Copolymers including L-histidine and hydrophobic moiety for preparation of nonbiofouling surface

A new type of copolymer composed of l-histidine (ampholyte) and n-butyl methacrylate (hydrophobic moiety) was developed for the preparation of nonbiofouling surfaces. The copolymer adsorbed onto resin surfaces and made the surface very hydrophilic. The hydrophilization effect was higher than that of bovine serum albumin (BSA). When polystyrene surfaces were coated with the copolymer, both the nonspecific adsorption of protein and the adhesion of cells were significantly reduced in comparison with BSA coating. The newly synthesized polymer is a new and useful candidate for the preparation of nonbiofouling surfaces.     

Association of protein Z and factor VII gene polymorphisms with risk of cerebral hemorrhage: a case–control and a family-based association study in a Chinese Han population

Protein Z (PZ) and factor (F) VII are two important factors in the clotting pathway which have similar structure, linked function and nearby gene sites. The aims of this study were to investigate whether the common variants of PZ and FVII genes are associated with the risk of cerebral hemorrhage (CH) and to explore the combined effects of PZ and FVII polymorphisms for CH risk. We performed genotyping analysis for two single-nucleotide polymorphisms (SNPs) of FVII (rs510317 and rs6046) and three SNPs of PZ (rs2273971, rs3024718 and rs3024731) both in a population-based case–control study and in a family-based association study. Case–control analysis found no evidence of significant association. But family-based association study revealed that the G allele of PZ rs2273971, and three haplotypes carrying the ‘G’ allele of PZ rs2273971: haplotype GA, CG and CGA of PZ and FVII genes, all had a significant effect on CH susceptibility (Z = 1.882, P = 0.049; Z = 1.922, P = 0.044; Z = 1.826, P = 0.047; Z = 1.977, P = 0.048, respectively). While, the A allele of PZ rs2273971, and four haplotypes carrying or crossing the ‘A’ allele of PZ rs2273971: haplotypes CA, ACAA, ACAT and ACAAT of PZ and FVII genes, may confer protection against CH (Z =?1.882, P = 0.049; Z =?2.000, P = 0.045; Z =?2.319, P = 0.020; Z =?2.002, P = 0.045; Z =?2.015, P = 0.043, respectively). This is a first family-based association study providing genetic evidences that PZ and FVII genes, especially PZ rs2273971 are involved in the development of CH in Han-Chinese families.     

Cell Sorting and Chondrogenic Aggregate Formation in Limb Bud Recombinants and in Culture

The cartilage pattern of the developing chick limb changes along the proximal-distal (PD) axis. It is assumed that these spatial changes are brought about by differences in the cellular properties of distal mesoderm, the progress zone (PZ). To examine whether these differences are actually maintained in the individual cells composing the PZ, we dissociated early (stage 20) and late (stage 25) PZ tissues into single cells, then mixed and recombined them with ectodermal jackets. The recombinants were grafted to limb bud stumps and allowed to develop into limb-like structures. Early PZ cells were distributed within whole cartilage elements along the PD axis of the limb-like structures, while cells from late PZ participated only in the formation of distal cartilage elements.
A difference in distribution pattern between the cells of early and late PZ in mixed culture was also observed. Cells of early PZ aggregated rapidly in patches and formed cartilage nodules, while the cells of late PZ distributed in regions surrounding these cell aggregates and gradually differentiated to cartilage cells. These results suggest that the cellular properties in the PZ concerning the rate of chondrogenic aggregate formation change during limb bud development, and that this change may relate to the cartilage pattern formation along the PD axis.     

Characterization of the protein Z-dependent protease inhibitor interactive-sites of protein Z

Background

Protein Z (PZ) has been reported to promote the inactivation of factor Xa (FXa) by PZ-dependent protease inhibitor (ZPI) by about three orders of magnitude. Previously, we prepared a chimeric PZ in which its C-terminal pseudo-catalytic domain was grafted on FX light-chain (Gla and EGF-like domains) (PZ/FX-LC). Characterization of PZ/FX-LC revealed that the ZPI interactive-site is primarily located within PZ pseudo-catalytic domain. Nevertheless, the cofactor function and apparent K_d of PZ/FX-LC for interaction with ZPI remained impaired ~ 6–7-fold, suggesting that PZ contains a ZPI interactive-site outside pseudo-catalytic domain. X-ray structural data indicates that Tyr-240 of ZPI interacts with EGF2-domain of PZ. Structural data further suggests that 3 other ZPI surface loops make salt-bridge interactions with PZ pseudo-catalytic domain. To identify ZPI interactive-sites on PZ, we grafted the N-terminal EGF2 subdomain of PZ onto PZ/FX-LC chimera (PZ-EGF2/FX-LC) and also generated two compensatory charge reversal mutants of PZ pseudo-catalytic domain (Glu-244 and Arg-212) and ZPI surface loops (Lys-239 and Asp-293).

Methods

PZ chimeras were expressed in mammalian cells and ZPI derivatives were expressed in Escherichia coli.

Results

The PZ EGF2 subdomain fusion restored the defective cofactor function of PZ/FX-LC. The activities of PZ and ZPI mutants were all impaired if assayed individually, but partially restored if the compensatory charge reversal mutants were used in the assay.

Conclusions

PZ EGF2 subdomain constitutes an interactive-site for ZPI. Data with compensatory charge reversal mutants validates structural data that the identified residues are part of interactive-sites.

General significance

Insight is provided into mechanisms through which specificity of ZPI–PZ–FXa complex formation is determined.     

Evaluation of protein Z plasma level in beta-thalassemia major patients in Ahvaz city in Iran

Objectives

Thrombotic episodes occurred frequently in beta-thalassemia major (BTM) patients, leading to hypercoagulability of plasma. Protein Z (PZ) is a vitamin-K-dependent anti-coagulation factor that plays a role in the human homeostatic process. The objective of the current study is to investigate the distribution pattern of PZ plasma concentrations between BTM patients and the normal population in Ahvaz city, the center of Khuzestan province, southwest of Iran.

Material and Methods

Forty confirmed BTM patients and 40 healthy volunteers were evaluated for complete blood count (CBC) indices and PZ plasma levels. CBC samples were measured using an automated cell counter, and PZ was assayed with an immunoassay method. Statistical analysis was conducted using SPSS software. The ROC curve and binary logistic regression estimated the sensitivity, specificity, and Odd’s ratio for PZ measurement.

Results

The mean±SD of the PZ plasma level in normal individuals was 1.68±0.63 μg/mL, and in BTM patients, it was 1.10±0.52 μg/mL. This shows a significant reduction of PZ in BTM patients statistically (CI = 0.99; p<0.001). Further, the mean±SD of the PZ plasma levels in BTM patients who received washed red blood cells was not significantly different from that of patients undergoing packed red blood cell therapy (CI = 0.95; p = 0.320). The area under the curve (AUC) for PZ was 0.759 (p = 0.00). The cut-off value = 1.4 μg/mL of the PZ plasma level had at least 70% sensitivity and specificity in BTM patients.

Discussion

Several epidemiologic studies have shown thromboembolism episodes in BTM patients. In the current study, PZ was reduced significantly in BTMs.

Conclusion

We noticed that BTMs have lower plasma PZ concentration might be predisposed to BTM.

     

Protein Z Exerts Pro-Angiogenic Effects and Upregulates CXCR4

Objective

Protein Z (PZ) is a vitamin K-dependent coagulation factor without catalytic activity. Evidence points towards PZ as an independent risk factor for the occurrence of human peripheral arterial disease. However, the role of PZ in ischemia-driven angiogenesis and vascular healing processes has not been elucidated so far.

Approach

Angiogenic potency of PZ was assessed in established in vitro assays using endothelial cells. PZ-deficient (PZ^−/−) mice and their wild-type littermates (PZ^+/+) were subjected to hindlimb ischemia. Furthermore, PZ^−/− mice were exposed to PZ expressing adenovirus (AdV-PZ) or control adenovirus (AdV-GFP). In an additional set of animals, PZ^−/− mice were exposed to AdV-PZ and AdV-GFP, each in combination with the CXCR4 antagonist AMD3100.

Results

In vitro, PZ stimulated migratory activity and capillary-like tube formation of endothelial cells comparable to SDF-1. PZ^−/− mice exhibited diminished hypoxia-driven neovascularization and reperfusion in post-ischemic hindlimbs, which was restored by adenoviral gene transfer up to levels seen in PZ^+/+ mice. The stimulatory impact of PZ on endothelial cells in vitro was abolished by siRNA targeting against PZ and PZ was not able to restore reduced migration after knock-down of CXCR4. The increased surface expression of CXCR4 on PZ-stimulated endothelial cells and the abrogated restoration of PZ^−/− mice via AdV-PZ after concomitant treatment with the CXCR4 antagonist AMD3100 supports the idea that PZ mediates angiogenesis via a G-protein coupled pathway and involves the SDF-1/CXCR4 axis. This is underlined by the fact that addition of the G-protein inhibitor PTX to PZ-stimulated endothelial cells abolished the effect of PZ on capillary-like tube formation.

Conclusions

The results of the current study reveal a role of PZ in ischemia-induced angiogenesis, which involves a G-protein coupled pathway and a raised surface expression of CXCR4. Our findings thereby extend the involvement of PZ from the coagulation cascade to a beneficial modulation of vascular homeostasis.     

Infrared spectroscopy with multivariate analysis potentially facilitates the segregation of different types of prostate cell

The prostate gland is conventionally divided into zones or regions. This morphology is of clinical significance as prostate cancer (CaP) occurs mainly in the peripheral zone (PZ). We obtained tissue sets consisting of paraffin-embedded blocks of cancer-free transition zone (TZ) and PZ and adjacent CaP from patients (n = 6) who had undergone radical retropubic prostatectomy; a seventh tissue set of snap-frozen PZ and TZ was obtained from a CaP-free gland removed after radical cystoprostatectomy. Paraffin-embedded tissue slices were sectioned (10-mum thick) and mounted on suitable windows to facilitate infrared (IR) spectra acquisition before being dewaxed and air dried; cryosections were dessicated on BaF(2) windows. Spectra were collected employing synchrotron Fourier-transform infrared (FTIR) microspectroscopy in transmission mode or attenuated total reflection-FTIR (ATR) spectroscopy. Epithelial cell and stromal IR spectra were subjected to principal component analysis to determine whether wavenumber-absorbance relationships expressed as single points in "hyperspace" might on the basis of multivariate distance reveal biophysical differences between cells in situ in different tissue regions. After spectroscopic analysis, plotted clusters and their loadings curves highlighted marked variation in the spectral region containing DNA/RNA bands ( approximately 1490-1000 cm(-1)). By interrogating the intrinsic dimensionality of IR spectra in this small cohort sample, we found that TZ epithelial cells appeared to align more closely with those of CaP while exhibiting marked structural differences compared to PZ epithelium. IR spectra of PZ stroma also suggested that these cells are structurally more different to CaP than those located in the TZ. Because the PZ exhibits a higher occurrence of CaP, other factors (e.g., hormone exposure) may modulate the growth kinetics of initiated epithelial cells in this region. The results of this pilot study surprisingly indicate that TZ epithelial cells are more likely to exhibit what may be a susceptibility-to-adenocarcinoma spectral signature. Thus, IR spectroscopy on its own may not be sufficient to identify premalignant prostate epithelial cells most likely to progress to CaP.     

Synthesis and in vitro muscarinic activities of a series of 1,3-diazacycloalkyl carboxaldehyde oxime derivatives

A series of 1,3-diazacycloalkyl carboxaldehyde oxime derivatives was synthesized and tested for muscarinic activity in receptor binding assays using [3H]-oxotremorine-M (OXO-M) and [3H]-pirenzepine (PZ) as ligands. Potential muscarinic agonistic or antagonistic properties of the compounds were determined using binding studies measuring their potencies to inhibit the binding of OXO-M and PZ. Preferential inhibition of OXO-M binding was used as an indicator for potential muscarinic agonistic properties; this potential was confirmed in functional studies on isolated organs.     

On Choosing Among Several Experimental Treatments and a Control

The two classical selection approaches in comparing experimental treatments with a control are combined to form an integrated approach. In this integrated approach, there is a preference zone (PZ) and an indifference zone (IZ), and the concept of a correct decision (CD) is defined differently in each of these zones. In the PZ, we are required to select the best treatment for a correct decision (CD₁) but in the IZ, we define any selected subset to be correct (CD₂) if it contains the best treatment among all the experimental treatments and the controlled treatment. We propose a single-stage procedure R to achieve the selection goals CD₁ and CD₂ simultaneously with certain probability requirements. It is shown that both the probability of a correct decision under PZ, P(CD₁ | PZ), and the probability of a correct decision under IZ, P(CD₂ | IZ), satisfy some monotonicity properties and the least favorable configuration in PZ and the worst configuration in IZ are derived by these properties. We also derive formulas for the probabilities of correct decision and provide a brief table to illustrate the magnitude of the procedure parameters and the common sample sizes needed for various probability requirements and configurations.     

Effector modeling of the action of ligands of the GABA receptor complex: functional interactions of muscimol and exogenous modulators of the complex]

In comparison with the literature data the pharmacological effects of muscimol (MS) on mice organism were studied under the condition of administration of the exogenic ligands of GABA-receptor complex GABA antagonist bicuculline (BC), chloride ionophor blocking agent pentylenetetrazole (PZ), phenazepam (BD), sodium barbital (BB) and GABA synthesis antagonist--thiosemicarbazide (TS). Antagonism of MS to the anticonvulsant effect of BB and BD at the administration of BC to the animal was observed, while no effect was observed at the administration of PZ. However at the administration of TS to the experimental animals MS exhibits anticonvulsant effect. The observed properties (partial agonism/partial reverse agonism) of MS at its effect to the whole system are probably determined by the level of endogenic GABA and its changes due to the administration of TS and functional state of GABA--receptor system modified with the exogenic ligand BC.     

Enhancer-trap detection of expression patterns corresponding to the polar coordinate system in the imaginal discs of Drosophila melanogaster

We have isolated three classes of enhancertrap lines of Drosophila in which lacZ expression patterns in the imaginal discs are consistent with the idea of a polar (radial and angular) coordinate system of positional information. In the first class (HZ76), a circular pattern was expressed transiently during the mid-third instar larval stage when the radial components of the coordinate are probably generated. The expression patterns of the second class (HZ84) were sector-shaped and circular in the leg disc, suggesting a correlation with both radial and angular coordinate values. The expression patterns found in the third class (PZ63 and PZ22) were circular and appeared to reflect radial positional values. Expression in the latter two classes always began in the presumptive dorsal region of the leg disc and gradually spread to the ventral region. These developmental profiles of expression suggested the existence of a centre that initiates patterned gene expression in the presumptive dorsal region of the leg disc. The PZ22 line showed transient expression during tarsal segmentation, suggesting its involvement in tarsal segment formation. We have cloned the PZ22 gene and partially determined its sequence. The deduced amino acid sequence contained a zinc finger motif found in DNA/RNA binding proteins. By in situ hybridization, we determined that the PZ22 gene was transcribed in the leg disc in a pattern identical to that of the lacZ expression. In addition, it was expressed transiently in the embryonic mesoderm during mesoderm segmentation. The PZ22 gene, therefore, may function both in mesodermal segmentation in the embryo and in tarsal segmentation in the imaginal disc.     

In vivo optical imaging of the viable epidermis around the nailfold capillaries for the assessment of heart failure severity in humans

Heart failure (HF) is among the socially significant diseases, involving over 2% of the adult population in the developed countries. Diagnostics of the HF severity remains complicated due to the absence of specific symptoms and objective criteria. Here, we present an indicator of the HF severity based on the imaging of tissue parameters around the nailfold capillaries. High resolution nailfold video capillaroscopy was performed to determine the perivascular zone (PZ) size around nailfold capillaries, and 2‐photon tomography with fluorescence lifetime imaging was used to investigate PZ composition. We found that the size of PZ around the nailfold capillaries strongly correlates with HF severity. Further investigations using 2‐photon tomography demonstrated that PZ corresponds to the border of viable epidermis and it was suggested that the PZ size variations were due to the different amounts of interstitial fluid that potentially further translates in clinically significant oedema. The obtained results allow for the development of a quantitative indicator of oedematous syndrome, which can be used in various applications to monitor the dynamics of interstitial fluid retention. We therefore suggest PZ size measured with nailfold video capillaroscopy as a novel quantitative sensitive non‐invasive marker of HF severity.      

Pentazocine-induced biphasic analgesia in mice.

Pentazocine (PZ) is well known to act as an opioid mixed agonist-antagonist analgesic. In the present study, we selected the mouse warm plate test condition of 51 +/- 0.5 degrees C instead of 55 +/- 0.5 degrees C to determine the analgesic action of PZ. As a result, i.c.v. PZ produced a biphasic antinociceptive response, while U-50,488H (U-50) and morphine (MRP) showed a monophasic response. Pretreatment with i.c.v. beta-FNA (mu antagonist) antagonized the initial response, whereas the delayed one was antagonized by pretreatment with nor-BNI (kappa antagonist). In addition, pretreatment with NTI (delta antagonist) significantly attenuated the initial response but not the delayed one. These results suggest that the initial and delayed responses may be mediated mainly by mu/delta and kappa receptors, respectively. With regards to the interaction between MRP and PZ, a low dose of PZ antagonized the analgesic action of MRP, while a high dose PZ plus MRP showed the additive effect. Furthermore, tolerance developed almost equally to both initial and delayed responses, indicating that tolerance to the kappa component of PZ may be developed as well as the mu component of action of PZ.     

Synthesis and in vitro muscarinic activities of a series of 3-(pyrazol-3-yl)-1-azabicyclo[2.2.2]octanes

A series of 3-(pyrazol-3-yl)-1-azabicyclo[2.2.2]octane derivatives C (Fig. 1) was synthesized and tested for muscarinic activity in receptor binding assays using [3H]-oxotremorine-M (OXO-M) and [3H]-pirenzepine (PZ) as ligands. Potential muscarinic agonistic or antagonistic properties of the compounds were determined using binding studies measuring their potencies to inhibit the binding of OXO-M and PZ. Preferential inhibition of OXO-M binding was used as an indicator for potential muscarinic agonistic properties; this potential was confirmed in functional studies on isolated organs.     

Protein Z-dependent protease inhibitor binds to the C-terminal domain of protein Z

Protein Z (PZ) is a multidomain vitamin K-dependent plasma protein that functions as a cofactor to promote the inactivation of factor Xa (fXa) by PZ-dependent protease inhibitor (ZPI) by three orders of magnitude. To understand the mechanism by which PZ improves the reactivity of fXa with ZPI, we expressed wild-type PZ, PZ lacking the gamma-carboxyglutamic acid domain (GD-PZ), and a chimeric PZ mutant in which both Gla and EGF-like domains of the molecule were substituted with identical domains of fXa. The ZPI binding and the cofactor function of the PZ derivatives were characterized in both binding and kinetic assays. The binding assay indicated that all PZ derivatives interact with ZPI with a similar dissociation constant (K(D)) of approximately 7 nm. However, the apparent K(D) for the chimeric PZ-mediated ZPI inhibition of fXa was elevated 6-fold on PC/PS vesicles and its capacity to function as a cofactor to accelerate the ZPI inhibition of fXa was also decreased 6-fold. The cofactor activity of GD-PZ was dramatically impaired; however, the deletion mutant exhibited a normal cofactor function in solution. A chimeric activated protein C mutant containing the Gla domain of fXa was susceptible to inhibition by ZPI in the presence of PZ. These results suggest that: (i) the ZPI interactive site of PZ is located within the C-terminal domain of the cofactor and (ii) a specific interaction between the Gla domains of PZ and fXa contributes approximately 6-fold to the acceleration of the ZPI inhibition of fXa on phospholipid membranes.     

贝那普利和氨氯地平对自发性高血压大鼠促胰液素、生长抑素表达的影响

目的：探讨贝那普利和氨氯地平对自发性高血压大鼠（SHR）促胰液素（PZ）、生长抑素（SS）表达的影响。方法：14周龄雄性SHR 45只随机分为3组（n=15）：SHR组、贝那普利组（盐酸贝那普利灌服0.90 mg·kg^-1·d^-1）、氨氯地平组（苯磺酸氨氯地平灌服0.45 mg·kg^-1·d^-1）。以Wistar-Kyoto大鼠（WKY）作为正常对照组（n=15）,SHR组及WKY组每日灌服等体积蒸馏水。干预8周后,采用免疫组化法和RT-PCR法检测十二指肠中PZ、胃窦中SS蛋白和mRNA表达的变化。结果：干预8周后,与WKY组比较,SHR组十二指肠PZ、胃窦SS蛋白和mRNA表达明显升高（P<0.05）;与SHR组比较,贝那普利组与氨氯地平组十二指肠PZ蛋白和mRNA、胃窦SS蛋白和mRNA表达均明显降低（P<0.05）;与贝那普利组比较,氨氯地平组十二指肠PZ蛋白和mRNA、胃窦中SS mRNA表达降低更为明显（P<0.05）。结论：SHR十二指肠中PZ和胃窦中SS调节失衡,贝那普利和氨氯地平的降压作用可能通过调节PZ和SS的表达而达到降压目的,而氨氯地平的调节作用较显著。     

The interchangeability of global positioning system and semiautomated video-based performance data during elite soccer match play

In elite-level soccer, player motion characteristics are commonly generated from match play and training situations using semiautomated video analysis systems and global positioning system (GPS) technology, respectively. Before such data are used collectively to quantify global player load, it is necessary to understand both the level of agreement and direction of bias between the systems so that specific interventions can be made based on the reported results. The aim of this report was to compare data derived from both systems for physical match performances. Six elite-level soccer players were analyzed during a competitive match using semiautomated video analysis (ProZone? [PZ]) and GPS (MinimaxX) simultaneously. Total distances (TDs), high speed running (HSR), very high speed running (VHSR), sprinting distance (SPR), and high-intensity running distance (HIR; >4.0 m·s(-1)) were reported in 15-minute match periods. The GPS reported higher values than PZ did for TD (GPS: 1,755.4 ± 245.4 m; PZ: 1,631.3 ± 239.5 m; p < 0.05); PZ reported higher values for SPR and HIR than GPS did (SPR: PZ, 34.1 ± 24.0 m; GPS: 20.3 ± 15.8 m; HIR: PZ, 368.1 ± 129.8 m; GPS: 317.0 ± 92.5 m; p < 0.05). Caution should be exercised when using match-load (PZ) and training-load (GPS) data interchangeably.