Contraction speed of the actomyosin cytoskeleton in the absence of the cell membrane

The contraction of the actomyosin cytoskeleton, which is produced by the sliding of myosin II along actin filaments, drives important cellular activities such as cytokinesis and cell migration. To explain the contraction velocities observed in such physiological processes, we have studied the contraction of intact cytoskeletons of Dictyostelium discoideum cells after removing the plasma membrane using Triton X-100. The technique developed in this work allows for the quantitative measurement of contraction rates of individual cytoskeletons. The relationship of the contraction rates with forces was analyzed using three different myosins with different in vitro sliding velocities. The cytoskeletons containing these myosins were always contractile and the contraction rate was correlated with the sliding velocity of the myosins. However, the values of the contraction rate were two to three orders of magnitude slower than expected from the in vitro sliding velocities of the myosins, presumably due to internal and external resistive forces. The contraction process also depended on actin cross-linking proteins. The lack of α-actinin increased the contraction rate 2-fold and reduced the capacity of the cytoskeleton to retain internal materials, while the lack of filamin resulted in the ATP-dependent disruption of the cytoskeleton. Interestingly, the myosin-dependent contraction rate of intact contractile rings is also reportedly much slower than the in vitro sliding velocity of myosin, and is similar to the contraction rates of cytoskeletons (different by only 2–3 fold), suggesting that the contraction of intact cells and cytoskeletons is limited by common mechanisms.

Effects of physical exercise and hydration on homocysteine concentrations in physically active male adults = Efectos del ejercicio físico y la hidratación sobre las concentraciones de homocisteína en varones físicamente activos

La presente tesis analiza el efecto del ejercicio físico agudo y la hidratación sobre las concentraciones de homocisteína total (tHcy) y su relación con los parámetros implicados en el metabolismo de la homocisteína como el folato, la vitamina B12, y la creatina en una muestra de varones jóvenes físicamente activos. El trabajo se basa en los resultados del estudio realizado en la Facultad de Ciencias de la Actividad Física y del Deporte de la Universidad Politécnica de Madrid. Para el cual se contó con un total de 29 voluntarios sanos físicamente activos de la Comunidad de Madrid. Los principales resultados de esta tesis son: a) Las concentraciones de tHcy aumentaron después del ejercicio agudo tanto tras una prueba de intensidad máxima (VO2max) como una submáxima (65 % of VO2max) en varones físicamente activos independientemente de las sus concentraciones basales de tHcy. b) Las concentraciones de tHcy disminuyeron 2 h después del ejercicio físico aeróbico submáximo tras aplicar un protocolo de hidratación con una bebida para deportistas. c) Un adecuado protocolo de hidratación durante el ejercicio físico agudo previno el aumento de las concentraciones de tHcy hasta 2 h después del ejercicio. d) Las concentraciones de tHcy aumentaron a las 6 h tras la finalización del ejercicio únicamente en los test en los que no se siguió un protocolo de hidratación durante el ejercicio físico. e) A las 24 h tras el ejercicio, las concentraciones de tHcy volvieron a los niveles basales independientemente de si se aplicó un protocolo de hidratación durante el ejercicio o no. f) Es necesario aclarar si existen mecanismos subyacentes relacionados con el riesgo cardiovascular debido al aumento transitorio de las concentraciones de tHcy inducidas por el ejercicio agudo. Se necesitan más estudios que analicen la relación entre las concentraciones de tHcy después del ejercicio físico agudo y la implicación de la creatina, vitamina B12 y folato como parámetros relacionados en el metabolismo de la homocisteína. El efecto agudo del ejercicio físico aumenta las concentraciones de tHcy por encima de los valores recomendados; sin embargo, un adecuado protocolo de hidratación mantiene las concentraciones a niveles basales y previene el posterior aumento en una muestra de varones adultos físicamente activos. ABSTRACT The current thesis analyzes the effect of exercise and hydration on total homocysteine (tHcy) concentrations and the relationship with the implicated parameters, like folate, vitamin B12, and creatine in physically active male adults. The work is based on the results of the study conducted at the Faculty of Physical Activity and Sport Sciences of the Technical University of Madrid. A total of 29 physically active voluntary healthy males from the Region of Madrid were recruited. The main outcomes of this thesis are: a) tHcy concentrations increased after acute exercise with both, maximal (VO2max) and submaximal (65 % of VO2max) tests in physically active male subjects independently of their baseline tHcy status. b) After 2 h of rehydration with a sport drink, tHcy concentrations, which had previously increased during an acute exercise, decreased significantly, although they didn´t recover to baseline values. c) An adequate hydration protocol during acute aerobic submaximal exercise prevents the increase of tHcy concentrations and maintains these concentrations at baseline up to 2 h post-exercise. d) Serum tHcy concentrations increased after submaximal exercise when the hydration protocol during exercise was not applied. Furthermore, tHcy concentrations reached maximal values 6 h after the end of exercise. e) At 24 h, tHcy concentrations recovered baseline values independently whether or not there was a hydration protocol during exercise. f) There is a need to clarify the underlying mechanisms related to cardiovascular risk due to the transient increase of tHcy concentrations induced by acute exercise. Further research analayzing the relationship between tHcy concentrations after acute exercise and the implication of creatine, vitamin B12 and folate as related parameters in the homocysteine metabolism is needed. Finally, tHcy concentrations increased above the recommended values after an acute aerobic submaximal exercise; nevertheless, a good hydration protocol maintains tHcy concentrations at baseline and prevents the further increase in a sample of physically active male adults.