recombinant human adamts4 protein  (R&D Systems)

Journal: Arthritis research & therapy

Article Title: Establishment of an ex vivo cartilage damage model by combined collagenase treatment and mechanical loading.

doi: 10.1186/s13075-025-03499-7

Figure Lengend Snippet: Fig. 3 External collagenase II and aggrecanase led to release of inflammatory markers, but mRNA expression of ACAN, COL2A1, SOX9, IL6, ADAMTS4, and ADAMTS5 did not change. (A, B) IL6 and IL-1β release in the medium were detected by ELISA (n = 4). (C) Release of Nitric oxide was detected by Griess reaction (n = 4). (D) RT-qPCR analysis of ACAN, COL2A1, SOX9, IL6, ADAMTS4, and ADAMTS5 (n = 5). Relative quantification of target mRNA was performed according to the 2−ΔΔCt method with RPLP0 as the endogenous control. Gene expression values were normalized to the average values of bovine carti lage at day 0. Triangles, dots or square of different color represent individual donors

Article Snippet: Recombinant human ADAMTS4 protein (aggrecanase 1) and recombinant human ADAMTS5 protein (aggrecanase 2) were purchased from R&D Systems.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Quantitative Proteomics, Control, Gene Expression

Journal: bioRxiv

Article Title: Adult oligodendrogenesis gates arcuate neuronal glucose sensing through remodelling of the blood-hypothalamus barrier via ADAMTS4

doi: 10.1101/2024.09.10.612227

Figure Lengend Snippet: (a) Representative images of Wisteria floribunda agglutinin (WFA) lectin labelling in the median eminence (ME) and arcuate nucleus (ARC) of myelin regulatory factor (MYRF) conditional knockout mice ( Myrf ­­ ) and controls ( Myrf ­­ ) six week after tamoxifen administration at postnatal day 60 and ( b ) associated quantification. Gene expression analysis of transcripts relating to ( c ) perineuronal net (PNN) structural components and ( d ) PNN-modifying enzymes expressed by oligodendrocyte lineage cells in the mediobasal hypothalamus of Myrf ­­ and Myrf ­­ mice. ( e ) (left) Hypothalamic cell populations from HypoMap ­­ . (right) Log-normalized expression of Adamts4 in the hypothalamic cell populations, demonstrating specific expression in OL populations. ( f ) Representative images of ADAMTS4 labelling in the ME-ARC of Myrf ­­ and Myrf ­­ mice. ( g) Representative images of Versican (VCAN) labelling in the ME-ARC of Myrf ­­ and Myrf ­­ mice and ( h ) associated quantification. Data presented as mean ± S.E.M. and analysed by unpaired student’s - or Welch’s – t test, *p<0.05, **p<0.01, scale bars represent 100 μm, n=5-10/group.

Article Snippet: Bilateral injections of aCSF (Tocris) or 4 ng recombinant human ADAMTS4 (R&D Systems 4307-AD; 2 ng/side in 100 nl) were performed at ZT5 using bevelled stainless steel 33-gauge injectors, extending 1 mm beyond the tip of the guide.

Techniques: Knock-Out, Gene Expression, Expressing

Journal: bioRxiv

Article Title: Adult oligodendrogenesis gates arcuate neuronal glucose sensing through remodelling of the blood-hypothalamus barrier via ADAMTS4

doi: 10.1101/2024.09.10.612227

Figure Lengend Snippet: ( a ) A disintegrin and metallopeptidase with thrombospondin motifs 4 (ADAMTS4) expression in the mediobasal hypothalamus (MBH) of ADAMTS4 knockout ( Adamts4 ­­ ) mice compared to controls ( Adamts4 ­­ ). Representative images of ( b ) versican (VCAN), ( d ) MECA32+/laminin+ fenestrated capillary loops and ( f ) albumin immunolabelling in Adamts4 ­­ and Adamts4 ­­ mice and associated quantifications ( c , e , g ). ( h ) Blood glucose and ( i ) plasma corticosterone levels in Adamts4 ­­ and Adamts4 ­­ mice at ZT2. ( j ) Blood glucose, ( k ) plasma corticosterone and ( l-m ) insulin sensitivity in C57BL/6J mice one hour after intra-hypothalamic aCSF or ADAMTS4 administration. Data presented as mean ± S.E.M. and analysed by unpaired student’s t-test or Mann-Whitney test, *p<0.05, **p<0.01, ***p<0.001, ****p<0.0001, n=8-18/group. Scale bars represent 100 μm.

Article Snippet: Bilateral injections of aCSF (Tocris) or 4 ng recombinant human ADAMTS4 (R&D Systems 4307-AD; 2 ng/side in 100 nl) were performed at ZT5 using bevelled stainless steel 33-gauge injectors, extending 1 mm beyond the tip of the guide.

Techniques: Expressing, Knock-Out, Clinical Proteomics, MANN-WHITNEY

Journal: bioRxiv

Article Title: Adult oligodendrogenesis gates arcuate neuronal glucose sensing through remodelling of the blood-hypothalamus barrier via ADAMTS4

doi: 10.1101/2024.09.10.612227

Figure Lengend Snippet: ( a ) Representative images of Wisteria floribunda agglutinin (WFA) lectin in the mediobasal hypothalamus (MBH) of ADAMTS4 knockout ( Adamts4 ­­ ) mice compared to controls ( Adamts4 ­­ ) and ( b ) associated quantification. Representative images of ( c ) MBP and ( e ) BCAS1 immunolabelling in the ME of Adamts4 ­­ and Adamts4 ­­ mice and ( d , f ) associated quantifications. Representative images of ( g ) VCAN and ( i ) laminin immunolabelling in the MBH of animals injected with aCSF or ADAMTS4 1 hour prior to sacrifice and ( h, j ) associated quantifications. Data presented as mean ± S.E.M. and analysed by unpaired student’s t-test or Mann-Whitney test, *p<0.05, ***p<0.001, ****p<0.0001, n=8-18/group. Scale bars represent 100 μm.

Article Snippet: Bilateral injections of aCSF (Tocris) or 4 ng recombinant human ADAMTS4 (R&D Systems 4307-AD; 2 ng/side in 100 nl) were performed at ZT5 using bevelled stainless steel 33-gauge injectors, extending 1 mm beyond the tip of the guide.

Techniques: Knock-Out, Injection, MANN-WHITNEY

Journal: bioRxiv

Article Title: Adult oligodendrogenesis gates arcuate neuronal glucose sensing through remodelling of the blood-hypothalamus barrier via ADAMTS4

doi: 10.1101/2024.09.10.612227

Figure Lengend Snippet: Representative images of A disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4), immunolabelling in the mediobasal hypothalamus (MBH) ( a ) one hour after intraperitoneal administration of vehicle (saline; 10 ml/kg) or insulin (0.75 U/kg) and ( b ) associated quantification. ( c ) Blood glucose levels in mice fasted for 6 hours in diet-induced-obese (DIO) mice four weeks after administration of vehicle (DIO-Veh) or streptozotocin (DIO-STZ). ( d ) ADAMTS4 immunolabelling in the MBH of DIO-Veh and DIO-STZ mice and ( e ) associated quantification. ( f ) Correlation between ME ADAMTS4 immunolabelling and blood glucose across multiple models, analysed by linear regression. ( g ) Working model by which ME OL lineage cells regulate glucose homeostasis. All data presented as mean ± S.E.M., scale bars represent 100 μm. Data analysed using an unpaired student’s- or Welch’s-t test, *p<0.05, **p<0.01, n=5-10/group.

Article Snippet: Bilateral injections of aCSF (Tocris) or 4 ng recombinant human ADAMTS4 (R&D Systems 4307-AD; 2 ng/side in 100 nl) were performed at ZT5 using bevelled stainless steel 33-gauge injectors, extending 1 mm beyond the tip of the guide.

Techniques: Saline

Journal: Communications Biology

Article Title: ADAMTS4 is a crucial proteolytic enzyme for versican cleavage in the amnion at parturition

doi: 10.1038/s42003-024-06007-w

Figure Lengend Snippet: a The illustration showing the presence of VCAN on the cell surface attached to hyaluronan and the cleavage of VCAN to versikine by ADAMTS4. AF, amnion fibroblasts; HA, hyaluronan; G1, N-terminal G1 Domain of VCAN. b Treatment of amnion fibroblasts with rhADAMTS4 (50 ng/mL; 24 h) decreased VCAN abundance in the cells, and increased versikine abundance in the conditioned culture medium ( n = 3). rhADAMTS4, recombinant human ADAMTS4. c The illustration showing human amnion tissue explant culture. d rhADAMTS4 (50 ng/mL; 24 h) treatment of amnion tissue explants increased VCAN cleavage to versikine ( n = 3). e Representative immunohistochemistry images showing decreased VCAN protein abundance in human amnion tissue explants with rhADAMTS4 (50 ng/mL; 24 h) treatment ( n = 3). Scale bars, 100 or 400 μm. f Representative images of transmission electron microscopy showing distorted collagen fibrils in the ECM of amnion tissue explants treated with rhADAMTS4 (50 ng/mL; 24 h). n = 3. Scale bars, 1 μm. g Representative images of Masson trichrome staining (blue) and Sirius red staining (red) showing unaltered total collagen abundance in amnion tissue explants treated with rhADAMTS4 (50 ng/ml; 24 h; n = 3), Scale bars, 625 μm.; ( h ). The average data of Masson staining and Sirius red staining images. OD, optical density. Data are mean ± SEM. Statistical analysis was performed with paired Student’s t test. * p < 0.05, ** p < 0.01, *** p < 0.001 vs. control.

Article Snippet: To study the effect of ADAMTS4 on VCAN degradation, fibroblasts were treated with rhADAMTS4 (50 ng/mL, R&D systems, Minneapolis, MN; #4307-AD-020) for 24 h. After treatment, fibroblasts were collected for measurement of cellular VCAN, and the conditioned culture medium was collected for measurement of versikine with Western blotting after concentration.

Techniques: Recombinant, Immunohistochemistry, Quantitative Proteomics, Transmission Assay, Electron Microscopy, Staining, Control

Journal: Communications Biology

Article Title: ADAMTS4 is a crucial proteolytic enzyme for versican cleavage in the amnion at parturition

doi: 10.1038/s42003-024-06007-w

Figure Lengend Snippet: a Increased ADAMTS4 abundance in conditioned culture medium (n = 4), and decreased VCAN abundance in amnion fibroblasts (n = 4) following siRNA-mediated knockdown of LRP1. b Representative confocal microscopic images showing ADAMTS4 endocytosis illustrated by co-localization of ADAMTS4 (green) and EEA1 (red), an early endocytosis marker, in the endosomes of human amnion fibroblasts treated with rhADAMTS4 (20 nM; 6 h). ADAMTS4 endocytosis was blocked by siRNA-mediated knockdown of LRP1 illustrated by the absence of ADAMTS4 in endosomes. n = 3. EEA1, early endosome antigen 1. NC, negative control. Scale bar, 10 or 50 μm. c Diagram illustrating the inhibitory effects of IL-1β and SAA1 on ADAMTS4 endocytosis in human amnion fibroblasts. IL-1β and SAA1 not only induced ADAMTS4 expression but also inhibited ADAMTS4 endocytosis by enhancing LRP1 shedding from the cell membranes. sLRP1, shedding LRP1. d and e Treatment of human amnion fibroblasts with IL-1β (10 ng/mL; 24 h; d) and SAA1 (100 ng/mL; 24 h; e) increased ADAMTS4 abundance in the conditioned culture medium and decreased VCAN abundance in the cells (n = 4). Data are mean ± SEM. Statistical analysis was performed with paired Student’s t-test. *p < 0.05, **p < 0.01, ***p < 0.001 vs. control (0) or NC.

Article Snippet: To study the effect of ADAMTS4 on VCAN degradation, fibroblasts were treated with rhADAMTS4 (50 ng/mL, R&D systems, Minneapolis, MN; #4307-AD-020) for 24 h. After treatment, fibroblasts were collected for measurement of cellular VCAN, and the conditioned culture medium was collected for measurement of versikine with Western blotting after concentration.

Techniques: Knockdown, Marker, Negative Control, Expressing, Control