(Ca2+-Mg2+)ATPase in Schistosoma mansoni: evidence for heterogeneity and resistance to praziquantel

Abstract

In mammals, Ca 2+ pumps use the chemical energy derived from the hydrolysis of ATP to remove Ca 2+ from the cytoplasm of a variety of cell types (L De Meis & G Inesi 1982, p. 141-169. In E Carafoli, Membrane Transport of Calcium, Academic Press, New York, AF Rega 1986, p. 13-20. In AF Rega, The Ca 2+ Pump of Plasma Membrane, CRC Press, Boca Raton). The pumps of plasma membranes (PMCA family) and those of endoplasmic or sarcoplasmic reticulum (SERCA family) (NM Green 1992 Ann NY Acad Sci 671: 104-169) have the same physiological functions, despite differences in their structure, subcellular localization and modulatory mechanisms. Thapsigargin, a plant-derived sesquiterpene lactone, inhibits the activity of all of the SERCA isoenzymes (SERCA 1 , SERCA 2 and SERCA 3 ) with equal potency, but has no effect on the ATPase or transport activities of PMCA pumps. In previous reports we demonstrated a (Ca 2+ -Mg 2+ )ATPase activity coupled to the active transport of Ca 2+ in subcellular fractions from Schistosoma mansoni (VMN Cunha et al. 1988 FEBS Lett 241: 65-68, 1992. The aim of the present work was, first, to investigate the subcellular localization of the (Ca 2+ -Mg 2+ )ATPase activities present in P 1 and P 4 fractions, using thapsigargin, cyclopiazonic acid (CPA) and tamoxifen as inhibitors of Ca 2+ pump. Second, to investigate if an ATPase responsible for the control of Ca 2+ homeostasis, could be involved in the molecular mechanism of praziquantel-induced contraction in S. mansoni.Male cercariae of S. mansoni (BH strain) were obtained from snails (Biomphalaria glabrata) previously infected with a single miracidium (Cunha 1988 loc. cit.). The subcellular fractions were obtained by homogenizing and centrifuging about 2000 worms to obtain four pellets (P 1 , P 2 , P 3 , P 4 ) sedimenting respectively at 300 x g av (5 min); 1000 x g av (10 min); 8000 x g av (10 min) and 100,000 x g av (1hr). About 25 mg of protein from P 1 or P 4 fractions were incubated for 1 hr at 37 o C in 0.5 ml of a mixture containing (unless otherwise stated) 5 mM Na 2 ATP, 0.3 mM EGTA, 10 mM NaN 3, 4 mM MgCl 2, 60 mM KCl, 5 mM A 23187 (calcimycin, a calcium ionophore), 50 mM HEPES-Tris buffer (pH 7.4) and 0 or 370 mM CaCl 2 (10 mM free Ca 2+ ), for the determination of ATPase activity. The (Ca 2+ -Mg 2+ )ATPase was calculated by subtracting the basal ATPase activity measured in the absence of calcium from the total activity measured in the presence of calcium.The effect of thapsigargin on (Ca 2+ -Mg 2+ )ATPase activity in the fractions P 1 and P 4 from S. mansoni is shown in Fig. 1. In fraction P 4 , the (Ca 2+ -Mg 2+ )ATPase activity was inhibited by the drug with a half-maximal effect (I 50 ) at 250 nM. At a saturating concentration of thapsigargin (3 mM), the activity was completely blocked (I max = 10 3 ± 5.1%, n = 3). Under the same conditions, about 20% of the (Ca 2+ -Mg 2+ )ATPase activity present in P 1 was resistant to thapsigargin (I max =78.3 ± 4.9%, n = 3; P<0.005, Student's ttest). There was no significant difference between fra...

Schistosomiasis - An Unusual Cause of Ureteral Obstruction: A Case History and Perspective

“…[116][117][118] The European Commission has established an International Initiative on Praziquantel Use to review reports of low efficacy in clinical trials in Senegal and Egypt, and reports of resistant S. mansoni strains isolated in the laboratory. 115,[119][120][121][122][123][124][125][126][127][128][129] While investigations suggest that no emergence of praziquantel resistance in S. haematobium has yet occurred, mathematical models predict that such resistance can be expected to occur as soon as 2010. 130 As a consequence new drugs are being actively investigated.…”

Formulation and Clinical Efficacy of Myrrh Extract in Hard Gelatin Capsules

“…For many of the Schistosoma species that infect humans, isoquinolin-4-one, or praziquantel (PZQ), is the only effective drug for treatment 1 . However, praziquantel has been in use for more than 20 years 2 , and concern is increasing the resistance has emerged in human parasites [3][4][5][6][7][8][9][10] . The presence of a gene or genes that confer resistance to PZQ cannot be excluded [11][12] .…”

“…Low efficacy of praziquantel was reported by many authors in several clinical trials in many countries including Egypt 3,[13][14][15] . Laboratory isolation of schistosome strains resistant to standard and high doses of the drug was reported 7,[11][12]16 . The role of praziquantel cannot be ignored as a possible etiological factor implicated in the process of carcinogenesis associated with schistosomiasis 17 .…”