nb100-2387  (Bio-Techne corporation)

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet: Expression of p21 and Zmpste24 in different regions of tibial cartilage from patients with varus undergoing TKA and the relationship between them (A) Schematic plot of plug drilling in different regions of the tibial plateau from patients with varus undergoing TKA. (B) Representative images of Safranine O/Fast green, H&E, Toluidine blue, and IHC staining of p21 and Zmpste24 in different regions. (C–E) Analysis of OARSI scores, p21 positive cells, and Zmpste24 positive cells of specimens in different regions. (F) Pearson correlation analysis and simple linear regression between the expression of p21 and Zmpste24. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Expressing, Immunohistochemistry

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet: Zmpste24 expression is downregulated in tibial cartilage in mice with DMM surgery (A) 3D reconstruction of the knee joints of Sham and DMM mice and representative section images of subchondral bone. (B) Analysis of trabecular bone parameters. BV/TV, percent bone volume; Tb.N, trabecular number; Tb.Th, trabecular thickness; Tb.Sp, trabecular separation. (C and D) H&E staining, Safranine O/Fast green, and corresponding quantitative analysis. Scale bar: 200 μm. (E and F) IF staining of p21, Col2a1, and Zmpste24 and positive cell proportions after treatment with DMM. (G) Pearson correlation analysis and simple linear regression between the expression of p21 and Zmpste24. Scale bar: 50 μm n = 6 in each group. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001, compared to Sham group. Black arrow: synovium.

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Expressing, Staining

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet: Excess cyclic tensile stretching induces Zmpste24 downregulated expression, nuclear membrane instability, and cell senescence in chondrocytes (A and B) mRNA expression of Zmpte24, p21, p16 INK4a , p53, Col2a1, Sox9, Mmp3, Mmp13, Col1a1, and Col10a1of chondrocytes experiencing 5% or 20% elongation of cyclic tensile stretch (C–F) Presentative images of cytoskeleton, IF staining of the nuclear membrane, SA-β-gal staining of chondrocytes after different periods of 5% or 20% CTS and corresponding quantitative analysis of nuclear blebbing cell proportions, SA-β-gal positive cell proportion, and Zmpste24 fluorescence. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Expressing, Membrane, Staining, Fluorescence

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet: Overexpression of Zmpste24 could rescue mechanical overloading-induced chondrocyte degeneration and senescence in vitro (A–D) Validation of Zmpste24 overexpression through qRT-PCR (B) Rescue effect of Zmpste24 overexpression on mRNA expression of Col2a1, Sox9, Mmp3, Mmp13, Col1a1, Col10a1, p16 INK4a , p21, and p53 (C and D) Rescue effect of Zmpste24 overexpression on protein expression of Col2a1, Sox9, Mmp3, Mmp13, p16 INK4a , p21, and p53 and corresponding analysis. (E–H) IF staining of Lamin A/C, Zmpste24 and ROS and corresponding cell proportion analysis. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, and ∗∗∗∗p < 0.0001.

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Over Expression, In Vitro, Biomarker Discovery, Quantitative RT-PCR, Expressing, Staining

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet: The role of Zmpste24 and nuclear membrane stability in cartilage degeneration and senescence under abnormal mechanical conditions

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Membrane

Journal: iScience

Article Title: Mechanical overloading leads to chondrocyte degeneration and senescence via Zmpste24-mediated nuclear membrane instability

doi: 10.1016/j.isci.2023.108119

Figure Lengend Snippet:

Article Snippet: For F-actin staining, the chondrocytes were incubated with SF488-labeled phalloidin for 30 min. For cell immunofluorescence, after fixation, permeabilization, and blocking with 10% goat serum, cells were incubated with primary antibody for Zmpste24 (NB100-2387, Novus) or Lamin A/C (4777, CST) overnight at 4°C.

Techniques: Recombinant, Staining, RNA Extraction, Software