MATERIALS AND METHODS: Cell viability assay using MTT, DNA fragmentation assay and real-time PCR were used to evaluate the cytotoxic effects of latex whole C-serum and its subfractions on the cell lines.
RESULTS: MTT assay revealed very low LC(50) values, 2.0 and 280 ng/ml, for DCS and DCP treatments, respectively. DCS was proven to be more potent compared to DCP, in conferring specific anti-proliferative effects on the cancer cell lines. The study also indicated that anti-proliferative activity of pre-heated C-serum fractions diminished significantly.
CONCLUSION: Although noteworthy cell death was reported, DNA fragmentation assay and real-time PCR confirmed that that induced by latex C-serum subfractions was not promoted via the classical apoptotic signalling pathway.
MATERIALS AND METHODS: The PBMC (n = 7) were cultured either in RPMI-1640 containing L-glutamine and 50 units/ml Penicillin-Streptomycin (BM) or in BM with either AuHS or FBS. Viability, proliferation and differentiation of PBMC were evaluated. Paracrine factors present in the secretomes (n = 6) were analysed using ProcartaPlex Human Cytokine panel (17 plex). Ingenuity Pathway Analysis (IPA) was performed to predict activation or inhibition of biological functions related to tissue regeneration.
RESULTS: The viability of PBMC that were cultured with FBS supplement was significantly reduced at 96 h compared to those at 0 and 24 h (P
OBJECTIVES: This study has aimed to establish optimum conditions to generate and characterize MSC from human umbilical cord (UC-MSC).
MATERIALS AND METHODS: To optimize MSC population growth, basic fibroblast growth factor (bFGF) was utilized in culture media. Effects of bFGF on expansion kinetics, cell cycle, survival of UC-MSC, cytokine secretion, expression of early stem-cell markers and immunomodulation were investigated.
RESULTS: bFGF supplementation profoundly enhanced UC-MSC proliferation by reducing population doubling time without altering immunophenotype and immunomodulatory function of UC-MSC. However, cell cycle studies revealed that bFGF drove the cells into the cell cycle, as a higher proportion of cells resided in S phase and progressed into M phase. Consistent with this, bFGF was shown to promote expression of cyclin D proteins and their relevant kinases to drive UC-MSC to transverse cell cycle check points, thus, committing the cells to DNA synthesis. Furthermore, supplementation with bFGF changed the cytokine profiles of the cells and reduced their apoptotic level.
CONCLUSION: Our study showed that bFGF supplementation of UC-MSC culture enhanced the cells' growth kinetics without compromising their nature.
MATERIALS AND METHODS: Diabetic ADSCs were treated with DFO and compared to normal and non-treated diabetic ADSCs for expression of HIF-1α, VEGF, FGF-2 and SDF-1, at mRNA and protein levels, using qRT-PCR, western blotting and ELISA assay. Activity of matrix metalloproteinases -2 and -9 were measured using a gelatin zymography assay. Angiogenic potential of conditioned media derived from normal, DFO-treated and non-treated diabetic ADSCs were determined by in vitro (in HUVECs) and in vivo experiments including scratch assay, three-dimensional tube formation testing and surgical wound healing models.
RESULTS: DFO remarkably enhanced expression of noted genes by mRNA and protein levels and restored activity of matrix metalloproteinases -2 and -9. Compromised angiogenic potential of conditioned medium derived from diabetic ADSCs was restored by DFO both in vitro and in vivo experiments.
CONCLUSION: DFO preconditioning restored neovascularization potential of ADSCs derived from diabetic rats by affecting the HIF-1α pathway.
MATERIALS AND METHODS: Adipose stromal cells were cultured in three different types of medium: (i) F12/DMEM (FD) supplemented with 10% FBS from passage 0 (P0) to P6; (ii) FD supplemented with 2% FBS at P6; and (iii) FD supplemented with 2% FBS plus 50 ng/ml of VEGF at P6. Morphological changes and growth rate of ASCs were recorded. Changes in stemness, angiogenic and endogenic genes' expressions were analysed using Real-Time PCR.
RESULTS: Adipose stromal cells changed from fibroblast-like shape when cultured in 10% FBS medium to polygonal when cultured in 2% FBS plus VEGF-supplemented medium. Their growth rate was lower in 2% FBS medium, but increased with addition of VEGF. Real-Time PCR showed that ASCs maintained most of their stemness and angiogenic genes' expression in 10% FBS at P1, P5 and P6, but this increased significantly in 2% FBS at P6. Endogenic genes expression such as PECAM-1, VE chaderin and VEGFR-2 decreased after serial passage in 10% FBS, but increased significantly at P6 in 2% FBS. Addition of VEGF did not cause any significant change in gene expression level.
CONCLUSION: Adipose stromal cells had greater angiogenic potential when cultured in reduced serum conditions. VEGF did not enhance their angiogenic potential in 2% FBS-supplemented medium.
MATERIALS AND METHODS: Anti-cancer activity of a tocotrienol-rich fraction (TRF) and a tocotrienol-enriched fraction (TEF) isolated from palm oil, as well as pure vitamin E analogues (α-tocopherol, α-, δ- and γ-tocotrienols) were studied using highly aggressive triple negative MDA-MB-231 cells and oestrogen-dependent MCF-7 cells, both of human breast cancer cell lines. Cell population growth was evaluated using a Coulter particle counter. Cell death mechanism, poly(ADP-ribose) polymerase cleavage and levels of NF-κB were determined using commercial ELISA kits.
RESULTS: Tocotrienols exerted potent anti-proliferative effects on both types of cell by inducing apoptosis, the underlying mechanism of cell death being ascertained using respective IC50 concentrations of all test compounds. There was marked induction of apoptosis in both cell lines by tocotrienols compared to treatment with Paclitaxel, which was used as positive control. This activity was found to be associated with cleavage of poly(ADP-ribose) polymerase (a DNA repair protein), demonstrating involvement of the apoptotic cell death signalling pathway. Tocotrienols also inhibited expression of nuclear factor kappa-B (NF-κB), which in turn can increase sensitivity of cancer cells to apoptosis.
CONCLUSION: Tocotrienols induced anti-proliferative and apoptotic effects in association with DNA fragmentation, poly(ADP-ribose) polymerase cleavage and NF-κB inhibition in the two human breast cancer cell lines.