). The host cells toxicity (LLC-MK2) was performed by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) reduction assay. The effect on epimastigote (24, 48 and 72h), trypomastigote (24h) and amastigotes (24 h) was evaluated. Flow cytometry assays were performed with 7-Aminoactinomycin D (7-AAD) and Annexin-PE, Dichlorofluorescein diaceteate (DCFH-DA) and Rhodamine123 (Rho123). Finally, molecular docking predicted interactions between chalcones and cruzain (TcCr) and trypanothione reductase (TcTR). The toxicity on host cells was reduced almost twenty times on chlorine substituted molecules. On epimastigote and trypomastigote forms, all substances presented similar effects. After treatment with molecule 3, it was observed a decrease in infected cells and intracellular amastigotes. Their effect is related to necrotic events, increase of cytoplasmic Reactive Oxygen Species (ROS) and mitochondrial dysfunction. Also, this effect might be associated with involvement of TcCr and TcTR enzymes. Therefore, the results showed that chlorine substitution on chalcones reduces the host cell's toxicity without compromising the effect on Trypanosoma cruzi Y strain forms, and it occurs over membrane damage, oxidative stress and possible interactions with TcCr and TcTR.

Preliminary Study for New Markers of Early Tubuloglomerular Injury and Renal Inflammation in Patients with Visceral Leishmaniasis Receiving Liposomal Amphotericin B Treatment.

Visceral leishmaniasis is treated with liposomal amphotericin B (L-AMB), which is associated with nephrotoxicity. Thus, we aimed to investigate nephrotoxicity through novel renal biomarkers in patients with visceral leishmaniasis during L-AMB use. Ours was a prospective study with 17 patients with visceral leishmaniasis treated with L-AMB during their hospital stay. Laboratory tests, renal parameters, urinary biomarkers (urinary kidney injury molecule 1, urinary monocyte chemoattractant protein 1 [uMCP-1], sodium-potassium-2 chloride cotransporter, sodium-hydrogen exchanger 3), and serum inflammatory biomarkers (MCP-1, interferon-γ, and IL-6) were evaluated in two periods: before and during L-AMB use. Glomerular filtration rate, creatinine, proteinuria, and albuminuria were similar before and during L-AMB use. IL-6 levels, aquaporin 2, and sodium-hydrogen exchanger 3 expression decreased, whereas uMCP-1 and urinary kidney injury molecule 1 levels increased during L-AMB treatment. In patients who developed acute kidney injury, uMCP-1 showed higher levels. L-AMB aggravated tubuloglomerular lesions, inflammation, and renal tubular disorders. Thus, patients treated with L-AMB need to be monitored for inflammatory and electrolyte disturbances to prevent acute kidney injury, longer length of hospital stay, higher public costs, and mortality.

(-)-α-Bisabolol as a protective agent against epithelial renal cytotoxicity induced by amphotericin B.

INTRODUCTION: Amphotericin B (AmB), used for systemic fungal infections, has a limited clinical application because of its high nephrotoxicity. Natural antioxidant and anti-inflammatory substances have been widely studied for protection against drug-induced nephrotoxicity. α-Bisabolol (BIS) has demonstrated a nephroprotective effect on both in vitro and in vivo models. AIMS: The aim of this work was to evaluate the effect of BIS against AmB-induced nephrotoxicity in vitro. MATERIAL AND METHODS: LLC-MK2 cells were pre- and post-treated with non-toxic BIS concentrations and/or AmB IC50 (13.97 µM). Cell viability was assessed by MTT [(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)] assay. Flow cytometry analyses were used to assess cell death mechanism, production of reactive oxidative stress (ROS) and mitochondrial transmembrane potential. Kidney Injury Molecule-1 (KIM-1) levels were measured via ELISA. KEY FINDINGS: The present work showed that BIS pretreatment (125; 62.5 and 31.25 µM) increased cell viability when compared to the group treated only with AmB IC50. AmB treatment induced both necrosis (7-AAD-labeled cells) and late apoptosis (AnxV-labeled). BIS was able to prevent the occurrence of these events. These effects were associated with a decrease of ROS accumulation, improving transmembrane mitochondrial potential and protecting against tubular cell damage, highlighted by the inhibition of KIM-1 release after BIS treatment. SIGNIFICANCE: BIS presented a potential effect on model of renal cytotoxicity induced by AmB, bringing perspectives for the research of new nephroprotective agents.

Tyramine exerts hypolipidemic and anti-obesity effects in vivo

Abstract Obesity and dyslipidemia are conditions often associated with cardiovascular risk, inflammation, oxidative stress, and death. Thus, a new approach has been highlighted to promote research and development of pharmacological tools derived from natural sources. Among the most widely studied groups of substances, polyphenols such as tyramine stand out. This study investigated hypolipidemic and anti-obesity properties of tyramine. Oral toxicity evaluation, models of dyslipidemia and obesity were used. To induce dyslipidemia, Poloxamer-407 (P-407) was administered intraperitoneally. In the hypercholesterolemic and obesity model, specific diet and oral tyramine were provided. After 24h of P-407 administration, tyramine 2 mg/kg (T2) decreased triglycerides (TG) (2057.0 ± 158.5 mg/dL vs. 2838 ± 168.3 mg/dL). After 48h, TG were decreased by T2 (453.0 ± 35.47 vs. 760.2 ± 41.86 mg/dL) and 4 mg/kg (T4) (605.8 ± 26.61 760.2 ± 41.86 mg/dL). T2 reduced total cholesterol (TC) after 24h (309.0 ± 11.17 mg/dL vs. 399.7 ± 15.7 mg/dL); After 48h, 1 mg/kg (T1) (220.5 ± 12.78 mg/dL), T2 (205.8 ± 7.1 mg/dL) and T4 (216.8 ± 12.79 mg/dL), compared to P-407 (275.5 ± 12.1 mg/dL). The treatment decreased thiobarbituric acid reactive substances and nitrite in liver, increased superoxide dismutase, reduced the diet-induced dyslipidemia, decreasing TC around 15%. Tyramine reduced body mass, glucose, and TC after hypercaloric feed. Treatment with 5 mg/L (0.46 ± 0.04 ng/dL) and 10 mg/L (0.44 ± 0.02 ng/dL) reduced plasma insulin (1.18 ± 0.23 ng/dL). Tyramine increased adiponectin at 5 mg/L (1.02 ± 0.02 vs. 0.83 ± 0.02 ng/mL) and 10mg/L (0.96 ± 0.04 ng/mL). In conclusion, tyramine has low toxicity in rodents, has antioxidant effect, reduces plasma triglycerides and cholesterol levels. However, further studies should be conducted in rodents and non-rodents to better understand the pharmacodynamic and pharmacokinetic properties of tyramine

Computational approach towards the design of artemisinin-thymoquinone hybrids against main protease of SARS-COV-2.

BACKGROUND: The sanitary emergency installed in the world, generated by the pandemic of COVID-19, instigates the search for scientific strategies to mitigate the damage caused by the disease to different sectors of society. The disease caused by the coronavirus, SARS-CoV-2, reached 216 countries/territories, where about 199 million people were reported with the infection. Of these, more than 4 million died. In this sense, strategies involving the development of new antiviral molecules are extremely important. The main protease (Mpro) from SARS-CoV-2 is an important target, which has been widely studied for antiviral treatment. This work aims to perform a screening of pharmacodynamics and pharmacokinetics of synthetic hybrids from thymoquinone and artemisin (THY-ART) against COVID-19. RESULTS: Molecular docking studies indicated that hybrids of artemisinin and thymoquinone showed a relevant interaction with the active fraction of the enzyme Mpro, when compared to the reference drugs. Furthermore, hybrids show an improvement in the interaction of substances with the enzyme, mainly due to the higher frequency of interactions with the Thr199 residue. ADMET studies indicated that hybrids tend to permeate biological membranes, allowing good human intestinal absorption, with low partition to the central nervous system, potentiation for CYP-450 enzyme inhibitors, low risk of toxicity compared to commercially available drugs, considering mainly mutagenicity and cardiotoxicity, low capacity of hybrids to permeate the blood-brain barrier, high absorption and moderate permeability in Caco-2 cells. In addition, T1-T7 tend to have a better distribution of their available fractions to carry out diffusion and transport across cell membranes, as well as increase the energy of interaction with the SARS-CoV-2 target. CONCLUSIONS: Hybrid products of artemisinin and thymoquinone have the potential to inhibit Mpro, with desirable pharmacokinetic and toxicity characteristics compared to commercially available drugs, being indicated for preclinical and subsequent clinical studies against SARS-CoV-2. Emphasizing the possibility of synergistic use with currently used drugs in order to increase half-life and generate a possible synergistic effect. This work represents an important step for the development of specific drugs against COVID-19.

Prescription Drug Overdose, Depression, and Other Mental Disorders in the Context of Kidney Disease.

Clinical Background: Mental disorders, especially depression, are associated with several comorbidities in the kidneys. Depression is the psychiatric disorder that mostly affects individuals with chronic kidney disease (CKD) and end-stage kidney disease. Epidemiology: The mainly prescribed drugs involved in overdose cases are opioids, benzodiazepines, and antidepressants. Antidepressants are the main psychiatric drugs that lead to kidney injury, mainly the second-generation ones. However, the prevalence of depression in dialysis patients varies from 22.8 to 39.3%. Therefore, psychiatric patients have 1.5-3 times more hospitalization compared to patients having only CKD. Challenges: Randomized clinical studies should be encouraged. Studies have shown an association between depression and progression of kidney disease. The mechanisms are not completely clear, but changes on neurotransmitter release and endocrine functions appear to be related to it. Additionally, the use of antidepressant and other psychoactive drugs can induce kidney injury. Hyponatremia induced by second-generation antidepressant drugs is an important feature and can be a risk factor for elderly or patients with comorbidities such as cerebral edema, brain damage or coma. Besides this class, drugs used for anxiety and bipolar disorders or sympathomimetic drugs of abuse can trigger acute kidney injury, possibly due to endothelial dysfunction and thromboembolic and ischemic events. Prevention and Treatment: The early detection of renal impairment and the prescription of nephroprotective strategies has been a clinical challenge. Some studies aim to describe the biochemical mechanisms involved and develop clinical management strategies for these patients. This chapter brings attention to this topic, discussing the major mechanisms and clinical features of kidney injury associated with mental illness, and the most relevant clinical strategies.

Protective Effect of Quercetin on Renal Tubular Cells and the Involvement with the Renin-Angiotensin-Aldosterone Axis

Abstract Ischemia-reperfusion (I/R) plays an important role in the process of acute kidney injury (AKI) due to the generation of reactive oxygen species (ROS). Substances of natural origin have been studied in the prevention of oxidative damage related to I/R. Quercetin is a flavonoid with antioxidant potential and modulate enzymes, such the inhibition of the Rennin-Angiotensin System (RAS). The aim of this study is to evaluate the nephroprotective effect of quercetin against the I/R and analyze the inhibition of RAS. Rhesus monkey Kidney Epithelial Cells (LLC-MK2 line) were submitted to an in vitro ischemia/reperfusion model. After the reperfusion cells were treated with quercetin, the cell viability was accessed by the MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) assay. Tubular cell damage was assessed by the Kidney Injury Molecule-1 (KIM-1) measurement. Oxidative stress was evaluated through Thiobarbituric Acid Reactive Substances (TBARS) and reduced glutathione (GSH). The evaluation of cell death and the mitochondrial depolarization were analyzed by flow cytometry. Quercetin prevents cell death reducing oxidative stress and preventing mitochondrial membrane depolarization. Molecular docking showed that quercetin prevents cell damage better than losartan and lisinopril, inhibitors of RAS. Quercetin has a potential to interact with type 1 angiotensin II receptor (AT1) with greater affinity through the formation of five hydrogen bonds of strong intensity.

Involvement of NADPH-oxidase enzyme in the nephroprotective effect of (-)-α-bisabolol on HK2 cells exposed to ischemia - Reoxygenation.

Acute Kidney Injury (AKI) is associated with high morbidity and mortality. Ischemia and reperfusion (I/R) are events that lead to AKI through hypoxia, reactive oxygen species (ROS) production, oxidative stress and apoptosis. We aimed to evaluate the mechanism of nephroprotection mediated by Bisabolol in human tubular kidney cells after injury by I/R in vitro. HK2 cells were exposed to I/R and treated with Bisabolol. Cell viability was accessed by MTT assay. Cells were submitted to flow cytometry to evaluate necrotic/apoptotic cells, reactive oxygen species production and mitochondrial transmembrane depolarization. TBARS and GSH were used as parameters of redox balance. Also, KIM-1 supernatant levels were measured. In order to identify an interaction between bisabolol and NOX4, molecular docking and enzymatic assays were performed. Expression of isoform NOX4 on treated cells was examined by western-blot. Finally, cells were visualized by scanning electron microscopy. Bisabolol improved cell viability and prevented cell death by apoptosis, indicated also by the decreased levels of KIM-1. It was observed a decrease on reactive oxygen species production and mitochondrial depolarization, with antioxidant regulation by increased GSH and decreased lipid peroxidation. It was also demonstrated that bisabolol treatment can inhibit NOX4. Finally, SEM images showed that bisabolol reduced I/R-induced cell damage. Bisabolol treatment protects HK2 cells against oxidative damage occasioned by I/R. This effect is related to inhibition of apoptosis, decrease on KIM-1 release, reactive oxygen species accumulation and mitochondrial dysfunction. Bisabolol inhibited NOX4 activity in the tubular cells, impairing reactive oxygen species synthesis.

Nanoencapsulation of benznidazole in calcium carbonate increases its selectivity to Trypanosoma cruzi.

Chagas disease is a public health problem, affecting about 7 million people worldwide. Benznidazole (BZN) is the main treatment option, but it has limited effectiveness and can cause severe adverse effects. Drug delivery through nanoparticles has attracted the interest of the scientific community aiming to improve therapeutic options. The aim of this study was to evaluate the cytotoxicity of benznidazole-loaded calcium carbonate nanoparticles (BZN@CaCO3) on Trypanosoma cruzi strain Y. It was observed that BZN@CaCO3 was able to reduce the viability of epimastigote, trypomastigote and amastigote forms of T. cruzi with greater potency when compared with BZN. The amount of BZN necessary to obtain the same effect was up to 25 times smaller when loaded with CaCO3 nanoparticles. Also, it was observed that BZN@CaCO3 enhanced the selectivity index. Furthermore, the cell-death mechanism induced by both BZN and BZN@CaCO3 was evaluated, indicating that both substances caused necrosis and changed mitochondrial membrane potential.