Identification of a novel 10.3 kb deletion causing α-thalassemia by third-generation sequencing: Pedigree analysis and genetic diagnosis.

Background: α-thalassemia is an inherited blood disorder caused by variants in the α-globin gene cluster. Identification of the pathogenic α-globin gene variants is important for the diagnosis and management of thalassemia.

Methods: Two suspected families from Xiantao, Hubei Province were recruited in this study.

Nuclear factor-κB regulates expression of platelet phospholipase C-β2 (PLCB2).

Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activation. Mechanisms regulating of PLC-β2 expression in platelets/MKs are unknown. Our studies in a patient with platelet PLC-β2 deficiency revealed the PLCB2 coding sequence to be normal and decreased platelet PLC-β2 mRNA, suggesting a defect in transcriptional regulation.

Effect of two administration routes of Shenmai Injection () on pulmonary gas exchange function after tourniquet-induced ischemia-reperfusion.

Objective: To compare the effect between nebulized and intravenous administration of Shenmai Injection () on pulmonary gas exchange function of patients following tourniquet-induced lower limb ischemia-reperfusion.

Methods: Thirty-eight patients scheduled for lower extremity surgery were randomized into three groups using the closed envelop method: Shenmai Injection was administered 30 min before tourniquet inflflation by nebulization [0.6 mL/kg in 10 mL normal saline (NS)] in the nebulization group or by intravenous drip (0.

Cross talk between serine/threonine and tyrosine kinases regulates ADP-induced thromboxane generation in platelets.

ADP-induced thromboxane generation depends on Src family kinases (SFKs) and is enhanced with pan-protein kinase C (PKC) inhibitors, but it is not clear how these two events are linked. The aim of the current study is to investigate the role of Y311 phosphorylated PKCδ in regulating ADP-induced platelet activation. In the current study, we employed various inhibitors and murine platelets from mice deficient in specific molecules to evaluate the role of PKCδ in ADP-induced platelet responses.

Distinct role of Pyk2 in mediating thromboxane generation downstream of both G12/13 and integrin αIIbβ3 in platelets.

Proline-rich tyrosine kinase 2 (Pyk2) is activated by various agonists in platelets. We evaluated the signaling mechanism and the functional role of Pyk2 in platelets by using pharmacological inhibitors and Pyk2-deficient platelets. We found that platelet aggregation and secretion in response to 2-methylthio-ADP (2-MeSADP) and AYPGKF were diminished in the presence of Pyk2 inhibitors or in Pyk2-deficient platelets, suggesting that Pyk2 plays a positive regulatory role in platelet functional responses.

Cardiotoxic and cardioprotective features of chronic β-adrenergic signaling.

Rationale: In the failing heart, persistent β-adrenergic receptor activation is thought to induce myocyte death by protein kinase A (PKA)-dependent and PKA-independent activation of calcium/calmodulin-dependent kinase II. β-adrenergic signaling pathways also are capable of activating cardioprotective mechanisms.

Objective: This study used a novel PKA inhibitor peptide to inhibit PKA activity to test the hypothesis that β-adrenergic receptor signaling causes cell death through PKA-dependent pathways and cardioprotection through PKA-independent pathways.

Protein kinase C isoform ε negatively regulates ADP-induced calcium mobilization and thromboxane generation in platelets.

Objective: Members of the protein kinase C (PKC) family are shown to positively and negatively regulate platelet activation. Although positive regulatory roles are extensively studied, negative regulatory roles of PKCs are poorly understood. We investigated the mechanism and specific isoforms involved in PKC-mediated negative regulation of ADP-induced functional responses.

[Role of vascular endothelial active facters in gas exchange impairment induced by tourniquet and the effect of shenmai injection].

Objective: To investigate the effect of Shenmai injection on vascular endothelial active facters nitric oxide (NO) and endothelin-1 (ET-1), and pulmonary gas exchange induced by tourniquet deflation in patients undergoing lower extremity surgery.

Method: Twenty-six patients scheduled for unilateral lower extremity surgery were randomly divided into 2 groups: control group (group C, n = 14) and Shenmai injection group (group SM, n = 12). All the patients agreed to a combined spinal-epidural anesthesia at the L2-L3 interspace and a radial artery catheter was placed for sampling.

G(12/13) signaling pathways substitute for integrin αIIbβ3-signaling for thromboxane generation in platelets.

Background: We have previously shown that ADP-induced TXA(2) generation requires signaling from αIIbβ3 integrin in platelets. Here we observed that, unlike ADP, protease-activated receptor (PAR)-mediated TXA(2) generation occurs independently of αIIbβ3. PAR agonists, but not ADP, activate G(12/13) signaling pathways.

Antagonism of P2Y₁₂ reduces physiological thromboxane levels.

Antiplatelet therapy for the management of patients with cardiovascular risks often includes a combination therapy of aspirin and clopidogrel, acting through inhibition of thromboxane generation and blockade of G(i)-coupled P2Y₁₂ receptor, respectively. We hypothesized that ADP acting through P2Y₁₂ regulates physiological thromboxane levels. The serum thromboxane levels in mice (n = 3) dosed with clopidogrel and prasugrel were decreased by 83.

BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism.

Though antiplatelet drugs are proven beneficial to patients with coronary heart disease and stroke, more effective and safer antiplatelet drugs are still needed. In this study we report the antiplatelet effects and mechanism of BF0801, a novel adenine derivative. BF0801 dramatically inhibited platelet aggregation and ATP release induced by ADP, 2MeSADP, AYPGKF, SFLLRN or convulxin without affecting shape change in vitro .

Mutational analysis of residues important for ligand interaction with the human P2Y(12) receptor.

The P2Y(12) receptor, a Gi protein-coupled receptor, plays a central role in platelet activation. In this study, we did a mutational analysis of residues possibly involved in the ligand interactions with the human P2Y(12) receptor. Mutant receptors were stably expressed in CHO-K1 cells with an HA-tag at the N-terminus.

Cbl-b is a novel physiologic regulator of glycoprotein VI-dependent platelet activation.

Cbl-b, a member of the Cbl family of E3 ubiquitin ligases, plays an important role in the activation of lymphocytes. However, its function in platelets remains unknown. We show that Cbl-b is expressed in human platelets along with c-Cbl, but in contrast to c-Cbl, it is not tyrosine-phosphorylated upon glycoprotein VI (GPVI) stimulation.

Regulation of plasmin-induced protease-activated receptor 4 activation in platelets.

Plasmin, a major extracellular protease, activates platelets through PAR4 receptors. Plasmin-induced full aggregation is achieved at lower concentrations (0.1 U/mL) in murine platelets as compared to human platelets (1 U/mL).

RhoA downstream of G(q) and G(12/13) pathways regulates protease-activated receptor-mediated dense granule release in platelets.

Platelet secretion is an important physiological event in hemostasis. The protease-activated receptors, PAR 1 and PAR 4, and the thromboxane receptor activate the G(12/13) pathways, in addition to the G(q) pathways. Here, we investigated the contribution of G(12/13) pathways to platelet dense granule release.

Differential regulation of threonine and tyrosine phosphorylations on protein kinase Cdelta by G-protein-mediated pathways in platelets.

Phosphorylation of activation loop threonine (Thr(505)) and regulatory domain tyrosine (Tyr(311)) residues are key regulators of PKC (protein kinase C) delta function in platelets. In the present study, we show that G(q) and G(12/13) pathways regulate the Thr(505) and Tyr(311) phosphorylation on PKCdelta in an interdependent manner. DiC8 (1,2-dioctanoylglycerol), a synthetic analogue of DAG (diacylglycerol), caused Thr(505), but not Tyr(311), phosphorylation on PKCdelta, whereas selective activation of G(12/13) pathways by the YFLLRNP peptide failed to cause phosphorylation of either residue.

Risk of thrombogenicity among nonionic radiocontrast agents.

Several contrast agents have been approved in the United States for radiographic imaging purposes. Most of the older ionic, high-osmolar contrast agents are no longer used because of their side effect profile. Therefore, newer nonionic, low or iso-osmolar contrast agents have been widely accepted as an alternative due to their improved tolerability and safety.

Characterization of a new peptide agonist of the protease-activated receptor-1.

A new peptide (TFRRRLSRATR), derived from the c-terminal of human platelet P2Y(1) receptor, was synthesized and its biological function was evaluated. This peptide activated platelets in a concentration-dependent manner, causing shape change, aggregation, secretion and calcium mobilization. Of the several receptor antagonists tested, only BMS200261, a protease activated receptor 1 (PAR-1) specific antagonist, totally abolished the peptide-induced platelet aggregation, secretion and calcium mobilization.

Relative contribution of G-protein-coupled pathways to protease-activated receptor-mediated Akt phosphorylation in platelets.

Protease-activated receptors (PARs) activate Gq and G(12/13) pathways, as well as Akt (protein kinase B [PKB/Akt]) in platelets. However, the relative contribution of different G-protein pathways to Akt phosphorylation has not been elucidated. We investigated the contribution of Gq and G(12/13) to Gi/Gz-mediated Akt phosphorylation downstream of PAR activation.

Molecular mechanism and functional implications of thrombin-mediated tyrosine phosphorylation of PKCdelta in platelets.

Thrombin has been known to cause tyrosine phosphorylation of protein kinase C delta (PKCdelta) in platelets, but the molecular mechanisms and function of this tyrosine phosphorylation is not known. In this study, we investigated the signaling pathways used by protease-activated receptors (PARs) to cause tyrosine phosphorylation of PKCdelta and the role of this event in platelet function. PKCdelta was tyrosine phosphorylated by either PAR1 or PAR4 in a concentration- and time-dependent manner in human platelets.

Rapid ubiquitination of Syk following GPVI activation in platelets.

Spleen tyrosine kinase (Syk) activation is a key intermediate step in the activation of platelets by the physiologic agonist collagen. We have found that Syk is rapidly ubiquitinated upon activation of platelets by collagen, collagen-related peptide (CRP), and convulxin. The Src family kinase inhibitors prevented Syk phosphorylation and its ubiquitination, indicating that the process is downstream of Src kinases.

Differential requirements for calcium and Src family kinases in platelet GPIIb/IIIa activation and thromboxane generation downstream of different G-protein pathways.

G(12/13) or G(q) signaling pathways activate platelet GPIIb/IIIa when combined with G(i) signaling. We tested whether combined G(i) and G(z) pathways also cause GPIIb/IIIa activation and compared the signaling requirements of these events. Platelet aggregation occurred by combined stimulation of G(i) and G(z) pathways in human platelets and in P2Y1-deficient and G alpha(q)-deficient mouse platelets, confirming that the combination of G(i) and G(z) signaling causes platelet aggregation.

Arg333 and Arg334 in the COOH terminus of the human P2Y1 receptor are crucial for Gq coupling.

The P2Y(1) ADP receptor activates G(q) and causes increases in intracellular Ca(2+) concentration through stimulation of PLC. In this study, we investigated the role of the amino acid residues in the COOH terminus of the human P2Y(1) receptor in G(q) activation. Stimulation of Chinese hamster ovary (CHO-K1) cells stably expressing the wild-type human P2Y(1) receptor (P2Y(1)-WT cells), P2Y(1)-DeltaR340-L373, or P2Y(1)-DeltaD356-L373 with 2-methylthio-ADP (2-MeSADP) caused inositol phosphate production.

Role of G protein-gated inwardly rectifying potassium channels in P2Y12 receptor-mediated platelet functional responses.

The role of the G(i)-coupled platelet P2Y(12) receptor in platelet function has been well established. However, the functional effector or effectors contributing directly to alphaIIbbeta3 activation in human platelets has not been delineated. As the P2Y(12) receptor has been shown to activate G protein-gated, inwardly rectifying potassium (GIRK) channels, we investigated whether GIRK channels mediate any of the functional responses of the platelet P2Y(12) receptor.

Plasmin-mediated activation of platelets occurs by cleavage of protease-activated receptor 4.

The activation of plasmin from its circulating precursor plasminogen is the mechanism of several clot-busting drugs used to clinically treat patients who have suffered a stroke; however, plasmin thus generated has been shown to activate platelets directly. There has been speculation as to whether plasmin interacts with the protease-activated receptors (PARs) because of its similarity in amino acid specificity with the classic platelet activator thrombin. We have investigated whether plasmin activates platelets via PAR activation through multiple complementary approaches.