Isolation of Trypanosoma Cruzi from the Blood of Infected Mice by Column Chromatography

López

Tanner

et al. 1995

Parasite Immunology

A substantial proportion of human peripheral blood mononuclear cells (PBMC) manifested a decreased capacity to express membrane interferon-gamma receptors (IFN-gamma R) when co-cultured with Trypanosoma cruzi. Among the lymphocytes, B cells accounted for the bulk of this effect, evidenced by a marked drop in the proportion of CD19+ or CD20+ cells expressing IFN-gamma R. Decreased IFN-gamma R expression by B lymphocytes was seen as early as 3 h after co-culture with T. cruzi and persisted for at least 24 h. The parasite had no detectable effect on CD19, CD20 or DR antigen expression by B lymphocytes. Neither the proportion of B cells expressing these markers nor the membrane density of these molecules varied significantly in the presence of T. cruzi. In PBMC cultures stimulated with Staphlyococcus aureus Cowan I (SACI), T. cruzi decreased the percentages of both IFN-gamma R+ and IFN-R+bright (cells expressing above-normal levels of surface IFN-gamma R) B lymphocytes. Cell-free filtrates of T. cruzi suspensions reproduced the suppressive effects of living parasites on IFN-gamma R expression by B cells. When T. cruzi present, the intracellular levels of IFN-gamma R molecules in resting or SACI-activated B lymphocytes, represented by fluorescence intensity, were well below control values, suggesting that decreased surface expression resulted from suppressed IFN-gamma R synthesis. Among T (CD3+) cells, 10.8% to 39.6% (7 donors) expressed surface IFN-gamma R and did so at a very low level. These percentages were also reduced by T. cruzi.(ABSTRACT TRUNCATED AT 250 WORDS)

Section: Parasitesmentioning

confidence: 99%

Trypanosoma cruz/‐induced decrease in the level of interferon‐7 receptor expression by resting and activated human blood lymphocytes

López

Tanner

et al. 1995

Parasite Immunology

“…Trypomastigotes were purified 10 to 12 days later from the blood by centrifugation through Isolymph (Gallard-Schlesinger, Carle Place, N.Y.) (5) followed by column chromatography through DEAEcellulose (15). The eluted trypanosomes were washed twice with Dulbecco's modified minimal essential medium (GIBCO, Grand Island, N.Y.) containing 100 IU of penicillin and 100 tg of streptomycin per ml and supplemented with 10% fetal bovine serum (Sigma Chemical Co., St. Louis, * Corresponding author.…”

mentioning

confidence: 99%

Inhibition of host cell invasion and intracellular replication of Trypanosoma cruzi by N,N'-bis(benzyl)-substituted polyamine analogs

Majumder

Antimicrob Agents Chemother

1993

We studied the effects of two N,N'-bis(benzyl)-substituted polyamine analogs on the capacities of Trypanosoma cruzi to invade and multiply within a mammalian host cell. At concentrations as low as 1 ,uM, these compounds reduced significantly the infectivity of the parasite for rat heart myoblasts in a time-dependent manner. Pretreatment of virulent T. cruzi trypomastigotes, but not myoblast pretreatment, reduced the level of infectivity. The inhibitory effects started to subside 3 h after removal of the drugs and were no longer detectable after 4 h. A significant decrease in the rate of intracellular amastigote multiplication was also seen when the drugs were added to myoblast cultures which had been previously infected with untreated T. cruzi.These results show that N,N'-bis(benzyl)-substituted polyamine analogs meet the two most important criteria for potential chemotherapeutic agents against T. cruzi infection, namely, inhibition of both host cell invasion and intracellular replication by this parasite.The unicellular parasite Trypanosoma cnrzi, the causative agent of Chagas' disease, requires an intracellular localization to multiply in mammalian hosts. This stresses the importance of developing drugs that can inhibit host cell invasion and subsequent intracellular parasite replication. Such drugs are badly needed given the lack of a consistently effective, nontoxic drug to cure Chagas' disease (9). There is evidence that both host cell invasion by T. cruzi trypomastigotes and intracellular multiplication of the amastigote form are impaired when production of the polyamines agmatine and putrescine is decreased by the specific arginine decarboxylase inhibitor DL-a-dlfluoromethylarginine (7,13).Polyamines play key roles in cell growth regulation, and polyamine synthesis inhibitors have been shown to affect some parasitic organisms (1,8,16). N,N'-Di-substituted tetraamines with the general formula RNH(CH2)XNH (CH2)YNH(CH2)NINHR have been reported to suppress the growth of several pathogenic protozoans (e.g., Leishmania donovani, Plasmodium falciparum, and Plasmodium berghei) in vitro and in infected animals (2-4) and could be suitable for exploring the role that polyamines and polyamine biosynthesis plays in T. cnuzi infection. In this work, we used two N,N'-bis(benzyl)-substituted polyamine analogs and studied their effects on T. cnuzi infectivity and replication using an in vitro model system of mammalian host cell infection.Stock CD1(ICR) Swiss mice (Charles River Laboratory, Portage, Mich.) were infected subcutaneously with 106 T. cruzi (Tulahen isolate) organisms. Trypomastigotes were purified 10 to 12 days later from the blood by centrifugation through Isolymph (Gallard-Schlesinger, Carle Place, N.Y.) (5) followed by column chromatography through DEAEcellulose (15). The eluted trypanosomes were washed twice with Dulbecco's modified minimal essential medium (GIBCO, Grand Island, N.Y.) containing 100 IU of penicillin and 100 tg of streptomycin per ml and supplemented with 10% fetal bovine serum (Sigma Chemi...

“…The trypomastigotes were purified by centrifugation over Isolymph (Gallard Schlesinger, Carle Place, NY) (27) followed by chromatography through a DEAE-cellulose column (28). The eluted parasites were concentrated by centrifugation (800 x g, 15 min, 200C) and resuspended at 3-5 x 106 organisms per ml in the desired medium.…”

mentioning

confidence: 99%

Arginine decarboxylase inhibitors reduce the capacity of Trypanosoma cruzi to infect and multiply in mammalian host cells.

Proc. Natl. Acad. Sci. U.S.A.

Wirth

McCann

et al. 1987