scholarly journals Effects of α-difluoromethylornithine on protein synthesis and synthesis of the variant-specific glycoprotein (VSG) in Trypanosoma brucei brucei

1988 ◽  
Vol 250 (1) ◽  
pp. 295-298 ◽  
Author(s):  
A J Bitonti ◽  
D E Cross-Doersen ◽  
P P McCann
Keyword(s):  
Protein Synthesis ◽  
Trypanosoma Brucei ◽  
Mechanism Of Action ◽  
Myristic Acid ◽  
Leucine Incorporation ◽  
Biochemical Mechanism ◽  
Trypanosoma Brucei Brucei ◽  
Apparent Decrease

Protein synthesis in Trypanosoma brucei brucei was rapidly inhibited during polyamine depletion by DL-alpha-difluoromethylornithine (DFMO) in vitro and in vivo. [3H]Leucine incorporation was depressed 30-40% by 24 h and 80-90% by 48 h of DFMO treatment. Concomitantly there was an apparent decrease in the synthesis of the variant-specific glycoprotein (VSG) in DFMO-treated trypanosomes, as measured by decreased incorporation of [3H]myristic acid into VSG. The discovery of decreased protein synthesis in T. b. brucei during DFMO treatment is noteworthy, because it was reported previously that protein synthesis was paradoxically stimulated 2-4-fold during DFMO treatment in these organisms. Decreased protein synthesis probably relates to the biochemical mechanism of action of DFMO on trypanosomes.

scholarly journals Cross-Resistance to Nitro Drugs and Implications for Treatment of Human African Trypanosomiasis

2010 ◽  
Vol 54 (7) ◽  
pp. 2893-2900 ◽  
Author(s):  
Antoaneta Y. Sokolova ◽  
Susan Wyllie ◽  
Stephen Patterson ◽  
Sandra L. Oza ◽  
Kevin D. Read ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Human African Trypanosomiasis ◽  
Parent Drug ◽  
Cross Resistance ◽  
Pharmacokinetic Studies ◽  
African Trypanosomiasis ◽  
Trypanosoma Brucei Brucei ◽  
Resistant Lines

ABSTRACT The success of nifurtimox-eflornithine combination therapy (NECT) for the treatment of human African trypanosomiasis (HAT) has renewed interest in the potential of nitro drugs as chemotherapeutics. In order to study the implications of the more widespread use of nitro drugs against these parasites, we examined the in vivo and in vitro resistance potentials of nifurtimox and fexinidazole and its metabolites. Following selection in vitro by exposure to increasing concentrations of nifurtimox, Trypanosoma brucei brucei nifurtimox-resistant clones designated NfxR1 and NfxR2 were generated. Both cell lines were found to be 8-fold less sensitive to nifurtimox than parental cells and demonstrated cross-resistance to a number of other nitro drugs, most notably the clinical trial candidate fexinidazole (∼27-fold more resistant than parental cells). Studies of mice confirmed that the generation of nifurtimox resistance in these parasites did not compromise virulence, and NfxR1 remained resistant to both nifurtimox and fexinidazole in vivo. In the case of fexinidazole, drug metabolism and pharmacokinetic studies indicate that the parent drug is rapidly metabolized to the sulfoxide and sulfone form of this compound. These metabolites retained trypanocidal activity but were less effective in nifurtimox-resistant lines. Significantly, trypanosomes selected for resistance to fexinidazole were 10-fold more resistant to nifurtimox than parental cells. This reciprocal cross-resistance has important implications for the therapeutic use of nifurtimox in a clinical setting and highlights a potential danger in the use of fexinidazole as a monotherapy.

Use of an in Vitro Protein Synthesizing System to Test the Mode of Action of Chloracetamides

Weed Science ◽  
1980 ◽  
Vol 28 (3) ◽  
pp. 334-340 ◽  
Author(s):  
Luanne M. Deal ◽  
J. T. Reeves ◽  
B. A. Larkins ◽  
F. D. Hess
Keyword(s):  
Protein Synthesis ◽  
Sand Culture ◽  
Leucine Incorporation ◽  
Insoluble Protein ◽  
In Vitro System ◽  
A Cell ◽  
Protein Incorporation ◽  
Vitro Protein

The effects of chloracetamides on protein synthesis were studied both in vivo and in vitro. Four chloracetamide herbicides, alachlor [2-chloro-2′,6′-diethyl-N-(methoxymethyl)acetanilide], metolachlor [2-chloro-N-(2-ethyl-6-methylphenyl)-N-(2-methoxy-1-methylethyl)acetamide], CDAA (N–N-diallyl-2-chloroacetamide), and propachlor (2-chloro-N-isopropylacetanilide) were tested for inhibition of [3H]-leucine incorporation into protein. Incorporation of3H-leucine into trichloroacetic acid (TCA)-insoluble protein was inhibited in oat (Avena sativaL. ‘Victory’) seedlings grown in sand culture and treated 12 h at 1 × 10−4M with these chloracetamides. The herbicides were also tested in a cell-free protein synthesizing system containing polyribosomes purified from oat root cytoplasm. These herbicides had no effect on the rates of polypeptide elongation nor on the synthesis of specific polypeptides when herbicides (1 × 10−4M) were added directly to the system. Polypeptide formation was inhibited 89% when 1 × 10−4M cycloheximide was added during translation. Cytoplasmic polyribosomes were isolated from oat roots treated 12 h with 1 × 10−4M herbicide. Translation rates and products were not altered when these polyribosomes were added to the in vitro system. Protein synthesis is inhibited when tested in an in vivo system; however, the inhibition does not occur during the translation of mRNA into protein.

scholarly journals In vitro and in vivo anti-trypanosomal potentials of Afrormosia laxiflora and Khaya senegalensis against Trypanosoma brucei brucei

2018 ◽  
Vol 39 (3) ◽  
pp. 269-284
Author(s):  
G.D. Chechet ◽  
J Yahaya ◽  
A.J. Nok
Keyword(s):  
Body Weight ◽  
Trypanosoma Brucei ◽  
Post Infection ◽  
Methanol Extract ◽  
Control Group ◽  
Phytochemical Analysis ◽  
Trypanosoma Brucei Brucei ◽  
Khaya Senegalensis

Animal African trypanosomiasis (AAT) also known as Nagana is a resurgent disease in Africa. Medicinal plants are being used in less developed countries for the treatment of various diseases including trypanosomiasis, due to the high cost of currently available drugs. Most of these plants have been useful sources of treatment of various diseases based on information obtained from folk medicine but have not been scientifically certified. Here, we investigated the in vitro and in vivo anti-trypanosomal potentials of the methanol extract of Aformorsia laxiflora and Khaya senegalensis against T. b. brucei. Phytochemical screening as well as LD50 of the plant extracts was carried out following standard procedures. Parasitemia was monitored daily while Packed Cell Volume was determined at three time points (days 1, 4 and 7) during the course of the infection. The phytochemical analysis showed the presence of saponins, alkaloids, flavonoids, antraquinones, resins and tanins. However, steriods/terpenoids were absent in K. senegalensis but present in A. laxiflora. The toxicity of methanol extract of both A. laxiflora and K. senegalensis was above 5000mg/kg body weight. Methanol extracts of A. laxiflora (leaves) and K. senegalensis (stem bark) showed promising trypanocidal potential in vitro against T. b. brucei at concentrations of 10, 15, 25mg/ml and 40 and 20mg/ml respectively. At these concentrations, both extracts immobilized the parasites within 55mins post-incubation. In general, A. laxiflora leaf extract demonstrated prophylactic activity against T. b. brucei in vivo at a dose of 500mg/Kg body weight particularly in group C animals where a delayed pre-patent period (6 days post-infection), extended survival (14 days post-infection) and significant (P<0.05) reduction in the parasite burden confirmed by an absence of anemia (PCV 47.00±0.8 %) was observed when compared to the infected untreated control group. K. senegalensis extract on the other hand did not show anti-trypanosomal activity in the treated groups (1, 2, and 3). Based on these observations, it was therefore deduced that the methanol extract of leaves of A. laxiflora possessed the ability to ameliorate the burden of the disease and could be a plausible candidate for drug development against the disease.Keywords: Trypanosoma brucei brucei, Afromosia laxiflora, Khaya senegalensis, anti-trypanosomal, in vitro, in vivo

THYROXINE STIMULATION OF AMINO ACID INCORPORATION INTO MITOCHONDRIAL PROTEIN: DIFFERENCES BETWEEN IN VIVO AND IN VITRO EFFECTS

1973 ◽  
Vol 72 (4) ◽  
pp. 684-696 ◽  
Author(s):  
Amirav Gordon ◽  
Martin I. Surks ◽  
Jack H. Oppenheimer
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Mitochondrial Protein ◽  
Leucine Incorporation ◽  
Mitochondrial Protein Synthesis ◽  
Amino Acid Incorporation ◽  
Acid Incorporation ◽  
Stimulation Of

ABSTRACT The in vivo and in vitro stimulation of rat hepatic mitochondrial protein synthesis by thyroxine (T4) was compared. In confirmation of Buchanan & Tapley (1966). T4 added to isolated mitochondria rapidly stimulated [14C] leucine incorporation into mitochondrial protein. The in vitro stimulation was reversed after T4 was removed by incubating the mitochondria with bovine serum albumin (BSA). The decrease in T4 stimulation of protein synthesis appeared proportional to the T4 removed by BSA. Thus, it appears probable that exchangeable T4 controls the in vitro system. In contrast, the increase in mitochondrial protein synthesis which was observed 3 to 4 days after pretreatment of hypothyroid rats with labelled and non-radioactive T4 was not reversed by BSA treatment. Moreover, mitochondrial radioactivity could not be extracted with albumin. The in vivo phenomenon does not, therefore, appear to be related to exchangeable hormone in the mitochondria. Furthermore, the estimated quantity of T4 associated with mitochondria after in vivo stimulation was at least two orders of magnitude less than that required to produce comparable stimulation of mitochondrial protein synthesis in vitro. These findings strongly suggest that in vitro and in vivo stimulation of amino acid incorporation by T4 may be mediated by different biochemical mechanisms.

scholarly journals An L-arginine-dependent mechanism mediates kupffer cell inhibition of hepatocyte protein synthesis in vitro

1989 ◽  
Vol 169 (4) ◽  
pp. 1467-1472 ◽  
Author(s):  
TR Billiar ◽  
RD Curran ◽  
DJ Stuehr ◽  
MA West ◽  
BG Bentz ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Peritoneal Macrophages ◽  
Metabolic Responses ◽  
Leucine Incorporation ◽  
Target Cells ◽  
Microbial Invasion ◽  
Nitrite Nitrate ◽  
Dependent Mechanism

The hepatic failure associated with severe sepsis is characterized by specific, progressive, and often irreversible defects in hepatocellular metabolism (1). Although the etiologic microbe can often be identified, the direct causes and mechanisms of the hepatocellular dysfunction are poorly understood. We have hypothesized that Kupffer cells (KC), which interact with ambient septic stimuli, respond by providing signals to adjacent hepatocytes (HC) in sepsis . Furthermore, we have provided evidence (2, 3) that KC activated by LPS from Gram-negative bacteria can induce profound changes in the function of neighboring HC in coculture. In our model, coculture of either KC (2) or peritoneal macrophages (Mφ)(3) with HC normally promotes HC protein synthesis ([(3)H]leucine incorporation). The addition of LPS or killed Escherichia colt' to such cocultures induces a profound decrease in HC protein synthesis, as well as qualitative changes ([(35)S]methionine, SDS-gel electrophoresis) in protein synthesis without inducing HC death (2, 3) . In this report we show that the inhibition in protein synthesis is mediated via an L-arginine-dependent mechanism. The metabolism of L-arginine by activated Mφ to substances with cytostatic and even lethal effects on target cells is a relatively recent discovery. After the description by Stuehr and Marletta (4, 5) that LPS- triggered Mφ produced nitrite/nitrate (NO(2)(-)/NO(3)(-)), Hibbs et al. (6, 7) and Iyengar et al. (8) demonstrated that L-arginine was the substrate for the formation of both these nitrogen end products and citrulline. A role for the arginine-dependent mechanism in Mφ tumor cytotoxicity (6, 7) and microbiostatic activity (9) has been suggested. However, the in vivo functions of this novel Mφ mechanism have not yet been defined, but it is possible that there are both physiologic as well as pathologic roles. Our in vitro results raise the possibility that some metabolic responses to microbial invasion maybe partially mediated by the L-arginine-dependent mechanism. What other metabolic responses are affected and the possible pathologic consequences remain to be studied.

scholarly journals In vitro assay and in vivo effect of artemisinin in Trypanosoma brucei brucei-infected Wistar rats

2021 ◽  
Vol 1 (3) ◽  
pp. 100061
Author(s):  
Kelvin Olutimilehin Jolayemi ◽  
Mohammed Mamman ◽  
Dahiru Sani ◽  
Magdalene Ogbonneya Okoronkwo ◽  
Abubakar Usman ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Wistar Rats ◽  
In Vitro Assay ◽  
Trypanosoma Brucei Brucei ◽  
Vitro Assay

scholarly journals In vitro and in vivo changes observed in Trypanosoma brucei brucei-infected rats treated with artesunate and/or diminazene aceturate

2021 ◽  
Vol 18 (4) ◽  
pp. 211-220 ◽  
Author(s):  
KO Jolayemi ◽  
M. Mamman ◽  
D. Sani ◽  
M.O. Okoronkwo ◽  
J. Amaje
Keyword(s):  
Trypanosoma Brucei ◽  
Wistar Rats ◽  
Standard Dose ◽  
Total Leucocyte Count ◽  
Post Inoculation ◽  
Trypanosoma Brucei Brucei ◽  
Diminazene Aceturate ◽  
Model Control

This study evaluated in vitro and in vivo antitrypanosomal effect of artesunate and/or diminazene aceturate in Wistar rats experimentally  infected with Trypanosoma brucei brucei. In vitro screening was carried out in triplicates using 50 μl of 0.2, 2 and 20 μg/μl of artesunate as test drug; diminazene aceturate, normal saline and trypanosome-infected blood served as controls in a 96-well microtitre plate, incubated at 37˚C for 5 minutes. Efficacy was observed over a period of 60 minutes for reduced or complete trypanosomal immobilization. Results showed concentration-dependent cessation of trypanosomal motility was significantly (p < 0.001) induced by artesunate when compared to the controls. Seventy Wistar rats of both sexes weighing between 190 and 210 g were randomly divided into 7 groups (5 males and 5 females) are used for in vivo study. Groups I and II served as normal control and model control respectively. Groups III to VII were infected with Trypanosoma brucei brucei (106 trypanosomes/ml) intraperitoneally. At peak parasitaemia (8 days post-infection), group III was treated with diminazene aceturate (3.5 mg/kg) intramuscularly once while groups IV, V, VI were treated with artesunate (200, 100, 50) mg/kg orally for 5 consecutive days and group VII was treated with combination of artesunate (50 mg/kg) orally and diminazene aceturate (1.75 mg/kg) intramuscularly for 5 days. Results indicated pre-patent period of 4 days and increase in levels of parasitaemia post-inoculation. PCV, Hb concentration, RBC count, MCV, MCHC and total leucocyte count decreased significantly (p < 0.05) between days 0and 8 in groups II to VII. Following treatment, significant increases (p < 0.05) were recorded except for groups II, IV, V and VI where the rats died. Thus, combination of artesunate (50 mg/kg) and half the standard dose of diminazene aceturate was able to reduce parasitaemia and ameliorate the anaemia elicited by the trypanosomes. Keywords: Artesunate, Diminazene Aceturate, Haematology, Trypanosoma brucei brucei, Wistar rats

scholarly journals Uptake of the antitrypanosomal drug 5′-{[(Z)-4-amino-2-butenyl]methylamino}-5′-deoxyadenosine (MDL 73811) by the purine transport system of Trypanosoma brucei brucei

1992 ◽  
Vol 283 (3) ◽  
pp. 755-758 ◽  
Author(s):  
T L Byers ◽  
P Casara ◽  
A J Bitonti
Keyword(s):  
Trypanosoma Brucei ◽  
Transport System ◽  
Time Course ◽  
Multidrug Resistant ◽  
Trypanosoma Brucei Brucei ◽  
Irreversible Inhibitor ◽  
Highly Active ◽  
Mouse Leukaemia

An irreversible inhibitor of S-adenosyl-L-methionine decarboxylase (AdoMetDC), 5′-([(Z)-4-amino-2-butenyl]methylamino)-5′-deoxyadenosine (MDL 73811), was found to cure Trypanosoma brucei brucei and multidrug-resistant T. b. rhodesiense infections in mice [Bitonti, Byers, Bush, Casara, Bacchi, Clarkson, McCann & Sjoerdsma (1990) Antimicrob. Agents Chemother. 34, 1485-1490]. Doses of this drug which resulted in a rapid clearance of parasites from T. b. brucei-infected rats resulted in plasma levels of 50-60 microM-MDL 73811 and an intratrypanosomal MDL 73811 concentration of 1.9 mM within 10 min of administration [Byers, Bush, McCann & Bitonti (1991) Biochem. J. 274, 527-533[. Based on this finding we speculated that MDL 73811, which is an adenosine analogue, is a substrate for the trypanosome active purine transport system. We now report evidence that supports this hypothesis. MDL 73811 uptake by T. b. brucei in vitro was time- and temperature-dependent and was saturable over a time course in which MDL 73811 metabolism was undetectable, suggesting that MDL 73811 uptake is a transport-mediated phenomenon. Inhibition of MDL 73811 uptake by purine nucleosides is consistent with the drug being a substrate for the trypanosome purine transport system. The accumulation of MDL 73811 by cultured L1210 mouse leukaemia cells was significantly less than by trypanosomes exposed to the same pharmacologically relevant concentrations of MDL 73811. Given that the half-life of MDL 73811 in the plasma of rats and mice is approx. 10 min, it seems likely that the existence of a highly active parasite transport system for MDL 73811 is crucial for the sensitivity of trypanosomes towards MDL 73811 in vivo, and that the absence of active transport of MDL 73811 by the host's cells may play a role in the selectivity of this drug.

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