opa1  (Proteintech)

Journal: Acta Pharmaceutica Sinica. B

Article Title: Rhynchophylline rewires DLAT lipoylation via conformational control to reverse mitochondrial bioenergetic collapse against dopaminergic neuronal injury

doi: 10.1016/j.apsb.2026.06.016

Figure Lengend Snippet: Rhy enhances mitochondrial function by promoting mitochondrial fusion. (A) Representative fluorescent images of mitochondria stained with Mito-tracker Red in PC12 cells. (B) Quantitative analysis of mitochondrial networks in PC12 cells using the MiNA plugin for ImageJ ( n = 5 images). (C) Mitochondrial morphology in primary midbrain neurons visualized by Mito-tracker Red. (D) Mitochondrial network parameters quantified by MiNA analysis in primary midbrain neurons ( n = 5 images). (E) High-resolution imaging of mitochondrial inner membranes using PKmito Orange (PKMO) microscopy. (F) Representative transmission electron microscopy (TEM) images of mitochondrial ultrastructure. (G) Mitochondrial membrane potential quantification by JC-1 ratio (Red/Green) in PC12 cells and primary midbrain neurons, n = 3 independent experiments. (H) Cellular ATP levels determined using ATPlite™ luminescence assay ( n = 3). (I) Western blotting analysis of DRP1, p-DRP1 Ser616 , p-DRP1 Ser637 , MFN2 and OPA1 levels, following 6-OHDA (150 μmol/L) and Rhy treatment (1, 2, 5 μmol/L), n = 3 independent experiments. The data are presented as mean ± SD. Multiple group comparisons were evaluated through one-way analysis of variance (ANOVA). P < 0.05 was considered statistically significant.

Article Snippet: Membrane were blocked with 5% non-fat milk in TBST (1 h, room temperature), and then incubated with primary antibodies (overnight, 4 °C) against DLAT (Proteintech, 13426-1-AP); DRP1 (Biodragon, RM8248); p-DRP1 Ser616 (Biodragon, BD-PP1318); p-DRP1 Ser637 (Biodragon, BD-PP0841); OPA1 (MCE, HY- P80255 ); MFN2 (Biodragon, BD-PT2740); GAPDH (Proteintech, 60004-1-Ig); TH (Santa Cruz, sc-14007); Lipoic acid (Santa Cruz, sc-101354); VMAT2 (Santa Cruz, sc-390,285); HA (Solarbio, K200003M), SIRT2 (Abways, CY5865); SIRT4(Proteintech, 66543-1-Ig); LIAS (Proteintech, 11577-1-AP); AMPK α (Biodragon, RM1159); p-AMPK α (Biodragon, RM0088); p-STAT3 (Abmart, TC52210); p-EGFR (Abmart, PC1644); KIF5B(Abclonal, A20928); KIF5C(Biodragon, BD-PN3289); KIF1B(Biodragon, BD-PN4193).

Techniques: Staining, Imaging, Microscopy, Transmission Assay, Electron Microscopy, Membrane, Luminescence Assay, Western Blot

Journal: Acta Pharmaceutica Sinica. B

Article Title: Rhynchophylline rewires DLAT lipoylation via conformational control to reverse mitochondrial bioenergetic collapse against dopaminergic neuronal injury

doi: 10.1016/j.apsb.2026.06.016

Figure Lengend Snippet: DLAT mediates Rhy-induced mitochondrial fusion regulation. (A) Experimental workflow for identifying potential targets of Rhy using thermal proteome profiling (TPP) strategy. (B) Volcano plot of TPP results and screening strategy for Rhy's anti-PD targets. (C) Schematic representation of the target screening and prioritization pipeline. (D) Western blotting analysis showing Rhy-regulated mitochondrial dynamics proteins (OPA1, p-DRP1 Ser637 ) in control versus DLAT-KO cells ( n = 3). (E, F) Representative confocal images (E) and quantification (F) of mitochondrial networks visualized by Mito-tracker Red ( n = 3 fields). (G) Super-resolution microscopy images of mitochondrial cristae structures labeled with PKmito Orange (PKMO), n = 3 images. (H, I) TMRE staining reveals mitochondrial membrane potential alterations ( n = 5 fields). The data are presented as mean ± SD. Multiple group comparisons were evaluated through one-way analysis of variance (ANOVA). P < 0.05 was considered statistically significant.

Article Snippet: Membrane were blocked with 5% non-fat milk in TBST (1 h, room temperature), and then incubated with primary antibodies (overnight, 4 °C) against DLAT (Proteintech, 13426-1-AP); DRP1 (Biodragon, RM8248); p-DRP1 Ser616 (Biodragon, BD-PP1318); p-DRP1 Ser637 (Biodragon, BD-PP0841); OPA1 (MCE, HY- P80255 ); MFN2 (Biodragon, BD-PT2740); GAPDH (Proteintech, 60004-1-Ig); TH (Santa Cruz, sc-14007); Lipoic acid (Santa Cruz, sc-101354); VMAT2 (Santa Cruz, sc-390,285); HA (Solarbio, K200003M), SIRT2 (Abways, CY5865); SIRT4(Proteintech, 66543-1-Ig); LIAS (Proteintech, 11577-1-AP); AMPK α (Biodragon, RM1159); p-AMPK α (Biodragon, RM0088); p-STAT3 (Abmart, TC52210); p-EGFR (Abmart, PC1644); KIF5B(Abclonal, A20928); KIF5C(Biodragon, BD-PN3289); KIF1B(Biodragon, BD-PN4193).

Techniques: Western Blot, Control, Super-Resolution Microscopy, Labeling, Staining, Membrane

Journal: Acta Pharmaceutica Sinica. B

Article Title: Rhynchophylline rewires DLAT lipoylation via conformational control to reverse mitochondrial bioenergetic collapse against dopaminergic neuronal injury

doi: 10.1016/j.apsb.2026.06.016

Figure Lengend Snippet: Rhy ameliorates mitochondrial dysfunction via suppression of the DLAT–SIRT4 axis in PD mice. (A) Representative transmission electron microscope images illustrating mitochondrial ultrastructure in substantia nigra neurons. (B, C) Protein levels of mitochondrial dynamics markers (DRP1, p-DRP1 Ser616 , p-DRP1 Ser637 , MFN2, OPA1) in SN tissues were analyzed by Western blotting ( n = 3). (D, E) Immunofluorescence co-staining demonstrating reduced DLAT–SIRT4 colocalization in substantia nigra following Rhy treatment ( n = 3). The data are presented as mean ± SD. Multiple group comparisons were evaluated through one-way analysis of variance (ANOVA). P < 0.05 was considered statistically significant.

Article Snippet: Membrane were blocked with 5% non-fat milk in TBST (1 h, room temperature), and then incubated with primary antibodies (overnight, 4 °C) against DLAT (Proteintech, 13426-1-AP); DRP1 (Biodragon, RM8248); p-DRP1 Ser616 (Biodragon, BD-PP1318); p-DRP1 Ser637 (Biodragon, BD-PP0841); OPA1 (MCE, HY- P80255 ); MFN2 (Biodragon, BD-PT2740); GAPDH (Proteintech, 60004-1-Ig); TH (Santa Cruz, sc-14007); Lipoic acid (Santa Cruz, sc-101354); VMAT2 (Santa Cruz, sc-390,285); HA (Solarbio, K200003M), SIRT2 (Abways, CY5865); SIRT4(Proteintech, 66543-1-Ig); LIAS (Proteintech, 11577-1-AP); AMPK α (Biodragon, RM1159); p-AMPK α (Biodragon, RM0088); p-STAT3 (Abmart, TC52210); p-EGFR (Abmart, PC1644); KIF5B(Abclonal, A20928); KIF5C(Biodragon, BD-PN3289); KIF1B(Biodragon, BD-PN4193).

Techniques: Transmission Assay, Microscopy, Western Blot, Immunofluorescence, Staining

Journal: Materials Today Bio

Article Title: A dual-responsive CO-releasing nanogel ameliorates retinal ischemia–reperfusion injury by restoring mitochondrial homeostasis and attenuating cGAS-STING pathway activation

doi: 10.1016/j.mtbio.2026.102974

Figure Lengend Snippet: COPN restores mitochondrial quality control and interrupts the ROS‒cGAS‒inflammation axis in OGD/R-treated R28 cells. a , TEM images showing changes in the mitochondrial ultrastructure of R28 cells subjected to various treatments (PBS, PDNs, COPN-L, and COPN-H) after OGD/R. Scale bar: 500 nm b , Quantitative analyses of mitochondrial length and number in R28 cells as indicated in a . c , Representative fluorescence images showing JC-1 staining of the mitochondrial Δψm: aggregates (red) indicate healthy mitochondria, whereas monomers (green) represent depolarized mitochondria. Nuclei were stained with DAPI (blue). Scale bar: 50 μm d , Intracellular ATP content assay revealing improved energy production after COPN treatment under OGD/R stress conditions. e , mRNA expression analysis of mitochondrial dynamic regulatory genes (Opa1, Mfn2, Drp1, and Fis1) and mitochondrial DNA transcription levels (mt-ND1 and mt-COX1). f , Quantitative analysis of the JC-1 fluorescence ratio (aggregates/monomers) and mitochondrial ROS levels (MitoSOX staining). g–h , Western blot analysis of proteins involved in mitochondrial autophagy (Pink1, Parkin, and P62) (g) and mitochondrial fusion/fission (Opa1, Mfn2, Mfn1, Drp1, and Fis1) (h) . i , Confocal fluorescence microscopy images of mitochondria (green, MitoTracker) and lysosomes (red, LysoTracker) in R28 cells after different treatments. Nuclei were stained with Hoechst (blue). Scale bar: 50 μm ∗ ∗∗∗P < 0.0001. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: The following primary antibodies were used: Drp1 (CST, #8570, 1:1000, ∼80 kDa), Fis1 (Proteintech, 10956-1-AP, 1:1000, ∼17 kDa), Mfn1 (Abcam, ab104274, 1:1000, ∼84 kDa), Mfn2 (CST, #9482, 1:1000, ∼86 kDa), Opa1 (CST, #80471, 1:1000, ∼100–120 kDa), Pink1 (CST, #6946, 1:1000, ∼63 kDa), cGAS (CST, #15102, 1:1000, ∼60 kDa), STING (CST, #13647, 1:1000, ∼42–45 kDa), TBK1 (CST, #3013, 1:1000, ∼84 kDa), p-TBK1 (CST, #5483, 1:1000, ∼84 kDa), COX IV (Abcam, ab14744, 1:2000, ∼17 kDa), β-actin (Proteintech, 66009-1-Ig, 1:5000, ∼43 kDa), and GAPDH (Proteintech, 60004-1-Ig, 1:5000, After washing, the membranes were incubated with HRP-conjugated secondary antibodies (goat anti-rabbit or goat anti-mouse IgG, CST, 1:5000) for 1 h at room temperature.

Techniques: Control, Fluorescence, Staining, Expressing, Western Blot, Microscopy