Ligustrazine suppresses platelet aggregation through inhibiting the activities of calcium sensors

Abstract Ligustrazine is widely used for the treatment of cardiovascular diseases in traditional Chinese medication. It has been reported that Ligustrazine decreases the concentration of intracellular calcium ions (Ca2+); however, the underlying mechanism remains unknown. In the present study, the effect of Ligustrazine on adenosine diphosphate (ADP)-induced platelet aggregation was evaluated using a turbidimetric approach. The changes in concentration of intracellular Ca2+ stimulated by ADP was measured using fluo-4, a fluorescent Ca2+ indicator dye. The mRNA expression of stromal interaction molecule l (STIM1) and Orai1, calcium sensor, was determined using real-time PCR. In addition, the protein expression of STIM1, Orai1, and serum/glucocorticoid-regulated protein kinase 1 (SGK1) was determined using Western blot analysis. The data demonstrated that Ligustrazine significantly suppressed platelet aggregation in a dose-dependent manner and reduced the concentration of intracellular Ca2+ triggered by ADP. Our data showed that Ligustrazine treatment inhibited the expression of STIM1 and Orai1 induced by ADP at both mRNA and protein levels, and suppressed the protein expression of SGK1. Taken together, our data indicated that Ligustrazine suppressed platelet aggregation by partly inhibiting the activities of calcium sensors, thereby suggesting that Ligustrazine may be a promising candidate for the treatment of platelet aggregation.

Effect of MiR-760 on Colon Cancer Metastasis by Inhibiting the Key Molecules of Calcium Ion Signal / 胃肠病学

Background: MicroRNAs (miRNAs) are considered as new biomarkers related to cancer treatment. Studies have shown that miR-760 plays an important role in the development of colorectal cancer. Aims: To investigate the regulatory effect and mechanism of miR-760 on proliferation, cycle, migration and invasion of colon cancer cells. Methods: qRT-PCR was used to detect the expression of miR-760 in colon cancer tissue and colon cancer cell lines HCT116, HT29, SW480 and SW620. The targeted relationship between miR-760 and STIM1 was verified by dual luciferase reporter gene test. Expression of STIM1 in colon cancer cells and tissue was detected by Western blotting and immunohistochemistry, respectively. CCK-8 assay was used to detect cell proliferation ability, flow cytometry was used to detect cell cycle, and Transwell assay was used to detect cell migration and invasion. The xenograft model of colon cancer in nude mice was constructed, and the effect of miR-760 on tumor growth was measured. Results: Expression of miR-760 was significantly decreased in colon cancer tissue and colon cancer cells, while expression of STIM1 was significantly increased in colon cancer tissue and colon cancer cells. MiR-760 inhibited expression of STIM1 by complementary binding to the 3'-UTR of STIM1. Overexpression of miR-760 inhibited colon cancer cell proliferation, S-phase arrest, migration and invasion, and up-regulation of STIM1 reversed the inhibitory effect. In the xenograft model of colon cancer in nude mice, overexpression of miR-760 down-regulated expression of STIM1 and inhibited tumor growth. Conclusions: MiR-760 is involved in the regulation of biological behavior of colon cancer cells by targeting STIM1, and plays a role in tumor inhibition of colon cancer.

Dose-and Time-Dependent Suppression of Rac1 and STIM1 in Acute Myeloid Leukaemia Cell Line Model

@#Introduction: Rac1 and STIM1 genes are emerging therapeutic targets for cancers. However, their roles in acute myeloid leukaemia (AML) are not well understood. The goal of this study was to evaluate the effects of dose and time on Rac1 and STIM1 knockdown in the AML cell line model (THP-1 cells). Methods: THP-1 cells were transfected with siRac1 at doses of 50, 100, and 200 nM or dsiSTIM1 at doses of 2, 5, and 10 nM. Expression level of Rac1 and STIM1 then were assessed at time points between 12 and 72 h post-transfection using real-time reverse transcription polymerase chain reaction. Results: Compared to the control, 87% Rac1 knockdown was attained with 50 nM siRac1 at 24 h post-transfection, and 70% STIM1 knockdown was achieved with 10 nM dsiSTIM1 at 48 h post-transfection. Conclusion: These results show that effective knockdown of Rac1 and STIM1 is possible, and therapy that includes Rac1 and STIM1 inhibitors eventually could provide a new and highly effective strategy for AML treatment.

Resveratrol reduces store-operated Ca2+ entry and enhances the apoptosis of fibroblast-like synoviocytes in adjuvant arthritis rats model via targeting ORAI1-STIM1 complex

BACKGROUND: Resveratrol was reported to trigger the apoptosis of fibroblast-like synoviocytes In adjuvant arthritis rats but the subcellular mechanism remains unclear. Since ER stress, mitochondrial dysfunction and oxidative stress were involved in the effects of resveratrol with imbalance of calcium bio-transmission, store operated calcium entry (SOCE), a novel intracellular calcium regulatory pathway, may also participate in this process. RESULTS: In the present study, Resveratrol was found to suppress ORAI1 expression of a dose dependent manner while have no evident effects on STIM1 expressive level. Besides, resveratrol had no effects on ATP or TG induced calcium depletion but present partly dose-dependent suppression of SOCE. On the one hand, microinjection of ORAI1 overexpressed vector in sick toe partly counteracted the therapeutic effects of resveratrol on adjuvant arthritis and serum inflammatory cytokine including IL-1, IL-6, IL-8, IL-10 and TNF-α. On the other hand, ORAI1 SiRNA injection provided slight relief to adjuvant arthritis in rats. In addition, ORAI1 overexpression partly diminished the alleviation of hemogram abnormality induced by adjuvant arthritis after resveratrol treatment while ORAI1 knockdown presented mild resveratrol-like effect on hemogram in rats model. CONCLUSION: These results indicated that resveratrol reduced store-operated Ca2+ entry and enhanced the apoptosis of fibroblast-like synoviocytes in adjuvant arthritis rats model via targeting ORAI1-STIM1 complex, providing a theoretical basis for ORAI1 targeted therapy in future treatment with resveratrol on rheumatoid arthritis.

Expression of miR-4687-5P and STIM1 in esophageal squamous cell carcinoma and their regulation mechanism / 临床与实验病理学杂志

Purpose To investigate the expression and the methylation status of miR-4687-5P and STIM1 gene in esophageal squamous cell cancer (ESCC) cell lines and ESCC tissue samples,in order to explore the correlation between miR-4687-5P and STIM1 expression,as well as whether they have a common expression regulation mechanism. Methods The qRTPCR and methylation specific PCR (MSP) methods were applied respectively to examine the expression and methylation of miR-4687-5P and STIM1 genes in ESCC cell lines (TE13, KYSE150,T. Tn) and ESCC samples,and further to analyze their correlation. Results The expression of miR-4687-5P and STIM1 genes in ESCC was significantly decreased,and consistent. The weak expression of miR-4687-5P and STIM1 genes was detected in three ESCC cell lines. After treated with 5-Aza-2'-deoxycytidine (5-Aza-Dc,a demethylation agent) ,the expression levels of these two genes were obviously increased. Meanwhile, the methylation bands were obviously weakened or disappeared. The promoter region of STIM1 gene was hypermethylated in ESCC tissues,and its methylation frequency was correlated with the expression of STIM1 and miR-4687-5P (P < 0. 01) . Conclusion miR-4687-5P and STIM1 genes are down-regulated in esophageal carcinoma,and the expression of miR-4687-5P may be regulated by the promoter of its host gene STIM1,and the hypermethylation may be one of the common mechanisms leading to down-regulatory expression of miR-4687-5P and STIM1 genes in ESCC.

Effect of STIM1 on survival and proliferation of breast cancer cells and its pre-liminary mechanism analysis / 中国免疫学杂志

Objective:To investigate the effect of STIM1 on the survival and proliferation of breast cancer cells and its preliminary mechanism analysis.Methods:Normal mammary epithelial cells MCF-10A as control,the expression of STIM1 in MCF7, HCC1569,MDA-MB-231 and BT549 breast cancer cells were detected by Western blot;STIM1 siRNA sequence(STIM1-siRNA group) were transfected into MDA-MB-231 cells,and the negative control group and the blank control group were set up,the protein expression of STIM1 in each group were detected after cells were transfected for 48 h;MTT method was used to detect cell activity cells were transfected for 24 h,48 h and 72 h;cell apoptosis was detected after cells were transfected for 48 h by flow cytometry;the mRNA expression of IL-6 and TNF-α were detected by RT-PCR;the expression of PCNA,Bcl-2,Bax,Caspase3,STAT3 and p-STAT3 protein were detected by Western blot.Results:The expression of STIM1 protein in breast cancer cells was significantly higher than that in MCF-10A cells (P<0.05);the expression of STIM1 protein in MDA-MB-231 cells transfected STIM1-siRNA was significantly lower than the control group(P<0.05);compared with the control group,cell viability in STIM1-siRNA group decreased significantly in cells were transfected for 48 h and 72 h,the apoptosis rate in 48 h was significantly increased,the expression of IL-6 and TNF-α mRNA sig-nificantly decreased,the expression of PCNA,Bcl-2 and p-STAT3 protein were significantly decreased,the expression of Bax and Caspase3 protein increased significantly.Conclusion:STIM1 gene is highly expressed in breast cancer cells.Inhibition of STIM1 expression by RNA interference can down regulate the activity of cancer cells,induce apoptosis,enhance immunity and by down regulation STAT3 signal.

Increased store-operated Ca²⁺ entry mediated by GNB5 and STIM1

Recent human genetic studies have shown that Gβ5 is related to various clinical symptoms, such as sinus bradycardia, cognitive disability, and attention deficit hyperactivity disorder. Although the calcium signaling cascade is closely associated with a heterotrimeric G-protein, the function of Gβ5 in calcium signaling and its relevance to clinical symptoms remain unknown. In this study, we investigated the in vitro changes of store-operated calcium entry (SOCE) with exogenous expression of Gβ5. The cells expressing Gβ5 had enhanced SOCE after depletion of calcium ion inside the endoplasmic reticulum. Gβ5 also augmented Stim1- and Orai1-dependent SOCE. An ORAI1 loss-of-function mutant did not show inhibition of Gβ5-induced SOCE, and a STIM1-ERM truncation mutant showed no enhancement of SOCE. These results suggested a novel role of GNB5 and Stim1, and provided insight into the regulatory mechanism of SOCE.

The mechanism research of STIM1 in breast cancer cells / 实用医学杂志

Objectives To explore the calcium signaling mechanism of STIM1 in breast cancer cells. Meth-ods After SiRNA interruption, Western blot and Transwell were used to measure protein expression of STIM1 and cell migration in MDA-MB-231 cells respectively. The relationship between STIM1 and SOCE calcium signaling were analysed by Laser confocal microscopy. Western blots were used to measure protein expression of FAK after si-lence STIM1. Results The numbers of cells without STIM1 were significantly lower than those cells with STIM1 by Transwell assay. STIM1 mediated SOCE in MDA-MB-231. Blocking SOCE might inhibite cells migration. Si-lence STIM1 did not affect the expression or activation of FAK in MDA-MB-231 cells. Conclusion STIM1 influ-ences cell migration through SOCE pathway in breast cancer cells, which is independent on the expression or activa-tion of FAK.

Expression of Orai1 and STIM1 protein in colon cancer and their clinical significance / 临床与实验病理学杂志

Purpose To investigate the expression of Orai1 and STIM1 in colon cancer and its clinical significance.Methods Immnohistochemistry was used to detect Orai1 and STIM1 protein expression level in 80 cases of colon cancer and 50 cases of normal colon epithelium.The relationships between Orai1 and STIM1 expression and prognosis were statistically analyzed.Result The expression levels of Orai1 and STIM1 in colon cancer were significantly higher than that in normal colon epithelium(P＜ 0.05).Positive expression of Orai1 and STIM1 was correlated with the TNM stage and lymph node metastasis (P ＜ 0.05),but not correlated with gender,age,and differentiation(P ＞0.05).There was a significant positive correlation between Orai1 and STIM1 expression(P =0.001,rs =0.349).Univariate analysis showed that the expression of Orai1 protein,STIM1 protein,TNM stage and lymph node metastasis were significant prognostic factors for colon cancer patients.Conclusion In colon cancer,both Orail and STIM1 proteins may have synergetic functions and promote the development of the tumor,and could be used as a novel biological indicator of anticancer therapy and prognosis.

Orai1 and Orai3 in Combination with Stim1 Mediate the Majority of Store-operated Calcium Entry in Astrocytes

Astrocytes are non-excitable cells in the brain and their activity largely depends on the intracellular calcium (Ca²⁺) level. Therefore, maintaining the intracellular Ca²⁺ homeostasis is critical for proper functioning of astrocytes. One of the key regulatory mechanisms of Ca²⁺ homeostasis in astrocytes is the store-operated Ca²⁺ entry (SOCE). This process is mediated by a combination of the Ca²⁺-store-depletion-sensor, Stim, and the store-operated Ca²⁺-channels, Orai and TrpC families. Despite the existence of all those families in astrocytes, previous studies have provided conflicting results on the molecular identification of astrocytic SOCE. Here, using the shRNA-based gene-silencing approach and Ca²⁺-imaging from cultured mouse astrocytes, we report that Stim1 in combination with Orai1 and Orai3 contribute to the major portion of astrocytic SOCE. Gene-silencing of Stim1 showed a 79.2% reduction of SOCE, indicating that Stim1 is the major Ca²⁺-store-depletion-sensor. Further gene-silencing showed that Orai1, Orai2, Orai3, and TrpC1 contribute to SOCE by 35.7%, 20.3%, 26.8% and 12.2%, respectively. Simultaneous gene-silencing of all three Orai subtypes exhibited a 67.6% reduction of SOCE. Based on the detailed population analysis, we predict that Orai1 and Orai3 are expressed in astrocytes with a large SOCE, whereas TrpC1 is exclusively expressed in astrocytes with a small SOCE. This analytical approach allows us to identify the store operated channel (SOC) subtype in each cell by the degree of SOCE. Our results propose that Stim1 in combination with Orai1 and Orai3 are the major molecular components of astrocytic SOCE under various physiological and pathological conditions.

Huangqi Danshen decoction attenuates isoproterenol-induced myocardial remodeling via STIM1, TRPC1, CaN and NFATc3 pathways in rats / 中国中药杂志

To investigate the inhibitory effect of Huangqi Danshen decoction (HDD) on isoproterenol (ISO)-induced myocardial remodeling and explore its effect on STIM1, TRPC1, CaN and NFATc3 expressions. ISO (2.5 mg•kg⁻¹•d⁻¹×14 d) was given by subcutaneous injection to establish myocardial remodeling models in rats, and then were randomly divided into control group, ISO model group, HDD5 group (HDD 5 g•kg⁻¹•d⁻¹+ISO), and HDD10 group (HDD 10 g•kg⁻¹•d⁻¹+ISO). After intervention for 4 weeks, the heart mass index (HW/BW) and the left ventricular mass index (LVW/BW) were calculated; the structure of myocardium was observed by echocardiography; the pathological changes of myocardium were observed by HE staining; levels of BNP, CaN and CaM kinases II in serum were detected by ELISA, and the protein expression levels of STIM1, TRPC1, p-CaN, p-NFATc3, and NFATc3 in left ventricular tissues were detected by Western blot. The results showed that the HW/BW and LVW/BW in ISO group were greater than those in HDD5 group and HDD10 group (P<0.05); Echocardiography showed that HDD inhibited ISO-induced increase in LVEDD and LVESD; ELISA results showed that HDD could significantly inhibit the increase of BNP, CaN and CaM kinases II levels in serum of rats with ISO-induced myocardial remodeling (P<0.01). Western blot results showed that STIM1, TRPC1, p-CaN, p-NFATc3 and NFATc3 expression levels were increased in the myocardial tissues of ISO group rats, and after HDD administration, the above expression levels were decreased in group ISO, HDD for myocardial tissue after administration of STIM1, TRPC1, p-CaN, p-NFATc3 and NFATc3 expression decreased (P<0.05). Our findings indicated that HDD can attenuate the myocardial remodeling induced by ISO, and its mechanism may be related to down-regulating the expression levels of STIM1, TRPC1, CaM kinases II, p-CaN/CaN and p-NFATc3/NFATc3.

Expression and function of store-operated calcium channels in human circulating fibrocytes / 中国病理生理杂志

AIM:To investigate the expression and function of store-operated calcium channels ( SOCC) in human circulating fibrocytes.METHODS:Peripheral blood mononuclear cells ( PBMCs) were isolated and cultured in ser-um-free media.After 7 d, the PBMCs differentiated into fibrocytes.RT-PCR and real-time PCR were performed to deter-mine the mRNA expression of ORAI1-3 and STIM1-2 in the fibrocytes.SOCC inhibitor SKF-96365 was used to elucidate the role of SOCC in the differentiation of fibrocytes.RESULTS:The results of real-time PCR showed that the mRNA ex-pression of ORAI1-3 and STIM1-2 was positive in cultured fibrocytes.SKF-96365 (10μmol/L) significantly inhibited the differentiation of fibrocytes.CONCLUSION:SOCC-related proteins ORAI1-3 and STIM1-2 are abundantly expressed in the fibrocytes, and may play an important role in the differentiation of these cells.

Expression of STIM1 gene in human hypopharyngeal carcinoma cell line FaDu and its effect on FaDu cell apoptosis / 局解手术学杂志

Objective To study the expression of STIM 1 gene in human hypopharyngeal carcinoma cell line FaDu and its effect on FaDu cell apoptosis .Methods Lentivirus infection was used to knock STIM 1 down in FaDu cells .Group STIM1-siRNA: the expression of STIM1 in FaDu cell was inhibited by STIM 1-siRNA lentivirus .Group control:FaDu cells were infected by negative control siRNA lentivirus . Real-Time PCR was applied to identify the efficacy of lenticirus infection and the expression of STIM 1 in FaDu cells.Western blot was used to identify the expression of STIM 1 protein after lenticirus infection .Flow cytometry assay was performed to detect the apoptosis of FaDu cells in the two groups.The data were statistically analyzed with SPSS 17.0 software.Results Compared with GAPDH (Ct=12.08 ±0.05),the expression of STIM1 in FaDu cells was significant expressed (Ct=22.21 ±0.05,P<0.001).Real-Time PCR analysis the relative mRNA expression of STIM1 in FaDu cells of control group and STIM 1-siRNA group were (1.00 ±0.08) and (0.12 ±0.01) respectively (P<0.001). Western blot showed that the expression of STIM 1 gene and protein in FaDu cells were inhibited significantly after STIM 1-siRNA lentiviral in-fection,which was in accordance with the results of Real-Time PCR analysis.Flow cytometry assay showed that the siRNA-mRNA group had a higher apoptosis percentage (9.81 ±0.56)% compared to the control group (4.36 ±1.32)%,with statistically significant difference (P<0.05).Conclusion STIM1 gene correlated significantly with FaDu cell apoptosis .It inhibits apoptosis of FaDu cells ,and it may be a potential diagnostic and therapeutic target for the hypopharyngeal carcinoma .

STIM1 promotes arterial smooth muscle cells proliferation by regulating Akt/mTOR pathway / 中国药理学通报

Aim To investigate the expression of stro-mal interaction molecule 1 (STIM1) in rat pulmonary arterial hypertension ( PAH ) tissues and effects of STIM1 on arterial muscle cells proliferation. Methods PAH was induced by a single intraperitoneal injec-tion of MCT at a dose of 60 mg·kg - 1 . The mRNA or protein expressions of STIM1 in monocrotaline-induced pulmonary hypertensive rats were measured by real-time PCR or Western blot, respectively. The arterial smooth muscle cells A7R5 were transiently transfected with STIM1 plasmids to prepare STIM1 overexpressed cells. Cell proliferations were detected by using CCK-8 kits. The expressions of Akt/ mTOR pathway molecules of A7R5 were measured by Western blot. Results The right ventricular systolic blood pressure ( RVSP) and right ventricular mass index ( RVMI ) were markedly elevated in MCT-treated rats (P < 0. 01) in comparison to control rats. The mRNA and protein ex-pression levels of STIM1 in monocrotaline-induced pul-monary hypertensive rats were 2. 19 and 1. 66 folds of control rats, respectively. STIM1 were transiently over-expressed in cultured A7R5. Cells transfected with STIM1 grew more quickly than non-transfected control. Overexpression of STIM1 significantly increased the phosphorylation of Akt, mTOR, p70-S6K, and 4E-BP1, but did not change their protein expression lev-els. Conclusion STIM1 are over-expressed in rat PAH tissues. Overexpression of STIM1 can promote ar-terial smooth muscle cells proliferation by regulating Akt/ mTOR pathway.

Regulation of autophagy by store-operated calcium channel in arterial smooth muscle cells / 中国药理学通报

Aim To investigate the effect of store-oper-ated calcium channel( SOCC) on autophagy in rat arte-rial smooth muscle cells A7 R5 . Methods Lentiviruses containing STIM1 or Orai1 gene were packaged in 293 T cells and then were used to infect rat arterial smooth muscle cells A7 R5 . The expression levels of STIM1 , Orai1 and Beclin 1 , a critical autophagy-regu-lating protein, of lentivirus-infected A7R5 cells, were detected by Western-blot. Autophagy in lentivirus-in-fected A7 R5 cells was induced by starvation or rapamy-cin, an inhibitor of mammalian target of rapamycin ( mTOR ) . Autophagy marker LC3 of these cells was detected by Western-blot. Results The constructions of vector pLV-STIM1 and pLV-Orai1 were confirmed by restriction enzymes digestion analysis. Compared with the control group, expressions of STIM1 or Orai1 protein was significantly increased after lentivirusLV-STIM1 and LV-Orai1infection, whereas the expressions of autophagy related protein Beclin-1 were down-regu-lated. Starvation or rapamycin stimulated A7R5 auto-phagy but overexpression of STIM1 or Orai1 significant-ly inhibited starvation or rapamycin induced autoph-agy. Conclusion Overexpression of store-operated calcium channel components STIM1 and/or Orai1 in rat arterial smooth muscle cells A7 R5 inhibit autoph-agy. This mechanism might contribute to the develop-ment of pulmonary arterial hypertension.

Orai1 Expression Is Closely Related with Favorable Prognostic Factors in Clear Cell Renal Cell Carcinoma

Store-operated calcium (Ca2+) entry (SOCE) is the principal Ca2+ entry route in non-excitable cells, including cancer cells. We previously demonstrated that Orai1 and STIM1, the molecular components of SOCE, are involved in tumorigenesis of clear cell renal cell carcinoma (CCRCC). However, a clinical relevance of Orai1 and STIM1 expression in CCRCC has been ill-defined. Here, we investigated the expression of Orai1 and STIM1 in CCRCC, and compared their expression with clinico-pathological parameters of CCRCC and the patients' outcome. Immunohistochemical staining for Orai1 and STIM1 was performed on 126 formalin fixed paraffin embedded tissue of CCRCC and western blot analysis for Orai1 was performed on the available fresh tissue. The results were compared with generally well-established clinicopathologic prognostic factors in CCRCC and patient survival. Membrane protein Orai1 is expressed in the nuclei in CCRCC, whereas STIM1 shows the cytosolic expression pattern in immunohistochemical staining. Orai1 expression level is inversely correlated with CCRCC tumor grade, whereas STIM1 expression level is not associated with tumor grade. The higher Orai1 expression is significantly associated with lower Fuhrman nuclear grade, pathologic T stage, and TNM stage and with favorable prognosis. The expression level of STIM1 is not correlated with CCRCC grade and clinical outcomes. Orai1 expression in CCRCC is associated with tumor progression and with favorable prognostic factors. These results suggest that Orai1 is an attractive prognostic marker and therapeutic target for CCRCC.

Inhibition of Store-Operated Calcium Entry Protects Endothelial Progenitor Cells from H₂O₂-Induced Apoptosis

Store-operated calcium entry (SOCE), a major mode of extracellular calcium entry, plays roles in a variety of cell activities. Accumulating evidence indicates that the intracellular calcium ion concentration and calcium signaling are critical for the responses induced by oxidative stress. The present study was designed to investigate the potential effect of SOCE inhibition on H₂O₂-induced apoptosis in endothelial progenitor cells (EPCs), which are the predominant cells involved in endothelial repair. The results showed that H₂O₂-induced EPC apoptosis was reversed by SOCE inhibition induced either using the SOCE antagonist ML-9 or via silencing of stromal interaction molecule 1 (STIM1), a component of SOCE. Furthermore, SOCE inhibition repressed the increases in intracellular reactive oxygen species (ROS) levels and endoplasmic reticulum (ER) stress and ameliorated the mitochondrial dysfunction caused by H₂O₂. Our findings provide evidence that SOCE inhibition exerts a protective effect on EPCs in response to oxidative stress induced by H₂O₂ and may serve as a potential therapeutic strategy against vascular endothelial injury.

Silence of STIM1 attenuates the proliferation and migration of EPCs after vascular injury and its mechanism

OBJECTIVE@#To investigate the effect of stromal interaction molecule 1(STIM1) knockdown on the proliferation and migration of endothelial progenitor cells (EPCs) after vascular injury and its mechanism.@*METHODS@#The rat bone marrow derived EPCs were divided into three groups: adenovirus negative control (group NSC), rat STIM1 adenovirus vector transfection group (group si/rSTIM1) and rat &human recombinant STIM1 adenovirus transfection group (group si/rSTIM1+hSTIM1). The STIM1 expressions in each group were detected by reverse transcription PCR after transfection; the cell proliferation was tested by [(3)H] thymidine incorporation assay ((3)H-TdR); Cell cycle was analyzed by flow cytometry; the cells' migration activity was detected by Boyden assay; Calcium ion concentration was detected by using laser confocal method.@*RESULTS@#48 h later after transfection, the expression level of STIM1 in si/rSTIM1 cells was significantly lower than that in NSC group (0.21 ± 0.12 vs 1.01 ± 0.01, P<0.05); EPCs that stayed in G1 phase in si/rSTIM1 group [(93.31 ± 0.24)%] were significantly more than that in NSC group [(78.03 ± 0.34)%, P<0.05]; EPCs' migration activity in si/rSTIM1 group (10.03±0.33) was significantly lower than that in NSC group: (32.11 ± 0.54, P<0.05); EPCs calcium ion concentration changes in EPCs in si/rSTIM1 group (38.03 ± 0.13) was significantly lower than that in NSC group (98.11 ± 0.34, P<0.05). While there was no significant difference between si/rSTIM1+hSTIM1 group and NSC group on the four indexes above.@*CONCLUSIONS@#Silence of STIM1 attenuates EPCs proliferation and migration after vascular injury, by mediating the calcium ion concentration in EPCs.

STIM1/Orai1 may be a new target for the prevention and treatment of ischemic cerebrovascular disease / 国际脑血管病杂志

In recent years,two important component proteins of the calcium release-activated calcium channel (CRAC) were identified fromDrosophila cells by RNA interference technique,including the calcitum sensor stromal interaction molecule 1 (STIM1) on the endoplasmic reticulum and the CRAC channel protein Orail on the cell membrane.Studies have shown that STIM 1 and Orail have regulatory effects on vascular smooth muscle cells,platelets,vascular endothelial cells and other cells.They play important roles in the aspects of vascular smooth muscle cell phenotypic modulation,hemostasis,thrombosis,and neovascularization.It shows that they both may be closely associated with ischemic cerebrovascular disease.This article reviews the advances in research on STIM1 and Orail proteins in ischemic cerebrovascular disease in order to investigate the possibility of STIM1/Orai1 as a new target in the prevention andtreatment of ischemic cerebrovascular

An ER Locating Protein Named RCN2 Interacts With STIM1-Orai1 Complex / 生物化学与生物物理进展

STIM1 is recognized as an ER Ca2+ sensor of calcium release-activated calcium (CRAC) channel that is constructed by membrane protein Orai1, However, this regulatory system may also be regulated by other proteins. Reticulocalbin 2 (RCN2) was purified and identified from STIM1-Orai1 complex. Confocal microscopy revealed that RCN2 co-localized with STIM1 in ER before and after Ca2+ store depletion. Single cell [Ca2+]I measurements of RCN2 EF hands mutant showed slight influence on SOC electrophysiological characters. Furthermore, a novel collar form aggregation of RCN2 surrounding STIM1 clusters suggested that RCN2 potentially plays a role of structure maintenance in STIM1 clustering.