Effects of variation in the human α2A‐ and α2C‐adrenoceptor genes on cognitive tasks and pain perception

Background: The mechanisms underlying interindividual variability in pain perception and cognitive responses are undefined but highly heritable. α_2C‐ and α_2A‐adrenergic receptors regulate noradrenergic activity and are important mediators of pain perception and analgesia. We hypothesized that common genetic variants in these genes, particularly the ADRA2C 322–325 deletion variant, affect pain perception or cognitive responses. Methods: We studied 73 healthy subjects (37 Caucasians and 36 African–Americans) aged 25.4 ± 4.6 years. Pain response to a cold pressor test was measured using a 10 cm visual analog scale and again on the next day, after three infusions of the selective α₂‐agonist dexmedetomidine. Standardized cognitive tests were administered at baseline and after each infusion. The contribution of ADRA2C deletion genotype, dexmedetomidine concentration, and other covariates to pain perception and cognitive responses was determined using multiple linear regression models. Secondary analysis examined the effects of ADRA2A and other ADRA2C variants on pain perception. Results: ADRA2C Del homozygotes had higher pain scores in response to cold at baseline (6.3 ± 1.8 cm) and after dexmedetomidine (5.6 ± 2.2 cm) than insertion allele carriers (4.6 ± 2.1 cm [baseline] and 3.8 ± 1.9 cm [after dexmedetomidine]; adjusted P‐values = 0.019 and 0.004, respectively). Cognitive responses were unrelated to ADRA2C Ins/Del genotype. None of the other ADRA2A and ADRA2C variants was significantly related to cold pain sensitivity before dexmedetomidine; after dexmedetomidine, ADRA2A rs1800038 was marginally associated (P = 0.03). Conclusion: The common ADRA2C del322–325 variant affected pain perception before and after dexmedetomidine but did not affect other cognitive responses, suggesting that it contributes to interindividual variability in pain perception.     

β2‐Glycoprotein I: evolution,structure and function

Summary. β₂‐Glycoprotein I (β₂‐GPI) is a protein that circulates in blood at high concentrations. The function of β₂‐GPI has long been an enigma. More than 20 years ago, it was discovered that β₂‐GPI is the major antigen for the circulating antibodies in the antiphospholipid syndrome. However, this knowledge has not advanced our understanding of the physiologic role of the protein. In recent years, new insights have suggested an important function of this protein in innate immunity. β₂‐GPI was found to scavenge lipopolysaccharide and was able to clear unwanted anionic cellular remnants such as microparticles from the circulation. The function of β₂‐GPI seems to depend on the structural conformation of the protein, and it has been established that β₂‐GPI can exist in at least two conformations. In this review, we will highlight and summarize the current knowledge on this protein.     

Antibacterial and β‐Lactamase Inhibitory Activity of Monocyclic β‐Lactams

Due to the widespread emergence of resistant bacterial strains, an urgent need for the development of new antibacterial agents with novel modes of action has emerged. The discovery of naturally occurring monocyclic β‐lactams in the late 1970s, mainly active against aerobic Gram‐negative bacteria, has introduced a new approach in the design and development of novel antibacterial β‐lactam agents. The main goal was the derivatization of the azetidin‐2‐one core in order to improve their antibacterial potency, broaden their spectrum of activity, and enhance their β‐lactamase stability. In that respect, our review covers the updates in the field of monocyclic β‐lactam antibiotics during the last three decades, taking into account an extensive collection of references. An overview of the relationships between the structural features of these monocyclic β‐lactams, classified according to their N‐substituent, and the associated antibacterial or β‐lactamase inhibitory activities is provided. The different paragraphs disclose a number of well‐established classes of compounds, such as monobactams, monosulfactams, monocarbams, monophosphams, nocardicins, as well as other known representative classes. Moreover, this review draws attention to some less common but, nevertheless, possibly important types of monocyclic β‐lactams and concludes by highlighting the recent developments on siderophore‐conjugated classes of monocyclic β‐lactams.     

Collagen‐mimetic peptides mediate flow‐dependent thrombus formation by high‐ or low‐affinity binding of integrin α2β1 and glycoprotein VI

Summary. Background: Collagen acts as a potent surface for platelet adhesion and thrombus formation under conditions of blood flow. Studies using collagen‐derived triple‐helical peptides have identified the GXX’GER motif as an adhesive ligand for platelet integrin α2β1, and (GPO)_n as a binding sequence for the signaling collagen receptor, glycoprotein VI (GPVI). Objective: The potency was investigated of triple‐helical peptides, consisting of GXX’GER sequences within (GPO)_n or (GPP)_n motifs, to support flow‐dependent thrombus formation. Results: At a high‐shear rate, immobilized peptides containing both the high‐affinity α2β1‐binding motif GFOGER and the (GPO)_n motif supported platelet aggregation and procoagulant activity, even in the absence of von Willebrand factor (VWF). With peptides containing only one of these motifs, co‐immobilized VWF was needed for thrombus formation. The (GPO)_n but not the (GPP)_n sequence induced GPVI‐dependent platelet aggregation and procoagulant activity. Peptides with intermediate affinity (GLSGER, GMOGER) or low‐affinity (GASGER, GAOGER) α2β1‐binding motifs formed procoagulant thrombi only if both (GPO)_n and VWF were present. At a low‐shear rate, immobilized peptides with high‐ or low‐affinity α2β1‐binding motifs mediated formation of thrombi with procoagulant platelets only in combination with (GPO)_n. Conclusions: Triple‐helical peptides with specific receptor‐binding motifs mimic the properties of native collagen I in thrombus formation by binding to both platelet collagen receptors. At a high‐shear rate, either GPIb or high‐affinity (but not low‐affinity) GXX’GER mediates GPVI‐dependent formation of procoagulant thrombi. By extension, high‐affinity binding for α2β1 can control the overall platelet‐adhesive activity of native collagens.     

A role for adhesion and degranulation‐promoting adapter protein in collagen‐induced platelet activation mediated via integrin α2β1

Summary. Background: Collagen‐induced platelet activation is a key step in the development of arterial thrombosis via its interaction with the receptors glycoprotein (GP)VI and integrin α₂β₁. Adhesion and degranulation‐promoting adapter protein (ADAP) regulates α_IIbβ₃ in platelets and α_Lβ₂ in T cells, and is phosphorylated in GPVI‐deficient platelets activated by collagen. Objectives: To determine whether ADAP plays a role in collagen‐induced platelet activation and in the regulation and function of α₂β₁. Methods: Using ADAP^?/? mice and synthetic collagen peptides, we investigated the role of ADAP in platelet aggregation, adhesion, spreading, thromboxane synthesis, and tyrosine phosphorylation. Results and Conclusions: Platelet aggregation and phosphorylation of phospholipase Cγ2 induced by collagen were attenuated in ADAP^?/? platelets. However, aggregation and signaling induced by collagen‐related peptide (CRP), a GPVI‐selective agonist, were largely unaffected. Platelet adhesion to CRP was also unaffected by ADAP deficiency. Adhesion to the α₂β₁‐selective ligand GFOGER and to a peptide (III‐04), which supports adhesion that is dependent on both GPVI and α₂β₁, was reduced in ADAP^?/? platelets. An impedance‐based label‐free detection technique, which measures adhesion and spreading of platelets, indicated that, in the absence of ADAP, spreading on GFOGER was also reduced. This was confirmed with non‐fluorescent differential‐interference contrast microscopy, which revealed reduced filpodia formation in ADAP^?/? platelets adherent to GFOGER. This indicates that ADAP plays a role in mediating platelet activation via the collagen‐binding integrin α₂β₁. In addition, we found that ADAP^?/? mice, which are mildly thrombocytopenic, have enlarged spleens as compared with wild‐type animals. This may reflect increased removal of platelets from the circulation.     

Integrin Ligands with α/β‐Hybrid Peptide Structure: Design,Bioactivity, and Conformational Aspects

Integrins are cell surface receptors for proteins of the extracellular matrix and plasma‐borne adhesive proteins. Their involvement in diverse pathologies prompted medicinal chemists to develop small‐molecule antagonists, and very often such molecules are peptidomimetics designed on the basis of the short native ligand‐integrin recognition motifs. This review deals with peptidomimetic integrin ligands composed of α‐ and β‐amino acids. The roles exerted by the β‐amino acid components are discussed in terms of biological activity, bioavailability, and selectivity. Special attention is paid to the synthetic accessibility and efficiency of conformationally constrained heterocyclic scaffolds incorporating α/β‐amino acid span.     

An αIIb mutation in patients with Glanzmann thrombasthenia located in the N‐terminus of blade 1 of the β‐propeller (Asn2Asp) disrupts a calcium binding site in blade 6

Summary. Background: Studies of Glanzmann thrombasthenia (GT)‐causing mutations has generated invaluable information on the formation and function of integrin αIIbβ₃. Objective: To characterize the mutation in four siblings of an Israeli Arab family affected by GT, and to analyze the relationships between the mutant protein structure and its function using artificial mutations. Methods and Results: Sequencing disclosed a new A97G transversion in the αIIb gene predicting Asn2Asp substitution at blade 1 of the β‐propeller. Alignment with other integrin α subunits revealed that Asn2 is highly conserved. No surface expression of αIIbβ₃ was found in patients’ platelets and baby hamster kidney (BHK) cells transfected with mutated αIIb and WT β₃. Although the αIIbβ₃ was formed, the mutation impaired its intracellular trafficking. Molecular dynamics simulations and modeling of the αIIbβ₃ crystal indicated that the Asn2Asp mutation disrupts a hydrogen bond between Asn2 and Leu366 of a calcium binding domain in blade 6, thereby impairing calcium binding that is essential for intracellular trafficking of αIIbβ₃. Substitution of Asn2 to uncharged Ala or Gln partially decreased αIIbβ₃ surface expression, while substitution by negatively or positively charged residues completely abolished surface expression. Unlike αIIbβ₃, αVβ₃ harboring the Asn2Asp mutation was surface expressed by transfected BHK cells, which is consistent with the known lower sensitivity of αVβ₃ to calcium chelation compared with αIIbβ₃. Conclusion: The new GT causing mutation highlights the importance of calcium binding domains in the β‐propeller for intracellular trafficking of αIIbβ₃. The mechanism by which the mutation exerts its deleterious effect was elucidated by molecular dynamics.