Journal: Purinergic Signalling

Article Title: Purinergic receptors are part of a signalling system for proliferation and differentiation in distinct cell lineages in human anagen hair follicles

doi: 10.1007/s11302-008-9108-0

Figure Lengend Snippet: Expression of P2Y 1 and P2Y 2 receptors in human anagen hair follicles. a P2Y 1 receptors were found in the outer root sheath ( ORS ) and bulb of anagen hair follicles in longitudinal section but not in the inner root sheath ( IRS ). Scale bar = 40 μm. b In transverse section, P2Y 1 receptors were only seen in the outer root sheath ( ORS ). Scale bar = 40 μm. c P2Y 2 receptors were found in the cortex ( CTX ). Scale bar = 40 μm. d Transverse section of anagen hair follicle: P2Y 2 receptors were seen in the cortex ( CTX ) but not in the central medulla ( M ), inner ( IRS ) or outer root sheaths ( ORS ), or in the surrounding adventitial layer ( A ). Scale bar = 40 μm. e , f Controls: the immunoreaction was abolished after preabsorption of the e P2Y 1 and f P2Y 2 receptor antibodies with the corresponding peptides, confirming the specificity of the immunoreaction. Scale bars = 100 μm

Article Snippet: Polyclonal anti-P2Y 1 and P2Y 2 antibodies were obtained from Alomone Labs (Jerusalem, Israel) and corresponded to the third extracellular loop of the P2Y 1 (AA 242–258) and P2Y 2 receptor (AA 227–244).

Techniques: Expressing

Journal: Purinergic Signalling

Article Title: Purinergic receptors are part of a signalling system for proliferation and differentiation in distinct cell lineages in human anagen hair follicles

doi: 10.1007/s11302-008-9108-0

Figure Lengend Snippet: Double labelling of P2Y 1 and P2Y 2 receptors with markers for cellular proliferation, and double labelling of P2X 5 receptors with markers for keratinocyte differentiation in anagen hair follicles. a Double labelling of P2Y 1 receptors ( red ) with Ki-67, a nuclear marker for proliferating cells ( green ), to show that P2Y 1 receptors are found in proliferating basal cells in the outer root sheath ( ORS ) and bulb region of the hair follicle in longitudinal section. Scale bar = 50 μm. b Transverse section: double labelling of P2Y 1 receptors ( red ) with Ki-67, a nuclear marker ( green ), to show that P2Y 1 receptors are found in proliferating cells in the outer root sheath ( ORS ) of the hair follicle. Scale bar = 50 μm. c Longitudinal section of anagen hair follicle through the dermal papilla ( DP ): double labelling of P2Y 2 receptors ( red ) with proliferating cell nuclear antigen (PCNA), a marker for proliferating cells ( green ), showed that P2Y 2 receptors were found in the cortex ( CTX ) and at the edge of the medulla ( M ) but not in the central medulla. P2Y 2 receptors were not found in the matrix ( Ma ), where cells were positive for PCNA. Scale bar = 75 μm. d Longitudinal section of anagen hair follicle: P2Y 2 receptors were absent from the keratinised cuticle ( Cu ) of the hair shaft. PCNA was also found in cells of the outer root sheath ( ORS ). Scale bar = 75 μm. e Double labelling of P2X 5 receptors ( red-brown ) with involucrin, a marker for differentiating cells ( green ). Involucrin was expressed both in the inner root sheath ( IRS ), cortex ( CTX ) and in the outermost edge of the medulla ( M ). P2X 5 receptors were expressed in the inner ( IRS ) and outer root sheaths ( ORS ) and in the medulla ( M ) and matrix cells ( Ma ). There was yellow colocalisation with P2X 5 receptors in the inner root sheath and in cells at the outermost edge of the medulla ( arrow ). The cortex only stained positive for involucrin, not P2X 5 receptors. Scale bar = 50 μm. f Transverse section: double labelling of P2X 5 receptors ( red ) with involucrin ( green ). Involucrin was expressed in the inner root sheath ( IRS ) and cortex ( CTX ) and colocalised ( yellow ) with P2X 5 receptor staining in the inner root sheath ( IRS ). Scale bar = 50 μm

Article Snippet: Polyclonal anti-P2Y 1 and P2Y 2 antibodies were obtained from Alomone Labs (Jerusalem, Israel) and corresponded to the third extracellular loop of the P2Y 1 (AA 242–258) and P2Y 2 receptor (AA 227–244).

Techniques: Marker, Staining

Journal: bioRxiv

Article Title: Platelet-specific P2Y 1 receptor deficient mice have suppressed leukocyte recruitment in response to lipopolysaccharide

doi: 10.1101/2024.11.04.621858

Figure Lengend Snippet: Tissue taken from pups of PRYC x PRYP parents was used in PCR to determine offspring homozygous for P2Y 1 - LoxP flanked allele and hemizygous for PF4- cre (test mice) and offspring homozygous for P2Y 1 - LoxP flanked allele and a non carrier for PF4- cre (control ‘wild type’ mice). Representative PCR for LoxP and cre is shown of a litter ( A ). Platelets were taken via cardiac puncture and stained for with anti-CD41-PE conjugated antibody (platelet marker) and anti-P2Y 1 -FITC conjugated antibody to elucidate P2Y 1 expression between ‘test’ (Plt-P2Y 1 -/- ) and control (WT) platelets ( B ). In other experiments, platelets were harvested from blood, and leukocytes harvested from bone marrow of donor mice and their ability to migrate to fMLP (30nM) was measured using a transwell system ( C ) platelets, co-incubated with 100nM ADP, ( D ) neutrophils. Data expressed as means +/-SEM. n=3 ( B ) or 5-6 ( C , D ) per group. Significant difference represented: * P <0.05, ** P <0.01.

Article Snippet: FITC-anti-P2Y 1 antibody (Cat #APR-021-F) was purchased from Alomone Labs. Stromatol (Cat. #321200S) was purchased from Mascia Brunelli, Italy.

Techniques: Control, Staining, Marker, Expressing, Incubation