In vitro immunocompatibility of a novel bioartificial liver reactor material: propylene-acidamide grafted polypropylene membrane

Abstract

To evaluate the immunocompatibility of a novel bioartificial liver bioreactor material: propylene-acidamide grafted polypropylene membrane (PP-g-AAm) in vitro on peripheral blood mononuclear cells (PBMCs). Fifty milliliters of peripheral blood were collected from 30 healthy young people. PBMCs were extracted and inoculated on the 24-well culture plate preset with sterilized PP-g-AAm membrane and ungrafted polypropylene (PP) membrane. Automatic biochemical analyzer was used to detect the lactic dehydrogenase (LDH) value of the PMBCs after 2 and 6 hours. The PMBCs on these 2 kinds of membrane were cultured for 6 hours and then added with lipopolysaccharide and cultured continually. Six, twelve, and thirty-six hours later the supernatant was collected. ELISA was used to detect the values of tumor necrosis factor (TNF)alpha, interleukin (IL)-1beta, and IL-6. Flow cytometry was used to detect the expression of CD69 antigen. Scanning electron microscopy was used to observe the adhesion of PMBCs on the materials. After 2 hours' epimembranous inoculation the LDH value was 43.50 U/L +/- 12.71 U/L in the PP group, significantly higher than that in the PP-g-AAm group (29.13 U/L +/- 8.74 U/L, P = 0.008) and the newly extracted PMBC group (0 h group, 19.89 U/L +/- 4.67 U/L, P = 0.000). However, the difference between the 0 h group and PP-g-AAm group (P = 0.080) was insignificant. After 6 hours' epimembranous inoculation the LDH value was 50.25 U/L +/- 13.38 U/L in the PP group, and 32.50 U/L +/- 9.21 U/L in the PP-g-AAm group (P = 0.001), both significantly higher than that in the 0 h group (P = 0.000, and P = 0.019). There was no significant difference between the values 2 hours and 6 hours after inoculation for the two groups. No expression of TNFalpha, IL-1beta, and IL-6 was found in the supernatant of the 2 groups without LPS stimulation. Expressions of TNFalpha, IL-1beta, and IL-6 could be found at a low level 12 hours after LPS stimulation for both groups and peaked 24 hours after LPS stimulation. The expressions of TNFalpha, IL-1beta, and IL-6 were lower in the PP-g-AAm group than in the PP group (P = 0.004, P = 0.003, and P = 0.022). The TNFalpha, IL-1beta, and IL-6 levels all decreased after 36 hours. The CD69 antigen expression rates were 17.20% +/- 3.45%and 12.02% +/- 2.44% respectively in the PP group and PP-g-AAm group, both significantly higher than that of the blank control group (3.38% +/- 1.30%, both P = 0.000). SEM showed that the PMBCs adhered on the PP-Aam membrane were significantly less then those adhered on the PP membrane. And the PMBCs adhered on the PP-g-AAm membrane were smaller and with less microvilli. PP-g-AAm membrane has weaker activation capability to PMBCs and has better immunocompatibility in comparison with the PP membrane.