Alkaloids from Prosopis juliflora leaves induce glial activation, cytotoxicity and stimulate NO production.

Abstract

Prosopis juliflora is used for feeding cattle and humans. Intoxication with the plant has been reported, and is characterized by neuromuscular alterations and gliosis. Total alkaloidal extract (TAE) was obtained using acid/basic-modified extraction and was fractionated. TAE and seven alkaloidal fractions, at concentrations ranging 0.03-30 microg/ml, were tested for 24h on astrocyte primary cultures derived from the cortex of newborn Wistar rats. The MTT test and the measure of LDH activity on the culture medium, revealed that TAE and fractions F29/30, F31/33, F32 and F34/35 were cytotoxic to astrocytes. The EC(50) values for the most toxic compounds, TAE, F31/33 and F32 were 2.87 2.82 and 3.01 microg/ml, respectively. Morphological changes and glial cells activation were investigated through Rosenfeld's staining, by immunocytochemistry for the protein OX-42, specific of activated microglia, by immunocytochemistry and western immunoblot for GFAP, the marker of reactive and mature astrocytes, and by the production of nitric oxide (NO). We observed that astrocytes exposed to 3 microg/ml TAE, F29/30 or F31/33 developed compact cell body with many processes overexpressing GFAP. Treatment with 30 microg/ml TAE and fractions, induced cytotoxicity characterized by a strong cell body contraction, very thin and long processes and condensed chromatin. We also observed that when compared with the control (+/-1.34%), the proportion of OX-42 positive cells was increased in cultures treated with 30 microg/ml TAE or F29/30, F31/33, F32 and F34/35, with values raging from 7.27% to 28.74%. Moreover, incubation with 3 microg/ml F32, 30 microg/ml TAE, F29/30, F31/33 or F34/35 induced accumulation of nitrite in culture medium indicating induction of NO production. Taken together these results show that TAE and fractionated alkaloids from P. juliflora act directly on glial cells, inducing activation and/or cytotoxicity, stimulating NO production, and may have an impact on neuronal damages observed on intoxicated animals.

The flavonoid rutin induces astrocyte and microglia activation and regulates TNF-alpha and NO release in primary glial cell cultures

“…Astrocyte primary cultures (APC) were prepared according to Cookson and Pentreath (1994), as described (Silva et al 2006). Briefly, cerebral hemispheres of newborn Wistar rat pups were isolated aseptically and meninges were removed.…”

Study of the distribution profile of piperidine alkaloids in various parts of Prosopis juliflora by the application of Direct Analysis in Real Time Mass Spectrometry (DART-MS)

“…1 It is known to contain piperidine alkaloids having medicinal properties. 2 Many alkaloids such as, juliprosopine, julifloricine, julifloridine, juliprosinine, 4 3′-oxojuliprosopine, sceojuliprosopinol, 3-oxojuliprosine and 3′-oxo-juliprosine 5 have been isolated from leaves and there in vitro biological activities demonstrated, which include anti-inflammatory, 2 antibacterial, 6,7 antifungal, 8 hemolytic, 9 allelopathic, 10 cytotoxic 11 and antitumoral. 12 Although the plant is reported to contain pharmacologically active piperidine alkaloids, limited work had been done to study the distribution profile of its metabolite.…”

Autophagy protects against neural cell death induced by piperidine alkaloids present in Prosopis juliflora (Mesquite)

“…A synergistic action of piperidine alkaloids has been suggested to be responsible for the neurotoxic damage observed in cattle and goats after intensive ingestion of Prosopis juliflora (Câmara et al 2009, Hughes et al 2006, Silva et al 2007, 2013, Tabosa et al 2000a, 2000b. Tabosa et al (2000a) found that the toxic activity observed in mice is chemically related to piperidine alkaloids such as juliprosopine present in the pods of this Leguminosae.…”

“…Tabosa et al (2000a) found that the toxic activity observed in mice is chemically related to piperidine alkaloids such as juliprosopine present in the pods of this Leguminosae. We have previously demonstrated that a fraction from leaves composed of the piperidine alkaloid juliprosopine, as the major constituent, amplifies the neurotoxic response induced by a total alkaloid extract (TAE) in neurons and glial cells in vitro (Silva et al 2007(Silva et al , 2013. This response was best characterized in a model of neuron/glial cell co-culture by neuronal death, astrogliosis, microgliosis, cytoskeleton disruption, mitochondrial damage and vacuolation induction (Silva et al 2013).…”