Therapeutic potential of hyaluronic acid/chitosan nanoparticles for the delivery of curcuminoid in knee osteoarthritis and an in vitro evaluation in chondrocytes.

Knee osteoarthritis (OA) is the main cause of leg pain in middle­aged and elderly individuals. Hyaluronic acid (HA), as well as curcuminoid, has been used in the treatment of knee OA. In the present study, HA/chitosan nanoparticles (CNPs) were prepared for the delivery of curcuminoid, in order to investigate whether HA and curcuminoid can act synergistically as a better treatment option. The knee OA model was established by the Hulth method, and a knee OA chondrocyte model was constructed by the co­induction of interleukin­1ß and tumor necrosis factor (TNF)­α. The drug loading capacity of HA/CNP for the delivery of curcuminoid was measured by an ultraviolet assay, and the cytotoxicity to chondrocytes was measured by an MTT assay. Collagen II was detected by immunofluorescence, and the expression levels of nuclear factor (NF)­κB and inflammation­related genes in cartilage tissue and chondrocytes were detected. Chondrocyte proliferation was determined by an EdU assay, and chondrocyte apoptosis was determined by flow cytometry. The Mankin pathological score of the Outerbridge classification was obtained. The results demonstrated that the optimum drug loading capacity of HA/CNP for the delivery of curcuminoid was 38.44%, with a good sustained release function. HA/CNP treatment resulted in inhibition of the NF­κB pathway, as well as the expression of matrix metalloproteinase (MMP)­1 and MMP­13, but it increased collagen II expression. HA/CNP for the delivery of curcuminoid significantly decreased the Outerbridge classification and Mankin pathological scores to close to normal until the 4th week. Furthermore, it was also observed that all the effects of HA/CNP on the delivery of curcuminoid were more prominent compared with the effects of HA or curcuminoid treatment individually. Taken together, these findings demonstrated that HA/CNP for the delivery of curcuminoid may suppress inflammation and chondrocyte apoptosis in knee OA via repression of the NF­κB pathway.

[Differential proteomic analysis and function study of human prostate carcinoma cells with different osseous metastatic tendency].

OBJECTIVE: To investigate the differential protein expression profiles of human prostatic carcinoma cells with different metastatic tendency and to screen the osseous metastasis associated proteins and investigate their function. METHODS: Proteomics and Western blotting were applied to screen and identify the differentially expressed proteins in the prostatic carcinoma cells of different lines: line T3B with high osseous metastasis potential, line P2-4 with high lymphatic metastasis potential, and their common parent cell line PC-3. The eukaryotic expression vector carrying human Pgenesil-1/HMGB1 siRNA was constructed and transfected into T3B cells by Lipofectamine 2000 and the positive clones was screened by G418. Pgenesil-1/HMGB1 siRNA/T3B, Pgenesil-1/T3B, and T3B cells were inoculated into the left ventricles of nude mice. Twelve weeks later the mice were killed. The number of osseous metastatic nodules and osseous metastasis inhibition rate were calculated. The mice metastatic tumor cells were identified by immunohistochemistry. RESULT: Six differential expressed proteins, correlated with cytoskeleton, transcriptional control, cellular metabolism, and phosphorylation were identified by proteomics and Western blotting. The recombinant plasmid Pgenesil-1/HMGB1 siRNA was successfully constructed. The HMGB1 expression of the T3B cells transfected with Pgenesil-1/HMGB1 siRNA was significantly lower than those of the other 2 groups (both P <0.05). The number of osseous metastatic nodules of the mice inoculated with Pgenesil-1/HMGB1 siRNA/T3B was significantly less than those of the other 2 groups (both P < 0.05). The metastatic osseous tumor cells were identified as the human prostatic carcinoma cells. CONCLUSION: Osseous metastasis associated proteins exist in prostatic carcinoma cells of the line with high osseous metastasis potential. HMGB1 is closely related to the osseous metastasis of human prostatic carcinoma cells. siRNA targeting HMGB1 specifically suppresses the expression of HMGB1 gene in the human prostatic carcinoma cells with high osseous metastasis potential and effectively inhibits the osseous metastasis.