Piplartine-Inspired 3,4,5-Trimethoxycinnamates: Trypanocidal, Mechanism of Action, and In Silico Evaluation.

Chagas disease (CD) is one of the main neglected tropical diseases that promote relevant socioeconomic impacts in several countries. The therapeutic options for the treatment of CD are limited, and parasite resistance has been reported. Piplartine is a phenylpropanoid imide that has diverse biological activities, including trypanocidal action. Thus, the objective of the present work was to prepare a collection of thirteen esters analogous to piplartine (1-13) and evaluate their trypanocidal activity against Trypanosoma cruzi. Of the tested analogues, compound 11 ((E)-furan-2-ylmethyl 3-(3,4,5-trimethoxyphenyl)acrylate) showed good activity with IC50 values = 28.21 ± 5.34 µM and 47.02 ± 8.70 µM, against the epimastigote and trypomastigote forms, respectively. In addition, it showed a high rate of selectivity to the parasite. The trypanocidal mechanism of action occurs through the induction of oxidative stress and mitochondrial damage. In addition, scanning electron microscopy showed the formation of pores and leakage of cytoplasmic content. Molecular docking indicated that 11 probably produces a trypanocidal effect through a multi-target mechanism, including affinity with proteins CRK1, MPK13, GSK3B, AKR, UCE-1, and UCE-2, which are important for the survival of the parasite. Therefore, the results suggest chemical characteristics that can serve for the development of new trypanocidal prototypes for researching drugs against Chagas disease.

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.

Quantum computational investigations and molecular docking studies on amentoflavone.

Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi, with approximately 6-7 million people infected worldwide, becoming a public health problem in tropical countries, thus generating an increasing demand for the development of more effective drugs, due to the low efficiency of the existing drugs. Aiming at the development of a new antichagasic pharmacological tool, the density functional theory was used to calculate the reactivity descriptors of amentoflavone, a biflavonoid with proven anti-trypanosomal activity in vitro, as well as to perform a study of interactions with the enzyme cruzain, an enzyme key in the evolutionary process of T-cruzi. Structural properties (in solvents with different values of dielectric constant), the infrared spectrum, the frontier orbitals, Fukui analysis, thermodynamic properties were the parameters calculated from DFT method with the monomeric structure of the apigenin used for comparison. Furthermore, molecular docking studies were performed to assess the potential use of this biflavonoid as a pharmacological antichagasic tool. The frontier orbitals (HOMO-LUMO) study to find the band gap of compound has been extended to calculate electron affinity, ionization energy, electronegativity electrophilicity index, chemical potential, global chemical hardness and global chemical softness to study the chemical behaviour of compound. The optimized structure was subjected to molecular Docking to characterize the interaction between amentoflavone and cruzain enzyme, a classic pharmacological target for substances with anti-gas activity, where significant interactions were observed with amino acid residues from each one's catalytic sites enzyme. These results suggest that amentoflavone has the potential to interfere with the enzymatic activity of cruzain, thus being an indicative of being a promising antichagasic agent.

Trypanocidal Mechanism of Action and in silico Studies of p-Coumaric Acid Derivatives.

Trypanosoma species are responsible for chronic and systemic infections in millions of people around the world, compromising life quality, and family and government budgets. This group of diseases is classified as neglected and causes thousands of deaths each year. In the present study, the trypanocidal effect of a set of 12 ester derivatives of the p-coumaric acid was tested. Of the test derivatives, pentyl p-coumarate (7) (5.16 ± 1.28 µM; 61.63 ± 28.59 µM) presented the best respective trypanocidal activities against both epimastigote and trypomastigote forms. Flow cytometry analysis revealed an increase in the percentage of 7-AAD labeled cells, an increase in reactive oxygen species, and a loss of mitochondrial membrane potential; indicating cell death by necrosis. This mechanism was confirmed by scanning electron microscopy, noting the loss of cellular integrity. Molecular docking data indicated that of the chemical compounds tested, compound 7 potentially acts through two mechanisms of action, whether by links with aldo-keto reductases (AKR) or by comprising cruzain (CZ) which is one of the key Trypanosoma cruzi development enzymes. The results indicate that for both enzymes, van der Waals interactions between ligand and receptors favor binding and hydrophobic interactions with the phenolic and aliphatic parts of the ligand. The study demonstrates that p-coumarate derivatives are promising molecules for developing new prototypes with antiprotozoal activity.

Antiparasitic effect of (-)-α-bisabolol against Trypanosoma cruzi Y strain forms.

Chagas disease is caused by Trypanosoma cruzi and affects about 7 million people worldwide. Benznidazole and nifurtimox have low efficacy and high toxicity. The present study was designed to identify the trypanocidal effect of (-)-α-Bisabolol (BIS) and investigate its mechanism. Epimastigotes and trypomastigotes were cultured with BIS and the viable cells were counted. BIS antiamastigote effect was evaluated using infected LLC-MK2 cells. MTT assay was performed to evaluate BIS cytotoxicity. Growth recovery was assessed to evaluate BIS effect after short times of exposure. BIS mechanism was investigated through flow cytometry, with 7-AAD and Annexin V-PE. DCFH-DA, rhodamine 123 (Rho123) and acridine orange (AO). Finally, enzymatic and computational assays were performed to identify BIS interaction with T. cruzi GAPDH (tcGAPDH). BIS showed an inhibitory effect on epimastigotes after all tested periods, as well on trypomastigotes. It caused cytotoxicity on LLC-MK2 cells at higher concentrations, with selectivity index (SeI) = 26.5. After treatment, infected cells showed a decrease in infected cells, the number of amastigotes per infected cell and the survival index (SuI). Growth recovery demonstrated that BIS effect causes rapid death of T. cruzi. Flow cytometry showed that BIS biological effect is associated with apoptosis induction, increase in cytoplasmic ROS and mitochondrial transmembrane potential, while reservosome swelling was observed at a late stage. Also, BIS action mechanism may be associated to tcGAPDH inhibition. Altogether, the results demonstrate that BIS causes cell death in Trypanosoma cruzi Y strain forms, with the involvement of apoptosis and oxidative stress and enzymatic inhibition.

Nephrotoxicity induced by the venom of Hypnale hypnale from Sri Lanka: Studies on isolated perfused rat kidney and renal tubular cell lines.

The hump-nosed pit viper Hypnale hypnale is responsible for a high number of snakebite cases in southwestern India and Sri Lanka. Although most patients only develop local signs and symptoms of envenoming, there is a growing body of evidence indicating that these envenomings may be associated with systemic alterations, including acute kidney injury. In this study we evaluated the renal toxicity of H. hypnale venom by using a perfused isolated rat kidney system and by assessing cytotoxicity in two different renal tubular cell lines in culture. The venom caused alterations in several renal functional parameters, such as reduction on perfusion pressure, renal vascular resistance, and sodium and chloride tubular transport, whereas glomerular filtration rate and urinary flow initially decreased and then increased after venom perfusion. In addition, this venom was cytotoxic to proximal and distal renal tubular cells in culture, with predominance of necrosis over apoptosis. Moreover, the venom affected the mitochondrial membrane potential and induced an increment in reactive oxygen species in these cells. Taken together, our results demonstrate a nephrotoxic activity of H. hypnale venom in these experimental models, in agreement with clinical observations.

Tailoring microstructural, drug release properties, and antichagasic efficacy of biocompatible oil-in-water benznidazol-loaded nanoemulsions.

This study explored the transition of lamellar-type liquid crystal (LLC) to biocompatible oil-in-water nanoemulsions able to modify benznidazole (BNZ) release and target the drug to cells infected with the T. cruzi parasite. Three cosolvents (2methylpyrrolidone [NMP], polyethylene glycol [POL], and propylene glycol [PRO] were tested to induce the transition of anisotropic LLC systems to isotropic nanoemulsions. Mixtures of soy phosphatidylcholine with sodium oleate stabilized the dispersions of medium chain triglyceride in water. Rheological measurements, polarized microscopy, and small angle X-ray scattering demonstrated that there is a phase transition from LLC to desired nanoemulsions. These small and narrow droplet-sized nanocarriers exhibited some advantages and promising features, such as the enhanced BNZ aqueous solubility and slow drug release rate. In vitro cell biocompatibility of formulations was assessed in the Vero E6 and SiHa cell lines. Drug-loaded nanoemulsions inhibited the epimastigote growth of the T. cruzi parasite (IC50 0.208 ±â€¯0.052 µg mL-1) and reduced its infective life form trypomastigote (IC50 0.392 ±â€¯0.107 µg mL-1). The oil-in-water nanoemulsions were demonstrated as promising biocompatible liquid drug delivery systems capable of improving the BNZ trypanocidal activity for the treatment of Chagas disease.

Visceral leishmaniasis-associated nephropathy in hospitalised Brazilian patients: new insights based on kidney injury biomarkers.

OBJECTIVE: To evaluate the usefulness of early acute kidney injury (AKI) biomarkers in clinical management of visceral leishmaniasis. METHODS: Prospective study with 50 hospitalised VL patients. AKI biomarkers, that is, serum and urinary neutrophil gelatinase-associated lipocalin (sNGAL, uNGAL, respectively), urinary kidney injury molecule-1 (uKIM-1) and urinary monocyte chemotactic protein-1 (uMCP-1), were quantified by immunoassay (ELISA). Also, interferon-gamma (INF-y) and C-reactive protein (CRP) were evaluated as inflammatory biomarkers possibly related to VL severity. RESULTS: VL patients had hyponatremia, hypoalbuminemia, hypergammaglobulinemia, haematologic and hepatic disorders. AKI was found in 46%, and one death (2%) occurred. The AKI group had significant longer hospital stay, lower levels of IFN-y and higher levels of CRP, more clinical renal abnormalities and higher levels of sNGAL, uNGAL, uKIM-1 and uMCP-1. Overall, sNGAL, uKIM-1 and uMCP-1 showed correlations with important clinical renal abnormalities, such as proteinuria, albuminuria, serum creatinine and glomerular filtration rate using adjusted correlations with CRP and IFN-y. Only sNGAL showed an early association with AKI development (OR = 2.78, 95% CI = 1.429-5.428, per each increase of 50 ng/ml), even after adjusting for clinical signals of VL severity and for immune biomarkers. Moreover, sNGAL showed a better performance in predicting AKI development (AUC-ROC = 0.81, 95% CI = 0.69-0.93; cut-off = 154 ng/ml, sensitivity = 82.6%, specificity = 74.1%, P < 0.001). CONCLUSIONS: Visceral leishmaniasis-associated nephropathy showed important proximal tubular injury and glomerular inflammation. Serum NGAL showed an early association with VL-associated nephropathy and may be used to improve clinical management strategies and decrease morbimortality in VL patients.

Effects of Tityus stigmurus (Thorell 1876) (Scorpiones: Buthidae) venom in isolated perfused rat kidneys.

Scorpions belonging to the Tityus genus are of medical interest in Brazil. Among them, Tityus stigmurus is the main scorpion responsible for stings in the Northeast region. After a sting, the scorpion venom distributes rapidly to the organs, reaching the kidneys quickly. However, there are few studies concerning the renal pathophysiology of scorpion poisoning. In this study, we evaluated the effects of T. stigmurus venom (TsV) on renal parameters in isolated rat kidneys. Wistar rats (n = 6), weighing 250-300 g, were perfused with Krebs-Henseleit solution containing 6 g/100 mL bovine serum albumin. TsV at 0.3 and 1.0 µg/mL was tested, and the effects on perfusion pressure (PP), renal vascular resistance (RVR), urinary flow (UF), glomerular filtration rate (GFR), and electrolyte excretion were analyzed. Effects were observed only at TsV concentration of 1.0 µg/mL, which increased PP (controlPP40' = 92.7 ± 1.95; TsVPP40' = 182.0 ± 4.70* mmHg, *p < 0.05), RVR (controlRVR40' = 3.28 ± 0.23 mmHg; TstRVR40' = 6.76 ± 0.45* mmHg, *p < 0.05), UF (controlUF50' = 0.16 ± 0.04; TstUF50' = 0.60 ± 0.10* mL/g/min,*p < 0.05), GFR and electrolyte excretion, with histological changes that indicate renal tubular injury. In conclusion, T. stigmurus venom induces a transient increase in PP with tubular injury, both of which lead to an augmented electrolyte excretion.

Involvement of Nitric Oxide on Bothropoides insularis Venom Biological Effects on Murine Macrophages In Vitro.

Viperidae venom has several local and systemic effects, such as pain, edema, inflammation, kidney failure and coagulopathy. Additionally, bothropic venom and its isolated components directly interfere on cellular metabolism, causing alterations such as cell death and proliferation. Inflammatory cells are particularly involved in pathological envenomation mechanisms due to their capacity of releasing many mediators, such as nitric oxide (NO). NO has many effects on cell viability and it is associated to the development of inflammation and tissue damage caused by Bothrops and Bothropoides venom. Bothropoides insularis is a snake found only in Queimada Grande Island, which has markedly toxic venom. Thus, the aim of this work was to evaluate the biological effects of Bothropoides insularis venom (BiV) on RAW 264.7 cells and assess NO involvement. The venom was submitted to colorimetric assays to identify the presence of some enzymatic components. We observed that BiV induced H2O2 production and showed proteolytic and phospholipasic activities. RAW 264.7 murine macrophages were incubated with different concentrations of BiV and then cell viability was assessed by MTT reduction assay after 2, 6, 12 and 24 hours of incubation. A time- and concentration-dependent effect was observed, with a tendency to cell proliferation at lower BiV concentrations and cell death at higher concentrations. The cytotoxic effect was confirmed after lactate dehydrogenase (LDH) measurement in the supernatant from the experimental groups. Flow cytometry analyses revealed that necrosis is the main cell death pathway caused by BiV. Also, BiV induced NO release. The inhibition of both proliferative and cytotoxic effects with L-NAME were demonstrated, indicating that NO is important for these effects. Finally, BiV induced an increase in iNOS expression. Altogether, these results demonstrate that B. insularis venom have proliferative and cytotoxic effects on macrophages, with necrosis participation. We also suggest that BiV acts by inducing iNOS expression and causing NO release.

Bothropoides pauloensis venom effects on isolated perfused kidney and cultured renal tubular epithelial cells.

Snake envenomation (Bothrops genus) is common in tropical countries and acute kidney injury is one of the complications observed in Bothrops snakebite with relevant morbidity and mortality. Here, we showed that Bothropoides pauloensis venom (BpV) decreased cell viability (IC50 of 7.5 µg/mL). Flow cytometry with annexin V and propidium iodide showed that cell death occurred predominantly by apoptosis and late apoptosis, through caspases 3 and 7 activation, mitochondrial membrane potential collapse and ROS overproduction. BpV reduced perfusion pressure, renal vascular resistance, urinary flow, glomerular filtration rate, percentage of sodium, chloride or potassium tubular transportation. These findings demonstrated that BpV cytotoxicity on renal epithelial cells might be responsible for the nephrotoxicity observed in isolated kidney.

L-Aminoacid Oxidase from Bothrops leucurus Venom Induces Nephrotoxicity via Apoptosis and Necrosis.

Acute renal failure is a common complication caused by Bothrops viper envenomation. In this study, the nefrotoxicity of a main component of B. leucurus venom called L-aminoacid oxidase (LAAO-Bl) was evaluated by using tubular epithelial cell lines MDCK and HK-2 and perfused kidney from rats. LAAO-Bl exhibited cytotoxicity, inducing apoptosis and necrosis in MDCK and HK-2 cell lines in a concentration-dependent manner. MDCK apoptosis induction was accompanied by Ca2+ release from the endoplasmic reticulum, reactive oxygen species (ROS) generation and mitochondrial dysfunction with enhanced expression of Bax protein levels. LAAO-Bl induced caspase-3 and caspase-7 activation in both cell lines. LAAO-Bl (10 µg/mL) exerts significant effects on the isolated kidney perfusion increasing perfusion pressure and urinary flow and decreasing the glomerular filtration rate and sodium, potassium and chloride tubular transport. Taken together our results suggest that LAAO-Bl is responsible for the nephrotoxicity observed in the envenomation by snakebites. Moreover, the cytotoxic of LAAO-Bl to renal epithelial cells might be responsible, at least in part, for the nephrotoxicity observed in isolated kidney.

Preliminary study on tubuloglomerular dysfunction and evidence of renal inflammation in patients with visceral leishmaniasis.

Visceral leishmaniasis (VL) is a re-emerging zoonosis of worldwide distribution. Monocyte chemotactic protein-1 (MCP-1) and malondialdehyde (MDA) are inflammation biomarkers that have never been investigated in VL. The aim of this study is to investigate the association between renal abnormalities and inflammation biomarkers in VL. This study is a preliminary prospective study with 16 VL adult patients evaluated before treatment compared with a group of 13 healthy volunteers and 5 VL patients evaluated after treatment. Urinary concentration and acidification tests were performed. MCP-1 and MDA were quantified in urine. Urinary concentration deficit was found in all VL patients before (100%) and four VL patients after (80%) treatment. Urinary acidification deficit was found in nine cases before (56.2%) and two cases after (40%) treatment. Urinary MCP-1 (374 ± 359 versus 42 ± 29 pg/mg creatinine, P = 0.002) as well as urinary MDA (5.4 ± 2.6 versus 2.0 ± 0.8 µmol/mL) showed significant differences between VL patients and controls. These data show that VL patients present urinary concentration and acidification deficit, which can persist even after specific treatment. Urinary MCP-1 and MDA are elevated in patients with VL, which suggests renal inflammation and incipient renal damage.

Proteinuria in adults with sickle-cell disease: the role of hydroxycarbamide(hydroxyurea) as a protective agent.

BACKGROUND: Renal abnormalities are often seen in sickle cell disease (SCD). OBJECTIVE: To investigate the role of hydroxycarbamide as a protective agent in sickle cell nephropathy. SETTING: Patients with SCD followed at a Hematology outpatients clinic. METHODS: Prospective study with 26 SCD patients. Renal function evaluation was performed and a comparison between patients and control group was done. Patients using hydroxycarbamide were compared to those not taking this drug. MAIN OUTCOME MEASURE: Effect of hydroxycarbamide on renal function. RESULTS: Patients mean age was 32.1 ± 9.9 years, and 16 (61 %) were males. Glomerular hyperfiltration was found in nine patients with SCD (34.6 %). GFR < 60 mL/min/1.73 m² was observed in three cases (11.5 %). Microalbuminuria (30-300 mg/day) was found in seven cases (27 %) and macroalbuminuria (>300 mg/dia) in one patient (3.8 %). All patients had urinary concentrating deficit, and inability to acidify urine was found in ten cases (38.4 %). The comparison of patients according to the use of hydroxycarbamide showed lower levels of serum creatinine in those using the drug (0.6 ± 0.1 vs. 0.8 ± 0.3 mg/dL, p = 0.03), as well as lower levels of 24 h-proteinuria (226 ± 16 vs. 414 ± 76 mg/dL, p = 0.0001), but not microalbuminuria (79 ± 15 vs. 55 ± 86 mg/dL, p = 0.35). CONCLUSION: SCD is associated with important renal abnormalities. Hydroxycarbamide seems to protect kidney function in SCD by decreasing proteinuria but not microalbuminuria.

Bothropoides insularis venom cytotoxicity in renal tubular epithelia cells.

Bothropoides insularis (jararaca-ilhoa) is a native endemic snake limited to the specific region of Queimada Island, on São Paulo coast. Several local and systemic effects have been described due to envenomation caused by it, such as edema, tissue necrosis, hemorrhage and acute renal failure. Our previous studies have shown that Bothropoides insularis venom (BinsV) demonstrated important functional and morphologic alterations in rat isolated kidney, especially decrease in tubular electrolyte transport, osmotic clearance and tubular necrosis. In order to elucidate the direct nephrotoxicity mechanism, the aim of the present study was to investigate BinsV cytotoxicity effect on renal epithelial cells. The treatment with BinsV over MDCK culture decreased cell viability in all concentrations tested with IC50 of 9 µg/mL. BinsV was able to induce membrane rupture and cell death with phosphatidilserine externalization. Furthermore, BinsV induced ROS overproduction and mitochondrial membrane potential collapse, as well as Bax translocation and caspases 3 and 7 expression. Therefore, these events might be responsible by BinsV-induced cell death caused by mitochondrial dysfunction and ROS overproduction in the direct cytotoxicity process.

Optical absorption of the antitrypanocidal drug benznidazole in water.

UV-vis optical absorption spectra of the antitrypanocidal drug benznidazole solvated in water were measured for various concentrations. The spectra show a prominent peak around 3.80 eV, while deconvolution of the UV-vis optical absorption spectra revealed six bands centered at 3.60, 3.83, 4.15, 4.99, 5.60, and 5.76 eV. Benznidazole electronic transitions were obtained after density functional theory (DFT) calculations within the polarized continuum (PCM) model for water solvation. Molecular geometry optimizations were carried out, and the measured absorption peaks were related to specific molecular orbital transitions obtained within the time dependent DFT (TD-DFT) with excellent agreement between theory and experiment.

Transcriptome analysis in venom gland of the predatory giant ant Dinoponera quadriceps: insights into the polypeptide toxin arsenal of hymenopterans.

BACKGROUND: Dinoponera quadriceps is a predatory giant ant that inhabits the Neotropical region and subdues its prey (insects) with stings that deliver a toxic cocktail of molecules. Human accidents occasionally occur and cause local pain and systemic symptoms. A comprehensive study of the D. quadriceps venom gland transcriptome is required to advance our knowledge about the toxin repertoire of the giant ant venom and to understand the physiopathological basis of Hymenoptera envenomation. RESULTS: We conducted a transcriptome analysis of a cDNA library from the D. quadriceps venom gland with Sanger sequencing in combination with whole-transcriptome shotgun deep sequencing. From the cDNA library, a total of 420 independent clones were analyzed. Although the proportion of dinoponeratoxin isoform precursors was high, the first giant ant venom inhibitor cysteine-knot (ICK) toxin was found. The deep next generation sequencing yielded a total of 2,514,767 raw reads that were assembled into 18,546 contigs. A BLAST search of the assembled contigs against non-redundant and Swiss-Prot databases showed that 6,463 contigs corresponded to BLASTx hits and indicated an interesting diversity of transcripts related to venom gene expression. The majority of these venom-related sequences code for a major polypeptide core, which comprises venom allergens, lethal-like proteins and esterases, and a minor peptide framework composed of inter-specific structurally conserved cysteine-rich toxins. Both the cDNA library and deep sequencing yielded large proportions of contigs that showed no similarities with known sequences. CONCLUSIONS: To our knowledge, this is the first report of the venom gland transcriptome of the New World giant ant D. quadriceps. The glandular venom system was dissected, and the toxin arsenal was revealed; this process brought to light novel sequences that included an ICK-folded toxins, allergen proteins, esterases (phospholipases and carboxylesterases), and lethal-like toxins. These findings contribute to the understanding of the ecology, behavior and venomics of hymenopterans.

Isolation, homology modeling and renal effects of a C-type natriuretic peptide from the venom of the Brazilian yellow scorpion (Tityus serrulatus).

Mammalian natriuretic peptides (NPs) have been extensively investigated for use as therapeutic agents in the treatment of cardiovascular diseases. Here, we describe the isolation, sequencing and tridimensional homology modeling of the first C-type natriuretic peptide isolated from scorpion venom. In addition, its effects on the renal function of rats and on the mRNA expression of natriuretic peptide receptors in the kidneys are delineated. Fractionation of Tityus serrulatus venom using chromatographic techniques yielded a peptide with a molecular mass of 2190.64 Da, which exhibited the pattern of disulfide bridges that is characteristic of a C-type NP (TsNP, T. serrulatus Natriuretic Peptide). In the isolated perfused rat kidney assay, treatment with two concentrations of TsNP (0.03 and 0.1 µg/mL) increased the perfusion pressure, glomerular filtration rate and urinary flow. After 60 min of treatment at both concentrations, the percentages of sodium, potassium and chloride transport were decreased, and the urinary cGMP concentration was elevated. Natriuretic peptide receptor-A (NPR-A) mRNA expression was down regulated in the kidneys treated with both concentrations of TsNP, whereas NPR-B, NPR-C and CG-C mRNAs were up regulated at the 0.1 µg/mL concentration. In conclusion, this work describes the isolation and modeling of the first natriuretic peptide isolated from scorpion venom. In addition, examinations of the renal actions of TsNP indicate that its effects may be related to the activation of NPR-B, NPR-C and GC-C.

Renal tubular dysfunction in sickle cell disease.

BACKGROUND/AIMS: Kidney abnormalities are one of the main chronic complications of sickle cell disease (SCD). The aim of this study is to investigate the occurrence of renal tubular abnormalities among patients with SCD. METHODS: This is a prospective study with 26 SCD adult patients in Brazil. Urinary acidification and concentration tests were performed using calcium chloride (CaCl2), after a 12h period of water and food deprivation. Fractional excretion of sodium (FENa), transtubular potassium gradient (TTKG) and solute free water reabsorption (TcH2O) were calculated. The SCD group was compared to a group of 15 healthy volunteers (control group). RESULTS: Patient`s average age and gender were similar to controls. Urinary acidification deficit was found in 10 SCD patients (38.4%), who presented urinary pH >5.3 after CaCl2 test. Urinary osmolality was significantly lower in SCD patients (355 ± 60 vs. 818 ± 202 mOsm/kg, p=0.0001, after 12h period water deprivation). Urinary concentration deficit was found in all SCD patients (100%). FENa was higher among SCD patients (0.75 ± 0.3 vs. 0.55 ± 0.2%, p=0.02). The TTKG was higher in SCD patients (5.5 ± 2.5 vs. 3.0 ± 1.5, p=0.001), and TcH2O was lower (0.22 ± 0.3 vs. 1.1 ± 0.3L/day, p=0.0001). CONCLUSIONS: SCD is associated with important kidney dysfunction. The main abnormalities found were urinary concentrating and incomplete distal acidification defect. There was also an increase in the potassium transport and decrease in water reabsorption, evidencing the occurrence of distal tubular dysfunction. .

Effects of Bixa orellana L. seeds on hyperlipidemia.

Bixa orellana L., urucum, or urucu, a native tropical tree of Central and South American rain forests is used to treat various diseases in popular medicine. In Ceará, Northeast of Brazil, the seeds of urucum have been used for the treatment of high lipid blood levels. The present study investigated the effects of the aqueous extract from Bixa orellana seeds (AEBO) in mice with hyperlipidemia induced by tyloxapol, fructose and ethanol. In hyperlipidemia induced by Triton WR1339, 400 and 800 mg/kg AEBO reduced triglycerides (TG) serum levels at 24 h and 48 h. In the study of hypertriglyceridemia induced by fructose, AEBO in doses of 400 mg/kg and 800 mg/kg reduced TG levels by 48.2% and 48.7%, respectively. Finally, the ethanol experimental model with 400 mg/kg AEBO promoted a reduction of 33.6% of TG levels, while the 800 mg/kg concentration reduced hypertriglyceridemia in 62.2%. In conclusion, the aqueous extract of the seeds of Bixa orellana was capable of reversing the hypertriglyceridemia induced by Triton, fructose and ethanol, demonstrating a hypolipidemic effect. However, further studies are necessary to discover the precise mechanism of action.