The effects of problem-solving training on two problem-solving tasks

The present study assessed the effects of problem-solving (D'Zurilla & Goldfried, 1971) training on two problem-solving tasks. Sixty-one subjects were assigned randomly to and trained in one of the following groups: general orientation (GO)-problem definition (PD)-generation of alternatives (GA)-decision making (DM), GO-PD-GA, GO-PD-DM, GO-GA-DM, GO, and a control group. The six groups did not differ significantly at pretesting on the measures of selecting the best alternative solution and describing problem-solving behaviors used to solve problems. Training was significantly effective on selecting the best alternative solution for all groups compared with the control group at the time of posttesting. The GO-PD-GA-DM and GO-PD-DM groups described significantly more problem-solving behaviors than the GO-PD-GA, GO-GA-DM, and GO groups. All five treatment groups performed significantly better than the control group at the time of post-testing. At follow-up 1 month after posttesting, groups trained with the PD component selected significantly better alternatives than the groups without PD training. Groups trained with the DM component described significantly more problem-solving behaviors at follow-up than the groups trained without the DM component.     

凋亡调控因子在凝血酶诱导海马神经元凋亡中的变化

目的探讨凝血酶（TM）对海马神经元凋亡调控凶子Bcl-2、Bax及c-jun氨基末端激酶（JNK）表达的影响，阐明TM诱导凋亡的机制。方法海马神经元经40U／mLTM作用0h、6h、12h、24h、48h、72h后终止培养，应用免疫细胞化学方法检测Bcl-2及Bax蛋白表达，Western blot检测磷酸化JNK（P—JNK）及JNK蛋白表达。结果TM作用6h后，Bcl-2蛋白表达增加，12h后开始下降，到48h达到最低值（P〈0．01），72h后Bcl-2蛋白表达开始回升。TM作用6h后，Bax蛋白表达增加，随作用时间的延长，Bax蛋白表达水平逐渐上升，到48h达到最高值（P〈0．01）。Bcl-2／Bax比值逐渐降低，在48h达最低。TM作用0h时P-JNK蛋白无表达，6h后开始表达，与0h组比较差异存在显著性（P〈0．011，12h时达高峰（P〈0．01），并持续到24h，48h后逐渐降低。0h组可见有少量JNK蛋白表达，6h后JNK蛋白表达显著增加（P〈0．01），随TM作用时间的延长，JNK蛋白表达无明显变化。结论TM可能是通过抑制Bcl-2的表达，增强Bax的表达，降低Bcl-2和Bax的比值，激活JNK信号传导通路导致海马神经元凋亡。     

胰高血糖素衍生物在大肠杆菌的克隆表达和纯化

 目的 通过基因工程途径获得胰高血糖素衍生物。 方法 根据胰高血糖素基因及凝血酶酶切位点序列，分段合成引物行 PCR 扩增，以 PCR 扩增得到的胰高血糖素-甘氨酸基因序列和 pET-30a 质粒转化 E.coli DH5α，获得重组质粒 pET-G。将重组质粒 pET-G 转化至 E.coli BL21(DE3)，重组菌株命名为 E.coli BL21[pET-G]。以异丙基-β-D-硫代半乳糖苷（IPTG）诱导表达，对表达产物进行SDS-Tricine- PAGE[十二烷基硫酸钠-三（羟甲基）甲基甘氨酸-聚丙烯酰胺凝胶电泳]分析和蛋白质印迹分析。对表达产物行Ni2+-NTA亲和层析纯化、凝血酶酶切和高效液相色谱（HPLC）纯化后，行 SDS-Tricine-PAGE分析和飞行质谱分析，并以 ELISA 方法检测其免疫活性。 结果 PCR 扩增获得的条带与预期的DNA表达片段大小一致，重组质粒 pET-G 测序结果与预期完全一致。SDS- Tricine-PAGE 和蛋白质印迹分析显示 E.coli BL21（DE3）的表达产物相对分子质量与预期相符。经亲和层析纯化、凝血酶酶切和 HPLC 纯化后得到了完整的重组胰高血糖素-甘氨酸衍生物，SDS-Tricine-PAGE 分析显示其相对分子质量约为 3500，飞行质谱分析相对分子质量为 3531，二者基本一致。ELISA 检测表明重组胰高血糖素-甘氨酸衍生物具有胰高血糖素免疫活性。 结论 采用基因工程技术在大肠杆菌中成功表达了胰高血糖素-甘氨酸衍生物，为通过体外酰胺化途径研制酰胺化胰高血糖素奠定了基础。     

A mathematical model of thrombin production in blood coagulation,Part I: The sparsely covered membrane case

This paper presents the first attempt to model the blood coagulation reactions in flowing blood. The model focuses on the common pathway and includes activation of factor X and prothrombin, including feedback activation of cofactors VIII and V by thrombin, and plasma inhibition of factor Xa and thrombin. In this paper, the first of two, the sparsely covered membrane (SCM) case is presented. This considers the limiting situation where platelet membrane binding sites are in excess, such that no membrane saturation or binding competition occurs. Under these conditions, the model predicts that the two positive feedback loops lead to multiple steady-state behavior in the range of intermediate mass transfer rates. It will be shown that this results in three parameter regions exhibiting very different thrombin production patterns. The model predicts the effect of flow on steady-state and dynamic thrombin production and attempts to explain the difference between venous and arterial thrombi. The reliance of thrombin production on precursor procoagulant protein concentrations is also assessed.     

Computational biology insights into genotype-clinical phenotype-protein phenotype relationships between novel SLC26A2 variants identified in inherited skeletal dysplasias

BackgroundPathogenic variants in the transmembrane sulfate transporter protein SLC26A2 are associated with different phenotypes of inherited chondrodysplasias. As limited data is published from India, in this study we sought to elucidate the molecular basis of inherited chondrodysplasias in an Indian cohort.MethodsMolecular screening of 32 fetuses with antenatally diagnosed lethal skeletal dysplasia was performed by next generation sequencing and Sanger sequencing. The genotype-protein phenotype characterization was done using computational biology techniques like homology modelling, stability and pathogenicity predictions.ResultsWe identified five rare autosomal recessive SLC26A2 [NM_000112.4] variants, including three homozygous c.796dupA(p.Thr266Asnfs*12), c.1724delA(p.Lys575Serfs*10), and c.1375_1377dup(p.Val459dup) and two heterozygous variants (c.532C > T(p.Arg178*)) and (c.1382C > T(p.Ala461Val)) in compound heterozygous form in a total of four foetuses. Genotype-protein phenotype annotations highlighted that the clinically severe achondrogenesis 1B causative c.796dupA(p.Thr266Asnfs*12) and c.1724delA(p.Lys575Serfs*10)variants impact SLC26A2 protein structure by deletion of the protein core and transmembrane STAS domains, respectively. In clinically moderate atelosteogenesis type 2 phenotype, the c.1382C > T(p.Ala461Val) variant is predicted to distort alpha helix conformation and alter the bonding properties and free energy dynamics of transmembrane domains and the c.532C > T(p.Arg178*) variant results in loss of both core transmembrane and STAS domains of the SLC26A2 protein. The c.1375_1377dup(p.Val459dup) variant identified in clinically milder atelosteogenesis type II-diastrophic dysplasia spectrum lethal phenotype is predicted to decrease the Qualitative Model Energy Analysis (QMean), which affects major geometrical aspects of the SLC26A2 protein structure.ConclusionWe expand the spectrum of SLC26A2 related lethal chondrodysplasia and report three novel variants correlating clinical severity and protein phenotype within the lethal spectrum of this rare dysplasia. We demonstrate the relevance of structural characterization to aid novel variant reclassification to provide better prenatal management and reproductive options to families with lethal antenatal skeletal disorder.     

Blood coagulation and fibrinolysis after long-duration treadmill exercise controlled by individual anaerobic threshold

For rehabilitation training it is recommended that the intensity of exercise should be clearly below the individual anaerobic threshold (IAT). We investigated blood coagulation, particularly endogenous thrombin potential (ETP) and fibrinolysis following a standardized treadmill (TR) ergometer test at 90% IAT for 60–120 min. Sixteen healthy male non-smokers underwent the TR test. Blood samples were taken after a 30-min rest, immediately after exercise, and 2 h after exercise completion. Extrinsic and intrinsic total (TTP_ex+in) and endogenous (ETP_ex+in) thrombin potential, prothrombin fragment 1+2 (F1+2), thrombin-antithrombin complex (TAT), plasmin-2-antiplasmin complex (PAP), D-dimer, tissue plasminogen activator antigen and activity (tPA-AG and tPA-ACT) and plasminogen activator inhibitor type 1 antigen and activity (PAI-1-AG and PAI-1-ACT) were measured. Immediately after TR, F1+2, TAT and TTP_ex+in were increased (P<0.05) while ETP_ex+in remained unchanged. In contrast, PAP, D-dimer, tPA-AG, tPA-ACT (P<0.05) were distinctly enhanced while PAI-1-ACT was decreased (P<0.05) immediately after exercise. The changes in tPA-AG, tPA-ACT, and PAI-1-ACT were reversed to nearly baseline while the enhancement in PAP and D-dimer was prolonged by more than 2 h after exercise. Long-duration exercise between 60 and 120 min controlled by IAT (90%) on a TR ergometer only implicates a small increase in thrombin generation markers and total (free and ₂-macroglubulin-bound thrombin), but not in endogenous (free) thrombin potential alone. In contrast, fibrinolysis is distinctly increased after this type of exercise. Endurance exercise with an intensity below 90% IAT and a duration below 2 h generates a more favourable condition for fibrinolysis than for blood coagulation in healthy young subjects. Data are given as mean (SD).     

Understanding the microbiome of diabetic foot osteomyelitis: insights from molecular and microscopic approaches

Objectives

Rigorous visual evidence on whether or not biofilms are involved in diabetic foot osteomyelitis (DFO) is lacking. We employed a suite of molecular and microscopic approaches to investigate the microbiome, and phenotypic state of microorganisms involved in DFO.

Sarcomeric determinants of striated muscle relaxation kinetics

Ca²⁺ is the primary regulator of force generation by cross-bridges in striated muscle activation and relaxation. Relaxation is as necessary as contraction and, while the kinetics of Ca²⁺-induced force development have been investigated extensively, those of force relaxation have been both studied and understood less well. Knowledge of the molecular mechanisms underlying relaxation kinetics is of special importance for understanding diastolic function and dysfunction of the heart. A number of experimental models, from whole muscle organs and intact muscle fibres down to single myofibrils, have been used to explore the cascade of kinetic events leading to mechanical relaxation. By using isolated myofibrils and fast solution switching techniques we can distinguish the sarcomeric mechanisms of relaxation from those of myoplasmic Ca²⁺ removal. There is strong evidence that cross-bridge mechanics and kinetics are major determinants of the time course of striated muscle relaxation whilst thin filament inactivation kinetics and cooperative activation of thin filament by cycling, force-generating cross-bridges do not significantly limit the relaxation rate. Results in myofibrils can be explained well by a simple two-state model of the cross-bridge cycle in which the apparent rate of the force generating transition is modulated by fast, Ca²⁺-dependent equilibration between off- and on-states of actin. Inter-sarcomere dynamics during the final rapid phase of full force relaxation are responsible for deviations from this simple model.     

Gene conversion-like sequence transfers in a mouse antibody transgene: antigen selection allows sensitive detection of V region interactions based on homology.

Gene conversion is important for antibody diversification in chickens, rabbits and cows. In mice, however, conversion events appear to be infrequent among endogenous antibody genes. DNA sequence transfer events that resemble gene conversions have been reported for a mouse H chain transgene (VVC(mu)) that contains two closely spaced homologous VDJ segments. Surprisingly, these reported VVC(mu) sequence transfers were found frequently among mouse B cells responding to immunization. Transgene sequence transfers could be occurring at high frequency in responding VVC(mu) B cells or could be occurring at lower frequency with subsequent amplification by preferential antigen selection. To distinguish these possibilities, we have analyzed a second transgene (InVVC(mu)) that is identical to VVC(mu) except that the two VDJ regions have been exchanged in position. We find that transgene sequence transfers are much less frequent among responding B cells in InVVC(mu) mice, demonstrating the importance of selection in the frequent transgene conversions observed in VVC(mu) mice. These results suggest that mice, like other species, can use gene conversion to diversify antibodies. Such diversification events are apparently infrequent, however, and might only be detected among endogenous Ig genes with a favorable arrangement of V genes and an antigenic stimulation that selects cells with conversions. For both VVC(mu) and InVVC(mu) mice, conversion-like sequence transfers are strongly correlated with somatic hypermutation. Based on these results, we hypothesize that, in mice, gene conversions represent infrequent alternative reactions of a homology-based DNA repair process that is central in the somatic hypermutational mechanism.     

A B220(-), CD19(-) population of B cells in the peripheral blood of quasimonoclonal mice

We describe a new population of non-naive B cells in the peripheral blood of quasimonoclonal (QM) mice. Surface Ig of switched isotypes is expressed, but not B220 nor CD19. These cells are larger and denser than naive B cells but smaller than blasts or plasma cells; they do not stain with syndecan, a marker for plasma cells. Telomerase, which is usually expressed in B cell blasts, was not present in this population. We sorted the switched, idiotype-positive, B220(-) B cells from the peripheral blood of QM mice and sequenced Ig H chain and lambda L chain cDNA. There were many point mutations but no V gene replacements, gene conversions or other type of diversifications. As they express switched isotypes and have mutated their Ig genes, cells in the B220(-), CD19(-) population must have been in an immune response and we suggest that it includes the memory B cell subset.