Experimental observation on the changes of hepatic stellate cells stimulated in vitro with tissue protein of Echinococcus multilocularis

doi:10.12140/j.issn.1000-7423.2023.04.007

Abstract

Abstract:

Objective To observe the changes in morphology and intracellular lipid droplets and ɑ-smooth muscle actin (α-SMA) expressed in the hepatic stellate cell (HSC)-LX2 stimulated with Echinococcus multilocularis tissue protein in vitro. Methods The E. multilocularis tissue was collected from infected mice under aseptic conditions, with which the tissue protein was extracted by ultrasonic-splitting. The protein extractant was added into HSC-LX2 culture in vitro at a final concentration of 0.125 mg/ml to stimulate for 24, 48, 72, 96 and 120 h, then the cells were stained with oil red O to observe the distribution of intracellular lipid droplets under light and fluorescence microscopes. The total protein of HSC-LX2 cells was extracted after stimulation for 48, 72, 96 and 120 h to detect the changes of relative expression of α-SMA using Western blotting. Results Under the light microscope, the morphology of HSC-LX2 changed gradually from polygonal to long spindle shape, and with prolonged stimulation with the E. multilocularis tissue protein, the number of intracellular lipid droplets increased, and lipid droplets distributed around the nuclei. The color of lipid droplets gradually changed from light yellow to bright red. After 120 h, almost all HSC-LX2 became long spindle-shaped. The number and volume of lipid droplets decreased, and the color changed from red to yellow. Under fluorescence microscopy, red fluorescent lipid droplets could be seen surrounding the nucleus, and the fluorescence signal did not decrease with the increase of irradiation times. Western blotting showed that the relative expression α-SMA in HSC-LX2 gradually increased with the prolongation of stimulation time, which peaked at 4.846 ± 0.741 after 96 hours and decreased to 2.420 ± 0.349 after 120 hours, but both were higher than that of the control group (t = 8.988, 7.045, both P < 0.01). Conclusion The E. multilocularis tissue protein can promote the activation of HSC-LX2 in vitro, inducing changes in cell morphology, the number and size of intracellular lipid droplets being raised first and then reduced, and the expression of intracellular α-SMA protein showing a similar changing trend as well.

Key words: Echinococcus multilocularis tissue protein, Hepatic stellate cells, Intracellular lipid droplets, α-SMA, In vitro

CLC Number:

R383.33

CAO Deping, LI Jiajing, SONG Mengwei, MO Gang. Experimental observation on the changes of hepatic stellate cells stimulated in vitro with tissue protein of Echinococcus multilocularis[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2023, 41(4): 440-445.

Figures/Tables 5

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