Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: STIM expression in the Xenopus oocyte and its downregulation by as-STIM injection. A) shows the RT-PCR amplification of products that corresponded to the size expected for either stim1 or stim2 in native oocytes (CNT); the corresponding amplicons were absent in oocytes from the same batch that had been injected with either as-STIM1 or as-STIM2 48 h before the assay. The rps2 amplicon indicates the reaction efficiency, and -RT and H 2 O lanes correspond to negative controls, either RNA without RT, or to the reaction mix without a cDNA template, respectively. B) STIM1 and STIM2 were identified by Western blot analysis in protein extracts from oocytes (Oo) or mouse brain (MB, positive control) using either NH-STIM1 (left panel) or COOH-STIM2 (right panel) as antibody. C) A similar analysis as in B was made for batches of oocytes injected with H 2 O as control (CNT), or with as-STIM1 or as-STIM2 48 h before the protein extraction, in which cases proteins were eliminated. (in all cases 10 oocytes per condition).

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Expressing, Injection, Reverse Transcription Polymerase Chain Reaction, Amplification, Western Blot, Positive Control, Protein Extraction

Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: Knockdown of STIM expression in oocytes co-injected with GPCR mRNA. A) RT-PCR amplification of stim1 , stim2 , or rps2 in batches of oocytes injected with H 2 O (CNT) or with cRNA (50 ng per oocyte) coding for either P2Y8 or M1 GPCR. In oocytes co-injected with as-STIM1 or as-STIM2 (50 ng per oocyte) together with P2Y8 or M1 cRNA, the corresponding STIM amplicon was downregulated. Control reactions illustrate specificity; rps2 amplicons are positive controls, and -RT and H 2 O lanes show negative controls. B) Similar groups of oocytes as in A) were assayed using the Western blot technique; in this case oocytes from the same donor injected with one GPCR mRNA (P2Y8 or M1) alone, or co-injected with as-STIM1, were tested with NH-STIM1, while as-STIM2-injected oocytes were probed with COOH-STIM2. In both as-STIM groups SERCA was used as gel-loading control. C) The graph shows the densitometric analysis of bands, summarizing the results obtained in different preparations of 10 oocytes per group and repeated in 3–5 frogs. Both PCR products and bands detected by Western blot (WB) were analyzed for batches of oocytes injected with H 2 O (CNT) or with either 50 ng as-STIM1 or as-STIM2 alone (native group). Similar analysis was made for batches of control oocytes injected with P2Y8 or M1 cRNA alone, and oocytes from the same frogs co-injected with either as-STIM or as-STIM together with the GPCR cRNA. Optical density units (ODU) for each band were normalized against the value obtained in the corresponding CNT conditions (*p < 0.01).

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Expressing, Injection, Reverse Transcription Polymerase Chain Reaction, Amplification, Western Blot

Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: I osc and T in responses activated by agonist stimulation. A) Strength of I osc elicited by first agonist application did not change by knockdown of STIM1 or STIM2, compared with that obtained in CNT oocytes; top traces are typical responses elicited by ACh, similar responses were obtained by FBS or ATP applications, and the graph shows the average I osc responses obtained in oocytes held at −60 mV. B) Record illustrating the activation of T in current obtained in an oocyte expressing the M1 receptor by a single ACh (100 μM) application for 40 s (acute protocol). Oocytes were held at −10 mV while being superfused with NR solution and stepped to −100 mV for 4 s every 40 s; sudden hyperpolarization generated T in current responses that follow consistent kinetics with a peak amplitude response at 280–360 s (c); after that the response was washed out with a similar time course. C) Shows the T in current during the steps from −10 to −100 mV indicated with letters in panel B) . D) A similar T in current response elicited in an oocyte from the same frog that was pre-incubated with 1 μM ACh for 4 h (long-lasting protocol), then monitored with the same electrical recording parameters and stimulated with 100 μM ACh. E) Shows the T in responses indicated with the same letters as in D) . In this protocol T in current was consistently activated from the beginning of the record, and a transient inhibition of the response was noted during application of the agonist ( b) ; after that, T in recovered and remained fully activated for a long period of time. Similar responses were obtained using oocytes expressing P2Y receptors and stimulating with ATP.

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Activation Assay, Expressing, Generated, Incubation, Inhibition

Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: Specific STIM knockdown by oocyte injection of as-STIM differentially decreased the T in current. A) Oocytes induced to express M1, P2Y8, or P2Y2 receptors were stimulated with either ACh or ATP (100 μM), and LPAR in native oocytes were stimulated by FBS (1:1000 dilution); the resulting T in currents (CNT, gray areas) were compared with the T in obtained in oocytes from the corresponding group that were also injected with 50 ng as-STIM1 (superimposed black traces); all responses were monitored 48–72 h after oocyte injection. B) The graph shows the results obtained using the different experimental conditions illustrated in A) . C) In a set of experiments similar to those shown in A) , T in currents were monitored, and the peak amplitudes of non-injected CNT oocytes were compared with those of oocytes injected (48–72 h before recording) with 50 ng as-STIM2 and stimulated with the agonists. D) The graph shows the results obtained using the different experimental conditions illustrated in C) . Bars correspond to the mean (± SEM) of the T in peak amplitude of 10–15 oocytes from 5–6 frogs (*p < 0.01, as-STIM vs. CNT).

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Injection

Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: Oocyte injection with COOH-STIM2 antibody produced a strong potentiation of T in current response. A) T in current responses were monitored in two conditions: non-loaded oocytes (CNT) and oocytes loaded with COOH-STIM2 antibody (ab-loaded). T in responses were elicited by ACh, FBS, or ATP application, depending on the receptor to be stimulated. In all cases, a strong potentiation of the response was observed in ab-loaded oocytes. B) Oocytes stimulated by ACh (M1) loaded with denatured COOH-STIM2 had control-like responses, while NH-STIM2 or NH-STIM1 loading did not produce T in potentiation. C) The graph shows the results obtained using the different experimental conditions illustrated in A and B ; each bar corresponds to the mean (± SEM) of the T in peak amplitude normalized against the CNT current of 10–15 oocytes from 3–6 frogs (*p < 0.01).

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Injection, Produced

Journal: BMC Physiology

Article Title: Differential role of STIM1 and STIM2 during transient inward ( T in ) current generation and the maturation process in the Xenopus oocyte

doi: 10.1186/s12899-014-0009-x

Figure Lengend Snippet: Effect of as-STIM2 on GVBD and oocyte membrane characteristics during maturation induced by progesterone. A) The maturation process promoted by progesterone (10 μM) was analyzed in uninjected oocytes, or in oocytes injected 72 h prior to the assay with either as-STIM1 or as-STIM2, and compared with control oocytes in the absence of progesterone. GVBD was quantified after 8–12 h in presence of progesterone (10 oocytes per group, repeated using 3 different frogs) and is normalized against the value observed in uninjected oocytes. B) Resting membrane potential was monitored 8–12 h after addition of progesterone in the same groups of oocytes (n = 3-5, repeated in 3 frogs) as in A) . C) The input membrane resistance (Rϕ) was estimated over the range from −80 to −20 mV in the different oocyte groups treated in the same conditions. Control groups, without progesterone, included both uninjected and antisense-injected oocytes. In all cases, values for as-STIM2-injected groups were different from as-STIM1-injected or uninjected groups (*p < 0.01).

Article Snippet: The antibody denoted NH-STIM1 (Alomone, Jerusalem, Israel) was directed against a region of the amino-terminus of the STIM1 protein, and the antibodies denoted NH-STIM2 (Alomone, Jerusalem, Israel) and COOH-STIM2 (ProSci Inc., Poway CA, USA) were against the amino and carboxy termini, respectively, of STIM2.

Techniques: Injection

Journal: Biochimica et biophysica acta

Article Title: Downregulation of STIM2 improves neuronal survival after traumatic brain injury by alleviating calcium overload and mitochondrial dysfunction.

doi: 10.1016/j.bbadis.2015.08.014

Figure Lengend Snippet: Fig. 2. Downregulation of STIM2 improves neuronal survival after TNI in vitro. After infection of Scr-shRNA, STIM1-shRNA or STIM2-shRNA for 48 h, the efficiency of downregulation was assessed for both the mRNA (A) and protein levels (B and C). After infection with lentivirus particle for 72 h and subjected to TNI for 24 h, the neuronal viability was assessed by LDH assay (D), and the neuron apoptosis was detected by TUNEL staining (E: red, TUNEL-positive; blue, Hoechst). The apoptotic index was then calculated (E). Neurons in the uninjured group were not subjected to lentiviral infection or injury. Data are means ± SEM; *, p b 0.05 vs. normal or uninjured group, and **, p b 0.05 vs. Scr-shRNA + TNI group, n = 4. Scale bars = 100 μm.

Article Snippet: The membraneswere then blockedwith 5% skimmilk and incubated at 4 °C overnight with the appropriate primary antibody: STIM1, 1:2000; STIM2, 1:1000; β-actin, 1:2000 (CST, Danvers, MA, USA).

Techniques: In Vitro, Infection, shRNA, Lactate Dehydrogenase Assay, TUNEL Assay, Staining

Journal: Biochimica et biophysica acta

Article Title: Downregulation of STIM2 improves neuronal survival after traumatic brain injury by alleviating calcium overload and mitochondrial dysfunction.

doi: 10.1016/j.bbadis.2015.08.014

Figure Lengend Snippet: Fig. 7. Downregulation of STIM2 reduced neuronal apoptosis, decreased brain lesion volume and improved neurological deficit recovery after TBI. Mice were injected with lentivirus particle, followed by CCI. The CCI injury zone (gray box) and microscope imaging zone (red dotted box) were shown in the sketch map of a coronal section of the mice brain (A). 3 d after TBI, the apoptotic rate was assessed (A and B). n = 3 for each group, scale bar = 100 μm. Lesion volume was measured 7 d post-TBI by cresyl violet staining (C). Neurological deficits were assessed using a standard rotarod test (D). n = 5 (sham group), 5 (Scr-shRNA group), and 6 (STIM2-shRNA group). *p b 0.05 vs. Scr-shRNA + CCI group.

Article Snippet: The membraneswere then blockedwith 5% skimmilk and incubated at 4 °C overnight with the appropriate primary antibody: STIM1, 1:2000; STIM2, 1:1000; β-actin, 1:2000 (CST, Danvers, MA, USA).

Techniques: Injection, Microscopy, Imaging, Staining, shRNA

Journal: The Korean Journal of Physiology & Pharmacology : Official Journal of the Korean Physiological Society and the Korean Society of Pharmacology

Article Title: Differential expression of ORAI channels and STIM proteins in renal cell carcinoma subtypes: implications for metastasis and therapeutic targeting

doi: 10.4196/kjpp.24.126

Figure Lengend Snippet: (A, C) FPKM for Orai3 according to normal and RCC grades (Stage I, II, III, and IV) in clear cell RCC (ccRCC) and papillary RCC (pRCC), respectively. Box whisker plots express the minimum, median, and maximum values of FPKM (expressed as log(expr + 1)), where FPKM was adjusted using a scaling factor instead of total mapped reads to remove a million units. ***p ≤ 0.001 of one-way ANOVA (normal vs . tumor stages). (B, D) Kaplan–Meler survival analysis comparing females (left) and males (right) in the low and high mRNA levels of Orai3 in ccRCC and pRCC, respectively. The cut-off value for Orai3 in ccRCC is 13.5, and in pRCC is 16.87. (E, G) Correlation analysis between gene copy number (log2-transformed values) of STIM2 and Orai3 in ccRCC (total n = 355) and pRCC (total n = 241), respectively. The linear regression line with a 95% confidence interval and the Spearman r correlation coefficient. (F, H) Kaplan–Meler survival analysis comparing males and females with high Orai3 and low STIM2 mRNA levels in ccRCC and pRCC, respectively. The cut-off value for Orai3 is 13, and for STIM2 is 4.5. p-values were determined by a log-rank test (B, D, F, and H).

Article Snippet: The primary antibodies used for immunoblotting were: β-actin (1:3,000, #sc-47778) and Cyclin D1 (1:200, #sc-246) from Santa Cruz Biotechnology; STIM1 (1:1,000, #S6072) from Sigma Aldrich; p-FAK (Tyr397) (1:1,000, #8556) and t-FAK (1:1,000, #13009) from Cell Signaling Technology Inc.; and STIM2 (1:500, #21192-1-AP) from ProteinTech.

Techniques: Whisker Assay, Transformation Assay

Journal: Frontiers in Physiology

Article Title: Store-operated calcium entry and the localization of STIM1 and Orai1 proteins in isolated mouse sinoatrial node cells

doi: 10.3389/fphys.2015.00069

Figure Lengend Snippet: List of antibody used in the study .

Article Snippet: STIM2 , ProSci (4123) , Polyclone , Rabbit , 1:400 , 1:1000.

Techniques: Staining, Western Blot, Transduction

Journal: Frontiers in Physiology

Article Title: Store-operated calcium entry and the localization of STIM1 and Orai1 proteins in isolated mouse sinoatrial node cells

doi: 10.3389/fphys.2015.00069

Figure Lengend Snippet: Expression of STIM and Orai molecules in mouse cardiac myocytes. (A) Total RNA was isolated from pacemaker cells (SAN), atrial myocytes (Atrium), ventricular myocytes (Ventricle) and spleen. Transcripts of Stim1, Stim2, Orai1, Orai2, and Orai3 in cardiac myocytes were compared with the spleen as positive control. The housekeeping gene Hypoxanthine–Guanine Phosphoribosyltransferase (HPRT1) was used as an internal control. (B) Protein levels of STIM1, STIM2, Orai1, and Orai3 were examined in total protein extracts from SAN region (SAN), atrium (Atrium), ventricle (Ventricle) and spleen and determined by Western Blot analysis. The nuclear protein histone was used as loading control. (C) Bar graph showing quantitative western blot analysis of STIM1 protein levels in spleen and heart with heart samples normalized to spleen as 1 ( n = 3 gels, P < 0.05, ANOVA). (D) Confocal images of an isolated pacemaker cell shows expression of HCN4 (green in color), a positive marker for pacemaker cells. The cell also positively stained with anti-Orai1 antibody (red in color). The nucleus is stained blue with DAPI. A merged image on the left shows the co-localization of HCN4 and Orai1 along the surface membrane (color in yellow). (E) Representative confocal images of an isolated pacemaker cell labeled with STIM1 (green in color) and DAPI (blue in color). When perfused with normal Ca 2+ containing solution, positive STIM1 staining was distributed both intracellularly and in the cell periphery as showed in the top panel. When perfused with Ca 2+ -free solution containing thapsigargin, STIM1 peripheral localization was enhanced, as shown in the bottom panel.

Article Snippet: STIM2 , ProSci (4123) , Polyclone , Rabbit , 1:400 , 1:1000.

Techniques: Expressing, Isolation, Positive Control, Control, Western Blot, Marker, Staining, Membrane, Labeling

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Protein expression of STIM2, but not STIM1, is increased in IPAH patients’ PASMC. (a) Photograph showing that PASMC isolated from normotensive control subjects (NPH) and IPAH patients are morphologically comparable. (b and c) Representative Western blot images ( left panels ) and summarized data ( right panels ) for STIM1 (b) and STIM2 (c) proteins in PASMC isolated from IPAH patients’ lung tissues or normotensive control patient lung tissues (NPH). β-tubulin was used as a loading control. Summarized data of STIM1 (b, right panel) or STIM2 (c, right panel) protein expression level (mean±SE) in IPAH-PASMC (n=6) and NPH-PASMC (n=6). Graph shows protein expression of STIM1 and STIM2 normalized to an average level in NPH-PASMC. * P <0.05 versus NPH. STIM2 protein expression level was increased in IPAH-PASMC, but STIM1 protein expression level was not changed in IPAH-PASMC

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Expressing, Isolation, Control, Western Blot

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Dose-dependent knockdown of STIM2 in IPAH patients’ PASMC with siRNA. (a) Representative Western blot image of STIM2 protein in IPAH-PASMC treated with siRNA against STIM2 (siSTIM2; with doses of 4, 8, and 16 μl) or control scrambled siRNA (indicated by ‘ C ’). β-tubulin (β-tub) was used as a loading control. (b) Quantification of STIM2 protein expression level in IPAH-PASMC treated with control scrambled siRNA (‘ C ’) or 4, 8, and 16 μl of siRNA-STIM2. Values were normalized to the β-tub level at the start and then normalized to the level of cells treated with control scrambled siRNA. The STIM2 protein expression level in IPAH-PASMC was decreased in a dose-dependent manner by siRNA targeting STIM2

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Knockdown, Western Blot, Control, Expressing

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Upregulated protein expression of STIM2 is necessary for enhanced SOCE in IPAH patients’ PASMC. (a) Representative records show CPA-induced changes in [Ca 2+ ] cyt in the absence or presence of extracellular Ca 2+ in NPH-PASMC (NPH, left panel), IPAH-PASMC (IPAH, middle panel), and IPAH-PASMC treated with siRNA targeting STIM2 (IPAH + siSTIM2, right panel). SOCE (indicated by the CPA-induced increase in [Ca 2+ ] cyt when extracellular Ca 2+ is restored) is induced by the passive depletion of SR Ca 2+ using 10 βM CPA. NPH-PASMC (left panel) and IPAH-PASMC (middle panel) were treated with scrambled siRNA as a control. (b) Summary data (mean±SE) showing changes in CPA-induced increase in [Ca 2+ ] cyt , immediately following the re-addition of Ca 2+ after store depletion (indicative of SOCE) in NPH-PASMC (black bar), IPAH-PASMC (red bar), and IPAH-PASMC, treated with siRNA-STIM2 (blue bar). ** P >0.01 (NS, not significant) versus NPH. (c) Western blot image of STIM2 protein expression in NPH-PASMC (NPH), IPAH-PASMC (IPAH), and IPAH-PASMC, treated with siRNA-STIM2 (IPAH + siSTIM2). NPH-PASMC (black) and IPAH-PASMC (red) were treated with scrambled siRNA

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Expressing, Control, Western Blot

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Knockdown of STIM2 mitigates enhanced proliferation in IPAH patients’ PASMC. (a) Summary data (mean±SE) showing cell numbers for NPH-PASMC and IPAH-PASMC before (0 hour) and after 72 hours of culture in the growth media. *** P <0.001 versus 0 hour. (b) Summary data showing the increase in cell numbers after 72 hours in NPH-PASMC (blue) and IPAH-PASMC (red). ** P <0.01 versus NPH. IPAH patients’ PASMC showed a significantly greater increase in cell number after 72 hours compared to NPH-PASMC. (c) Summary data showing the increase in cell number after 72 hours in NPH-PASMC (left panel) and IPAH-PASMC (right panel) treated with either scrambled siRNA (siC) or siRNA against STIM2 (siS2). Decreasing the protein expression level of STIM2 did not affect the increase in cell number after 72 hours in NPH-PASMC (left panel), but signifi cantly inhibited the increase in cell number after 72 hours in IPAH-PASMC (right panel). ** P <0.01 versus siC. (d) Summary data showing the total number of IPAH-PASMC cultured in growth media at 0, 24, 48, and 72 hours. The cells were treated with scrambled siRNA (square symbols) or siRNA against STIM2 (circle symbols). IPAH-PASMC treated with siRNA-STIM2 had a slower rate of proliferation than IPAH-PASMC treated with scrambled siRNA. The growth curves in IPAH-PASMC treated with scrambled siRNA and siRNA against STIM2 were signifi cantly different

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Knockdown, Expressing, Cell Culture

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Dose-dependent overexpression of STIM2 in NPH-PASMC transiently transfected with siRNA. (a) Representative Western blot image of STIM2 protein in NPH-PASMC. STIM2 was overexpressed using 1, 2, or 4 μg of DNA. Vector DNA was used as a control (‘ C ’). β-tubulin (β-tub) was used as a loading control. (b) Quantifi cation of STIM2 protein expression level in NPH-PASMC transfeted with vector DNA (‘ C ’) and 1, 2, or 4 μg of STIM2-DNA. Values were normalized to β-tubulin. STIM2 protein expression level was increased in a dose-dependent manner in NPH-PASMC transfected with STIM2

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Over Expression, Transfection, Western Blot, Plasmid Preparation, Control, Expressing

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Overexpression of STIM2 is not sufficient to enhance SOCE in normal PASMC. (a) Representative records showing changes in [Ca 2+ ] cyt before, during, and after the application of CPA (10 °M), in the absence or presence of extracellular Ca 2+ in the vector control (left panel) and STIM2- transfected (right panel) NPH-PASMC. SOCE was induced by the passive depletion of SR Ca 2+ , using CPA. (b) Summary data (mean±SE) showing changes in [Ca 2+ ] cyt immediately following the addition of CPA in the absence of extracellular Ca 2+ , which reflects SR Ca 2+ release (SR Ca 2+ Release, left panel), in control (black) and STIM2-transfected (red) NPH-PASMC. There was no difference in magnitude of SR Ca 2+ release between the two groups. Summary data (mean±SE) showing changes in [Ca 2+ ] cyt immediately following the re-addition of Ca 2+ after CPA-induced store depletion (indicative of SOCE, right panel) in control (black) and STIM2-overexpressing PASMC counting the cells at time 0 and 72 hours. Transfection (red). There was no difference in magnitude of SOCE between the two groups

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Over Expression, Plasmid Preparation, Control, Transfection

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Overexpression of STIM2 does not increase NPH-PASMC proliferation. (a) Summary data for NPH-PASMC proliferation after 72 hours. Cell number was counted at time zero (0 hours: white bar) and 72 hours (72 hours: black bar) and shown as the multiple of 10 4 . After 72 hours, both vector control and STIM2-overexpressing NPH-PASMC increased in cell number. * P < 0.05 versus open bars (0 hour). (b) Summary data (mean±SE) showing the increase in cell number after 72 hours in NPH-PASMC. STIM2-overexpressing PASMC (STIM2) did not show a greater increase in cell number after 72 hours compared to control (vector). Cell counts were repeated four times (n=4)

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Over Expression, Plasmid Preparation, Control

Journal: Pulmonary Circulation

Article Title: STIM2 contributes to enhanced store-operated Ca 2+ entry in pulmonary artery smooth muscle cells from patients with idiopathic pulmonary arterial hypertension

doi: 10.4103/2045-8932.78106

Figure Lengend Snippet: Hypoxia increases protein expression of STIM2 and Orai2 in rat PASMC. (a) and (b). Representative Western blot images (left panels) and summarized data (mean±SE) showing protein levels (right panels) for Orai2 (a) and STIM2 (b) protein in rat PASMC exposed to normoxia (Nor, room air supplemented with 5% CO 2 ) or hypoxia (Hyp, 3% O 2 and 5% CO 2 balanced in N 2 for 48 hours). β-tubulin (β-tub) was used as a loading control. Values were normalized to μ-tubulin. Hypoxia increased the protein expression of Orai2 and STIM2 in rat PASMC. ** P <0.01 versus Nor

Article Snippet: The STIM2 plasmid was ordered from Addgene, as provided by Dr. Tobais Meyer.

Techniques: Expressing, Western Blot, Control