antagonist 4  (MedChemExpress)

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet: P2ry14 knockout in Nf1fl/fl; Dhh+ mice restores BNB morphology, reduces fibrinogen deposition and increases Cav1 expression (A) β-galactosidase immunofluorescent staining of 7-month-old Nf1fl/fl;Dhh+ and P2ry14 −/− ;Nf1fl/fl ; Dhh+ sciatic nerve shows P2RY14 knockdown (dotted lines denote the perineurium; Nf1 fl/fl DhhCre n = 3; P2ry14 −/− ; Nf1fl/flDhh + n = 3). (B) Nerve semi-thin sections and electron micrographs of 7-month Nf1fl/fl ; Dhh+ and P2RY14 −/− ; Nf1fl/fl ; Dhh+ saphenous nerve showing perineurium structure (arrows denote perineurium). (C) Electron micrograph showing TJs (arrow) in 7-month Nf1fl/fl ; Dhh+ and P2RY14 −/− ; Nf1fl/fl ; Dhh+ saphenous nerve ( Nf1 fl/fl Dhh+ n = 3; P2RY14 Nf1 fl/fl Dhh+ n = 3). (D) Quantification of TJs per field of view (∗∗ p = 0.0021; ∗∗∗ p = 0.0007; one-way Anova) ( WT n = 3; Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (E) Anti-Glut1 immunofluorescent staining of 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve (arrows indicate Glut1+ endoneurial capillaries). ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (F) Anti fibrin/fibrinogen and anti-CD31 staining in 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve. Arrows point to CD31 + endoneurial capillaries, some of which are shown at increased magnification below boxed regions of the images ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh n = 3). (G) Quantification of CD31 + endoneurial vessels in 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve (p = n.s.; un-paired t-test) ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh n = 3). (H) RNAseq showing Cav1 expression in nerve from 4-month-old WT , P2ry14 −/− , Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh+ sciatic nerve normalized to control wild-type mice. ( WT n = 3, P2ry14 −/− n = 3, Nf1fl/fl ; Dhh+ n = 3 and P2ty14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (I) Anti-Caveolin immunofluorescent staining within 7-month WT and Nf1fl/fl ; Dhh+ sciatic nerve shows that in WT nerves, Cav-1 expression is increased and localized to both the perineurium and the endoneurium. Cav-1 expression is reduced in Nf1 fl/flDhh+ sciatic nerve and rescued by genetic deletion of P2ry14. ( WT n = 3, Nf1fl/fl ; Dhh+ n = 3 and P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (J) Quantification of caveolae per field of view at the EM level in 7-month saphenous nerves (p = n.s. One-way Anova) ( WT n = 3; Nf1fl/fl ; Dhh+ n = 3; P2ry14 −/− ; Nf1 fl/fl Dhh+ n = 3). (K) anti-pPKA substrate immunofluorescent staining in 7-month WT and Nf1fl/fl ; Dhh+ sciatic nerve shows that in WT nerves, pPKA expression is localized to the perineurium (between dotted lines) as well as the endoneurium. Staining is reduced in Nf1fl/fl ; Dhh+ perineurium and endoneurium and rescued by genetic deletion of P2RY14. ( WT n = 3; Nf1fl/fl ; Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). All stats Student’s t test (Unpaired t-test) in all panels, n = numbers of individuals.

Article Snippet: P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) , TOCRIS , Cat #: 4862.

Techniques: Knock-Out, Expressing, Staining, Knockdown, Control

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet: Pharmacological inhibition of P2RY14 restores nerve p-PKA and Cav1 expression (A) Illustration of experimental design to treat mice with the P2RY14 inhibitor (PPTN) drug via osmotic minipump delivery to 4-month WT and Nf1fl/fl ; Dhh+ mice. (B) Anti- p -PKA-substrate immunofluorescent sciatic nerve staining of 4-month WT , Nf1fl/fl ; Dhh+ (Vehicle) and Nf1fl/fl ; Dhh+ (PPTN treated) mice showing that p -PKA expression is rescued to WT levels in PPTN treated mice. (C) Anti-fibrinogen immunofluorescent staining with antibody is not changed with treatment. (D) anti-caveolin immunofluorescent staining shows rescue by PPTN treatment in double mutants. ( WT n = 3, Nf1fl/fl ; Dhh+ (Vehicle) n = 3, and Nf1fl/fl ; Dhh+ (PPTN treated) n = 3).

Article Snippet: P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) , TOCRIS , Cat #: 4862.

Techniques: Inhibition, Expressing, Staining

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet:

Article Snippet: P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) , TOCRIS , Cat #: 4862.

Techniques: Recombinant, Expressing, Microarray, Software

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet: P2ry14 knockout in Nf1fl/fl; Dhh+ mice restores BNB morphology, reduces fibrinogen deposition and increases Cav1 expression (A) β-galactosidase immunofluorescent staining of 7-month-old Nf1fl/fl;Dhh+ and P2ry14 −/− ;Nf1fl/fl ; Dhh+ sciatic nerve shows P2RY14 knockdown (dotted lines denote the perineurium; Nf1 fl/fl DhhCre n = 3; P2ry14 −/− ; Nf1fl/flDhh + n = 3). (B) Nerve semi-thin sections and electron micrographs of 7-month Nf1fl/fl ; Dhh+ and P2RY14 −/− ; Nf1fl/fl ; Dhh+ saphenous nerve showing perineurium structure (arrows denote perineurium). (C) Electron micrograph showing TJs (arrow) in 7-month Nf1fl/fl ; Dhh+ and P2RY14 −/− ; Nf1fl/fl ; Dhh+ saphenous nerve ( Nf1 fl/fl Dhh+ n = 3; P2RY14 Nf1 fl/fl Dhh+ n = 3). (D) Quantification of TJs per field of view (∗∗ p = 0.0021; ∗∗∗ p = 0.0007; one-way Anova) ( WT n = 3; Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (E) Anti-Glut1 immunofluorescent staining of 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve (arrows indicate Glut1+ endoneurial capillaries). ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (F) Anti fibrin/fibrinogen and anti-CD31 staining in 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve. Arrows point to CD31 + endoneurial capillaries, some of which are shown at increased magnification below boxed regions of the images ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh n = 3). (G) Quantification of CD31 + endoneurial vessels in 7-month Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh sciatic nerve (p = n.s.; un-paired t-test) ( Nf1 fl/fl Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh n = 3). (H) RNAseq showing Cav1 expression in nerve from 4-month-old WT , P2ry14 −/− , Nf1fl/fl ; Dhh+ and P2ry14 −/− ; Nf1fl/fl ; Dhh+ sciatic nerve normalized to control wild-type mice. ( WT n = 3, P2ry14 −/− n = 3, Nf1fl/fl ; Dhh+ n = 3 and P2ty14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (I) Anti-Caveolin immunofluorescent staining within 7-month WT and Nf1fl/fl ; Dhh+ sciatic nerve shows that in WT nerves, Cav-1 expression is increased and localized to both the perineurium and the endoneurium. Cav-1 expression is reduced in Nf1 fl/flDhh+ sciatic nerve and rescued by genetic deletion of P2ry14. ( WT n = 3, Nf1fl/fl ; Dhh+ n = 3 and P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). (J) Quantification of caveolae per field of view at the EM level in 7-month saphenous nerves (p = n.s. One-way Anova) ( WT n = 3; Nf1fl/fl ; Dhh+ n = 3; P2ry14 −/− ; Nf1 fl/fl Dhh+ n = 3). (K) anti-pPKA substrate immunofluorescent staining in 7-month WT and Nf1fl/fl ; Dhh+ sciatic nerve shows that in WT nerves, pPKA expression is localized to the perineurium (between dotted lines) as well as the endoneurium. Staining is reduced in Nf1fl/fl ; Dhh+ perineurium and endoneurium and rescued by genetic deletion of P2RY14. ( WT n = 3; Nf1fl/fl ; Dhh+ n = 3; P2ry14 −/− ; Nf1fl/fl ; Dhh+ n = 3). All stats Student’s t test (Unpaired t-test) in all panels, n = numbers of individuals.

Article Snippet: 4-month-old neurofibroma mice (Nf1 fl/fl Dhh+) were treated with the P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) (TOCRIS: Cat #.

Techniques: Knock-Out, Expressing, Staining, Knockdown, Control

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet: Pharmacological inhibition of P2RY14 restores nerve p-PKA and Cav1 expression (A) Illustration of experimental design to treat mice with the P2RY14 inhibitor (PPTN) drug via osmotic minipump delivery to 4-month WT and Nf1fl/fl ; Dhh+ mice. (B) Anti- p -PKA-substrate immunofluorescent sciatic nerve staining of 4-month WT , Nf1fl/fl ; Dhh+ (Vehicle) and Nf1fl/fl ; Dhh+ (PPTN treated) mice showing that p -PKA expression is rescued to WT levels in PPTN treated mice. (C) Anti-fibrinogen immunofluorescent staining with antibody is not changed with treatment. (D) anti-caveolin immunofluorescent staining shows rescue by PPTN treatment in double mutants. ( WT n = 3, Nf1fl/fl ; Dhh+ (Vehicle) n = 3, and Nf1fl/fl ; Dhh+ (PPTN treated) n = 3).

Article Snippet: 4-month-old neurofibroma mice (Nf1 fl/fl Dhh+) were treated with the P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) (TOCRIS: Cat #.

Techniques: Inhibition, Expressing, Staining

Journal: iScience

Article Title: NF1-dependent disruption of the blood-nerve-barrier is improved by blockade of P2RY14

doi: 10.1016/j.isci.2024.110294

Figure Lengend Snippet:

Article Snippet: 4-month-old neurofibroma mice (Nf1 fl/fl Dhh+) were treated with the P2RY14 inhibitor (PPTN-HCl:4-[4-(4-Piperidinyl]-7-[4-(trifluoromethyl)phenyl]-2-naphthalenecarboxyliv acid hydrochloride) (TOCRIS: Cat #.

Techniques: Recombinant, Gene Expression, Microarray, Software