Abstract
The influence of L-carnitine in vitro on the lysosomal cysteine proteinase activity and stability of the lysosomal membrane of the liver homogenates of intact sexually Mature female rats of Wistar line weighing 280-330 g were studied. In the experimental groups isolated lysosomes were incubated in vitro in a solution of L-carnitine during 1, 2 and 4 hours, in the control groups in vitro incubation was carried out in a medium of isolating solution. The activity of ca-thepsins B, L and H was investigated by spectrofluorimetric method of Barrett & Kirschke in two fractions - lysosomal and outside of lysosomes. The activity of acid phosphatase was used as the main marker of a membrane labilization. In vitro incubation of lysosomes showed that carnitine at a concentration of 5 mM increases the total activity of cathepsin B in a one-hour incubation at 73,2% (p=0,008), cathepsin L in a two- and four-hour incubation - at 77,7% (p=0,005) and 42,3% (p=0,013) respectively, and reduces the overall activity of the cathepsin H in a one-hour incubation at 200,0% (p=0,008), in a two-hour - by 67,9% (p=0,05), in a four-hour -27,1% (p=0,02). In addition, incubation in 5 mM L-carnitine solution leads to an increase of unsedimentable activity and fall sedimentaries activity for cathepsin L in a two-hour, and for acid phosphatase - in a two - and four-hour exposure. 5 mM L-carnitine in one - and two-hour incubation stabilizes lysosomal membrane (whereas increase in incubation time up to 4 hours leads to its damage) and increases the selective permeability of the lysosomal membrane for the studied cathepsins, to the greatest extent - for cathepsin H.
Highlights
The influence of L-carnitine in vitro on the lysosomal cysteine proteinase activity and stability of the lysosomal membrane of the liver homogenates of intact sexually Mature female rats of Wistar line weighing 280-330 g were studied
In vitro incubation of lysosomes showed that carnitine at a concentration of 5 mМ increases the total activity of cathepsin B in a one-hour incubation at 73,2% (p=0,008), cathepsin L in a two- and four-hour incubation – at 77,7% (p=0,005)
Incubation in 5 mM L-carnitine solution leads to an increase of unsedimentable activity and fall sedimentaries activity for cathepsin L in a two-hour, and for acid phosphatase – in a two – and four-hour exposure. 5 mM L-carnitine in one – and two-hour incubation stabilizes lysosomal membrane and increases the selective permeability of the lysosomal membrane for the studied cathepsins, to the greatest extent – for cathepsin H
Summary
Активность катепсинов В, L и Н изучалась спектрофлуориметрическим методом по Barrett & Kirschke в двух фракциях – лизосомальной и внелизосомальной. При in vitroинкубации лизосом было обнаружено, что L-карнитин в концентрации 5 мМ увеличивает общую активность катепсина B при одночасовой инкубации на 73,2% (р=0,008), катепсина L при двух- и четырехчасовой инкубации – на 77,7% (р=0,005) и 42,3% (р=0,013) соответственно, а также снижает общую активность катепсина Н при одночасовой инкубации на 200,0% (р=0,008), при двухчасовой – на 67,9% (р=0,05), при четырехчасовой – на 27,1% (р=0,02). Инкубация в 5 мМ растворе L-карнитина приводит к росту неседиментируемой активности и падению седиментируемой активности для катепсина L при двухчасовом, а для кислой фосфатазы – при двух- и четырехчасовом воздействии. Также 5 мМ L-карнитин при одно- и двухчасовой инкубации стабилизирует лизосомальную мембрану, тогда как увеличение времени инкубации до 4-х часов приводит к её повреждению, и повышает избирательную проницаемость лизосомальной мембраны для исследуемых катепсинов, в наибольшей степени – для катепсина Н. Ключевые слова: В, L, Н катепсины, L-карнитин, стабильность лизосомальной мембраны
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