Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: Relative HSP abundances in whole skeletal muscle homogenates from young adults and older adults pre and post HIT exercise. Representative Westen blots of (A) HSP72, HSP27, and αB-crystallin and (B) phosphorylated HSP27 Ser15 (pHSP27 Ser15) and pαB-crystallin Ser59 in whole muscle homogenates from the vastus lateralis of the same individuals. Calibration curves of mixed muscle homogenates are indicated and were used to determine the relative number of given proteins (see Methods). Stain-free gels are indicative of total protein loading, and molecular weights are indicated by markers collected under white light capture without moving the membrane between that and chemiluminescence detection. Relative abundances of (C) HSP72, (D) HSP27, (E) pHSP27 Ser15, (F) αB-crystallin, and (G) pαB-crystallin Ser59 from young (circle) and older adults Pre (square) and older adults Post (triangle) HIT exercise are shown relative to average Old (pre) on a given gel (data are presented as mean ± SD). Individuals indicated by the number of symbols ( n : 5–7), with the same color assigned to the same individual and consistent across all graphs. * p ≤ 0.05 indicates Brown-Forsye and Welch’s and post hoc analysis using Games-Horwell. HIT = high-intensity training; HSP = heat shock protein; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques: Staining, Membrane

Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: HSP abundances in type I and II skeletal muscle fibers from young and older adults. (A, C, and F) The MHC isoform present was determined in individual muscle fiber segments from the vastus lateralis and, following pooling into type I and type II groups from a given biopsy, were analyzed by Westen blotting. Westen blots of (A) HSP72, (C) HSP27 and pHSP27 Ser15, (F) αB-crystallin and pαB-crystallin Ser59, with MHC isoforms in groups of fibers. Stain-free gels are indicative of total protein loading, and molecular weights are indicated by markers collected under white light capture without moving the membrane between that and chemiluminescence detection. Calibration curves of mixed muscle homogenates are indicated. Relative protein abundances of (B) HSP72, (D) HSP27, (E) pHSP27 Ser15, (G) αB-crystallin, and (H) pαB-crystallin Ser59 in fibers from young (circle) and older adults (square) type I fibers (no outline) and type II fibers (outline). All fibers are expressed relative to the average older adult’s type I fibers. The same color is assigned to the same individual and is consistent with (data are presented as mean ± SD). * p < 0.05 and ** p < 0.01, mixed effect model Univariant using either Tukey’s or Games-Horwell’s multiple comparison test (see Methods). HIT = high-intensity training; HSP = heat shock protein; MHC = myosin heavy chain; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques: Staining, Membrane, Comparison

Journal: Journal of Sport and Health Science

Article Title: Exercise attenuates stress-related signaling as sensed by higher phosphorylation of small heat shock proteins in skeletal muscle from older individuals

doi: 10.1016/j.jshs.2025.101111

Figure Lengend Snippet: HSP abundances in type I and II skeletal fibers from older adults pre- and post HIT exercise. Relative protein abundances of (A and B) HSP72, (C and D) HSP27, (E and F) pHSP27 Ser15, (G and H) αB-crystallin, and (I and J) pαB-crystallin Ser59 in fibers from old pre and old post HIT exercise. All fibers are expressed relative to the average old pre type I fibers or relative pre type II depending on fiber type. The same color is assigned to the same Individual, consistent in both graphs and all figures. * p < 0.05 and ** p < 0.01 indicated significant difference in paired t -test (except pHSP27 Ser15 Wilcoxon match-pair rank test). Representative blots are shown in . HIT = high-intensity training; HSP = heat shock protein; pαB-crystallin Ser59 = phospho-αB-crystallin at Serine59; pHSP27 Ser15 = phospho-HSP27 at Serine15.

Article Snippet: Details of antibodies used are as follows: HSP72 (1 in 500 mouse monoclonal, SMC100A; StressMarq Biosciences, Victoria, Canada); HSP27 (1 in 1000 mouse monoclonal, G3.1 ab2790; Abcam, Cambridge, UK); pHSP27 Ser15 (1 in 2000 monoclonal rabbit, ab76313; Abcam), pHSP27 Ser82 (1 in 2000 polyclonal mouse, ADI-SPA-524; Enzo Biochem, Farmingdale, NY, USA), αB-crystallin (1 in 1000 mouse monoclonal, SPA-222; StressGen Biotechnologies), pαB-crystallin Ser59 (1 in 1000 rabbit polyclonal, SPA-227; StressGen Biotechnologies).

Techniques:

Journal: iScience

Article Title: Synthetic zipper mediated pre-targeting system for near-infrared photoimmunotherapy

doi: 10.1016/j.isci.2025.114558

Figure Lengend Snippet: The pre-targeting complex of NIR-PIT induces the release of ICD markers (A) Flow cytometric histograms show cell surface expression of calreticulin, HSP70, and HSP90 compared to untreated controls in SKOV3 cells with scFv-tisotumab-Zip2-Zip1-SNAP-IR700. (B) OVCAR3 cells treated with scFv-Sacituzumab-Zip2-Zip1-SNAP-IR700. (C) IGROV1 cells treated with scFv-Farletuzumab-Zip2-Zip1-SNAP-IR700. Only viable cells were included in the analysis. Bar graphs show the mean fluorescence intensity (MFI) of cell surface calreticulin, HSP70, and HSP90. Data are represented as mean ± SD from three biological replicates. Cells exposed to NIR light irradiation without incubation with NIR-PIT agents were used as control. Statistical significance was determined by a one-way ANOVA and Dunnett's test. ∗∗∗ p ≤ 0.001, ∗∗∗∗ p ≤ 0.0001.

Article Snippet: After 24 h NIR light irradiation, cells were harvested and incubated with 0.50 μg calreticulin (488-conjugated, Clone 681233, R&D Systems #IC38981G-100UG) antibody or 2 μL HSP70 (FITC-Conjugated, Miltenyi Biotech #130-105-548) antibody or 1 μg HSP90 (APC conjugated, H9010, Invitrogen #MA5-45102) antibody for 30 min.

Techniques: Expressing, Fluorescence, Irradiation, Incubation, Control

Journal: Redox Biology

Article Title: Combination of YAP inhibition and photodynamic therapy induces dual DNA damage and activates STING pathway to enhance immunotherapy in uveal melanoma

doi: 10.1016/j.redox.2025.103965

Figure Lengend Snippet: HANP/VP induces ICD via PDT in vitro. (A) Schematic illustration of ICD induction by HANP/VP. n = 3/group. (B) Apoptosis was analyzed by Annxin V/PI staining after corresponding treatments. After HANP/VP combined laser irradiation treatment, the proportions of early apoptosis and late apoptosis were much higher than those in the free VP combined laser irradiation-treated group, n = 3/group. (C) Chemiluminescence was used to detect ATP release after corresponding treatments. After HANP/VP combined laser irradiation treatment, the extracellular ATP concentration was much higher than that in the VP combined laser irradiation treatment group, n = 3/group. (D) Immunofluorescence staining was performed to assess the expression of HMGB1 (purple) after corresponding treatments. The level of nuclear HMGB1 expression was reduced after HANP/VP combined laser irradiation, n = 3/group. (E) The expression level of HMGB1 and HSP70 in cell culture supernatants was detected by Western blot after corresponding treatments. Compared with the VP with Laser group, the levels of HMGB1 and HSP70 in the cell culture supernatant were much higher after HANP/VP combined laser irradiation, n = 3/group. (F) Immunofluorescence staining was used to detect ecto-CRT expression (purple) after corresponding treatments. After HANP/VP combined laser irradiation, significant ecto-CRT signals appeared at a higher level than the VP combined laser irradiation treatment, and no ecto-CRT signals were observed in the control group, n = 3/group. (G) The proportion of ecto-CRT-positive cells was analyzed by flow cytometry after corresponding treatments. After HANP/VP combined laser irradiation treatment, the proportion of ecto-CRT-positive cells was significantly higher than that in the VP combined laser irradiation treatment group (30.23 ± 12.13 %), and there were almost no ecto-CRT-positive cells in the Control group, n = 3/group. Statistical significance in panels (C – G) was determined by one-way ANOVA (∗∗ p < 0.01, ∗∗∗ p < 0.001).

Article Snippet: The membranes were blocked in 5 % skim milk; incubated overnight at 4 °C with Abs against YAP1 (1:1000; Proteintech), STING (1:2000; CST), p-STING (1:1000; CST), TBK1 (1:2000; CST), p -TBK1 (1:1000; CST), IRF-3 (1:2000; CST), p -IRF-3 (1:1000; CST), vinculin (1:3000; Proteintech), HMGB1 (1:000; Proteintech), and HSP70 (1:1000; Proteintech); and then incubated with appropriate secondary Abs for 1 h at 25 °C.

Techniques: In Vitro, Staining, Irradiation, Concentration Assay, Immunofluorescence, Expressing, Cell Culture, Western Blot, Control, Flow Cytometry