Journal: Asian Journal of Andrology

Article Title: Decreased AKAP4/PKA signaling pathway in high DFI sperm affects sperm capacitation

doi: 10.4103/aja202329

Figure Lengend Snippet: The DNA sequences of target gene

Article Snippet: SPA17 rabbit polyclonal antibody, phosphorylated protein kinase B (p-AKT) rabbit polyclonal antibody, and AKT rabbit polyclonal antibody were purchased from Proteintech (Rosemont, CA, USA).

Techniques:

Journal: Asian Journal of Andrology

Article Title: Decreased AKAP4/PKA signaling pathway in high DFI sperm affects sperm capacitation

doi: 10.4103/aja202329

Figure Lengend Snippet: Relative expression of HSPA2, PCNA, SPA17 protein in the L-DFI and H-DFI groups. ( a ) Western blot analysis and ( b ) densitometric ratio histogram of HSPA2, PCNA and SPA17 with 4 semen samples of L-DFI (L-1 to L-4) and H-DFI (H-1 to H-4) groups each. Western blot analysis of SPA17 with 4 semen samples of L-DFI and H-DFI groups each, ( c ) L-9 to L-12, and H-9 to H-12, and ( d ) L-5 to L-8, and H-5 to H-8 in the L-DFI and H-DFI groups. We selected 12 semen samples in the L-DFI (L-1 to L-12) and H-DFI (H-1 to H-12) groups each. ** P < 0.01. Data is presented as mean ± standard deviation. HSPA2: heat shock protein A2; PCNA: proliferating cell nuclear antigen; SPA17: sperm protein antigen 17; H-DFI: high sperm DNA fragmentation index; L-DFI: low sperm DNA fragmentation index; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: SPA17 rabbit polyclonal antibody, phosphorylated protein kinase B (p-AKT) rabbit polyclonal antibody, and AKT rabbit polyclonal antibody were purchased from Proteintech (Rosemont, CA, USA).

Techniques: Expressing, Western Blot, Standard Deviation

Journal: Asian Journal of Andrology

Article Title: Decreased AKAP4/PKA signaling pathway in high DFI sperm affects sperm capacitation

doi: 10.4103/aja202329

Figure Lengend Snippet: Relative expressions of PKA-associated protein in L-DFI and H-DFI groups. Western blot analysis of PKA-associated protein ( a ) with 3 semen samples of L-DFI (L-13 to L-15) and H-DFI (H-13 to H-15) groups, respectively, and ( b ) with 4 semen samples of L-DFI (L-15 to L-19) and H-DFI (H-15 to H-19) groups, respectively. Histograms demonstrated the densitometric ratio of ( c ) PKARII, ( d ) AKAP4, ( e ) SPA17, ( f ) PI3K, ( g ) p-AKT, ( h ) AKT, and ( i ) p-ERK in L-DFI and H-DFI groups. We selected 7 semen samples in the L-DFI (L-13 to L-19) and H-DFI (H-13 to H-19) groups, respectively. * P < 0.05. Data are presented as mean ± standard deviation. PKA: protein kinase A; AKAP: a-kinase anchoring proteins; PI3K: phosphoInositide-3 kinase; AKT: protein kinase B; p-AKT: phosphorylated protein kinase B; p-ERK: extracellular signal-regulated kinase of phosphorylation; H-DFI: high sperm DNA fragmentation index; L-DFI: low sperm DNA fragmentation index; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: SPA17 rabbit polyclonal antibody, phosphorylated protein kinase B (p-AKT) rabbit polyclonal antibody, and AKT rabbit polyclonal antibody were purchased from Proteintech (Rosemont, CA, USA).

Techniques: Western Blot, Standard Deviation, Phospho-proteomics

Journal: Asian Journal of Andrology

Article Title: Decreased AKAP4/PKA signaling pathway in high DFI sperm affects sperm capacitation

doi: 10.4103/aja202329

Figure Lengend Snippet: The protein expression of SPA17, AKAP4 and PKARII after H 2 O 2 treatment of semen from the L-DFI group. Western blot analysis of SPA17, AKAP4 and PKARII ( a ) after 20 µmol l −1 and 40 µmol l −1 H 2 O 2 treatment, and ( b ) after 40 µmol l −1 and 80 µmol l −1 H 2 O 2 treatment. Histograms demonstrated the densitometric ratio of ( c ) AKAP4, ( d ) PKARII, and ( e ) SPA17. * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001. Data are presented as mean ± standard deviation. SPA17: sperm protein antigen 17; PKA: protein kinase A; AKAP: A-kinase anchoring proteins; L-DFI: low sperm DNA fragmentation index; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: SPA17 rabbit polyclonal antibody, phosphorylated protein kinase B (p-AKT) rabbit polyclonal antibody, and AKT rabbit polyclonal antibody were purchased from Proteintech (Rosemont, CA, USA).

Techniques: Expressing, Western Blot, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: Reciprocal Regulatory Interaction between TRPV1 and Kinin B1 Receptor in a Rat Neuropathic Pain Model

doi: 10.3390/ijms21030821

Figure Lengend Snippet: Cellular distribution of B1R in the ipsilateral Dorsal Horn (iDH). Representative microphotographs by immunofluorescence of B1R (green, B – E ) relative to GFAP (red, B1 ), Iba-1 (red, C1 ), CGRP (red, D1 ) and IB4+ (red, E1 ) in the spinal cord (SC) of PSNL rats. Control spinal cord for B1R (A) expression exhibits very weak-to-no signal. Expression of CGRP ( A1 ), GFAP ( A2 ) and Iba-1 ( A3 ) is moderate in the spinal cord of control animals. B1R immunolabelling is colocalized with astrocytes (arrows, GFAP, B2 ) and non-peptide fibers (arrows, IB4+, E2 , E3 ), but not with microglia (Iba-1, C2 ) and peptide fibers (CGRP, D2 ) in the spinal dorsal horn of PSNL rats. E2 is enlarged in E3 as indicated by the white square and arrow. Scale bar = 120 µm (all panels, 10×), except 60 µm ( E3 , 20×).

Article Snippet: In order to block non-specific labeling, samples were incubated with 5% serum in PBS-T (5 mL normal donkey serum, 0.5 mL 20% Triton X-100, 10 mL 10× PBS, 84.5 mL ddH2O) in a humidified chamber at RT for 1 h. Then, sections were incubated overnight at 4 °C with the blocking buffer containing one of the following primary antibodies: mouse monoclonal anti-TRPV1, VR-1 (sc-398417, Santa Cruz Biotechnology Inc, Dallas, USA) for TRPV1 immunostaining [ ], chicken polyclonal anti-glial fibrillary acid protein, GFAP (ab4674, Abcam, Toronto, ON, Canada) for astrocytes immunostaining [ ], mouse monoclonal anti-ionized calcium binding adapter molecule, Iba-1 (MA5-27726, TermoFisher scientific, Rockford, AZ, USA) for microglia immunostaining [ ], rabbit monoclonal anti-calcitonin gene related peptide IgG, CGRP (sp-17, Abcam, Toronto, ON, Canada) for peptide sensory fibers immunostaining [ ], fluorescein labeled GSL I isolectin B4, IB4+ (FL-1201, Vector laboratories, Burlingame, CA, USA) for non-peptide sensory fibers immunostaining [ ], polyclonal Goat anti-IL1-β IgG (PA5-46956, Polyclonal Goat IgG, TermoFisher scientific, Rockford, AZ, USA) to identify IL-1β [ ] and a polyclonal rabbit antiserum for B1R (Biotechnology Research Institute, Montreal, QC, Canada) for kinin B1R immunostaining [ , ].

Techniques: Immunofluorescence, Expressing

Journal: International Journal of Molecular Sciences

Article Title: Reciprocal Regulatory Interaction between TRPV1 and Kinin B1 Receptor in a Rat Neuropathic Pain Model

doi: 10.3390/ijms21030821

Figure Lengend Snippet: Cellular distribution of TRPV1 in the ipsilateral Dorsal Horn (iDH). Representative microphotographs by immunofluorescence of TRPV1 (red, A – D ) relative to IB4+ (green, A1 ), CGRP (green, B1 ) GFAP (green, C1 ), Iba-1 (green, D1 ) in the spinal cord (SC) of PSNL rats. TRPV1 colocalizes weakly with peptide (CGRP, B2 ), and more abundantly with non-peptide fibers (IB4+, A2 ) and astroglia (GFAP, C2 C3 ), but not with microglia ( D2 ). The same markers are shown in iDH of control rats. Scale bar = 120 µm (all panels, 10×), 60 µm ( C – C2 and D – D2 , 20×), 75 µm ( C3 , 60×).

Article Snippet: In order to block non-specific labeling, samples were incubated with 5% serum in PBS-T (5 mL normal donkey serum, 0.5 mL 20% Triton X-100, 10 mL 10× PBS, 84.5 mL ddH2O) in a humidified chamber at RT for 1 h. Then, sections were incubated overnight at 4 °C with the blocking buffer containing one of the following primary antibodies: mouse monoclonal anti-TRPV1, VR-1 (sc-398417, Santa Cruz Biotechnology Inc, Dallas, USA) for TRPV1 immunostaining [ ], chicken polyclonal anti-glial fibrillary acid protein, GFAP (ab4674, Abcam, Toronto, ON, Canada) for astrocytes immunostaining [ ], mouse monoclonal anti-ionized calcium binding adapter molecule, Iba-1 (MA5-27726, TermoFisher scientific, Rockford, AZ, USA) for microglia immunostaining [ ], rabbit monoclonal anti-calcitonin gene related peptide IgG, CGRP (sp-17, Abcam, Toronto, ON, Canada) for peptide sensory fibers immunostaining [ ], fluorescein labeled GSL I isolectin B4, IB4+ (FL-1201, Vector laboratories, Burlingame, CA, USA) for non-peptide sensory fibers immunostaining [ ], polyclonal Goat anti-IL1-β IgG (PA5-46956, Polyclonal Goat IgG, TermoFisher scientific, Rockford, AZ, USA) to identify IL-1β [ ] and a polyclonal rabbit antiserum for B1R (Biotechnology Research Institute, Montreal, QC, Canada) for kinin B1R immunostaining [ , ].

Techniques: Immunofluorescence