Computational protocol: Thymosin beta-4 regulates activation of hepatic stellate cells via hedgehog signaling

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[…] Cell proliferation was measured with a CellTiter Proliferation Assay (Promega, Madison, WI, USA) as described in previous report. Briefly, cells were plated at a density of 1 × 104 cells per well in 96-well plates with the indicated treatment and time. After adding the MTS reagent, the plates were incubated in a CO2 incubator at 37 °C until the color developed. Absorbance was then measured at a wavelength of 490 nm using a Glomax multi-detection system (Promega). Standard wound healing assays were performed by growing cells to a confluent monolayer, and making a manual scratch using a 200 μL pipette tip. Then, floating cells were washed out and fresh medium was added. As the incubation time passed, the width of scratch was narrow and it was recorded by taking photographs (×20) using an Olympus inverted microscope (Olympus Optical Co., Ltd.) from 24, 36 and 48 hours after the scratch. Empty area in each time point was quantified with NIH image J version 1.49 analysis software (Rasband, W.S., ImageJ, U.S. National Institutes of Health, Bethesda, Maryland, USA, http://imagej.nih.gov/ij/, 1997–2012) and compared with that in the initiation of cell migration. [...] Results are expressed as the mean ± s.e.m. Statistical significances were determined by the unpaired two-sample Student’s t-test. Differences were considered as significant when P-values are <0.05. Statistical analyses were performed using IBM SPSS Statistics 21 software (Release version 21.0.0.0, IBM Corp., Armonk, NY, USA). […]

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