厌氧发酵污泥中磷酸(酯)酶活性

在消化器超负荷进料即将出现消化障碍时,污泥中磷酸酶活性,特别是酸性磷酸酶活性明显增高,它比其它检测指标(如挥发性脂肪酸的积累、pH值的下降等)约提前出现10天,可用它作为厌氧消化障碍的监测指标,也可作为厌氧消化负荷大小的度量。

5-脂氧合酶激活蛋白及磷酸二酯酶4D基因多态性与脑梗死易患性的分子流行病学研究

目的:分析5-脂氧合酶激活蛋白(ALOX5AP)及磷酸二酯酶4D(PDE4D)基因多态性与脑梗死(CI)易患性的分子流行病学特征。方法:收集CI患者396例作为病例组,另选取300名健康体检者作为对照组,运用PCR检测技术和基质辅助激光解析分析2组受检者ALOX5AP基因中SG13S89、SG13S114和PDE4D基因中SNP83的多态性,分析ALOX5AP及PDE4D基因多态性与CI的关系。结果:病例组SG13S89中AG基因型、A等位基因频率均高于对照组(P<0.05)。多元logistic逐步回归分析显示,调整年龄、高血压及糖尿病等因素的影响后,SG13S89中AG基因型是增加CI发生的危险性因素(P<0.05);AT型及SAD型CI患者SG13S89基因中的基因型及基因频率差异均无统计学意义(P>0.05);年龄分层后分析发现,女性是增加SG13S89基因中的AG基因型发生脑卒中的危险因素(P<0.05)。结论:SG13S89中的AG基因型和A等位基因使得CI的频率高于其他等位基因,可能的机制是通过介导的血管炎性反应实现,而未发现SG13S114及SNP83基因的多态性与CI有关。

厌氧消化活性污泥中磷酸(酯)酶的活力分析方法研究

本文叙述了厌氧发酵污泥中微生物磷酸(酯)酶活性分析条件的研究,其中包括酶的提取,酶反应的最佳 pH,最适温度,底物的最佳浓度,底物的饱和常数和酶浓度对酶反应速率的影响等。最后提出了一套检测厌氧污泥中磷酸(酯)酶活性的详细分析程序,并以此作为预测厌氧发酵障碍的一项生化指标,实验得到了良好的效果。

6 Kinds of Isozymes after Long-term Subculture of Emmenopterys henryi Oliv.

[Objective] The aim was to study the changes of zymography in 6 kinds of isozymes after long-term subculture of Emmenopterys henryi Oliv.. [Method] Non-denaturing polyacrylamide gel electrophoresis was used to analyze isozyme patterns such as esterase (EST),acid phosphatase (ACP),ATP enzyme (ATPase),amylase (AMY),superoxide dismutase (SOD) and peroxidase (POD) in long-term subculture callus of Emmenopterys henryi Oliv. [Result] The research showed that there were differences among the 6 kinds of isozymes in embryogenic callus and non-embryogenic callus of Emmenopterys henryi Oliv.,and both levels could be taken as the basis for identification,the EST,ACP,and POD of non-embryogenic callus were significantly higher than embryogenic callus. The browning of non-embryogenic callus was non-level in the AMY,SOD and POD isozymes when was compared with normal non-embryogenic callus,while the EST,ACP and ATPase isozymes decreased; When the browning of embryogenic callus was contrasted with normal embryogenic callus,EST isozyme increased and the other 5 kinds of enzymes decreased. [Conclusion] The study provided theoretical basis for research morphological difference and browning of long-term tissue culture of Emmenopterys henryi Oliv..

Dynamic Effects of PAEs on Soil Urease and Phosphatase

[Object] The study aimed to supply a reference for evaluating ecotoxicology of soil contaminated with phthalate acid esters(PAEs).[Method] The dynamic effects of DBP and DEHP on activities and kinetics parameters of urease and phosphatase in agro-soil contaminated artificially with DBP and DEHP were studied.[Result] The activities of urease and phosphatase were both inhibited significantly by higher contents of DBP and DEHP in soils compared with CK.The inhabitations increased with increasing DBP and DEHP c...

Application of PDE3 and Brilliant Cresyl Blue in In-vitro Culture of Sheep Oocyte

[Objective] The aim of this study was to increase the viability of sheep oocytes in vitro by using phosphodiesterase type 3（PDE 3） inhibitor milrinone combined with brilliant cresyl blue（BCB） staining.[Method] The differences between BCB tested and morphologically selected oocytes,as well as the effect of them on embryo development were compared;and then suitable inhibitive time of milrinone to sheep oocytes in vitro was studied and used in BCB-oocytes for in vitro embryo production（IVEP）.[Result] The BCB＋ oocytes percentage in A-and B-level sheep oocytes was 64.42%,which was extremely significantly higher than that in C-level（17.0%）.The maturing rate,cleavage rate and blastocyst rate of BCB＋ oocytes（86.16%,85.29% and 34.40%） of was significantly higher than those of BCB-oocytes（50.94%,36.19% and 6.73%）.The best time for PDE 3 inhibitor delaying the sheep oocyte mature in vitro was 6 h.In addition,the rate of embryo development in vitro could be significantly increased by inhibiting the BCB-oocytes for 6 h with Milrinone.[Conclusion] The study will provide reference for improving the efficiency of sheep oocytes culture in vitro.

Serum arylesterase and paraoxonase activity in patients with chronic hepatitis

AIM： To investigate the relationship between serum paraoxonase （PON1）, AST, ALT, GGT, and arylesterase （AE） activity alterations and the degree of liver damage in patients with chronic hepatitis. METHODS： We studied 34 chronic hepatitis patients and 32 control subjects, aged between 35 and 65 years, in the Department of Infection and Clinical Microbiology at the Firat University School of Medicine. Blood samples were collected from subjects between 8：00 and 10：00 a.m. following a 12-h fast. Baseline and salt-stimulated PON1 activities were measured by the hydrolysis of paraoxon. Phenyl acetate was used as the substrate and formed phenol was measured spectrophotometrically at 270 nm after the addition of a 10-fold diluted serum sample in AE activity measurements. RESULTS： The results of this investigation revealed that the levels of AE activity decreased from 132±52 to 94± 36 （29%）, baseline PON1 activity from 452±112 to 164 ±67 （64%）, salt-stimulated PON1 activity from 746± 394 to 294±220 （61%）, HDL from 58.4±5.1 to 47.2±5.6 （200）, triglyceride from 133±51.2 to 86±34.0 （350）, while a slight increase in the level of LDL （from 163± 54.1 to 177.3±56.0; 9%） and significant increases in the levels of AST （from 29±9.3 to 98±44）, ALP （from 57.2±13.1 to 91±38.1）, ALT （from 27.9±3.32 to 89± 19.1）, GGT （from 24.3±2.10 to 94±48.2）, total bilirubin （from 0.74±0.02 to 1.36±0.06; 84%） and direct bilirubin （from 0.18±0.01 to 0.42±0.04; 133%） were detected. However, the levels of albumin, total protein, cholesterol, and uric acid were almost the same in chronic hepatitis and the control subjects.CONCLUSION： Low PON1 and AE activity may contribute to the increased liver dysfunction in chronic hepatitis patients by reducing the ability of HDL to retard LDL oxidation and might be clinically useful for monitoring the disease of chronic hepatitis.

Leptin transiently antagonizes ghrelin and long-lastingly orexin in regulation of Ca^(2+) signaling in neuropeptide Y neurons of the arcuate nucleus

AIM： To explore the mechanism for interactions of leptin with ghrelin and orexin in the arcuate nucleus （ARC） activating neuropeptide Y （NPY） neurons during physiological regulation of feeding, METHODS： Single neurons from ARC of adult rats with matured feeding function were isolated. [Ca2＋]i was measured to monitore their activities. The time course of leptin effects on ghrelin-induced versus orexin-induced [Ca2＋]i increases in NPY neurons was studied. RESULTS： Administration of ghrelin or orexin-A at 101~ mol/L increased cytosolic Ca2~ concentration （[Ca2＋]~） in NPY neurons isolated from the ARC of adult rats. Upon administration of leptin at 10^-14-10^-12 mol/L, ghrelin-induced [Ca2＋]i increases were initially （〈 10 min） inhibited but later restored, exhibiting a transient pattern of inhibition. In contrast, orexin-induced [Ca2＋]i increases were inhibited by leptin in a long- lasting manner. Furthermore, a prior administration of leptin inhibited orexin action but not ghrelin action to increase ICa 2＋li, CONCLUSION： Leptin counteracted ghrelin effects transiently and orexin effects long-lastingly in NPY neurons. The transient property with which leptin counteracts ghrelin action in NPY neurons may allow the fasting-associated increase in ghrelin levels to activate NPY neurons in the presence of physiological leptin and to stimulate feeding.

Phosphodiesterase-3 inhibitor （cilostazol） attenuates oxidative stress-induced mitochondrial dysfunction in the heart

Background Cilostazol is a type 3 phosphodiesterase inhibitor which has been previously demonstrated to prevent the occurrence of tachyarrhythmia and improve defibrillation efficacy. However, the mechanism for this beneficial effect is still unclear. Since cardiac mito-chondria have been shown to play a crucial role in fatal cardiac arrhythmias and that oxidative stress is one of the main contributors to arr-hythmia generation, we tested the effects of cilostazol on cardiac mitochondria under severe oxidative stress. Methods Mitochondria were isolated from rat hearts and treated with H2O2 to induce oxidative stress. Cilostazol, at various concentrations, was used to study its protective effects. Pharmacological interventions, including a mitochondrial permeability transition pore （mPTP） blocker, cyclosporine A （CsA）, and an inner membrane anion channel （IMAC） blocker, 4＇-chlorodiazepam （CDP）, were used to investigate the mechanistic role of cilostazol on cardiac mitochondria. Cardiac mitochondrial reactive oxygen species （ROS） production, mitochondrial membrane potential change and mi-tochondrial swelling were determined as indicators of cardiac mitochondrial function. Results Cilostazol preserved cardiac mitochondrial function when exposed to oxidative stress by preventing mitochondrial depolarization, mitochondrial swelling, and decreasing ROS produc-tion. Conclusions Our findings suggest that cardioprotective effects of cilostazol reported previously could be due to its prevention of car-diac mitochondrial dysfunction caused by severe oxidative stress.

Inhibitory effects of antisense phosphorothioate oligodeoxynucleotides on pancreatic cancer cell Bxpc-3 telomerase activity and cell growth in vitro

AIM： To investigate the effect of telomerase hTERT gene antisense oligonucleotide （hTERT-ASO） on proliferation and telomerase activity of pancreatic cancer cell line Bxpc-3. METHODS： MTT assay was used to detect the effect of different doses of hTERT-ASO on proliferation of Bxpc-3 cell for different times. To study the anti-tumor activity, the cells were divided into there groups： Control group （pancreatic cancer cell Bxpc-3）; antisense oligonucleotide （hTERT-ASO） group; and nosense oligonucleotide group decorated with phosphorothioate. Telomerase activity was detected using TRAP-PCR-ELISA. Cell DNA distribution was examined using flow cytometry assay. Cell apoptosis was observed by transmission electron microscope in each group. RESULTS： After treatment with 6 mmollL hTERT- ASO, cell proliferation was inhibited in dose- and time- dependent manner. The telomerase activity decreased after treatment with hTERT-ASO for 72 h. Flow cytometry showed the cell number of G0/G1 phase increased from 2.7% to 14.7%, the cell number of S phase decreased from 72.7% to 51.0%, and a sub-G1 stage cell apoptosis peak appeared in front of G1 stage. CONCLUSION： Telomerase antisense oligodeoxy- nucleotide can inhibit the proliferation of pancreatic cancer cell line Bxpc-3 and decrease the telomerase activity and increase cell apoptosis rate in vitro.

A New (2+1)-Dimensional Integrable Equation

A new nonlinear partial differential equation （PDE） in 2＋1 dimensions is obtained from the mKP equation by means of an asymptotically exact reduction method based on Fourier expansion and spatio-temporal resealing. In order to demonstrate integrability property of the new equation, the corresponding Lax pair is obtained by applying the reduction technique to the Lax pair of the mKP equation.

An Enzymologic Study on Bone Marrow Cells in Patients with Aplastic Anemia Treated by Supplementing the Kidney and Removing Blood Stasis

The content of cytochrome oxidase (CCO), succinate dehydrogenase (SDH) and neutrophil alkaline phosphatase (NAP) in bone marrow cells in 68 cases of aplastic anemia before and after treatment was determined by computerized graphical analysis and compared with that of normal volunteers (control group). The significantly lowered CCO and SDH levels and the markedly increased NAP content before treatment (P<0.01) became approximately normal after that of supplementing the kidney and removing blood stasis (P >0.05).

Phosphatase of Regenerating Liver and Its Association with Tumors

Protein kinases and protein phosphatases play key roles in regulating functions of diverse proteins which control numerous essential events in eukaryotes, such as transcriptional regulation, apoptosis, cell cycle progression, protein degradation and protein trafficking. Protein kinases transfer a phosphate from ATP to a specific residue（s）, typically at serine, threonine, or tyrosine, in proteins, while phosphatases remove phosphoryl groups from proteins. Such a phosphorylation-dephosphorylation cycle can be regarded as a molecular ＂on-off＂ switch. It is estimated that the human genome contains over 2,000 kinases and 1,000 phosphatases. A very large number of protein kinases has been identified and studied in detail, and more and more, information concerning protein phosphatases has emerged recently.

Isolation and functional characterization of the C-terminus of rice phos-phatidylinositol 4-kinase in vitro

A partial rice (Oryza sativa L.) cDNA clone. OsPMK1c, was isolated through screening of a cDNA library constructed from tillering materials. OsPI4Klc encoded a peptide of 608 amino acids with a calculated molecular mass of 68.4 kDa. The OsPI4Klc peptide shared high homology and possessed the highly conserved domains present in most isolated cloned PI4-kinases, i.e. a lipid kinase unique (LKU) domain and a catalytic (CAT) domain. A region with similarity to pleckstrin homology (PH) domain was present in OsPI4K1c as well. Further comparison with genomic sequences in databases revealed that OsPI4K1c is located at the 3'-end of a putative rice PI 4-kinase coding gene OsPI4K1, and its coding region corresponded to the C-terminal half of OsPI4Kl protein. Twelve exons (49-562 bp in size) and 11 introns (77-974 bp in size) were identified in OsPI4K1c. The recombinant protein expressed in Escherichia coli phosphorylates phosphatidylinositol at the D4 position of the inositol ring. OsPI4K1 transcript levels were detected in a low but constitutive manner in shoot, stem, leaf, spike and root tissues and did not change upon treatment with different hormones, calcium and jasmonic acid (JA). However, treatment with salicylic acid (SA) elevated the mRNA level of the OsPI4K1 gene, which suggested the involvement of OsPI4K1 in wounding responses.

THE REGULATORY EFFECT OF NUCLEOSIDE DIPHOSPHATE KINASE ON G－PROTEIN AND G－PROTEIN MEDIATED PHOSPHOLIPASE C

The effect of nucleoside diphosphate kinase (NDPK) on the activrty of guanine nucleotide regulatory protein (G-protein) mediated phospholipase C(PLC) and on the [35S]GTPTτS binding of G-protein was investigated in this work in order to demonstrate the mechanism behind the regulation of G-protein and its effector PLC by NDPK. The stimulation of PLC in turkey erythrocyte membrane by both GTP and GTPτS indicated that the PLC stimulation was mediated by G-protein. NDPK alone stimulated PLC activity. as well as the stimulation in the presence of GTP and GDP, in a dose-dependent manner. However. NDPK inhibited GTPτS-stimulated PLC. Furthermore, NDPK inhibited [35S]GTPτS binding of purified Gi-protein in a non-competitive manner. A hypothesis implying an important role of direct interaction of G-protein and NDPK in the regulation of their functions is suggested and discussed.

Purification of γ-Subunits Phosphodiesterase （γ-PDE6） Protein

Phosphodiesterase （PDE） exist various forms for the different structure and property, which is one of key components of light receptors in retinal rod cells. In this paper, cloning the DNA of T-subunits Phosphodiesterase （γ-PDE6） from cDNA library, constructing the corresponding recombinant plasmid, and purification of the protein after initial expression was studied.

EFFECTS OF NOVEL PHOSPHODIESTERASE 4 INHIBITORS,ARIFLO AND SB242126A, ON ENDOTHELIN-1-INDUCED CONTRACTILITY OF ISOLATED HUMAN MYOMETRIUM

Objective To investigate the effects of novel selective phosphodiesterase4 (PDE4) inhibitors,Ariflo and SB242126A, on the endothelin-1 (ET-1) - induced contractility occurring in nonpregnant human myome-trium specimens. Methods Contractile responses to Ariflo and SB242126A were recorded cumulatively on isola-ted human longitudinal myometrium specimens obtained through surgical operations. Results Ariflo andSB242126A could inhibit both the frequency and amplitude of spontaneous contractions of myometrium (pD2 =8. 6and 7. 6,n =4) and ET-1 -induced contractions in a concentration-dependent manner (pD2 = 7. 7 and 8. 1 ,n =5) ,with a potency similar to that of Rolipram. Conclusion Ariflo and SB242126A have an obvious inhibitory effecton endothelin-1-induced contractility of isolated human myometrium. The finding suggested that PDE4 inhibitorsmight have clinical potential in treating preterm labour and dysmenorrhoea.

Inhibition of protein tyrosine phosphatase 1B activity by triterpenes isolated from Aceriphyllum rossii

An organic layer prepared from the seed of Aceriphyllum rossii was studied to identify the active compounds for protein tyrosine phosphatase 1B（PTP1B） inhibition.Bioassay guided fractionation resulted in the isolation of PTP1B inhibitory activity of triterpenes（1-4）.These four compounds were identified as aceriphyllic acid C（1）,aceriphyllic acid D（2）,aceriphyllic acid E（3） and aceriphyllic acid F（4）.The isolated 1-4 compounds inhibited PTP1B with IC50 values ranged from（2.1±1.5） μmol/L to（11.2±2.5） μmol/L.Kinetic analysis of PTP1B inhibition by aceriphyllic acid C（1） and aceriphyllic acid D（2） suggested that oleanane-type triterpenes inhibited PTP1B activity in a mixed-type manner.

Identification of novel cyclic nucleotide phosphodiesterase gene cDNAs in the brain of guinea pig

Objective Cyclic nucleotide phosphodiesterase（PDE）is a critical component of the nitric oxide（NO）signaling pathway and plays critical roles in cognition and learning,Parkinson’s disease,attention deficit hyperactivity disorder, psychosis and depression.The PDEs in the brain of guinea pig have not yet been reported.The present study aimed to detect the unknown Pde cDNAs in the brain of guinea pig.Methods Reverse transcription polymerase chain reaction（RT-PCR）and sequence comparison analysis were performed to detect the expression of Pde cDNAs and to assess the identity rates of cDNA and amino acid sequences between guinea pig and human or mouse,respectvely.The RT-PCR primers were located on the conserved region of human PDE and mouse Pde cDNAs.Results Eleven novel Pde cDNAs were detected in the brain of guinea pig（Cavia porcellus）,including CpPde1a,CpPde1b,CpPde2a,CpPde4a,CpPde4d,CpPde5a,CpPde6c,CpPde7b, CpPde8a,CpPde9a,and CpPde10a.The identity rates of the Pde cDNA sequences between guinea pig and human ranged from 83.8%to 94.3%,and those of the amino acid sequences ranged from 91.9%to 100%.The identity rates of Pde cDNA sequences between guinea pig and mouse ranged from 84.6%to 92.1%,and those of amino acid sequences ranged from 91.2% to 99.2%.The average identity rate of the 11 Pde cDNA sequences between guinea pig and human was significantly higher（P 0.01）than that between guinea pig and mouse.The putative partial amino acid sequences of guinea pig contained at least one of the conserved domains of human and mouse PDE proteins.Conclusion These results indicate that the brainexpressed Pde genes are identified in guinea pig,which lays the foundation for further investigating the physiological roles of PDE proteins in the brain.

Inhibition of protein tyrosine phosphatase 1B by triterpenes isolated from Potentilla discolor Bge

Seven oleanene triterpenes were isolated from the roots of Potentilla discolor Bge and their structures were identified as 3-oxoolean-12-en-27-oic acid（1）, gypsogenic acid（2）, 3α-hydroxyolean-12-en-27-oic acid（3）, 3β-hydroxyolean-12-en-27-oic acid（4）, aceriphyllic acid A（5）, aceriphyllic acid A methyl ester（6）, and oleanolic acid（7）. Compounds 1–7 inhibited protein tyrosine phosphatase 1B（PTP1B） activity, with IC50 values ranging from（7.5±0.5） to（22.7±0.5） μmol/L. Among the isolates, compounds 1, 2, 3 and 7 from the Potentilla discolor Bge were found to exhibit selective PTP1 B inhibitory activity.