Different RBC aggregating properties of the Aalpha2Bbeta2gammaA2 and Aalpha2Bbeta2gammaAgamma' subpopulations of human fibrinogen

Human fibrinogen (TF) has been separated into two fractions: F1 - homodimers with respect to the gamma chain, and F2 - heterodimers composed of gammaA and gamma' polypeptides. Their rouleaux-inducing properties were as follows: (1) both, at the same concentration of 0.8%, were less effective than TF; (2) F1 produced larger rouleaux even under static conditions of a hemocytometer where F2 was silent; (3) F2 induced the process when a suspension was gently sheared between microscopic slides. Since the synthetic peptide gamma'(414-427) inhibited the rouleau formation in a mixture with F2, the C-terminal amino acids of the gamma' polypeptide probably bind the molecule to the cell. The inhibition was feebly visible in the native ratio of F1/F2, implicating a compensatory effect of F1.     

Characterization of two types of fetal hemoglobin: alpha 2G gamma 2 and alpha 2A gamma 2

The effect of differences in G gamma and A gamma fractions of fetal hemoglobin (HbF) on the kinetics of polymerization of HbS-HbF mixtures was studied. We also examined their effect on oxygen affinity, surface hydrophobicity, mechanical stability, and solubility of HbF. Differences in G gamma:A gamma ratio did not affect the polymerization of mixtures of HbF and HbS, suggesting that the inhibitory effect of HbF on the polymerization of HbS is independent of the G gamma:A gamma ratio of HbF and is totally dependent on the fraction of HbF in the mixture. The oxygen equilibrium curve of HbF was not affected by differences in the ratios of G gamma and A gamma in HbF. In contrast, surface hydrophobicity, mechanical stability, and solubility of HbF were affected by differences in the G gamma:A gamma ratio. The higher the G gamma:A gamma ratio, the smaller the elution volume on a TSK Gel SW hydrophobic column in high phosphate buffer. The mechanical stability of HbF was also dependent on the ratio of G gamma:A gamma; stability was greater at higher fractions of A gamma. Differences in the G gamma:A gamma ratio also affected solubility of HbF: HbF containing the higher fraction of G gamma was the more soluble. These data indicate that although alanine at the 136th position of the gamma chains has a stronger surface hydrophobicity than does glycine, this difference does not affect either the polymerization of HbS or the oxygen affinity of HbF.     

Structure-function of Hb Marseille-Long Island [alpha 2 beta 2 N-methionyl-2(NA2) His----Pro

Suspensions of red cells containing Hb Marseille-Long Island showed decreased oxygen affinity and low interaction with 2,3-diphosphoglycerate. Oxygen equilibrium studies of the purified component confirmed these abnormalities. Oxidation rate measurements of carbonmonoxy-Hb Marseille and carbonmonoxy-Hb A by ferricyanide showed an increased rate for the former, suggesting an increased dissociation constant for carbon monoxide. Nuclear Magnetic Resonance spectra in the high field region revealed small changes in the proximal region of the heme pocket. These results indicated that the mutation causes a perturbation at a distance from the mutation site.     

Classification and characterization of hereditary types 2A, 2B, 2C, 2D, 2E, 2M, 2N, and 2U (unclassifiable) von Willebrand disease.

All variants of type 2 von Willebrand disease (VWD) patients, except 2N, show a defective von Willebrand factor (VWF) protein (on cross immunoelectrophoresis or multimeric analysis), decreased ratios for VWF:RCo/Ag and VWF:CB/Ag and prolonged bleeding time. The bleeding time is normal and FVIII:C levels are clearly lower than VWF:Ag in type 2N VWD. High resolution multimeric analysis of VWF in plasma demonstrates that proteolysis of VWF is increased in type 2A and 2B VWD with increased triplet structure of each visuable band (not present in types 2M and 2U), and that proteolysis of VWF is minimal in type 2C, 2D, and 2E variants that show aberrant multimeric structure of individual oligomers. VWD 2B differs from 2A by normal VWF in platelets, and increased ristocetine-induced platelet aggregation (RIPA). RIPA, which very likely reflects the VWF content of platelets, is normal in mild, decreased in moderate, and absent in severe type 2A VWD. RIPA is decreased or absent in 2M, 2U, 2C, and 2D, variable in 2E, and normal in 2N. VWD 2M is usually mild and characterized by decreased VWF:RCo and RIPA, a normal or near normal VWF multimeric pattern in a low resolution agarose gel. VWD 2A-like or unclassifiable (2U) is distinct from 2A and 2B and typically featured by low VWF:RCo and RIPA with the relative lack of high large VWF multimers. VWD type 2C is recessive and shows a characteristic multimeric pattern with a lack of high molecular weight multimers, the presence of one single-banded multimers instead of triplets caused by homozygosity or double hereozygosity for a mutation in the multimerization part of VWF gene. Autosomal dominant type 2D is rare and characterized by the lack of high molecular weight multimers and the presence of a characteristic intervening subband between individual oligimers due to mutation in the dimerization part of the VWF gene. In VWD type 2E, the large VWF multimers are missing and the pattern of the individual multimers shows only one clearly identifiable band, and there is no intervening band and no marked increase in the smallest oligomer. 2E appears to be less well defined, is usually autosomal dominant, and accounts for about one third of patients with 2A in a large cohort of VWD patients.     

Experimental Investigation of Radiation Shielding Competence of Bi2O3-CaO-K2O-Na2O-P2O5 Glass Systems

The gamma-ray shielding features of Bi₂O₃-CaO-K₂O-Na₂O-P₂O₅ glass systems were experimentally reported. The mass attenuation coefficient (MAC) for the fabricated glasses was experimentally measured at seven energy values (between 0.0595 and 1.33 MeV). The compatibility between the practical and theoretical results shows the accuracy of the results obtained in the laboratory for determining the MAC of the prepared samples. The mass and linear attenuation coefficients (MACs) increase with the addition of Bi₂O₃ and A4 glass possesses the highest MAC and LAC. A downward trend in the linear attenuation coefficient (LAC) with increasing the energy from 0.0595 to 1.33 MeV is found. The highest LAC is found at 1.33 MeV (in the range of 0.092–0.143 cm⁻¹). The effective atomic number (Z_eff) follows the order B1 > A1 > A2 > A3 > A4. This order emphasizes that increasing the content of Bi₂O₃ has a positive effect on the photon shielding proficiencies owing to the higher density of Bi₂O₃ compared with Na₂O. The half value layer (HVL) is also determined and the HVL for the tested glasses is computed between 0.106 and 0.958 cm at 0.0595 MeV. The glass with 10 mol% of Bi₂O₃ has lower HVL than the glasses with 0, 2.5, 5, and 7.5 mol% of Bi₂O₃. So, the A4 glass needs a smaller thickness than the other glasses to shield the same radiation. As a result of the reported shielding parameters, inserting B₂O₃ provides lower values of these three parameters, which in turn leads to the development of superior photons shields.     

First-order antiferromagnetic transitions of SrMn2P2 and CaMn2P2 single crystals containing corrugated-honeycomb Mn sublattices

SrMn₂P₂ and CaMn₂P₂ are insulators that adopt the trigonal CaAl₂Si₂-type structure containing corrugated Mn honeycomb layers. Magnetic susceptibility χ and heat capacity versus temperature T data reveal a weak first-order antiferromagnetic (AFM) transition at the Néel temperature TN=53(1) K for SrMn₂P₂ and a strong first-order AFM transition at TN=69.8(3) K for CaMn₂P₂. Both compounds exhibit isotropic and nearly T-independent χ(T≤TN), suggesting magnetic structures in which nearest-neighbor moments are aligned at ≈120° to each other. The ³¹P NMR measurements confirm the strong first-order transition in CaMn₂P₂ but show critical slowing down above TN for SrMn₂P₂, thus also evidencing second-order character. The ³¹P NMR measurements indicate that the AFM structure of CaMn₂P₂ is commensurate with the lattice whereas that of SrMn₂P₂ is incommensurate. These first-order AFM transitions are unique among the class of (Ca, Sr, Ba)Mn₂ (P, As, Sb, Bi)₂ compounds that otherwise exhibit second-order AFM transitions. This result challenges our understanding of the circumstances under which first-order AFM transitions occur.

The Mn-based 122-type pnictides AMn2Pn2 (A= Ca, Sr, Ba; Pn = P, As, Sb, Bi) have received attention owing to their close stoichiometric 122-type relationship to high-Tc iron pnictides. The undoped Mn pnictides are local-moment antiferromagnetic (AFM) insulators like the high-Tc cuprate parent compounds (1–3). The BaMn2Pn2 compounds crystallize in the body-centered tetragonal ThCr2Si2 structure as in AFe2As2 (A = Ca, Sr, Ba, Eu), whereas the (Ca,Sr)Mn2Pn2 compounds crystallize in the trigonal CaAl2Si2-type structure (4). Recently, density-functional theory (DFT) calculations for the 122 pnictide family have suggested that the trigonal 122 transition-metal pnictides that have the CaAl2Si2 structure might compose a new family of magnetically frustrated materials in which to study the potential superconducting mechanism (5, 6). It had previously been suggested on theoretical grounds that CaMn₂Sb₂ is a fully frustrated classical magnetic system arising from proximity to a tricritical point (7–9).The electrical resistivity ρ and heat capacity Cp versus temperature T of single-crystal CaMn₂P₂ were reported in ref. 10. The compound is an insulator at T = 0 and undergoes a first-order transition of some type at 69.5 K. The Raman spectrum of CaMn₂P₂ at T = 10 K showed new peaks compared to the spectrum at 300 K, whereas the authors’ single-crystal X-ray diffraction measurements showed no difference in the crystal structure at 293 and 40 K. They suggested that the results of the two types of measurements could be reconciled if a superstructure formed below 69.5 K (10). The authors’ magnetic susceptibility χ(T) measurements below 400 K revealed no evidence for a magnetic transition.Here we report the detailed properties of trigonal CaMn₂P₂ and SrMn₂P₂ (11) single crystals. We present the results of single-crystal X-ray diffraction (XRD), electrical resistivity ρ in the ab plane (hexagonal unit cell) versus temperature T, isothermal magnetization versus applied magnetic field M(H), magnetic susceptibility χ(T), heat capacity Cp(H,T), and ³¹P NMR measurements. We find from Cp(T), χ(T), and NMR that CaMn₂P₂ exhibits a strong first-order AFM transition at TN=69.8(3) K whereas SrMn₂P₂ shows a weak first-order transition at TN=53(1) K but with critical slowing down on approaching TN from above as revealed from NMR, a characteristic feature of second-order transitions. Thus, remarkably, the AFM transition in SrMn₂P₂ has characteristics of both first- and second-order transitions. The χ(T) data also reveal the presence of strong isotropic AFM spin fluctuations in the paramagnetic (PM) state above TN up to our maximum measurement temperatures of 900 and 350 K for SrMn₂P₂ and CaMn₂P₂, respectively. This behavior likely arises from spin fluctuations associated with the quasi–two-dimensional nature of the Mn spin layers (12) together with possible contributions from magnetic frustration. Our single-crystal XRD data at room temperature and high-resolution synchrotron XRD data at T = 20 K for SrMn₂P₂ and CaMn₂P₂ demonstrate conclusively that there is no structure change of either compound on cooling below their respective TN.Our studies of SrMn₂P₂ and CaMn₂P₂ thus identify the only known members of the class of materials with general formula AMn2Pn2 containing Mn2+ spins S = 5/2 that exhibit first-order AFM transitions, where A = Ca, Sr, or Ba and the pnictogen Pn= P, As, Sb, or Bi. In particular, only second-order AFM transitions are found in CaMn2As2 (13), SrMn2As2 (13–15), CaMn2Sb2 (8, 9, 16–19), SrMn2Sb2 (16, 19), and CaMn2Bi2 (20).     

Identification of SH2B2beta as an inhibitor for SH2B1- and SH2B2alpha-promoted Janus kinase-2 activation and insulin signaling

The SH2B family has three members (SH2B1, SH2B2, and SH2B3) that contain conserved dimerization (DD), pleckstrin homology, and SH2 domains. The DD domain mediates the formation of homo- and heterodimers between members of the SH2B family. The SH2 domain of SH2B1 (previously named SH2-B) or SH2B2 (previously named APS) binds to phosphorylated tyrosines in a variety of tyrosine kinases, including Janus kinase-2 (JAK2) and the insulin receptor, thereby promoting the activation of JAK2 or the insulin receptor, respectively. JAK2 binds to various members of the cytokine receptor family, including receptors for GH and leptin, to mediate cytokine responses. In mice, SH2B1 regulates energy and glucose homeostasis by enhancing leptin and insulin sensitivity. In this work, we identify SH2B2beta as a new isoform of SH2B2 (designated as SH2B2alpha) derived from the SH2B2 gene by alternative mRNA splicing. SH2B2beta has a DD and pleckstrin homology domain but lacks a SH2 domain. SH2B2beta bound to both SH2B1 and SH2B2alpha, as demonstrated by both the interaction of glutathione S-transferase-SH2B2beta fusion protein with SH2B1 or SH2B2alpha in vitro and coimmunoprecipitation of SH2B2beta with SH2B1 or SH2B2alpha in intact cells. SH2B2beta markedly attenuated the ability of SH2B1 to promote JAK2 activation and subsequent tyrosine phosphorylation of insulin receptor substrate-1 by JAK2. SH2B2beta also significantly inhibited SH2B1- or SH2B2alpha-promoted insulin signaling, including insulin-stimulated tyrosine phosphorylation of insulin receptor substrate-1. These data suggest that SH2B2beta is an endogenous inhibitor of SH2B1 and/or SH2B2alpha, negatively regulating insulin signaling and/or JAK2-mediated cellular responses.     

Identification of CYP2C9*2 allele in HepG2 cell line

Background Hepatoma is caused by many factors including alcohol, chemicals, viral infection, and chronic inflammation. Cytochrome P450 polymorphism plays an important role in its pathogenesis. CYP2D6, CYP2E1, and CYP1A1 have been identified to be related with hepatic carcinogenesis and tumor size and stage. However, no studies have been performed on CYP2C9, a major CYP in the liver and hepatoma. Aim of the study To identify if there is polymorphism of CYP2C9 in a HepG2 cell line. Methods A pair of primers was used to clone CYP2C9 exon 3 region and subsequently sequenced. The sequence was compared to normal CYP2C9 for identification of any mutation. Results A point mutation was identified. It was located in the amino acid number 144 of CYP2C9 protein with the change of normal amino acid arginine into cysteine, which is the same as identified in poor metabolism patients as homozygous CYP2C9*2. Conclusions There is a mutation (CYP2C9*2/ CYP2C9*2) in a HepG2 cell line. Thus, polymorphism of CYP2C9 may also be involved in the carcinogenesis of hepatoma as CYPs2D6 and 2E1.     

CYP2J2基因和EPHX2基因多态性与缺血性脑卒中的遗传易感性

目的 研究我国汉族人群细胞色素P450表氧化酶2J2(CYP2J2),可溶性环氧化物水解酶2(EPHX2)基因多态性与缺血性脑卒中的关系.方法 选择200例缺血性脑卒中患者和350例健康人群,采用PCR-RFLP技术分析两个基因CYP2J2 G-50T,EPHX2 G860A多态性的基因型.结果 仅EPHX2 860A等位基因频率在缺血性脑卒中组与对照组比较有显著性差异.多元Logistic回归分析表明,携带EPHX2 860A等位基因的人群患缺血性脑卒中相对风险率下降50%(OR=0.5).当个体同时携带CYP2J2-50GG和EPHX2 860A (A 示A等位基因)联合基因型时,其患缺血性脑卒中相对风险率下降53.9%(OR=0.461).结论 虽然EPHX2 860A等位基因与缺血性脑卒中有相关性并且为缺血性脑卒中一个独立的保护因子,但联合基因型CYP2J2-50GG/EPHX2 860A 的协同作用对缺血性脑卒中有更强的保护作用.     

The effects of changes of maternal PaO2 and PaCO2 on the fetal PaO2 and PaCO2--in vivo study

Umbilical PaO2 and PaCO2 were continuously monitored in vivo in acute fetal lamb preparations with a semipermeable membrane connected to a mass spectrometer. The response time of this system (0 to 90% of final value) was 36 sec. In seven pregnant sheep (128--135 days gestation) the maternal inspired mixture was abruptly changed and the following changes in fetal PaO2 and PaCO2 were observed: (1) 100% O2 to room air: PaO2 decreased from 21.5 +/- 0.8 (mean +/- SEM) to 14 +/- 1.1 mm Hg at a rate of 1.63 +/- 0.33 mm Hg/min. Following return to 100% O2 the PaO2 returned to 21 +/- 1.1 mm Hg at a rate of 2.44 +/- 0.4 mm Hg/min. (2) 100% O2 to 12% O2/10% CO2: after 6 min the PaO2 fell from 19.3 +/- 1.3 to 6.3 +/- 0.3 mm Hg at a rate of 4.65 mm Hg/min and the PaCO2 rose from 37 +/- 8 to 70 +/-5 mm Hg. At 100% O2 the PaO2 returned to 19 +/- 1.0 mm Hg at a rate of 11.76 +/- 0.086 mm Hg, the PaCO2 to 39 +/- 7 mm Hg. (3) 100% O2 to 90% O2/10% CO2. The PaO2 and PaCO2 increased by 4.7 and 22 mm Hg, respectively. The changes of fetal PaO2 and PaCO2 occurred after 1 minute of changing in maternal inspired mixture except in the transition from 12% O2/10% CO2 to 100% O2 (34 +/- 12 sec). Following the reinstitution of 100% the fetal PaO2 and PaCO2 returned to their previous values within 4 and 16 min, respectively.     

Effects of neuraminidase on O2 consumption and release of O2 and H2O2 from phagocytosing human polymorphonuclear leukocytes

Neuraminidase type X (NMD-type-X, Sigma Chemical Co., St. Louis, Mo.), which is obtained from a further purification of neuraminidase type V (NMD-type-V, Sigma), markedly enhanced the release of O2- and H2O2 from phagocytosing human polymorphonuclear leukocytes (PMN). In contrast, O2 consumption by NMD-type-X-treated PMN was identical to that of untreated PMN. Morphological observations suggested that the enhancement of O2- and H2O2 release was caused by excessive release of the oxygen metabolites into the extracellular medium from incompletely formed phagocytic vacuoles as was observed with cytochalasin-B-treated cells. Our observations are in contrast to the previous reports of Tsan et al. that showed complete inhibition of both O2- and H2O2 release from phagocytosing PMN by the treatment with NMD-type-V.     

采用重组侧链二氧酸脱氢酶复合物E2亚单位检测M2抗体及其临床意义

目的 探讨用重组表达的侧链二氧酸脱氢酶复合物E2亚单位（BCOADC—E2）检测M2抗体，以早期诊断原发性胆汁性肝硬化（PBC）。方法 重组表达BCOADC—E2，采用酶联免疫吸附试验（ELISA）和免疫印迹法（IBT）来检测PBC患者的M2抗体，并以健康体检者、自身免疫病患者、其他非胆汁性肝硬化患者为对照组。结果 60例肝病患者血清中检测出抗BCOADC—E2抗体阳性33例，阴性者为27例，阳性率为55％，而健康体检者和疾病对照组血清中M2抗体检测均为阴性。结论 采用重组表达的人侧链二氧酸脱氢酶复合物E2亚单位检测M2抗体，有一定的特异性，对于早期诊断PBC有一定的辅助参考作用。     

Cross-species recognition of SARS-CoV-2 to bat ACE2

The coronavirus disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has emerged as a major threat to global health. Although varied SARS-CoV-2–related coronaviruses have been isolated from bats and SARS-CoV-2 may infect bat, the structural basis for SARS-CoV-2 to utilize the human receptor counterpart bat angiotensin-converting enzyme 2 (bACE2) for virus infection remains less understood. Here, we report that the SARS-CoV-2 spike protein receptor binding domain (RBD) could bind to bACE2 from Rhinolophus macrotis (bACE2-Rm) with substantially lower affinity compared with that to the human ACE2 (hACE2), and its infectivity to host cells expressing bACE2-Rm was confirmed with pseudotyped SARS-CoV-2 virus and SARS-CoV-2 wild virus. The structure of the SARS-CoV-2 RBD with the bACE2-Rm complex was determined, revealing a binding mode similar to that of hACE2. The analysis of binding details between SARS-CoV-2 RBD and bACE2-Rm revealed that the interacting network involving Y41 and E42 of bACE2-Rm showed substantial differences with that to hACE2. Bats have extensive species diversity and the residues for RBD binding in bACE2 receptor varied substantially among different bat species. Notably, the Y41H mutant, which exists in many bats, attenuates the binding capacity of bACE2-Rm, indicating the central roles of Y41 in the interaction network. These findings would benefit our understanding of the potential infection of SARS-CoV-2 in varied species of bats.

The coronavirus disease 2019 (COVID-19), caused by infection with the novel coronavirus (CoV) severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a major threat to global health with an increasing number of infected cases globally, and the end to this pandemic is still full of uncertainties (1–3). Seven CoVs have been reported to infect humans: SARS-CoV, Middle East respiratory syndrome coronavirus (MERS-CoV), NL-63, OC43, 229E, HKU1, and the newly emerging SARS-CoV-2. Among the SARS-CoV-2 proteins, the spike (S) protein, which consists of an N-terminal S1 subunit and a C-terminal S2 subunit, is critical for the recognition of host cell receptors and serves as the key determinant of host specificity for CoVs (4). The C-terminal domain of the S1 subunit, also known as the receptor binding domain (RBD), binds to the human angiotensin-converting enzyme 2 (hACE2), the receptor for SARS-CoV and the human coronavirus NL-63 (5–9). Monoclonal antibodies (mAbs) that block the binding between SARS-CoV-2 RBD and ACE2 could efficiently inhibit virus infection in host cells expressing ACE2 (10, 11). Animal studies revealed that a single dose of these neutralizing mAbs showed promising therapeutic efficacy in reducing both viral load and pathological lung damage in hACE2 transgenic mice or rhesus macaques (10, 11). Recently, the structures of the complex between SARS-CoV-2 S protein (or RBD) with hACE2 have been determined, showing similar binding mode to that of SARS-CoV but with enhanced affinity (5–7).Bats are considered as the reservoir host animals of SARS-CoV-2, and several SARS-CoV-2-related CoVs have been identified from bats (12, 13). The binding assay showed that bat ACE2 (bACE2) from Rhinolophus macrotis (bACE2-Rm) bound to SARS-CoV-2 RBD efficiently (14). The entry capacity of SARS-CoV-2 through ACE2 orthologs from 46 bat species was evaluated by virus-host receptor binding and infection assays. The results indicated that although some bACE2 receptors could mediate SARS-CoV-2 entry, there are many bACE2 receptors that do not yet support SARS-CoV-2 entry (15). To date, the genome most closely related to SARS-CoV-2 is RaTG13, which was identified from a Rhinolophus affinis sampled from Yunnan Province in 2013 and had a 92.9% amino acid identity in the S gene (13). RmYN02 was also a coronavirus identified from bat with 93.3% nucleotide identity with SARS-CoV-2 at the scale of the complete virus genome (12). Yongyi Shen and colleagues (16) reported a coronavirus isolated from Malayan pangolin and shared 90.7% amino acid identity with SARS-CoV-2 in the S proteins. In addition, a coronavirus that showed 97.4% amino acid identity with SARS-CoV-2 in the RBD region was identified from another batch of Malayan pangolin (17). Bioinformatics analysis indicated that bats are the primary reservoir for the SARS-CoV-2 lineage (18). However, the origin of SARS-CoV-2 is full of uncertainties and whether or not there is an intermediate host is still unknown. Moreover, more than 1,400 species of bats have been identified and extensive species diversity among different species of bats has resulted in varied susceptibility for CoVs. Research into the mechanisms of viral entry and virus–host interaction would not only benefit our understanding of this virus but is also important for the design of antivirals and vaccines. However, the structure of bACE2 has not been determined, and the molecular basis of the binding between SARS-CoV-2 S protein and bACE2 has not been well studied.Here, we report that SARS-CoV-2 RBD can bind to bACE2-Rm with substantially lower affinity than that to hACE2, and infection of host cells carrying bACE2-Rm was also investigated with pseudotyped or wild SARS-CoV-2. Interaction mechanisms between SARS-CoV-2 RBD and bACE2-Rm were elucidated by determining the structure of the SARS-CoV-2 RBD and bACE2-Rm complex. The results of this study would broaden our understanding of the receptor binding mechanisms for SARS-CoV-2.