Progress of STIM1, a dynamic calcium signal transducer, #br# in cardiovascular diseases

Stromal interaction molecule 1 (STIM1) is a transmembrane protein of the endoplasmic reticulum (ER), a Ca²⁺ transducer in ER that activates the store-operated calcium channel. Through Orai1 protein, STIM1 adjusts the intracellular and extracellular calcium concentration. This way is called a store-operated Ca²⁺ entry. STIM1 plays a key role in phenotypic transformation of vascular smooth muscle cells, proliferation of endothelial cells, myocardial hypertrophy and myocardial fibrosis to regulate lots of cardiovascular diseases, such as atherosclerosis, congestive heart failure and systemic hypertension. STIM1 is closely related to cardiovascular diseases through calcium signal. The research progress of STIM1 in cardiovascular diseases is mainly discussed in this article.     

Calcium-sensing receptor modulates pulmonary artery tension through G-protein-PLC-IP3 pathways

AIM: To observe the role of calcium-sensing receptor (CaSR) in the regulation of pulmonary artery tension. METHODS: The intracellular calcium concentration ([Ca²⁺]_i) was detected by laser-scanning confocal microscopy, and the pulmonary artery tension was determined by the pulmonary arterial ring technique. RESULTS: Increased levels of [Ca²⁺]_o or Gd³⁺ (an agonist of CaSR) induced the increase in [Ca²⁺]_i and pulmonary artery constriction in a concentration-dependent manner. Additionally, the effects of Ca²⁺ and Gd³⁺ were inhibited by U73122 and D609 (specific inhibitor of PLC), and 2-APB and heparin (specific antagonist of IP₃ receptor). However, U73343 (U73122 inactive analogue) did not take effect. CONCLUSION: CaSR may be involved in the regulation of pulmonary artery tension by increasing [Ca²⁺]_i through G-protein-PLC-IP₃ pathway.     

Effects of tetrandrine and fructose-1, 6-diphosphate on the elevated intrasynaptosomal [Ca2+]i induced by excitatory amino acids

AIM: To study the effects of tetrandrine(Tet) and fructose-1, 6-diphosphate(FDP) on the elevated intrasynaptosomal [Ca²⁺]_i induced by excitatory amino acids(EAA). METHODS: A rapid method for preparing synaptosomes was used, and intrasynaptosomal free calcium([Ca²⁺]_i) was measured by using the fluorescent indicator quin-2. RESULTS: L-glutamate(Glu, 100 μmol/L), aspartate(Asp, 100 μmol·L^-1), N-methy1-D-aspartate(100 μmol/L) and Glu(50 μmol/L) plus Asp(50 μmol/L) all elevated intrasynaptosomal [Ca²⁺]_i in a dose-dependent manner. Pretreatment with Tet(10, 30, 60 μmol/L), FDP(15, 30, 75, 150 μmol/L), MK-801(10, 20 μmol/L) and Tet(15, 30 μmol/L) plus FDP(15, 30 μmol/L) all attenuated the increase in intrasynaptosomal [Ca²⁺]_i induced by EAAs mentioned as above in a dose-dependent manner, and the effect of Tet plus FDP was most significant. CONCLUSION: Both Tet and FDP inhibited a rise in intrasynaptosomal [Ca²⁺]_i induced by EAAs, which may be one of mechanisms that Tet and FDP pretect cerebral tissues against ischemia injury.     

Salvia miltiorrhiza antagonized the alterations of contraction and intracellular calcium of rat ventricular cardiomyocytes induced by anoxia and reoxygenation

AIM:To study the effect of salvia miltiorrhiza (SM) on cell contraction and intracellular calcium of enzymatically isolated rat ventricular myocytes during normoxia and anoxia/reoxygenation.METHODS:Contraction and intracel ular calcium were determined with video tracking system and spectrofluorometric method,and the chemical anoxic method was employed. RESULTS:The ±dL/dt_max, dL of cell contraction and the amplitude of [Ca²⁺]_i in the cardiomyocytes following SM treatment were decreased in a dose-dependent manner. During anoxia, the ±dL/dt_max, dL and amplitude of [Ca²⁺]_i were decreased, while the diastolic Ca²⁺ level was elevated compared with control group. All the contractile parameters and the diastolic Ca²⁺ level were back toward pretreatment values during reoxygenation, but could not return to control level. After the treatment with SM (3 g/L), ±dL/dt_max and dL of cell contraction and the amplitude of [Ca²⁺]_i were higher and the diastolic Ca²⁺ level was lower than those in anoxia/reoxygenation group. CONCLUSION:SM antagonized effects of anoxia and reoxygenation on cell contraction and intracellular calcium in isolated ventricular myocytes.     

Role of calcium ion in the cyotoxicity and DNA fragment induction in HL-60 cells by 6F isolated from Pteris semipinnata L.

AIM: To investigate the changes of cytosolic free calcium concentration([Ca²⁺]_i) and expression of Bcl-2 in HL-60 cells treated by 6F isolated from Pteris semipinnata L.(PSL), and to discuss the relations between calcium ion and cytotoxicity and DNA fragment induction effects of 6F. METHODS: HL-60 cells were used as in vitro model. [Ca²⁺]_i was measured on fluorescent spectrophotometry using Fura-2/AM as Ca²⁺ indicator. Bcl-2 expressing level was measured by flow cytometry. Tetrazolium salt(MTT) and diphenylamine staining methods were applied for cytotoxicity assay and DNA fragmentation detection, respectively. RESULTS: [Ca²⁺]_i increased obviously in a dose and time dependent manner after treated HL-60 cells with 6F. 6F decreased the expressing level of Bcl-2. Adding 2 mmol/L Ca²⁺ to the medium, or 1 mmol/L EDTA to chelate Ca²⁺, or 4 μmol/L calcium ionophore A 23187 to increase the concentration of cytosolic Ca²⁺, the DNA fragment induction by 6F was not affected, whereas the cytotoxicity of 6F was enhanced. 250 μmol/L Zn²⁺ attenuated the DNA fragment induction, and the cytotoxicity of 6F against HL-60 cells was enhanced significantly. CONCLUSION: It was speculated that the decreased expressing of Bcl-2 by compound 6F was related to increased [Ca²⁺]_i in HL-60 cells, and DNA fragment induction was possibly catalyzed by Ca²⁺ - independent DNase.     

Role of potassium channels in the regulation of intracellular free Ca2+ concentration of pulmonary artery smooth muscle cells in rats

AIM: To investigate the role of potassium channels in the regulation of intracellular free calcium concentration ( [Ca²⁺]_i) of pulmonary artery smooth muscle cells (PASMCs) in rats. METHODS: The fluorescence Ca²⁺ indicator Fura-2/AM was used to observe [Ca²⁺]_i of rat PASMCs in normal and chronic hypoxic condition. The influences of potassium channels on PASMCs proliferation were assessed by MTT assay. RESULTS: 1. In normoxic condition, [Ca²⁺]_i was (156.91±8.60) nmol/L, and in hypoxic condition, [Ca²⁺]_i was (294.01±16.81) nmol/L. 2. In normoxic condition, the voltage-dependent K⁺-channel antagonist 4-aminopyridine (4AP), but not the Ca²⁺-activated K⁺-channel antagonist tetraethylammonium (TEA) and the ATP-sensitive K⁺-channel antagonist glibenclamide (Glib) increased [Ca²⁺]_i. 3. In hypoxic condition, 4AP and TEA caused the rise in [Ca²⁺]_i , but Glib had no effect on [Ca²⁺]_i. 4. MTT assay showed that 4AP increased the value of absorbing light degree (A value) in normoxic and hypoxic condition (0.582±0.062,0.873±0.043,respectively, P＜0.01), TEA increased A value only in hypoxic condition, and Glib had no effect on the proliferation of PASMCs. CONCLUSIONS: K_V plays an important role in the regulation of [Ca²⁺]_i and proliferation of PASMCs. K_Ca serves as distinct responsive roles in the regulation of proliferation of PASMCs in hypoxic condition. K_ATP has no effect on [Ca²⁺]_i and proliferation of PASMCs in normoxic and hypoxic conditions.     

Amiloride slows down calpain-mediated ABCA1 degradation through inhibition of hypoxia-induced NHE1 expression

AIM: To examine the effects of hypoxia on sodium-hydrogen exchange 1(NHE1) expression, intracellular Ca²⁺ concentration ([Ca²⁺]_i) and calpain activity, and to explore the effect of amiloride on adenosine triphosphate-binding cassette transporter A1(ABCA1) degradation and its calpain-related mechanism. METHODS: RAW264.7 cells were exposed to hypoxia for 0 h, 12 h, 24 h and 48 h. The cell viability was measured by MTT assay and the expression of NHE1 at mRNA and protein levels was detected by real-time PCR and Western blot. [Ca₂₊]_i was analyzed by flow cytometry. Calpain activity was assessed by the method of Suc-LLVY-aminoluciferin. Furthermore, the protein levels of ABCA1 in the RAW264.7 cells exposed to hypoxia for 24 h were determined after 6 h or 12 h treatment with NHE1 inhibitor amiloride in the presence of cycloheximide. ABCA1 protein levels and calpain activity were detected after 12 h incubation with calpain inhibitor ALLN or intracellular calcium-chelating agent BAPTA. RESULTS: Hypoxia inhibited the cell viability in a time-dependent manner. Hypoxia up-regulated the mRNA and protein expression of NHE1, and increased [Ca²⁺]_i and calpain activity. Hypoxia increased the degradation of ABCA1 and amiloride slowed down the ABCA1 degradation. ALLN or BAPTA increased ABCA1 protein level and decreased calpain activity. CONCLUSION: NHE1 inhibitor amiloride attenuates the calpain-mediated degradation of ABCA1, indicating that hypoxia-induced NHE1 might, at least in part, participate in the ABCA1 degradation.     

Effect of interleukin-2 on intracellular calcium levels in rat ventricular myocytes during anoxia and reoxygenation

AIM: To investigate the effect of interleukin-2(IL-2) on the intracellular calcium in electrically stimulated adult rat ventricular myocytes during anoxia and reoxygenation. METHODS: The isolated cardiac ventricular myocytes were exposed to 5 min anoxia followed by 10 min reoxygenation. Chemical anoxia was introduced by Krebs-Henseleit(K-H) solution containing 10^-3 mol/L sodium dithionite. The spectrofluorometric method was used to verify intracellular calcium transient with fura-2/AM as calcium fluorescence probe. RESULTS: It was shown that during anoxia, the amplitude of Ca²⁺ transient was decreased, diastolic [Ca²⁺]_i, time to peak and time to relaxation of Ca²⁺ transient were increased. All the parameters were got back but did not returned to the pre-anoxia level during reoxygenation. IL-2 at 2×10⁵ U/L administrated during anoxia aggravated the effect of rexoxygenation on [Ca²⁺]_i transient. Pretreatment with a specific κ opioid antagonist, nor-BNI(10^-8 mol/L), abolished the effect induced by IL-2 during anoxia on the [Ca²⁺]_i transients, whereas specific δ opioid antagonist, naltrindole(10^-6 mol/L), did not cancel the effect. CONCLUSION: It is concluded that administration of IL-2 during anoxia aggravated the effect of reoxygenation on the [Ca²⁺]_i transients of isolated ventricular myocytes, which was mediated by cardiac κ opioid receptor pathway.     

Qishen-Yiqi dripping pills attenuate atherosclerosis by regulating Ca2+ homeostasis via TRPC1/STIM1 pathway

AIM:To explore the therapeutic effect of Qishen-Yiqi dripping pills (QS) on atherosclerosis (AS) and the mechanism. METHODS:AS rat model was established by high-fat diet, and SD rats were randomly divided into normal control group, AS model group, low-dose, middle-dose and high-dose QS groups, and positive group (n=6 each). After administration for 12 weeks, serum samples were collected to detect the serum lipid and Ca²⁺ levels. HE staining was used evaluated the histopathological changes of arterial tissue. The serum levels of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) were measured by ELISA. The nitric oxide (NO) level was detected by nitrate reductase method. The protein levels of transient receptor potential channel protein 1 (TRPC1), stromal interaction molecule 1 (STIM1) and endothelial NO synthase (eNOS) were determined by Western blot. RESULTS:QS significantly reduced the arterial damage via inhibiting the formation of atherosclerotic plaque and attenuated intimal thickening and vascular stenosis. Compared with AS group, the serum levels of total cholesterol (TC), triglyceride (TG) and low-density lipoprotein cholesterol (LDL-C) were decreased significantly and the levels of high-density lipoprotein cholesterol (HDL-C) were increased significantly in high-dose QS group (P<0.05). The serum levels of IL-1β, IL-6 and TNF-α in high-dose QS group were lower than those in AS group (P<0.05). Compared with AS group, the serum Ca²⁺ level was lowered and the arterial tissue NO level was elevated in QS groups (P<0.05). Compared with AS rats, the protein levels of TRPC1 and STIM1 were decreased significantly and the protein level of eNOS was increased significantly in the rats treated with QS (P<0.05). CONCLUSION:QS regulate calcium homeostasis via TRPC1/STIM1 pathway, increase the production of NO and inhibit the inflammatory responses, thus exerting anti-AS effect.     

Effects of Ca2+on the releases of ET-1, NO and PGI2 from bovine coronary artery endothelial cells during hypoxia

AIM: To explore the mechanism of ET-1, NO and PGI² release from coronary artery endothelial cells(CAEC) induced by acute hypoxia. METHODS: Bovine coronary artery endothelial cells were cultured and [⁴⁵ Ca²⁺] was used to investigate the difference of calcium uptake between normoxia group and hypoxia group(3% O₂). The contents of ET-1, NO and PGI² in media of normoxia group, hypoxia group and hypoxia + verapamil group were measured 24 h after hypoxia. RESULTS: [ ⁴⁵ Ca²⁺] uptake by CAEC in hypoxia group was 1.9 times more than normoxia group(P< 0.01). Hypoxia + verapamil group released more PGI², ET-1 and less NO than hypoxia group(P< 0.05). CONCLUSION: Changes of ET-1, NO and PGI² releases during hypoxia may be caused by the inflow of Ca²⁺ into coronary artery endothelial cells.     

Effects of cimetidine on platelet function and thrombosis

AIM:To observe the effects of cimetidine(Cim) on platelet function and thrombosis. METHODS:After incubated with Cimin vitro, rat platelets were activated with ADP or thrombin. The platelet aggregation, platelet malondialdehyde(MDA) formation, platelet intracellular free calcium( [Ca²⁺]_i), and thromboxane B₂ (TXB₂) were measured. The effects of Cim on electric-induced thrombosis in rat carotid artery were examined. RESULTS:Cim potentiated ADP induced platelet aggregation, increased the thrombin induced [Ca²⁺]_i and MDA formation, decreased TXB₂. Also, Cim shortened the duration of electric-stimulated occlution time in rat carotid artery. CONCLUSION:Cim increased platelet function and accelerated thrombosis.     

Heterogeneity of basal intracellular calcium concentration and its relations to the reactivity in mouse peritoneal macrophages

AIM: To investigate the heterogeneity of basal intracellular free calcium concentration([Ca²⁺]_i) in peritoneal macrophages(PM) and whether it is relative to the reactivity of PM at the single cell level. METHODS:[Ca²⁺]_iimplicated stimulated were measured by fluorescent microscopic imaging system after loading with fluorescent probe fura-2/AM. Superoxide(O₂^-)produced by single PM was determined by modified NBT test. RESULTS: The values of basal[Ca²⁺]_idetermined in 392 PMs of 7 mice showed normal distribution [(54±24) nmol/L, n=392] with wide range(less than 20 nmol/L to more than 100 nmol/L), among which about 50% were in the range of 40-60 nmol/L. When stimulated with PMA or fMLP,[Ca²⁺]_iwas increased, the peak values were positively correlated with the basal[Ca²⁺]_iin one mouse(PMA stimulated cells: r=0.52, P<0.01, n=58; fMLP stimulated cells:r=0.59, P<0.01, n=44. Both of the experiments were repeated in 3 mice, the results in the other 2 mice were similar). The O₂^- in PMA stimulated PMs were also positively correlated with the basal i(r=0.42, P<0.01, n=43, repeated in 4 mice, the results in the other 3 mice were similar). CONCLUSION: Basal[Ca²⁺]_iin murine PM is heterogeneous, and the value of basal[Ca²⁺]_iis tightly correlated to the reactivity of PM stimulated by proinflammatory factors.     

Salidroside increases release of intracellular free Ca2+ from sarcoplasmic reticulum in cultured rat cardiomyocytes

AIM: To observe the effects of salidroside on intracellular free Ca²⁺ concentration in cultured rat cardiomyocytes. METHODS: Primarily cultured cardiomyocytes of neonatal rats were divided into control group, different concentrations of salidroside groups and verapamil pretreatment+different concentrations of salidroside groups. The fluorescent intensity of intercellular free calcium concentration ([Ca²⁺]_i) in cultured cardiomyocytes of newborn rats loaded with fluo-3/AM(5 μmol/L) was measured by laser scanning confocal microscopy. RESULTS: Salidroside at concentrations of 15 mg/L, 30 mg/L and 60 mg/L elevated [Ca²⁺]_i in cultured rat cardiomyocytes with the peak values of 574.08±4.65, 591.86±3.64 and 618.66±4.27, respectively (all P<0.01), indicating that the effect of salidroside on the level of [Ca²⁺]_i was dose-dependent. In the presence of verapamil in D-Hanks solution, salidroside also elevated the fluorescent intensity of [Ca²⁺]_i in cardiomyocytes from 357.74±3.13, 387.17±2.37 and 391.43±1.34 to 480.86±3.98, 496.70±3.08 and 522.18±3.19, respectively (all P<0.01). CONCLUSION: Salidroside increases the release of [Ca²⁺]_i from sarcoplasmic reticulum in cultured rat cardiomyocytes.     

Effects of glucose and Mg2+ in the neurons damaged by glutamate

AIM and METHODS: To observe the effects of glucose-free and Mg²⁺-free in the extracellular fluid on the changes of [Ca ²⁺]_i in the cerebro-cortical neurons damaged by 1mmol/L glutamate using laser confocal scanning microscope. RESULTS: Both frequency and amplitude of neuronal calcium oscillation induced by glutamate were lowered in glucose-free and Mg²⁺-free buffers. The basic [Ca²⁺]_i concentration was lowered in the former case , but it was elevated in the latter case. CONCLUSION: Mg²⁺-free aggravates [Ca²⁺]_i overload induced by 1mmol/L glutamate ,under certain conditions the glucose-free might resist damage role of glutamate and Mg²⁺-free.     

Hydrogen sulfide inhibits adenosine triphosphate-induced activation and IL-1β releases in rat microglia

AIM: To investigate the effects of sodium hydrosulfide (NaHS), a donor of hydrogen sulfide (H₂S), on the membrane permeability, intracellular Ca²⁺ concentration ([Ca²⁺]_i) and the release of IL-1β induced by adenosine triphosphate (ATP) in rat microglia, and to explore the effect of H₂S on ATP-P2X purinergic signaling pathway and the molecular mechanism of its neuroprotective effect. METHODS: Rat microglia in logarithmic growth phase were used in the study. The[Ca²⁺]_i was detected by Fura-2/AM staining. Fluorescent dye YO-PRO-1 was used to observe the membrane permeability. Interleukin-1β (IL-1β) was measured by rat IL-1β ELISA kits. RESULTS: The YO-PRO-1 fluorescence intensity was obviously elevated by ATP induction in a dose-dependent manner in the rat microglia, but this effect was counteracted by NaHS pretreatment (P<0.05).[Ca²⁺]_i rapidly increased and then decreased slowly, forming a stable platform for a long time when rat microglia were treated with ATP. Ca²⁺ spike activity induced by ATP had no change, but the platform disappeared (P<0.05) after NaHS pretreatment. The ATP and LPS together facilitated the release of IL-1β, but the phenomenon was inhibited by NaHS (P<0.05). CONCLUSION: Hydrogen sulfide may decrease the membrane permeability, calcium inflow and IL-1β release in rat microglia activated by high dose of ATP. The cytoprotection of hydrogen sulfide may be mediated by purinergic signaling pathway.     

ROS mediates regulation of intracellular Ca2+ induced by angiotensin II in primarily cultured medullary neurons

AIM: To investigate the role of reactive oxygen species (ROS) in the regulation of intracellular Ca²⁺ induced by angiotensin II (Ang II) in the primarily cultured medullary neurons. METHODS: Primarily cultured medullary neurons were prepared from 14-day-old embryos of Sprague-Dawley rats in the study. The identification of medullary neurons was assessed by double-labeling immunofluorescence. To explore the role of ROS, mainly the superoxide (O₂^·), the O₂^·generation was measured using the fluorogenic probe dihydroethidium (DHE). To determine intracellular free calcium concentration ([Ca²⁺]_i), the neurons were loaded with the Ca²⁺-specific dye Fura-2/AM. The cell viability after adding Ang II was also examined using CCK-8 assay. RESULTS: Most of the cultured cells were medullary neurons, more than 80% of which were glutamate positive neurons. Ang II (5 μmol/L) increased the level of ROS within 10 min in the medullary neurons. Ang II at 5 μmol/L induced a significant[Ca²⁺]_i increase in the medullary neurons, and the effect of Ang II occurred rapidly and reached a peak within 20 min after administration. The level of[Ca²⁺]_i started to decline after washout. The Ca²⁺ elevation induced by Ang II was significantly decreased by apocynin or TEMPOL. No significant difference in the cell viability between control group and 5 μmol/L Ang II treatment group was observed. CONCLUSION: ROS is involved in the regulation of[Ca²⁺]_i induced by Ang II in the primarily cultured medullary neurons, suggesting a potential intracellular signaling mechanism involved in the Ang II-mediated oxidant regulation of central neural control of blood pressure.     

Sodium Ferulate protects human aortic smooth muscle cells against oxidized Lipoprotein(a)

AIM: To investigate the influences of native and oxidized lipoprotein(a) on human arterial smooth muscle cell (SMC) proliferation, change of intracellular free calcium concentration ( [Ca²⁺]_i) and the protective effect of sodium ferulate(SF). METHODS: Lp(a) was oxidized by Cu²⁺ and the extent of oxidation was assessed by the MDA content.Human SMC were incubated in culture media with SF for 12 h, then exposed to Lp(a) and oxidized-Lp(a) , respectively. MTT colorimetry and flow cytometry were used to evaluated the proliferation of SMC and flurorescent indicator Fura-2/AM was used to determined [Ca²⁺]_i. RESULTS: ox-Lp(a) significantly promoted proliferation of SMC and increased [Ca²⁺]_i compared with Lp(a). SF(40,80 mg/L) remarkedly inhibited SMC proliferation and decreased the rising of [Ca²⁺]_i induced by ox-Lp(a) in a dose-dependent manner, but no effect on SMC proliferation and the increase in [Ca²⁺]_i induced by Lp(a).CONCLUSION: ox-Lp(a) induces the strong growth-promoting effect in SMC through increasing in [Ca²⁺]_i, which might be one of the cellular mechanisms responsible for the higher atherogenic potential of ox-Lp(a) compared with Lp(a), and this process can be prevented by inhibiting of oxidation by SF.     

Antagonistic effects of cyproheptadine and anisodamine on [Ca2+]i elevation induced by TNFα in endothelial cell strains

AIM: To study the effects of cyproheptadine (Cyp) and anisodamine (Ani) on the changes of intracellular free Ca²⁺ concentration ([Ca²⁺]_i) induced by tumor necrosis factor (TNFα) in single endothelial cells, and to explore the mechanisms of TNFα mediated shock and antishock actions of Cyp and Ani. METHODS: Human umbilical vein endothelial cell strains (ECV304) were seed in 35 mm tissue culture dish with 2 mL DMEM culture medium. The cultured cells were loaded by Fluo-3/AM. The spatial distribution and the dynamic changes of [Ca²⁺]_i in single endothelial cell was determined by laser scanning confocal microscopy (LSCM). RESULTS: [Ca²⁺]_i in single endothelial cell after stimulation of TNFα rapidly increased in a dose-dependent manner and approached the peak value within 60 seconds, afterwards, decreased and kept above the basal level. The confocal scanning image showed that [Ca²⁺]_i elevation was more obvious in nuclear than in cytoplasma, and decreased slowly. Cyp (3×10^-5, 6×10^-5 mol/L) and Ani (2×10^-5, 4×10^-5 mol·L^-1) markedly inhibited TNFα (1.2×10^-9 mol·L^-1)-induced [Ca²⁺]_i elevation. CONCLUSIONS: TNFα markedly induces elevation of [Ca²⁺]_i in single endothelial cell, it may be an important mechanism of TNFα-induced shock and tissue injury. Cyp and Ani obviously suppress TNFα-induced [Ca²⁺]_i elevation, which probably is one of the mechanisms of their antishock effects.     

Effects of oxidized HDL on the levels of MCP-1, ICAM-1 and free calcium in cultured human umbilical venous endothelial cells

AIM:To study the effects of oxidized high-density lipoprotein (oxHDL) on the expression of monocyte chemoattractant protein-1(MCP-1) and intercellular adhesion molecule-1(ICAM-1) and intracellular free calcium concentration ([Ca²⁺]i) level in cultured human umbilical venous endothelial cells(HUVECs). METHODS:The MCP-1 protein content in the medium of conditioned HUVEC was measured by ELISA, and the ICAM-1 on HUVECs was detected by indirect immunofluorescence, and [Ca²⁺]i was determined by Fluo-3/AM, the injury of cells was observed by scanning electron microscopy (SEM).RESULTS:oxHDL could induce the expression of MCP-1 and ICAM-1 in HUVECs. In oxHDL group (HUVECs were incubated with 100 mg protein/L oxHDL for 24 h), the levels of MCP-1, ICAM-1 and [Ca²⁺]i increased by 160%, 60% and 70% respectively compared with the control group (P＜0.01). When HUVECs were incubated with 300 mg protein/L oxHDL for 24 h, cells were injured obviously. CONCLUSION:By inducing the expression of ICAM-1 and MCP-1 in endothelial cells, oxHDL may promote monocyte-endothelium adhesion and monocyte migration to intima, it may promote atherosclerosis as oxidized low-density lipoprotein (oxLDL).     

Effect of chronic hypoxia on [Ca2+]iin pulmonary artery endothelial cells and smooth muscle cells under acute hypoxia

AIM AND METHODS: Using Ca²⁺-sensitive fluorescent probe Fura-2,we measured the changes of [Ca²⁺]_iin cultured rat pulmonary artery endothelial cells (PAEC) and porcine pulmonary artery smooth muscle cells (PASMC) from normoxic (NC group) or chronic hypoxic group (CH group) when they were exposed to acute hypoxia. RESULTS: The increase in [Ca²⁺]_iin 6th passage of PASMC caused by acute hypoxia in CH group was significantly lower than that in the same passage of NC group (P<0.05).On the contrary, in PAEC, the acute hypoxia induced increase in _i, which was significantly higher in 5th passage of CH group than that in NC group (P<0.05). CONCLUSION: The decrease of the elevation of [Ca²⁺]_icaused by acute hypoxia in PASMC of CH group indicated that it functioned to lower the constrictive response to hypoxia.The intensive increase in [Ca²⁺]_icaused by acute hypoxia in PAEC of CH group might lead to more relaxing factors derived from PAEC,which results in a decrease in HPV.