Journal: bioRxiv

Article Title: Microsecond pulse electrical stimulation modulates cell migration

doi: 10.1101/2022.10.23.513372

Figure Lengend Snippet: (a) A graphical illustration showed the effects of μsPEF (pulse width:20 μs, frequency: 10 Hz, duration: 5 s) on the skin wound, promoting cell migration and extracellular matrix remodeling. (b) Representative time-lapse images showing the fibroblasts morphology after μsPEF (i.e., 750 and 1500 V/cm) treatment of different intensity at different time points (i.e., 0, 1 and 2 h). Detached cells are marked by yellow arrow. Scale bar, 50 μm. (c) The line graph representing the cell migration average speed per 1 h from 0 to 12 h after different intensity μsPEF (i.e., 750 and 1500 V/cm) treatment (blue circle: control; orange square: 750 V/cm; pink triangle: 1500 V/cm). Results are presented as mean ± standard deviation with 95% CI (n CTRL =42 cells, n 750 v/cm =48 cells, n 1500 v/cm =47 cells); * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 versus control by one-way ANOVA for multiple comparisons. (d) Box plot showing the average cell migration speed with different intensity μsPEF (i.e., 750 and 1500 V/cm) and control treatments in 24 h. Results are presented as mean ± standard deviation with 95% CI (n CTRL =470 cells, n 750 V/cm =979 cells, n 1500 V/cm =535 cells). (e) Effects of μsPEF on the secretion of COLA2. The level of collagen type I α2 in cellular supernatants was measured after 48 h. The concentration of COLA2 was 0.109 ± 0.018 ng/mL in the control group, 0.257 ± 0.058 ng/mL in the 750 V/cm group and 0.363 ± 0.034 ng/mL in the 1500 V/cm group. Results are presented as mean ± standard deviation with 95% CI (n=5); * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 versus control by one-way ANOVA for multiple comparisons. (f) Effects of μsPEF on the expression of FGF2. The level of FGF2 in cellular supernatants was measured after 48 h. The concentration of FGF2 was 44.139 ± 0.360 ng/mL in the control group, 48.012 ± 1.488 ng/mL in the 750 V/cm group and 48.523 ± 1.944 ng/mL in the 1500 V/cm group. Results are presented as mean ± standard deviation with 95% CI (n=3); ns=0.7329, * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 versus control by one-way ANOVA for multiple comparisons.

Article Snippet: Then, the cells were cultured in serum-free medium for 48 h. The content of type I α collagen and basic fibroblast growth factor (FGF-2) in the supernatant were measured using commercially available Human COL1A2 (Collagen Type I Alpha 2) ELISA Kit (Elabscience, Wuhan, China) and Human bFGF/FGF2 (Basic Fibroblast Growth Factor) ELISA Kit (Elabscience, Wuhan, China) according to the manufacturer’s protocol respectively.

Techniques: Migration, Control, Standard Deviation, Concentration Assay, Expressing

Journal: Communications Biology

Article Title: Selectivity matters: selective ROCK2 inhibitor ameliorates established liver fibrosis via targeting inflammation, fibrosis, and metabolism

doi: 10.1038/s42003-023-05552-0

Figure Lengend Snippet: a C57BL/6 female mice were treated with vehicle, GV101 (30, 100, and 150 mg/kg) or KD025 (150 mg/kg) by oral gavage on Week 6 after starting TAA in drinking water and continued for 6 weeks. b Hydroxyproline was assessed in the median lobe of liver collected on week 6 (normal W6 and TAA W6 with 5 mice/group) or harvest day on Week 12 (normal W9, TAA W9 and treatments with 10 mice/group). c KD025 dose was lowered to 100 mg/kg after 4 weeks of treatment due to toxicities. Representative images of Picrosirius Red (PSR) staining are shown for each group. d The levels of leptin, insulin, IL-1β and IL-17 in serum collected on Week 12 were measured by ELISA. e Liver tissue lysates were prepared in RIPA buffer and levels of pAMPK, pAkt, pCofilin, and pSTAT3 were determined by Western Blot. Graphs represent mean +/− s.e.m. Unpaired t-test statistical analysis was performed: * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001.

Article Snippet: On day 1, cells were seeded in 24-well plates at density of 50,000 cell/well in 1 ml culture medium; On day 2, cells was switched to starve EMEM containing 0.5% FBS; On day3, cells were treated with indicated concentration of ROCK inhibitors, incubated for 1 h, and then TGF-β1 (2.5 ng/mL) was applied to the cells; On day 5 (48 h after treatment), cultured medium was collected for Col1α1 ELISA (human Pro-Collagen 1α1 DuoSet ELISA kit, R&D systems), cells were collected for RNA extraction or for western analysis.

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Western Blot

Journal: Communications Biology

Article Title: Selectivity matters: selective ROCK2 inhibitor ameliorates established liver fibrosis via targeting inflammation, fibrosis, and metabolism

doi: 10.1038/s42003-023-05552-0

Figure Lengend Snippet: Human subcutaneous preadipocytes were induced adipogenesis with a medium containing differentiation cocktail (DM) for 7 days in the presence of indicated inhibitors. Differentiated cells were stained at day 8 by Oil Red O ( a ), quantification of lipid accumulation was measured by absorbance at 492 nM of extracted Oil Red O, all the absorbance was normalized to vehicle (DMSO/DM) treated differentiated cells ( a ), total RNA was extracted at day 8 and Glut4 gene expression was accessed by real-time RT-PCR ( b ). Human subcutaneous preadipocytes were treated with indicated inhibitors for 2 h before whole cell extracts were collected for Western blot analysis of phosphorylated AMPK and β-Actin ( c ). MRC-5 cells were pre-treated with various concentration of indicated inhibitors for 1 h before TGF-β1 stimulation for 48 h. Whole cell extracts were subjected to Western blot for the expression of α-SMA, pCofilin and β-actin ( d ). Western blots were quantified and normalized to the β-actin, and values are indicated under the corresponding immunoblots. Col1α levels in supernatants were measured by ELISA. All the Col1α levels were normalized with that of vehicle (DMSO/TGF-β1) treated cells ( d ), gene expression of α-SMA, CTGF, Col3α levels were measured by real-time RT-PCR. All the levels were first normalized with internal 18s control and then were normalized with the level of vehicle (DMSO/TGF-β1) treated cells ( e ). The data ( a , d ) are representative of three repeated experiments, ( b , e ) are average of three repeated experiments.

Article Snippet: On day 1, cells were seeded in 24-well plates at density of 50,000 cell/well in 1 ml culture medium; On day 2, cells was switched to starve EMEM containing 0.5% FBS; On day3, cells were treated with indicated concentration of ROCK inhibitors, incubated for 1 h, and then TGF-β1 (2.5 ng/mL) was applied to the cells; On day 5 (48 h after treatment), cultured medium was collected for Col1α1 ELISA (human Pro-Collagen 1α1 DuoSet ELISA kit, R&D systems), cells were collected for RNA extraction or for western analysis.

Techniques: Staining, Gene Expression, Quantitative RT-PCR, Western Blot, Concentration Assay, Expressing, Enzyme-linked Immunosorbent Assay, Control

Journal: Communications Biology

Article Title: Selectivity matters: selective ROCK2 inhibitor ameliorates established liver fibrosis via targeting inflammation, fibrosis, and metabolism

doi: 10.1038/s42003-023-05552-0

Figure Lengend Snippet: PBMCs were treated with indicated doses of GV101 or KD025 1.5 h before stimulation with LPS. After 24 h, TNF, IL-23, and IL-10 secretion in supernatants was analyzed by ELISA and normalized ( n = 12 except GV101 0.3 and 0.6 μM: n = 4) ( a ) and Western blots for the indicated proteins were performed on whole cell extracts ( b ). Human ( c , e ) or murine primary Kupffer cells ( d ) were treated with indicated doses of GV101 or KD025 1.5 h before stimulation with LPS and 24 h before collection of Kupffer cells supernatants for analysis of IL-6 and TNF secretion by ELISA followed by normalization ( c : n = 8 except GV101/KD025 2.5 μM, n = 5 ; d : n = 4), and preparation of human Kupffer cell extracts for Western blots analysis of the indicated proteins ( e ). All experiments represent at least 4 independent repeats. Graphs represent mean +/− s.e.m. In ( a , c , d ), one way ANOVA was performed followed by multiple comparisons Sidak tests to allow two-by-two comparisons. ns: not significant, * p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; **** p ≤ 0.0001. In ( b , e ), Western blots were quantified and normalized to the LPS-stimulated conditions, and values are indicated under the corresponding immunoblots.

Article Snippet: On day 1, cells were seeded in 24-well plates at density of 50,000 cell/well in 1 ml culture medium; On day 2, cells was switched to starve EMEM containing 0.5% FBS; On day3, cells were treated with indicated concentration of ROCK inhibitors, incubated for 1 h, and then TGF-β1 (2.5 ng/mL) was applied to the cells; On day 5 (48 h after treatment), cultured medium was collected for Col1α1 ELISA (human Pro-Collagen 1α1 DuoSet ELISA kit, R&D systems), cells were collected for RNA extraction or for western analysis.

Techniques: Enzyme-linked Immunosorbent Assay, Western Blot

Journal: International journal of cosmetic science

Article Title: Moisturizing and antioxidant factors of skin barrier restoring cream with shea butter, silkflo and vitamin E in human keratinocyte cells.

doi: 10.1111/ics.13014

Figure Lengend Snippet: FIGURE 5 Effect of MZ on procollagen type 1 level. Values are expressed as average of triplicate experiment and are represented as mean ± SEM. * represent significant difference from H2O2 induced group (p < 0.05), # represent significant difference from untreated cells.

Article Snippet: Estimation of type I procollagen The level of type I procollagen was measured using a human type I procollagen ELISA kit (Elabscience Biotechnology, USA).

Techniques: