MRI Arthrograms Diagnostic Radiology (Outpatient) Welcome: We would like to welcome you to OSF St Joseph Medical Center Diagnostic Radiology Department. You have been scheduled to have an MRI Arthrogram. This is a diagnostic test using fluoroscopy (a form of x-ray). The information contained in this pamphlet will help you understand the test that has been scheduled. Wh
Dell'aglim. pontignano 2002VI Seminario Nazionale per Dottorandi in Farmacologia e Scienze Affini
Siena, Certosa di Pontignano, 23 - 26 Settembre 2002
IN VITRO STUDIES ON THE MECHANISM OF ACTION OF TWO COMPOUNDS WITH
ANTIPLASMODIAL ACTIVITY: ELLAGIC ACID AND 3,4,5-TRIMETHOXYPHENYL-(6’-O-
Dell’Agli M., 2° anno di corso del Dottorato in Scienze Farmacotossicologiche, Farmacognostiche e
Biotecnologie Farmacologiche, XVI ciclo. Durata del dottorato in anni: 4. Sede di servizio: Dipartimento di
Scienze Farmacologiche, Università di Milano.
Malaria remains a major problem of public health worldwide mainly due to the increase in parasite resistance
to available drugs. The identification of new biochemical targets and the search for new antimalarial
compounds is essential (1). Several drug target can be identified in the metabolism of intraerythrocytic
parasites: one is the proteolytic cleavage or haemoglobin and another one is the detoxification of haem
leading to the formation of malaria pigment or haemozoin (1,2). Proteinases are essential to a variety of
viruses and other pathogenic microrganisms, including malaria parasites (1). In particular Plasmodium
falciparum degrades haemoglobin in a digestive vacuole (DV) by means of aspartic proteinases (plasmepsin
I-X) and one cysteine proteinase (falcipain) . Plasmepsins I and II appear to initiate cleavage of the
haemoglobin tetramers, while falcipain digests the fragmented substrate (3).
It was previously shown that the methanolic extract of the bark of Tristaniopsis calobuxus Brongniart & Gris
(Myrtaceae) inhibited parasite growth in vitro. From the extracts, two compounds with in vitro
antiplasmodial activity were isolated and identified as ellagic acid and 3,4,5-trimethoxyphenyl-(6’-O-
galloyl)-O-β-D-glucopyranoside (TMPGG) (4).
The present study was performed to investigate the mechanism of action of ellagic acid and TMPGG. Two
hypotheses were taken into consideration: first, we analysed the ability of ellagic acid to interfere with the
formation of β-haematin in vitro, using a spectrophotometric microtitre-based method recently developed (5).
Secondly, since T. calobuxus bark extract was shown to inhibit other proteases (6), we investigated whether
the compounds could inhibit plasmepsins, using recombinant plasmepsin II as target (7). Ellagic acid
inhibited β-haematin formation in a dose-dependent manner. At the highest dose ellagic acid inhibited β-
haematin formation by 65.6% whereas the inhibition by the reference compound chloroquine was complete.
The IC50s, namely the molar equivalent of test compound able to inhibit β-haematin formation by 50%, were
1.67 ± 0.81 and 5.42 ± 1.3 for chloroquine and ellagic acid, respectively. When tested on recombinant
plasmepsin II, both ellagic acid and TMPGG inhibited the enzyme in a concentration dependent manner:
IC50s were 4.02 ±drslt 1.26 and 35 ± 8.74 µM for ellagic acid and TMPGG, respectively (mean ± sd). These
doses are 10 fold higher than those needed to inhibit the parasite growth in vitro (0.5 µM and 3.2 µM for
ellagic acid and TMPGG, respectively) (4). The potency ratio between the two compounds remained similar,
equal to 10.
The further step of this study will be to investigate the mode of enzyme inhibition at the molecular level.
HPLC-MS of the protein will be performed to obtain data as regards the binding of the inhibitor to the
protein. Preliminary results have been obtained by analysis of the recombinant proenzyme and its activated
form by HPLC-MS before and after trypsin digestion.
The molecular weight of proplasmepsin II was 43580 Da, which is very close to that calculated on the basis
of the amminoacid sequence (43583 Da)(8). After 1 and 3 hours of activation, at pH 4,7 we obtained two
forms of the activated protein, one with molecular weight 38185 Da and the other of 37093 Da. The
quantitative proportion of the two forms was dependent of the time of activation.
These experiments will be repeated after incubation of the protein with ellagic acid.
1. Olliaro, P. L., Gottlieb, M., and Wirth, D. F., 1996. Parasitol. Today 12, 413-414.
2. Olliaro, P. L., and Goldberg, D. E., 1995. Parasitol. Today 11, 294-297.
3. Coombs, G.H., Goldberg, D.E., Klemba, M., Berry, C., Kay, J and Mottram, J.C. Trends Parasitol.
4. Verotta, L., Dell'Agli, M., Giolito, A., Guerrini, M., Cabalion, P., and Bosisio, E., 2001. J. Nat. Prod. 64, 5. Parapini, S., Basilico, N., Pasini, E., Egan, T. J., Olliaro, P., Taramelli, D., and Monti, D., 2000. Exp. 6. Bosisio, E., Mascetti, D., and Cabalion, P., 2000. Pharm. Biol. 38, 18-24. 7. Moon, R. P., Tyas, L., Certa, U., Rupp, K., Bur, D., Jaquet, C., Matile, H., Loetscher, H., Grueninger- Leitch, F., Kay, J., Dunn, B. M., Berry, C., and Ridley, R. G., 1997. Eur. J. Biochem. 244, 552-560. 8. Berry, C., Humphreys, J.M., Matharu, P., Granger, R., Horrocks, P., Moon, R.P., Certa, U., 9. Ridley, R.G., Bur, D. and Kay, J., 1999, FEBS Letters, 149-154.
European Journal of Neurology 2011, 18: 1115–1121E F N S G U I D E L I N E S / C M E A R T I C L ETreatment of medication overuse headache – guideline of theEFNS headache panelS. Eversa and R. JensenbaDepartment of Neurology, University of Mu¨nster, Mu¨nster, Germany; and bDanish Headache Center, Department of Neurology, GlostrupHospital, University of Copenhagen, Copenhagen, DenmarkBack