The relationship between triglyceride, cholesterol and lipoprotein levels, and immune responses to hepatitis B vaccine.

Cholesterol homeostasis disorder and hypertriglyceridemia, as common metabolic conditions, have rarely been reported to affect the immune responses to the hepatitis B vaccine. Our study found that higher high-density lipoprotein (HDL) level showed a significant relationship with positive anti-HBs results (cOR = 1.479, 95% CI: 1.150, 1.901, p = 0.002; aOR = 1.304, 95% CI: 1.006, 1.691, p = 0.045), especially in individuals aged 18- to 40-year-old, female, smoking more than 100 cigarettes in life, and drinking more than 12 times every year. Lower low-density lipoprotein (LDL) level was associated with a negative anti-HBs result among participants aged 18- to 40-year-old, and participants who were obese. Higher level of HDL and lower level of LDL may be protective factors of better immune effect of hepatitis B vaccine. More research should be conducted to investigate the influence of the cholesterol level on the immune responses to the hepatitis B vaccine, and more in-depth research should be performed to uncover the mechanism.

Reconstruction and analysis of potential biomarkers for hypertrophic cardiomyopathy based on a competing endogenous RNA network.

Hypertrophic cardiomyopathy (HCM) is a common heritable cardiomyopath. Although considerable effort has been made to understand the pathogenesis of HCM, the mechanism of how long noncoding RNA (lncRNA)-associated competing endogenous RNA (ceRNA) network result in HCM remains unknown. In this study, we acquired a total of 520 different expression profiles of lncRNAs (DElncRNAs) and 371 messenger RNAs (mRNA, DEGs) by microarray and 33 microRNAs (DEmiRNAs) by sequencing in plasma of patients with HCM and healthy controls. Then lncRNA-miRNA pairs were predicted using miRcode and starBase and crossed with DEmiRNAs. MiRNA-mRNA pairs were retrieved from miRanda and TargetScan and crossed with DEGs. Combined with these pairs, the ceRNA network with eight lncRNAs, three miRNAs, and 22 mRNAs was constructed. lncRNA RP11-66N24.4 and LINC00310 were among the top 10% nodes. The hub nodes were analyzed to reconstruct a subnetwork. Furthermore, quantitative real-time polymerase chain reaction results showed that LINC00310 was significantly decreased in patients with HCM. For LINC00310, GO analysis revealed that biological processes were enriched in cardiovascular system development, sprouting angiogenesis, circulatory system development, and pathway analysis in the cGMP-PKG signaling pathway. These results indicate that the novel lncRNA-related ceRNA network in HCM and LINC00310 may play a role in the mechanism of HCM pathogenesis, which could provide insight into the pathogenesis of HCM.

Prevalence of myopia: A large-scale population-based study among children and adolescents in weifang, china.

Purpose: This study aimed to determine the prevalence of myopia among school-aged children and adolescents at the whole city level of Weifang, China. Methods: This study was a large scale school-based cross-sectional study among children and adolescents aged 5-20 years old. Participants were selected by the school-based registration system in 2020. All the subjects underwent spherical equivalent (SE) error with non-cycloplegic autorefraction measurement. Myopia was defined as an SE refraction of ≤-0.75 diopters (D) and graded into low myopia (-0.75 to -3.00 D), moderate myopia (-3.01 to -5.99 D), and high myopia (≤-6.00 D). Results: A total of 1,059,838 participants were eligible for this survey and 1,013,206 (95.6% participation rate) were selected through data quality control, which comprised 17 districts/counties and 1,254 schools, including 861 elementary schools, 313 middle schools, and 80 high schools. The mean age of participants was 11.57 ± 3.36 years (5-20 years), and the male-to-female ratio was 1.11. The whole city-level prevalence of total myopia was 75.35%. The prevalence of total myopia among the students in the Hanting District was 45.47%, but ≈82.37% of students living in Changyi have myopia. The overall prevalence of low myopia in elementary, middle, and high school students was 48.56, 47.30, and 31.62%, respectively, while high myopia (SE ≤ -6.00 D) prevalence was 1.12, 8.89, and 20.12%, respectively. The overall prevalence of myopia increased fastest in children aged 7-9 years old. The prevalence of high myopia was 7.59% for girls and 6.43% for boys, respectively (p < 0.001). The prevalence of myopia increased with increasing age and grade, but SE decreased with increasing age and grade. Conclusions: The current investigation demonstrated a high proportion of myopia among school students in the city of Weifang, and gradually increased with age, and the prevalence of myopia was the highest in Changyi areas. The high myopia prevalence for girls was higher than that in boys.

MicroRNA expression profiles in familial hypertrophic cardiomyopathy with myosin-binding protein C3 (MYBPC3) gene mutations.

Familial hypertrophic cardiomyopathy (FHCM) is an autosomal dominant inherited disease caused by mutations in genes encoding cardiac sarcomere proteins. MicroRNAs (miRNAs) play an important role in the pathogenesis of FHCM. In the present study, we aimed to determine the miRNA profile in FHCM patients with myosin-binding protein C3 (MYBPC3) gene mutations. We recruited three FHCM patients and age- and sex-matched controls. The three probands all had hypertrophic obstructive cardiomyopathy with severe myocardial hypertrophy, and two of the three had a history of sudden cardiac death, representing a "malignant" phenotype. We then compared the miRNA expression profiles of three FHCM patients carrying MYBPC3 gene mutations with those of the normal control group using miRNA sequencing technology. Differentially expressed miRNAs were verified using real-time polymerase chain reaction (qPCR). Target genes and signaling pathways of the identified differentially expressed miRNAs were predicted using bioinformatics analysis. A total of 33 significantly differentially expressed miRNAs were detected in the peripheral blood of the three probands, of which 28 were upregulated, including miR-208b-3p, and 5 were downregulated. Real-time PCR confirmed the upregulated expression of miR-208b-3p in FHCM patients (P < 0.05). Bioinformatics analysis showed that miR-208b-3p was mainly enriched in 79 target genes including UBE2V2, MED13, YBX1, CNKSR2, GATA4, andSOX5/6, et al. Gene ontology (GO) analysis of target genes showed that miR-208b was mainly involved in the processes of negative regulation of transcription from RNA polymerase II promoter, and regulation of transcription, DNA templated. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis revealed that the target genes regulated by miR-208b-3p were mainly involved in the Wnt signaling pathway. These findings suggest that FHCM patients with MYBPC3 gene mutations have a specific miRNA expression profile, and that miR-208b-3p is significantly upregulated in cardiac hypertrophy. Our results also indicate that miRNA-208b-3p activates the Wnt signaling pathway through its target gene to promote cardiac hypertrophy.

Molecular Mechanisms of Iron Mediated Programmed Cell Death and Its Roles in Eye Diseases.

Ferroptosis, a newly identified, iron-dependent type of programmed cell death, is active in several diseases, such as heart disease, brain damage, and cancer. Its main characteristics commonly involve excess iron accumulation, elevated lipid peroxides and reactive oxygen species, and reduced levels of glutathione and glutathione peroxidase 4 levels. The effects of ferroptosis in eye diseases cannot be underestimated, with ferroptosis becoming a research target in ocular disorders and emerging evidence from a series of in vivo and in vitro researches into ferroptosis revealing its role in eye conditions. However, no report provides comprehensive information on the pathophysiology of ferroptosis in eye diseases and its possible treatments. In the current review, we present an up-to-date overview of ferroptosis biology and its involvement in the pathological processes of ocular diseases. Furthermore, we pose several outstanding questions and areas for future research in this topic. We deem ferroptosis-associated cell death a pivotal new field of scientific study in ocular diseases and consider it a new therapeutic target in the treatment of some eye disorders.

Association of maternal diabetes with attention deficit/hyperactivity disorder (ADHD) in offspring: A meta-analysis and review.

AIMS: Researches on the relationship between maternal pregestational or gestational diabetes and attention deficit/hyperactivity disorder (ADHD) in offspring provided inconsistent findings; therefore, we performed an updated and comprehensive literature review and meta-analysis to evaluate the available evidence. METHOLDS: Relevant articles in Pubmed, Web of Science, Cochrane, Embase, and Wanfang database published until January 2019 were searched without language restriction. We performed a meta-analysis about maternal pregestational and gestational diabetes and risk of ADHD in offspring using odds ratio (OR), relative risk (RR), hazard ratio (HR) and 95% confidence interval (95% CI) extracted from each study. RESULTS: Seven articles were included in this study and a total of 3,169,529 participants were accumulated. We found maternal pregestational diabetes increased the risk of ADHD in offspring by 44% (95% CI was 1.32-1.57). CONCLUSIONS: Maternal pregestational diabetes is a potential adverse risk of ADHD in offspring. Considering the limited amount of reliable information availabe. In the future, more in-depth and detailed researches, especially population-based prospective cohort studies, are needed to explore this topic more comprehensively.

Is low body mass index a risk factor for semen quality? A PRISMA-compliant meta-analysis.

BACKGROUND: Male infertility has become a worldwide public health problem. However, the effect of low body mass index (BMI) is still controversial. METHODS: Relevant articles in Pubmed, Embase, Web of science, and Wanfang database published until September 2017 were searched without language restriction. We performed a meta-analysis about low BMI and semen parameters containing total sperm count, concentration, semen volume, and sperm motility (overall and progressive), including 709 men with low BMI and 14,622 men with normal BMI. RESULTS: Thirteen studies were included in this meta-analysis and a total of 15,331 individuals were accumulated. We pooled data from these articles and found standardized weighted mean differences in semen parameters (total sperm count and semen volume) showed significant difference between low BMI and normal BMI. CONCLUSIONS: This systematic review with meta-analysis has confirmed that there was a relationship between low BMI and semen quality, which suggesting low BMI may be a harmful factor of male infertility. Yet lacking of the raw data may influence the accuracy of the results. Further researches are needed to identify the role of underweight in male sterility.

Sphingosine kinase regulates oxidized low density lipoprotein-mediated calcium oscillations and macrophage survival.

We recently reported that oxidized LDL (oxLDL) induces an oscillatory increase in intracellular calcium ([Ca(2+)](i)) levels in macrophages. Furthermore, we have shown that these [Ca(2+)](i) oscillations mediate oxLDL's ability to inhibit macrophage apoptosis in response to growth factor deprivation. However, the signal transduction pathways by which oxLDL induces [Ca(2+)](i) oscillations have not been elucidated. In this study, we show that these oscillations are mediated in part by intracellular mechanisms, as depleting extracellular Ca(2+) did not completely abolish the effect. Inhibiting sarco-endoplasmic reticulum ATPase (SERCA) completely blocked [Ca(2+)](i) oscillations, suggesting a role for Ca(2+) reuptake by the ER. The addition of oxLDL resulted in an almost immediate activation of sphingosine kinase (SK), which can increase sphingosine-1-phosphate (S1P) levels by phosphorylating sphingosine. Moreover, S1P was shown to be as effective as oxLDL in blocking macrophage apoptosis and producing [Ca(2+)](i) oscillations. This suggests that the mechanism in which oxLDL generates [Ca(2+)](i) oscillations may be 1) activation of SK, 2) SK-mediated increase in S1P levels, 3) S1P-mediated Ca(2+) release from intracellular stores, and 4) SERCA-mediated Ca(2+) reuptake back into the ER.

A comparative study of alpha-adrenergic receptor mediated Ca(2+) signals and contraction in intact human and mouse vascular smooth muscle.

In many vascular smooth muscle cells, physiological and pharmacological agonists initiate oscillatory fluctuations in intracellular Ca(2+) to initiate and maintain vasoconstriction. These oscillations are supported by the underlying cellular ultrastructure, particularly the close apposition between the plasma membrane (PM) and superficial sarcoplasmic reticulum (SR), the so-called PM-SR junctions, which are important for SR Ca(2+) refilling. We hypothesize that the disappearance of PM-SR junctions during aging and/or disease is directly related to the disappearance of agonist-induced Ca(2+) oscillations. We compared phenylephrine-mediated Ca(2+) signals and contraction in human and murine smooth muscle cells in small mesenteric arteries and also employed electron microscopy to examine the cytoplasmic distribution of the SR. Phenylephrine elicited tonic contractions in both types of vessels, asynchronous Ca(2+) oscillations in the mouse mesenteric smooth muscle cells, but only single transient Ca(2+) signals in the human mesenteric smooth muscle cells. While nifedipine inhibited 90% of the phenylephrine-induced tonic contraction in mouse mesenteric arteries, it only slightly attenuated tonic contraction in human mesenteric arteries, although the nifedipine-resistant component was abolished by the Rho-kinase blocker 1-(5-Isoquinolinylsulfonyl)homopiperazine dihydrochloride (HA-1077). Furthermore, superficial SR was found to be abundant in the mouse vessels and many PM-SR junctions were observed, but the smooth muscle of human mesenteric arteries had far less peripheral SR and was almost devoid of PM-SR junctions. As PM-SR junctions are essential for the maintenance of Ca(2+) oscillations, the change in Ca(2+) signalling pattern in the relatively old human patients was due to impaired SR refilling.

Endothelin-1-mediated wave-like [Ca2+]i oscillations in intact rabbit inferior vena cava.

Endothelin-1 (ET1) is an endogenous vasoconstrictor released by the vascular system to regulate the contractility of vascular smooth muscle cells (VSMC). It is implicated in the pathogenesis of hypertension and diabetic vasculopathy. In rabbit inferior vena cava (IVC), 10 nM ET1 induces tonic contraction mainly via type A endothelin receptor activation. Using confocal imaging of Fluo-3 loaded in thein situ VSMC within the intact IVC, we found that ET1 elicited [Ca2+]i oscillations with an average frequency of 0.31 +/- 0.01 Hz. These [Ca2+]i oscillations occurred as repetitive Ca2+ waves traveling along the longitudinal axis of the cells with an average velocity of 29 +/- 3 microm/s. The Ca2+ waves were not synchronized between neighboring VSMC nor were they propagated between them. Nifedipine (10 microM) inhibited the tonic contraction by 27.0 +/- 5.0% while SKF96365 (50 microM) abolished the remaining contraction. In a parallel Ca2+ study, nifedipine reduced the frequency of the oscillations to 0.22 +/- 0.01 Hz while SKF96365 abolished the remaining [Ca2+]i oscillations. Subsequent application of 25 mM caffeine elicited no further Ca2+ signal. Thus, we conclude that ET1 stimulates tonic contraction in the rabbit IVC by inducing [Ca2+]i oscillations and that stimulated Ca2+ entry through both the L-type voltage-gated Ca2+ channels and a nifedipine-resistant and SKF96365-sensitive pathway is crucial for the maintenance of [Ca2+]i oscillations and tonic contraction.

Acetylcholine-induced asynchronous calcium waves in intact human bronchial muscle bundle.

Calcium (Ca2+) is an important activator of the contractile machinery in airway smooth muscle (ASM). While agonist-induced Ca2+ signals are well characterized in animal ASM, little is known about what occurs in adult human ASM. In this study, we examined the Ca2+ signal elicited by acetylcholine (ACh) in smooth muscle cells of the intact human bronchial muscle strips obtained from fresh surgical specimens in relation to muscle contraction. We found that ACh induces repetitive Ca2+ waves that spread along the longitudinal axis of individual cells in the intact human bronchial smooth muscle strips. These Ca2+ waves display no apparent synchronization between neighboring cells, and their generation precedes force development. Comparison of the ACh concentration dependence of tissue contraction and selected parameters of the asynchronous Ca2+ waves (ACW) reveals that the graded force generation by ACh-stimulated human bronchial muscle strips is achieved by differential recruitment of cells to initiate Ca2+ waves and by enhancement of the frequency of ACW once the cells are recruited. Furthermore, pharmacologic characterization shows that the ACW are produced by repetitive cycles of SR Ca2+ release via ryanodine-sensitive channels followed by SR Ca2+ reuptake by sarco(endo)plasmic reticulum Ca2+ ATPase. Extracellular Ca2+ entry involving receptor-operated channels/store-operated channels, reverse-mode Na+/Ca2+ exchange, and to a lesser extent L-type voltage-gated Ca2+ channels is required to maintain the ACW. These findings for the first time demonstrate the occurrence and the role of ACW in excitation-contraction coupling in adult human ASM.

Mechanism of ACh-induced asynchronous calcium waves and tonic contraction in porcine tracheal muscle bundle.

Stimulation of the tracheal muscle bundle by acetylcholine (ACh) results in the generation of asynchronous repetitive Ca2+ waves (ACW) in intact tracheal smooth muscle (TSM) cells. We showed previously that ACW underlie cholinergic excitation-contraction coupling in porcine TSM and that Ca2+ entry through the L-type voltage-gated Ca2+ channel (VGCC) contributes partially to maintenance of the ACW. However, the mechanism of the ACW remains undefined. In this study, we pharmacologically characterized the mechanism of ACh-induced ACW in the intact porcine tracheal muscle bundle. We found that inhibition of receptor-operated channels/store-operated channels (ROC/SOC) by SKF-96365 completely abolished the nifedipine-insensitive component of ACh-mediated ACW and tonic contraction. Blockade of Na+/Ca2+ exchange with KB-R7943 or 2',4'-dichlorobenzamil or removal of extracellular Na+ resulted in nearly complete inhibition of the nifedipine-insensitive component of ACh-mediated ACW and tonic contraction. Inhibition of the sarco(endo)plasmic reticulum Ca2+-ATPase by cyclopiazonic acid abolished the ongoing ACW. Application of 2-aminoethoxydiphenyl borate (2-APB) or xestospongin C to inhibit the inositol 1,4,5-trisphosphate-sensitive sarcoplasmic reticulum (SR) Ca2+ release channels produced no effect on ACh-mediated ACW and tonic contraction. However, pretreatment with caffeine or ryanodine inhibited ACh-induced ACW. Furthermore, application of procaine or tetracaine prevented the generation and abolished the ongoing ACh-mediated ACW and tonic contraction. Collectively, these results indicate that the ACh-stimulated ACW in porcine TSM are produced by repetitive cycles of Ca2+ release from SR through 2-APB- and xestospongin C-insensitive Ca2+ release channels, and plasmalemmal Ca2+ entry involving reverse-mode Na+/Ca2+ exchange, ROC/SOC, and L-type VGCC is required to refill the SR via SERCA to support the ongoing ACW.

Asynchronous calcium waves in smooth muscle cells.

Asynchronous Ca2+ waves or wave-like [Ca2+]i oscillations constitute a specialized form of agonist-induced Ca2+ signaling that is observed in a variety of smooth muscle cell types. Functionally, it is involved in the contractile regulation of the smooth muscle cells as it signals for tonic contraction in certain smooth muscle cells while causing relaxation in others. Mechanistically, repetitive Ca2+ waves are produced by repetitive cycles of sarcoplasmic reticulum Ca2+ release followed by Ca2+ uptake. Plasmalemmal Ca2+ entry mechanisms are important for providing the additional Ca2+ necessary to maintain proper refilling of the sarcoplasmic reticulum Ca2+ store and support ongoing Ca2+ waves. In this paper, we will review the phenomenon of asynchronous Ca2+ waves in smooth muscle and discuss the scientific and clinical significance of this new understanding.

Rearrangement of the close contact between the mitochondria and the sarcoplasmic reticulum in airway smooth muscle.

The mitochondria and the sarcoplasmic reticulum (SR) are two major intracellular calcium-storing organelles that exhibit close functional interaction with each other. Close spatial association is believed to be important for their functional interaction. In this study, we have characterized the spatial relationship between the SR and the mitochondria in porcine tracheal smooth muscle cells (TSMC) under different conditions. By examining the cross-section of unstimulated TSMC with electron microscopy, we found that 99.4 +/- 0.5% of the mitochondria seen on random cross-sections were situated within 30 nm of the SR and that 82.2 +/- 6.7% of the mitochondria were completely enveloped by the SR network. Overall, 48.0 +/- 3.5% of the mitochondrial outer membrane was within 30 nm with the SR. After stimulation of the TSMC with acetylcholine (ACh) or 80 mM [K(+)] solution 97.0 +/- 2.1% and 98.6 +/- 1.4% of the mitochondria observed were situated within 30 nm of the SR, respectively. However, the proportion of the mitochondria that was completely enveloped by the SR was significantly reduced to 12.2 +/- 5.9% in ACh-stimulated cells and 9.7 +/- 6.6% in 80 mM [K(+)] stimulated cells. The percentage of mitochondrial membrane closely associated with the SR was correspondingly lower at 10.1 +/- 1.0% during ACh stimulation and 10.8 +/- 0.9% during 80 mM [K(+)] stimulation. During smooth muscle cell stimulation, the SR appears to unwrap from the mitochondria and extend into the cytoplasm while maintaining close contact with the mitochondria over a smaller area. Such static and dynamic components of the close spatial association between the mitochondria and the SR may serve as a structural basis for the selective and efficient Ca(2+) trafficking between the two organelles in TSMC.

Calyculin-A disrupts subplasmalemmal junction and recurring Ca2+ waves in vascular smooth muscle.

Excitation-contraction coupling (E-C coupling) in phenylephrine(PE)-stimulated rabbit inferior vena cava (IVC) depends on the generation of asynchronous recurring Ca2+ waves in the in situ vascular smooth muscle cells (VSMC). Previous studies by our group have implicated a putative non-selective cationic store-operated channel and the reverse-mode Na+-Ca2+ exchange in refilling of the intracellular Ca2+ store via the sarco/endoplasmic reticulum ATPase (SERCA) and the maintenance of the recurring Ca2+ waves. We hypothesize that for the proper functioning of these three Ca2+ translocators in the process of SR refilling, the plasma membrane (PM) and the underlying superficial sarcoplasmic reticulum (SR) form specialized PM-SR junctions, which are essential for the maintenance of the recurring Ca2+ waves. In order to test this hypothesis, calyculin-A, a serine/threonine phosphatase inhibitor that has been demonstrated to result in the disruption of the PM-SR junctions was used. In the control rabbit IVC, electron microscopy of the in situ VSMC indicates that 14.2+/-0.7% of the PM is closely apposed by the prominent superficial SR network, forming numerous flattened PM-SR junctional cytoplasmic spaces. In the control IVC stimulation with 5 microM PE resulted in sustained recurring Ca2+ waves with a frequency of 0.42+/-0.02 Hz. In calyculin-A treated rabbit IVC, a concentration-dependent dissociation of the superficial SR and loss of PM-SR junctions was observed. This progressive loss of the PM-SR junctions occurs over the same concentration range as the inhibition of PE-induced recurring Ca2+ waves. These findings offer support for the hypothesis that the presence of the PM-SR junctions is required for the generation of asynchronous recurring Ca2+ waves, which underlie excitation-contraction coupling in the VSMC of the rabbit IVC.

Organellar junctions promote targeted Ca2+ signaling in smooth muscle: why two membranes are better than one.

Numerous cellular processes are regulated by fluctuations in the concentration of a single cation, Ca(2+). To accomplish this feat, cells have developed mechanisms that target Ca(2+) signals to specific effectors in both space, by strategically localizing effectors and ion-transporting molecules, and time, by encoding the regulation of the frequency of Ca(2+) oscillations. With an emphasis on smooth muscle, we have analyzed how the interaction of Ca(2+) transporters located on closely apposing membranes of the plasma membrane, sarcoplasmic reticulum and mitochondria provides the structural foundation for site-specific and time-specific Ca(2+) signaling. These junctional membrane complexes can either control the concentration of Ca(2+) in the microdomain that surrounds an effector molecule or deliver Ca(2+) from the translocator on one membrane to a second translocator on the opposing membrane without significant diffusion into the bulk cytosol, an event we term 'linked Ca(2+) transport'.

Relationship between asynchronous Ca2+ waves and force development in intact smooth muscle bundles of the porcine trachea.

Fluctuations in intracellular calcium concentration ([Ca2+]i) constitute the main link in excitation-contraction coupling (E-C coupling) in airway smooth muscle cells (ASMC). It has recently been reported that ACh induces asynchronous recurring Ca2+ waves in intact ASMC of murine bronchioles. With the use of a novel technique allowing us to simultaneously measure subcellular [Ca2+]i and force generation in ASMC located within an intact tracheal muscle bundle, we examined a similar pattern of Ca2+ signaling in the trachea. We found that application of ACh resulted in the generation of recurring intracellular Ca2+ waves progressing along the longitudinal axis of the ribbon-shaped intact ASMC. These Ca2+ waves were not synchronized between neighboring cells, and induction of wave-like [Ca2+]i oscillations was temporally associated with development of force by the tracheal muscle bundle. By comparing the concentration dependence of force generation and the parameters characterizing the [Ca2+]i oscillations, we found that the concentration-dependent increase in ACh-induced force development by the tracheal smooth muscle bundle is achieved by differential recruitment of intact ASMC to initiate Ca2+ waves and by enhancement in the frequency of [Ca2+]i oscillations and elevation of interspike [Ca2+]i once the cells are recruited. Our findings demonstrate that asynchronous recurring Ca2+ waves underlie E-C coupling in ACh-induced contraction of the intact tracheal smooth muscle bundle. Furthermore, in contrast to what was reported in enzymatically dissociated ASMC, Ca2+ influx through the L-type voltage-gated Ca2+ channel was not an obligatory requirement for the generation of [Ca2+]i oscillations and development of force in ACh-stimulated intact ASMC.

Proteinase-activated receptor-2 (PAR2): vascular effects of a PAR2-derived activating peptide via a receptor different than PAR2.

We studied the actions of the proteinase-activated receptor-2-activating peptide (PAR2-AP) trans-cinnamoyl-LIGRLO-amide (tc-LI) in femoral (FA), renal, and small mesenteric (MA) arterial vessels from C57BL/6 [PAR2 (+/+)] and PAR2 (-/-) mice. The actions of tc-LI were compared with those of the parent PAR2-AP Ser-Leu-Ile-Gly-Arg-Leu-amide (SLIGRL-amide; SLI-NH2). Either SLI-NH2 or tc-LI (0.1-10 microM) induced relaxation of either 9,11-dideoxy-9alpha,11alpha-methanoepoxy-prosta-5Z,13E-dien-1-oic acid (U46619)- or cirazoline-precontracted FA from PAR2 (+/+) in endothelium-intact preparations but did not relax vessels from PAR2 (-/-) mice. This FA relaxation by SLI-NH2 and by tc-LI was inhibited by 1) pretreatment with a combination of L-N(G)-nitroarginine methyl ester (L-NAME) and 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (ODQ), 2) precontraction with 30 mM KCl, or 3) removal of the endothelium. In contrast, tc-LI caused an L-NAME/ODQ/indomethacin-resistant relaxation of MA from PAR2 (+/+) mice. In contrast with SLI-NH2, tc-LI (>30 microM) contracted arteries from both PAR2 (-/-) and PAR2 (+/+) mice. Pretreatment of tissues with a combination of cyclopiazonic acid plus caffeine reduced significantly tc-LI-induced contractions, whereas nifedipine, CdCl2, and Ca2+-free conditions did not. Inhibitors of vascular muscarinic, alpha1-adrenergic, neurokinin, thromboxane A2, histamine, angiotensin II, or endothelin-1 receptors failed to inhibit contractions by 50 microM tc-LI. At resting tension, SLI-NH2 (>10 microM) contracted all arteries in an endothelium-independent manner but only from PAR2 (+/+) mice. We conclude that the endothelium-dependent vasodilation initiated by SLI-NH2 and tc-LI, but not the endothelium-independent contraction initiated by tc-LI, are due to the activation of PAR2. Indeed, the data from PAR2 (-/-) mice indicate that tc-LI, in addition to activating PAR2, is an agonist of vascular smooth muscle contraction via a receptor different than PAR2.