Identification and pharmacological induction of autophagy in the larval stages of Echinococcus granulosus: an active catabolic process in calcareous corpuscles

Biology and Systematics of Echinococcus

“…Their function may be that of a buffering system or a source of inorganic ions, CO 2 and phosphates (Smyth, 1969), but their transitory nature suggests an association with cell death (Thompson, 1995) and recently they have been shown to be associated with autophagy and catabolic processes (Loos et al, 2014). Within 3e4 days after infection, the lateral excretory canals of the young worm are clearly evident and by the end of the first week a posterior excretory bladder is seen (Smyth and Davies, 1974a).…”

Metformin promotes autophagy in Echinococcus granulosus larval stage

“…Echinococcus granulosus protoscoleces were removed aseptically from hydatid cysts of infected cattle presented for routine slaughter at the Liminal abattoir (official number: 3879) located in the Southeast of Buenos Aires, Argentina. Viable protoscoleces with conserved morphology (n=3,000) were cultured in 24-well culture plates using medium 199 (Gibco) supplemented with glucose (4 mg/ml) and antibiotics (penicillin, streptomycin and gentamicin 100 µg/ml) under normal atmospheric conditions as we previously reported [7]. Additionally, E. granulosus murine cysts (n= [10][11][12][13][14][15][16][17][18][19][20] were incubated in Leighton tubes under the same culture conditions as mentioned for protoscoleces [9].…”

“…In vitro protoscolex treatments were performed with 1, 5 and 10 mM Met and with 10 and 100 µM rapamycin for different periods according to the experiment. Otherwise, in vitro metacestode treatments were performed with 10 mM Met for 48 h. For further molecular assays, parasites were washed with sterile and RNAse-free PBS Finally, to allow the development of vesicularised protoscoleces and microcysts, protoscoleces were cultured in medium 199 supplemented with antibiotics (penicillin, streptomycin and gentamicin; 100 μg/ml), glucose (4 mg/ml), insulin (1.2 U ml-1) and 15% FBS as we already reported [7]. The complete process was followed every day under an inverted light microscope.…”

“…They act as potent transcriptional activators of some autophagy genes by binding to the conserved consensus core recognition motif TTGTTTAC [16,17]. As in other invertebrates, a single Eg-FoxO transcription factor was identified in E. granulosus, as well as conserved consensus sequences for FoxO binding in autophagy genes [7]. The regulation of FoxO is achieved by changes at the protein levels, the subcellular localization and the transcriptional activity, which occurs through post-translational modifications, including phosphorylation, acetylation and ubiquitination [18].…”

“…A well-orchestrated program including over 30 autophagyrelated (Atg) genes controls autophagy, which can be activated by nutrient starvation and subsequent inhibition of mechanistic target of rapamycin (mTOR) signaling or by induction of the unfolded protein response (UPR) as a result of the accumulation of misfolded proteins aggregates [5,6]. Recently, we have reported the identification of the core autophagic machinery, the occurrence of TORC1 (Target of Rapamycin Complex 1)-controlled events, and the correlation between autophagy and the activation of UPR by bortezomib in E. granulosus larval stage [7,8]. In addition, we have demonstrated that both larval forms of E. granulosus are susceptible to metformin (Met), an anti-hyperglycemic drug that indirectly activates Eg-AMPK (AMP-activated protein kinase), as a consequence of cellular energy charge depletion [9,10].…”