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Hs27 and HUVEC cells were grown in 60 mm dishes and HUES cells in 24-well plates.
To evaluate the nominal concentration-effect relationship for the cytotoxic action of [H.sub.2][O.sub.2], human somatic cells (Hs27 and HUVEC) and embryonic stem (HUES3 and HUES7) cell lines were exposed to rising concentrations of [H.sub.2][O.sub.2] between 4 and 768 [micro]M during 72 h and cell viability was analysed by AlamarBlue[R] reduction and normalized to the nontreated control samples of each cell line (Figure 1(a)).
To determine the AGE status of the cells under oxidative stress conditions, we quantified specific protein-bound AGEs by slot blot analysis in three independent experiments of treated HUES3 and Hs27 cell lines, with specific antibodies against pentosidine, argpypyrimidine, and [N.sup.[epsilon]]-carboxymethyl-lysine (CML).
To confirm if the RAGE levels decrease during differentiation in basal conditions, we next examined the mRNA RAGE levels by quantitative real-time PCR in ES cell lines, in their differentiated progenies and on the somatic cell lines Hs27 and HUVEC.
U-2 OS (HTB-96) human osteosarcoma, MCF-7 (HTB-22) human breast adenocarcinoma, HeLa (CCL-2) human cervical cancer, Ca Ski (CRL-1550) human cervical cancer, and Hs27 (CRL 1634) human foreskin fibroblast cell lines were all obtained from the American Type Culture Collection (ATCC), USA.
MCF-7, HeLa, and Hs27 cells were maintained in DMEM.
The cell lines used for this study are Hs27, U-2 OS, MCF7, HeLa, and Ca Ski.
However, in both the human cervical cancer cell lines tested, Ca Ski and HeLa, as well as on Hs27 cell line, 40 nm nanoparticles were shown to be more toxic compared to 20 nm sized nanoparticles as shown in Figures 8, 9 and 10, respectively.
Hs27 human dermal fibroblasts were cultured in DMEM until reaching 85% confluence.
In our previous studies, NF3 has been demonstrated to be able to promote human skin fibroblast cell line Hs27 cell proliferation and cell cycle progression, and microarray analysis revealed two intracellualr signaling pathways, Wnt and angiogenesis, that may contribute to NF3 effects on fibroblasts (Tam et al.
Human skin fibroblast cell line Hs27 was purchased from ATCC (American Type Culture Collection, Manassas, VA, USA).
Hs27 fibroblasts cells were treated with or without NF3 at 4 mg/ml for 24 h.
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- HS1 binding protein
- HS1-associating protein X-1
- HS1-binding protein 1