Production of amastigotes from metacyclic trypomastigotes of Trypanosoma cruzi

Attempts to recreate all the developmental stages of Trypanosoma cruzi in vitro have thus far been met with partial success. It is possible, for instance, to produce trypomastigotes in tissue culture and to obtain metacyclic trypomastigotes in axenic conditions. Even though T. cruzi amastigotes are known to differentiate from trypomastigotes and metacyclic trypomastigotes, it has only been possible to generate amastigotes in vitro from the tissue-culture-derived trypomastigotes. The factors and culture conditions required to trigger the transformation of metacyclic trypomastigotes into amastigotes are as yet undetermined. We show here that pre-incubation of metacyclic trypomastigotes in culture (MEMTAU) medium at 37ºC for 48 h is sufficient to commit the parasites to the transformation process. After 72 h of incubation in fresh MEMTAU medium, 90 percent of the metacyclic parasites differentiate into forms that are morphologically indistinguishable from normal amastigotes. SDS-PAGE, Western blot and PAABS analyses indicate that the transformation of axenic metacyclic trypomastigotes to amastigotes is associated with protein, glycoprotein and antigenic modifications. These data suggest that (a) T. cruzi amastigotes can be obtained axenically in large amounts from metacyclic trypomastigotes, and (b) the amastigotes thus obtained are morphological, biological and antigenically similar to intracellular amastigotes. Consequently, this experimental system may facilitate a direct, in vitro assessment of the mechanisms that enable T. cruzi metacyclic trypomastigotes to transform into amastigotes in the cells of mammalian hosts

Early and late molecular and morphologic changes that occur during the in vitro transformation of Trpanosoma cruzi metacyclic trypomastigotes to amastigotes

The amastigogenesis primary of T. cruzi occurs naturally when metacyclic trypomastigotes transform into amastigotes within the cells of the mammalian host. The in vitro study of the macromolecular changes that occur over several days during the transformation process should provide significant indications of how the parasite adapts to the mammalian host environment. We show here that metacyclic trypomastigotes pre-incubated at 37 degrees C in a protein-rich medium reach a high degree of transformation to amastigotes when re-incubated in the fresh medium. Giemsa-stained smears show that during the pre-incubation phase, the metacyclic trypomastigotes undergo lengthening at the posterior end and a thinning out of the entire body. SDS-PAGE analysis of polypeptides and glycopeptides or Western blot with stage-specific antisera analyses indicate that the in vitro primary amastigogenesis is associated with abrupt changes in protein, glycoprotein, and stage-specific antigens that occur simultaneously during the first 24 hours of pre-incubation. Since the differentiating system consists of a rich media at 37 degrees C, temperature and medium constitution must trigger a macromolecular differentiation to amastigotes that precedes the morphological transformation by several days. This transformation is associated with the rearrangement of stage-specific antigens and takes place when the culture medium is changed.

Comparación de los niveles de leptina sérica en las diferentes fases del ciclo mestrual de mujeres normopeso con los de mujeres obesas de la Escuela de Bioánalis de la Universidad de Carabobo / Comparison of the levels of serum peptin in the various phases of the mestrual cycle in normal weight and obese women from the School of Medical Technology at the University of Carabobo

La leptina es hormona que se sintetiza en el tejido adiposo, en la placenta y en el tracto gastroinstestinal, mantiene una relación con el eje hipotalámico-pituitario-ovárico y participa en la regulación del peso corporal. El presente estudio tuvo como objetivo comparar los niveles de leptina sérica (por RIA) de diez (10) mujeres normopeso (IMC: 20,1 ± 1,19 kg/m²) con los de (8) mujeres obesas (IMC: 30,7 ± 2,94 kg/m²), para conocer si en ambos grupos la leptina sigue el mismo patrón de fluctuaciones en el ciclo mestrual, encontrándose en las mujeres normopeso niveles de leptina más bajos en la fase folicular (12,38 ± 4,39 S), intermedios en la mitad del ciclo (15,27 ± 7,68 S) y m s altos en la fase lúctea (17,33 ± 6,79 S). Hubo diferencia significativa entre los valores de leptina en el ciclo mestrual de los grupos, siendo en las obesas más altos y no presentaron el mismo patrón de fluctuaciones de las normopeso, pues se encontraron valores intermedios en la fase folicular (53,97 ± 30,97 S), más altos en la fase media (56,75 ± 27,25 S) y más bajos en la fase lúctea (52,86 ± 22,18 S), además, hubo correlación positiva entre el IMC y los niveles de leptina, siendo mayor en mujeres obesas. Se concluye que posiblemente los valores de leptina están relacionados con la cantidad de tejido graso y que las diferencias en el patrón de fluctuaciones en el ciclo mestrual de obesas y normopeso se deba a que los requerimientos energéticos de cada grupo son diferentes