Study Of The Production Of MWNTs-PEG Loaded With LBP And The Effect Of MWNTs-peg To HepG2 Cells

Objective:To research the penetrability in hepatocellular carcinoma Hep G2 cells and the inhibition in cells proliferation of walled carbon nanotubes targeting drug delivery system with drug-loaded function.Methods:1.Oxidating and acylating the multiwalled carbon nanotube to get a modified one which was loaded to PEG after acylation to get a functional multiwalled carbon nanotube. Then represent the morphology and size of MWCNTs-PEG.2.Labeling the MWCNT-PEG with FITC by π-π bond adsorption. Calcuate the drug loading capacity and test the fluorescence intensity in solution with the method of fluorescence spectrophotometry. According to the fluorescence intensity to ascertain the leakage degree of FITC.3.Loading IV, the precursor of LBP which is a common anti-cancer drug,to animated MWCNT-PEG through π-π bond to make LBP-MWCNT-PEG. Test the in vitro release of the complex and draw out its release curve.4.Testing the intake of Hep G2 cells cultured in vitro to MWCNT-PEG labeled by FITC and the average fluorescence intensity of FITC in cells through LSCM.5.Putting LBP-MWCNT-PEG into DMEM, controlled with LBP, and intervening the Hep G2 cells for 24 h,48h and 72 h separately. Inhibition of the complex to Hep G2 cells proliferation is assessed by MTT assay. And compare the different inhibition effects along with the change of time.Results:1.Viewing through the TEM, before and after reaction there is no obviously change in MWCNT, and the morphological structure of wall can be clearly observed,which can be proved that the wall structure is not destructed.Water-soluble appearance evenly distributed in light grey. The particle size after rehydration is about 165 nm,and the change is unobviously in two weeks.2.Through the calculation of drug loadings of FITC is 43.2%. According to FITC-MWCNT-PEG leakage test the complex leakage rate will not more than 2.0%in PBS(PH 7.4) within 36 h.3.Through the calculation of drug loadings of LBP is 43.2%. According to in vitro release test the complex leakage rate is more than 90% in PBS(PH5.0) within12 d and less than 10% in PBS(PH9.0). It shows that LBP-MWCNT-PEG is easily released in acidic PH solution,slowly released in neutral PH solution and hardly released in alkaline PH solution which proved that LBP-MWCNT-PEG is sensitive to acid.4.The greenish and bluish fluorescence are overlaped in LSCM which testifies taht MWCNT-PEG can get into cells. And the mean intensity of fluorescence is743.41±121.36.5.MTT assay shows that the intensity of inhibition to proliferation of Hep G2 cells in LBP group is higher than in LBP-MWCNT-PEG group while the inhibition of LBP-MWCNT-PEG is improved with time increasing.The intensity of inhibition to proliferation in LBP-MWCNT-PEG has already higher than LBP after 48 h drug action with drug concentration in 50ug/ml.72 h later, LBP-MWCNT-PEG inhibition to proliferation further increased while LBP’s is not significantly.Conclusions:1.MWCNT modified by PEG is well soluble and stable. In the foundation of MWCNT-PEG,FITC-MWCNT-PEG label probe is made.After loading to LBP the antitumor drug,drug loading model LBP-MWCNT-PEG is successfully made.The complex made in this method owns stability and PH targeting property which can beput into further research in cellular level.2.MWCNT-PEG labeled by FITC can enter cells is proved in the cellular uptake test. And it also shows that the MWCNT-PEG is with great cell penetrability.3.LBP-MWCNT-PEG has the inhibition to proliferation of Hep G2 cells which proves that it is targeting and has sustained release effects.