Protective action of N-acetyl-L-cysteine associated with a polyvalent antivenom on the envenomation induced by Lachesis muta muta (South American bushmaster) in rats

In this study, we examined the potential use of N-acetyl-L-cysteine (NAC) in association with a polyvalent antivenom and as stand-alone therapy to reduce the acute local and systemic effects induced by Lachesis muta muta venom in rats. Male Wistar rats (300–350 g) were exposed to L. m. muta venom (1.5 mg/kg – i.m.) and subsequently treated with anti-Bothrops/Lachesis serum (antivenom:venom ratio 1:3 ‘v/w’ – i.p.) and NAC (150 mg/kg – i.p.) separately or in association; the animals were monitored for 120 min to assess changes in temperature, locomotor activity, local oedema formation and the prevalence of haemorrhaging. After this time, animals were anesthetized in order to collect blood samples through intracardiac puncture and then euthanized for collecting tissue samples; the hematological-biochemical and histopathological analyses were performed through conventional methods. L. m. muta venom produced pronounced local oedema, subcutaneous haemorrhage and myonecrosis, with both antivenom and NAC successfully reducing the extent of the myonecrotic lesion when individually administered; their association also prevented the occurrence of subcutaneous haemorrhage. Venom-induced creatine kinase (CK) release was significantly prevented by NAC alone or in combination with antivenom; NAC alone failed to reduce the release of hepatotoxic (alanine aminotransferase) and nephrotoxic (creatinine) serum biomarkers induced by L. m. muta venom. Venom induced significant increase of leucocytes which was also associated with an increase of neutrophils, eosinophils and monocytes; antivenom and NAC partially reduced these alterations, with NAC alone significantly preventing the increase of eosinophils whereas neither NAC or antivenom prevented the increase in monocytes. Venom did not induce changes in the erythrogram parameters. In the absence of a suitable antivenom, NAC has the potential to reduce a number of local and systemic effects caused by L. m. muta venom

Amyloid seeds formed by cellular uptake, concentration, and aggregation of the amyloid-beta peptide

One of the neuropathological hallmarks of Alzheimer's disease (AD) is the amyloid plaque, primarily composed of aggregated amyloid-beta (Aβ) peptide. In vitro, Aβ1–42, the major alloform of Aβ found in plaques, self-assembles into fibrils at micromolar concentrations and acidic pH. Such conditions do not exist in the extracellular fluid of the brain where the pH is neutral and Aβ concentrations are in the nanomolar range. Here, we show that extracellular soluble Aβ (sAβ) at concentrations as low as 1 nM was taken up by murine cortical neurons and neuroblastoma (SHSY5Y) cells but not by human embryonic kidney (HEK293) cells. Following uptake, Aβ accumulated in Lysotracker-positive acidic vesicles (likely late endosomes or lysosomes) where effective concentrations (>2.5 μM) were greater than two orders of magnitude higher than that in the extracellular fluid (25 nM), as quantified by fluorescence intensity using laser scanning confocal microscopy. Furthermore, SHSY5Y cells incubated with 1 μM Aβ1–42 for several days demonstrated a time-dependent increase in intracellular high molecular weight (HMW) (>200 kDa) aggregates, which were absent in cells grown in the presence of Aβ1–40. Homogenates from these Aβ1–42-loaded cells were capable of seeding amyloid fibril growth. These results demonstrate that Aβ can be taken up by certain cells at low physiologically relevant concentrations of extracellular Aβ, and then concentrated into endosomes/lysosomes. At high concentrations, vesicular Aβ aggregates to form HMW species which are capable of seeding amyloid fibril growth. We speculate that extrusion of these aggregates may seed extracellular amyloid plaque formation during AD pathogenesis