p2rx2  (Alomone Labs)

Journal: Journal of Clinical Medicine

Article Title: The Expression of F2RL1, P2RX2, P2RX3 and P2RY2 in the Esophagus of Patients with Gastroesophageal Reflux Disease and Their Relationship to Reflux Symptoms—A Pilot Study

doi: 10.3390/jcm14061884

Figure Lengend Snippet: The expression of F2RL1 , P2RX2 , P2RX3 and P2RY2 in healthy controls ( n = 9) and patients with GERD ( n = 53). The Mann–Whitney test was used to compare the values; ## p < 0.01 vs. respective control.

Article Snippet: The reaction mixture consisted of cDNA, TaqManTM Gene Expression Master Mix, TaqManTM Gene Expression Assays ( F2RL1: Hs00608346_m1, P2RX2: Hs00247255_m1, P2RX3: Hs01125554_m1 and P2RY2: Hs00925146_m1) and RNase-free water, with a final volume of 10 μL.

Techniques: Expressing, MANN-WHITNEY, Control

Journal: Journal of Clinical Medicine

Article Title: The Expression of F2RL1, P2RX2, P2RX3 and P2RY2 in the Esophagus of Patients with Gastroesophageal Reflux Disease and Their Relationship to Reflux Symptoms—A Pilot Study

doi: 10.3390/jcm14061884

Figure Lengend Snippet: The expression of F2RL1 ( A ), P2RX2 ( B ), P2RX3 ( C ) and P2RY2 ( D ) in healthy controls ( n = 9) and GERD patients with NERD ( n = 32–36) or ERD ( n = 12–15). The Kruskal–Wallis test was used to compare the values.

Article Snippet: The reaction mixture consisted of cDNA, TaqManTM Gene Expression Master Mix, TaqManTM Gene Expression Assays ( F2RL1: Hs00608346_m1, P2RX2: Hs00247255_m1, P2RX3: Hs01125554_m1 and P2RY2: Hs00925146_m1) and RNase-free water, with a final volume of 10 μL.

Techniques: Expressing

Journal: Journal of Clinical Medicine

Article Title: The Expression of F2RL1, P2RX2, P2RX3 and P2RY2 in the Esophagus of Patients with Gastroesophageal Reflux Disease and Their Relationship to Reflux Symptoms—A Pilot Study

doi: 10.3390/jcm14061884

Figure Lengend Snippet: Heatmap showing the relation between the expression of F2RL1 , P2RX2 , P2RX3 and P2RY2 and DIS score ( n = 24–30). The Spearman rank correlation test was used to analyze the association between the expression of genes and DIS score vs. respective control.

Article Snippet: The reaction mixture consisted of cDNA, TaqManTM Gene Expression Master Mix, TaqManTM Gene Expression Assays ( F2RL1: Hs00608346_m1, P2RX2: Hs00247255_m1, P2RX3: Hs01125554_m1 and P2RY2: Hs00925146_m1) and RNase-free water, with a final volume of 10 μL.

Techniques: Expressing, Control

Journal: Journal of Clinical Medicine

Article Title: The Expression of F2RL1, P2RX2, P2RX3 and P2RY2 in the Esophagus of Patients with Gastroesophageal Reflux Disease and Their Relationship to Reflux Symptoms—A Pilot Study

doi: 10.3390/jcm14061884

Figure Lengend Snippet: Correlations between the expression of F2RL1 ( A ), P2RX2 ( B ), P2RX3 ( C ) and P2RY2 ( D ) and HRQL in patients with GERD. The Spearman rank test was used to analyze the association between the expression of genes and GERD-HRQL vs. respective control.

Article Snippet: The reaction mixture consisted of cDNA, TaqManTM Gene Expression Master Mix, TaqManTM Gene Expression Assays ( F2RL1: Hs00608346_m1, P2RX2: Hs00247255_m1, P2RX3: Hs01125554_m1 and P2RY2: Hs00925146_m1) and RNase-free water, with a final volume of 10 μL.

Techniques: Expressing, Control

Journal: The Journal of Reproduction and Development

Article Title: AVPV Kiss1 neuron-specific knockdown of purinergic P2X2 receptor suppresses LH surge and ovulation in Kiss1-Cre rats

doi: 10.1262/jrd.2024-046

Figure Lengend Snippet: DNA sequences of shRNAs against rat P2rx2

Article Snippet: To examine the effects of AVPV kisspeptin neuron-specific knockdown of P2rx2 mRNA expression on E2-induced LH surge, Kiss1 - Cre female rats were bilaterally injected with AAV-U6- P2rx2 -shRNA-flox-CMV-EGFP (AAV- P2rx2 -shRNA-EGFP, 3.1 × 10 13 viral genome/ml; n = 8) or AAV-U6-scrambled-shRNA-flox-CMV-EGFP (AAV-scrambled-shRNA-EGFP, 2.5 × 10 13 viral genome/ml; n = 4) into the AVPV region through a cannula (C315I, Plastics One, Roanoke, VA, USA) with its tip at 0.12 mm posterior to the bregma, ± 0.5 mm from the midline, and 8.2 mm below the surface of the skull according to the coordinates of a rat brain atlas [ ].

Techniques: Sequencing, shRNA

Journal: The Journal of Reproduction and Development

Article Title: AVPV Kiss1 neuron-specific knockdown of purinergic P2X2 receptor suppresses LH surge and ovulation in Kiss1-Cre rats

doi: 10.1262/jrd.2024-046

Figure Lengend Snippet: Evaluation of the efficacy of siRNA in suppressing P2rx2 mRNA expression in the mHypo-A51 cell line. A: mHypoA-51 cells were transfected with one of three P2rx2 siRNA constructs (siRNA#1, #2, or #3) or negative-control siRNA (siRNA-NC). The cells were incubated for 24 or 48 h and then treated with E2 for 4 h prior to collection of the cells to analyze P2rx2 mRNA levels. P2rx2 mRNA expression levels were measured by real-time RT-PCR. B: P2rx2 mRNA expression levels in mHypo-A51 cells 24 or 48 h after the transfection of P2rx2 siRNA or siRNA-NC. Opti-MEM, non-transfected control. Values are the mean ± SEM. Numbers in each column indicate the number of samples used. Circles on the bar charts indicate the individual data. Different letters indicate statistically significant differences ( p < 0.05; one-way ANOVA followed by the Bonferroni test). C: The target sequence of the rat P2rx2 shRNA corresponding to the mouse P2rx2 target sequence of siRNA#2. The base-mismatches are highlighted in magenta. The sequences are shown from 5' to 3', left to right. siRNA, small interfering RNA; SEM, standard error of the mean; ANOVA, analysis of variance.

Article Snippet: To examine the effects of AVPV kisspeptin neuron-specific knockdown of P2rx2 mRNA expression on E2-induced LH surge, Kiss1 - Cre female rats were bilaterally injected with AAV-U6- P2rx2 -shRNA-flox-CMV-EGFP (AAV- P2rx2 -shRNA-EGFP, 3.1 × 10 13 viral genome/ml; n = 8) or AAV-U6-scrambled-shRNA-flox-CMV-EGFP (AAV-scrambled-shRNA-EGFP, 2.5 × 10 13 viral genome/ml; n = 4) into the AVPV region through a cannula (C315I, Plastics One, Roanoke, VA, USA) with its tip at 0.12 mm posterior to the bregma, ± 0.5 mm from the midline, and 8.2 mm below the surface of the skull according to the coordinates of a rat brain atlas [ ].

Techniques: Expressing, Transfection, Construct, Negative Control, Incubation, Quantitative RT-PCR, Control, Sequencing, shRNA, Small Interfering RNA

Journal: The Journal of Reproduction and Development

Article Title: AVPV Kiss1 neuron-specific knockdown of purinergic P2X2 receptor suppresses LH surge and ovulation in Kiss1-Cre rats

doi: 10.1262/jrd.2024-046

Figure Lengend Snippet: AVPV kisspeptin neuron-specific knockdown of P2rx2 suppressed E2-induced LH surge in OVX Kiss1 - Cre female rats. A: AAV vectors carrying P2rx2 -specific or scrambled shRNA were injected into the bilateral AVPV regions of Kiss1 - Cre female rats. The area outlined by the dotted line indicates the Cre-dependent expression of shRNAs in AVPV kisspeptin neurons of Kiss1 - Cre rats. The AAV vectors were designed to express either P2rx2 -specific shRNA or non-specific scrambled shRNA under the human U6 promoter in a Cre-dependent manner, along with an EGFP reporter under a separate CMV promoter. The shRNA is conditionally expressed in AVPV kisspeptin neurons through Cre-mediated deletion, which excises the stop cassette positioned between the sense (shRNA S.) and antisense (shRNA A.S.) sequences. Fourteen days after the injection of AAV-U6- P2rx2 -shRNA-flox-CMV-EGFP (AAV- P2rx2 -shRNA-EGFP, n = 8) or AAV-U6-scrambled-shRNA-flox-CMV-EGFP (AAV-scrambled-shRNA-EGFP, n = 4), the Kiss1 - Cre rats were ovariectomized (OVX) and implanted with diestrus levels of estradiol-17β (low E2) for 5 days and proestrus levels of E2 (high E2) for 2 days. Blood samples were collected from freely moving rats every 1 h from 1000 h to 2100 h. B: Images of brain sections of representative Kiss1 - Cre rats transfected with AAV-scrambled-shRNA-EGFP or AAV- P2rx2 -shRNA-EGFP showing EGFP expression in the AVPV-preoptic region but not in the ARC. Arrowheads indicate magnified EGFP-expressing cells in the insets. 3V, third ventricle; scale bars, 100 µm. C: Representative images of Kiss1 -expressing (magenta) and EGFP-immmunopositive (green) cells in the AVPV of Kiss1-Cre rats injected with scrambled shRNA (left) or P2rx2 shRNA (right). Insets show magnified images of Kiss1 -expressing EGFP-immunopositive cells indicated by arrowheads. D: The number of Kiss1 -expressing (magenta) and Kiss1 -expressing EGFP-immunopositive (striped) cells in the AVPV of Kiss1-Cre rats injected with scrambled shRNA or P2rx2 shRNA. The number of Kiss1 -expressing cells, as well as the number of Kiss1 -expressing EGFP-immunopositive cells in the AVPV of OVX + high E2 Kiss1-Cre rats in the P2rx2 shRNA group (n = 8) and scrambled shRNA group (n = 4), were not significantly different (P = 0.116 and P = 0.054, respectively; Student’s t -test). E: Plasma LH profiles in OVX + high E2 Kiss1 - Cre rats treated with AVPV administration of P2rx2 shRNA (n = 8, indicated by red circles) or scrambled shRNA (n = 4, indicated by blue circles). Values with * in the P2rx2 -shRNA-treated group were significantly lower than scrambled-shRNA-treated controls at each time point (P < 0.05, Two-way repeated measures ANOVA followed by simple main effects). F: The AUC of plasma LH levels was significantly lower (* P = 0.0129, Student’s t -test) in Kiss1 - Cre rats injected with P2rx2 shRNA compared to the corresponding value of the control Kiss1 - Cre rats injected with scrambled shRNA. Values represent the mean ± SEM. Numbers in each column indicate the number of animals used. Circles on the bar chart indicate the individual data. AVPV, anteroventral periventricular nucleus; LH, luteinizing hormone; AAV, adeno-associated virus; shRNA, short hairpin RNA; EGFP, enhanced green fluorescent protein; ARC, arcuate nucleus; AUC, area under the curve; ANOVA, analysis of variance; SEM: standard error of the mean.

Article Snippet: To examine the effects of AVPV kisspeptin neuron-specific knockdown of P2rx2 mRNA expression on E2-induced LH surge, Kiss1 - Cre female rats were bilaterally injected with AAV-U6- P2rx2 -shRNA-flox-CMV-EGFP (AAV- P2rx2 -shRNA-EGFP, 3.1 × 10 13 viral genome/ml; n = 8) or AAV-U6-scrambled-shRNA-flox-CMV-EGFP (AAV-scrambled-shRNA-EGFP, 2.5 × 10 13 viral genome/ml; n = 4) into the AVPV region through a cannula (C315I, Plastics One, Roanoke, VA, USA) with its tip at 0.12 mm posterior to the bregma, ± 0.5 mm from the midline, and 8.2 mm below the surface of the skull according to the coordinates of a rat brain atlas [ ].

Techniques: Knockdown, shRNA, Injection, Expressing, Transfection, Clinical Proteomics, Control, Virus

Journal: The Journal of Reproduction and Development

Article Title: AVPV Kiss1 neuron-specific knockdown of purinergic P2X2 receptor suppresses LH surge and ovulation in Kiss1-Cre rats

doi: 10.1262/jrd.2024-046

Figure Lengend Snippet: AVPV kisspeptin neuron-specific knockdown of P2rx2 disrupted spontaneous LH surge, ovulation, and estrous cyclicity in ovary-intact Kiss1 - Cre female rats. A: After the AVPV injection of AAV- P2rx2- shRNA-EGFP or AAV-scrambled-shRNA-EGFP, vaginal smears were monitored daily for at least 21 days, and the blood samples were collected to examine plasma LH levels on the day of proestrus or corresponding day and ovulated oocytes were collected on the day after the blood sampling. B: Estrous cyclicity from 8 days before AAV injection until at least 3 weeks after AAV scrambled shRNA or P2rx2 shRNA injection in Kiss1 - Cre rats. Four out of five ovary-intact control Kiss1 - Cre rats showed normal 4-day estrous cycles throughout the experimental period, while three of four ovary-intact AVPV kisspeptin neuron-specific P2rx2 knocked-down Kiss1 - Cre rats showed persistent diestrus. E, estrus; P, proestrus; D, diestrus. Individual plasma LH profiles measured in ovary-intact rats treated with either AVPV scrambled shRNA (C) or P2rx2 shRNA (D). E: The AUC of plasma LH levels in Kiss1-Cre rats that were in proestrus on the day of blood sampling in scrambled shRNA- (animal #1–#4, n = 4) or P2rx2 shRNA-treated Kiss1 - Cre rat (#7, n = 1). F: AVPV administration of P2rx2 shRNA blocked ovulation in the ovary-intact Kiss1 - Cre rat (#7, n = 1), whereas control rats (#1–#4, n = 4) showed ovulation at estrus. G: Images of ovaries in representative Kiss1 - Cre rats with AVPV injection of scrambled shRNA (#4, left) or P2rx2 shRNA (#7, right). Scale bar, 2 mm. Arrowheads, corpora lutea. H: Ovarian weights (total of left and right) of the Kiss1 - Cre rat injected with P2rx2 shRNA (#4, n = 1) and the control rats (#1–#4, n = 4) injected with scrambled shRNA. Values are represented as mean ± SEM. Circles on the bar charts indicate the individual data. AVPV, anteroventral periventricular nucleus; LH, luteinizing hormone; AAV, adeno-associated virus; shRNA, short hairpin RNA; EGFP, enhanced green fluorescent protein; SEM, standard error of the mean.

Article Snippet: To examine the effects of AVPV kisspeptin neuron-specific knockdown of P2rx2 mRNA expression on E2-induced LH surge, Kiss1 - Cre female rats were bilaterally injected with AAV-U6- P2rx2 -shRNA-flox-CMV-EGFP (AAV- P2rx2 -shRNA-EGFP, 3.1 × 10 13 viral genome/ml; n = 8) or AAV-U6-scrambled-shRNA-flox-CMV-EGFP (AAV-scrambled-shRNA-EGFP, 2.5 × 10 13 viral genome/ml; n = 4) into the AVPV region through a cannula (C315I, Plastics One, Roanoke, VA, USA) with its tip at 0.12 mm posterior to the bregma, ± 0.5 mm from the midline, and 8.2 mm below the surface of the skull according to the coordinates of a rat brain atlas [ ].

Techniques: Knockdown, Injection, shRNA, Clinical Proteomics, Sampling, Control, Virus