A Chinese Multi-Specialty Delphi Consensus to Optimize RAASi Usage and Hyperkalaemia Management in Patients with Chronic Kidney Disease and Heart Failure.

Objective Variations are present in common clinical practices regarding best practice in managing hyperkalaemia (HK), there is therefore a need to establish a multi-specialty approach to optimal renin-angiotension-aldosterone system inhibitors (RAASi) usage and HK management in patients with chronic kidney disease (CKD) & heart failure (HF).This study aimed to establish a multi-speciality approach to the optimal use of RAASi and the management of HK in patients with CKD and HF. Methods A steering expert group of cardiology and nephrology experts across China were convened to discuss challenges to HK management through a nominal group technique. The group then created a list of 41 statements for a consensus questionnaire, which was distributed for a further survey in extended panel group of cardiologists and nephrologists across China. Consensus was assessed using a modified Delphi technique, with agreement defined as "strong" (≥75% and <90%) and "very strong" (≥90%). The steering group, data collection, and analysis were aided by an independent facilitator. Results A total of 150 responses from 21 provinces across China were recruited in the survey. Respondents were comprised of an even split (n=75, 50%) between cardiologists and nephrologists. All 41 statements achieved the 75% consensus agreement threshold, of which 27 statements attained very strong consensus (≥90% agreement) and 14 attained strong consensus (agreement between 75% and 90%). Conclusion Based on the agreement levels from respondents, the steering group agreed a set of recommendations intended to improve patient outcomes in the use of RAASi therapy and HK management in China.

Clinical Diagnosis and Treatment of Male Breast Cancer:Analysis of 24 Cases.

Objective To summarize our experiences in the clinical diagnosis and treatment of male breast cancer(MBC).Methods The clinical date of 24 MBC patients treated in our hospital from January 2006 to December 2012 were retrospective analyzed.Results The average age of these 24 patients was(55.7±2.1) years.All the patients received surgical treatment,and the surgical procedures were simple excision of breast lesion in 6 patients,breast resection alone in 5 patients,and modified radical mastectomy in 13 patients(bilateral in 1 case).The pathological diagnoses included invasive ductal carcinoma in 18 cases,papillary carcinoma in 4 cases,mucinous adenocarcinoma in 1 case,and malignant solitary fibrous tumor in 1 case.Twenty patients received chemotherapy,7 received radiotherapy,and 15 received endocrine therapy after operation.The 5-year survival rate was 54.2%.Conclusions The incidence of MBC is low.This malignancy is mainly seen in elderly individuals,with relatively long disease course,poor prognosis,and high risk of metastasis.MBC is mainly treated by surgery,and adjuvant chemotherapy,radiotherapy,and endocrine therapy may be applied,if appropriate,after the operation.

[Effect of Kuanxiong aerosol on coronary heart disease angina patients: a multicenter randomized controlled clinical study].

OBJECTIVE: To evaluate the anginal attack-relieving efficacy and safety of Kuanxiong Aerosol (KA) in patients with coronary heart disease (CHD). METHODS: A total of 780 patients confirmatively diagnosed as CHD angina from November 2011 to December 2012 in 13 medical centers in the mainland area were assigned to 2 groups by blocked randomization, the treatment group (376 cases) and the control group (374 cases). When the angina attacked, patients in the treatment group received sublingual spray three times, 0.6 mL each time, while those in the control group sublingually dissolved Nitroglycerin Tablet (NT), 0.5 mg each tablet. The effective rate of angina relief, efficacy of electrocardiogram (ECG), and the incidence of adverse reactions were observed. RESULTS: The 3 min and 5 min remission rates of angina attack were 53.72% (202/376) and 94.41% (355/376) in the treatment group, and 47.86% (179/374) and 90.64% (339/374) in the control group. The 95% confidence interval (CI) of the difference between the 2 groups of 3 min and 5 min remission rates of angina attacks were [(-1.84%, 12.32%) and (-1.33%, 6.85%) respectively, P > 0.05]. The total improvement rates of ST-T changes in the treatment group and the control group after treatment were 74.07% and 73.13% respectively (P > 0.05). The adverse reaction rate was 9.31 (35/376 cases) in the treatment group and 22.46% (84/374 cases) in the control group (P < 0.01). CONCLUSION: KA was not inferior to NT in relieving anginal attacks and improving ischemic ECG changes, and had obviously less adverse reaction.

Highly selective luminescent sensing of fluoride and organic small-molecule pollutants based on novel lanthanide metal-organic frameworks.

Two novel isostructural lanthanide metal-organic frameworks (Ln-MOFs), [Ln2(BPDC)(BDC)2(H2O)2]n (Ln = Eu (1) and Tb (2)), have been successfully synthesized via a mixed ligand approach using 2,2'-bipyridine-3,3'-dicarboxylic acid (H2BPDC) and 1,4-benzenedicarboxylic acid (H2BDC) under hydrothermal conditions. Structural analysis shows that two lanthanide ions are 4-fold linked by two κ(1)-κ(1)-µ2 carboxylates from BDC(2-) and the other two κ(2)-κ(1)-µ2 carboxylates from BPDC(2-) to form a binuclear core. The binuclear units are further connected by BDC(2-) and BPDC(2-) to build a three-dimensional framework possessing tfz-d topology with the short (Schläfli) vertex symbol {4(3)}2{4(6)·6(18)·8(4)}. Moreover, isostructural doped Ln-MOFs [Eu(2x)Tb2(1-x)(BPDC)(BDC)2(H2O)2]n (x = 0.1 (1a), 0.3 (1b), 0.5 (1c), 0.7 (1d), and 0.9 (1e)) were also successfully synthesized. Thermal gravimetric analyses (TGA) reveal high thermal stability of these Ln-MOFs. Luminescent measurements indicate that the characteristic sharp emission bands of Eu(3+) and Tb(3+) ions are simultaneously observed in 1a-e. Further luminescent studies reveal that 1, 2, and 1a not only display a high-sensitivity sensing function with respect to fluoride but also exhibit significant solvent-dependent luminescent response to small-molecule pollutants, such as formaldehyde, acetonitrile, and acetone.

[Reference value of plasma N-terminal-pro-B-type natriuretic peptide in middle-aged and elderly Chinese].

OBJECTIVE: To establish the normal ranges for plasma N-terminal-pro-B-type natriuretic peptide (NT-proBNP) of middle-aged and elderly ( ≥ 40 years) healthy subjects in China. METHODS: A total of 5133 subjects (2170 men and 2963 women) from the cohort of Shanghai Heart Health Study (SHHS) were included in this study. Plasma NT-proBNP was measured by electrochemiluminescence immunoassay. The reference values (2.5th- 97.5th quartiles) were determined using both empiric and quantile regression methods. RESULTS: Plasma NT-proBNP values were higher in women than in men at all respective age groups (all P < 0.01) , and natural log-transformed NT-proBNP values increased in proportion with age for both genders and there was a lineal correlation between natural log-transformed NT-proBNP values and age (all P < 0.01) . Quantile regression derived normal reference values for NT-proBNP in male were 4.5-86.8 ng/L in the 40-44 years old group, 5.4-108.5 ng/L in the 45-49 years old group, 6.6-135.5 ng/L in the 50-54 years old group, 7.9-169.4 ng/L in the 55-59 years old group, 9.6-211.7 ng/L in the 60-64 years old group, 11.7-264.6 ng/L in the 65-69 years old group, 14.2-330.7 ng/L in the 70-74 years old group, and 18.1-429.2 ng/L in the ≥ 75 years old group. The reference values in female for NT-pro-BNP in respective age group were 8.5-141.8 ng/L, 10.4-166.6 ng/L, 12.8-195.7 ng/L, 15.7-229.9 ng/L, 19.3-270.1 ng/L, 23.7-317.3 ng/L, 29.1-372.8 ng/L, and 35.7-451.9 ng/L. CONCLUSION: This study preliminarily establishes the normal ranges of plasma NT-proBNP in middle-aged and elderly ( ≥ 40 years) Chinese.

Status and change pattern of kidney transplantation: one center research.

OBJECTIVE: To retrospectively investigate the distribution in kidney transplantation for fifteen years in Peking Union Medical College Hospital. METHODS: We conducted a descriptive research counting up the number of patients who received kidney transplantation each year in our hospital during 1995 and 2010. RESULTS: The first kidney transplantation in our hospital occurred in the 1960s. The number of kidney transplantation increased until reaching a maximum of 47 grafts in 2001; since then the number fell. CONCLUSIONS: With the decreased number of kidney transplantation, we have realized the shortage of transplantable organs is very serious. The continuing transplant shortage requires major efforts to expand the donor pool. Donation after cardiac death offers the potential to enlarge the donor pool, but we need to strictly control the criteria for potential donors.

[Association between the urinary microalbumin/creatinine ratio and cardiovascular risk factors among elderly community subjects].

OBJECTIVE: To determine the value of the urinary microalbumin/creatinine ratio (UACR) and the relationship between UACR and traditional cardiovascular risk factors among elderly community subjects. METHODS: A representative population in Shanghai rural district aged more than 65 years who participated in the heart health survey of the key projects in the national science and technology pillar program in the eleventh five-year plan period of China were sampled via a clustered complex sampling method. A midstream collection from the first morning void collected was used to measure the urinary microalbumin, the urinary creatinine and the UACR. Baseline information including traditional cardiovascular risk factors were obtained by standard questionaire to analyze the distribution status of UACR in the population with or without the risk factors. RESULTS: (1) There were 1718 subjects (721 males) of (73.3 ± 5.5) years included in this study. (2) The prevalence of with at least one cardiovascular risk factor was 78.00% in this cohort, the top there risk factors were dyslipidemia (61.06%), hypertension (44.59%) and diabetes (13.80%). (3) The median (the lower quartile-the upper quartile) of the UACR of the population without cardiovascular diseases and risk factors was 13.81 (6.03 - 26.51) µg/mg. The level of UACR was significantly higher in females than that in males [17.12 (7.28 - 33.28) µg/mg vs. 5.49 (2.92 - 9.76) µg/mg, P < 0.01]. (4) The level of UACR in population with hypertension, diabetes or dyslipidemia was 16.27 (6.65 - 42.00) µg/mg, 26.27 (10.92 - 76.65) µg/mg and 16.39 (6.98 - 41.03) µg/mg respectively, all exceeding that of the healthy group (P < 0.05 or P < 0.01). (5) The levels of UACR increased in proportion to the increase of cardiovascular risk factor numbers, the UACR of the population with 0, 1, 2, 3 and 4 cardiovascular risk factors were 13.81 (6.03 - 26.51) µg/mg, 15.76 (6.79 - 36.44) µg/mg, 13.82 (5.68 - 34.43) µg/mg, 16.47 (6.07 - 50.56) µg/mg and 18.63 (11.26 - 83.09) µg/mg, respectively. The population with 4 cluster of cardiovascular risk factors posed the higher level of UACR than that of population with 0 cardiovascular risk factors (P < 0.05). CONCLUSIONS: The three most common risk factors of cardiovascular diseases among the elderly community subjects aged more than 65 years are dyslipidemia, hypertension and diabetes, all of which are related to the elevation of UACR.

Efficacy and safety of Shexiang Baoxin pill (MUSKARDIA) in patients with stable coronary artery disease: a multicenter, double-blind, placebo-controlled phase IV randomized clinical trial.

BACKGROUND: The Shexiang Baoxin Pill (MUSKARDIA) has been used for treating coronary artery disease (CAD) and angina for more than 30 years in China. Nevertheless, methodologically sound trials on the use of MUSKARDIA in CAD patients are scarce. The aim of the study is to determine the effects of MUSKARDIA as an add-on to optimal medical therapy (OMT) in patients with stable CAD. METHODS: A total of 2674 participants with stable CAD from 97 hospitals in China were randomized 1:1 to a MUSKARDIA or placebo group for 24 months. Both groups received OMT according to local tertiary hospital protocols. The primary outcome was the occurrence of a major adverse cardiovascular event (MACE), defined as a composite of cardiovascular death, non-fatal myocardial infarction (MI), or non-fatal stroke. Secondary outcomes included all-cause mortality, non-fatal MI, non-fatal stroke, hospitalization for unstable angina or heart failure, peripheral revascularization, angina stability and angina frequency. RESULTS: In all, 99.7% of the patients were treated with aspirin and 93.0% with statin. After 2 years of treatment, the occurrence of MACEs was reduced by 26.9% in the MUSKARDIA group (MUSKARDIA: 1.9% vs. placebo: 2.6%; odds ratio = 0.80; 95% confidence interval: 0.45-1.07; P  = 0.2869). Angina frequency was significantly reduced in the MUSKARDIA group at 18 months (P = 0.0362). Other secondary endpoints were similar between the two groups. The rates of adverse events were also similar between the two groups (MUSKARDIA: 17.7% vs. placebo: 17.4%, P = 0.8785). CONCLUSIONS: As an add-on to OMT, MUSKARDIA is safe and significantly reduces angina frequency in patients with stable CAD. Moreover, the use of MUSKARDIA is associated with a trend toward reduced MACEs in patients with stable CAD. The results suggest that MUSKARDIA can be used to manage patients with CAD. TRIAL REGISTRATION: chictr.org.cn, No. ChiCTR-TRC-12003513.

Survival Benefit and Genetic Profile of Pemetrexed as Initial Chemotherapy in Selected Chinese Patients With Advanced Lung Adenocarcinoma.

Objective: This study investigated survival in selected Chinese patients with advanced lung adenocarcinoma who received initial chemotherapy with pemetrexed. We also explored the relationship between genetic biomarkers and pemetrexed efficacy. Methods: We retrospectively collected patients (n = 1,047) enrolled in the Chinese Patient Assistance Program from multiple centers who received pemetrexed alone or combined with platinum as initial chemotherapy and continued pemetrexed maintenance therapy for advanced lung adenocarcinoma from November 2014 to June 2017. The outcomes were duration of treatment (DOT) and overall survival (OS). Clinical features were analyzed for their influence on the treatment effect and prognosis. Next-generation sequencing (NGS) was performed to identify genetic biomarkers associated with the efficacy of pemetrexed. Results: The median DOT was 9.1 months (95% CI: 8.5-9.8), and the median OS was 26.2 months (95% CI: 24.2-28.1). OS was positively correlated with DOT (r = 0.403, P < 0.001). Multivariable analysis showed that smoking status and Eastern Cooperative Oncology Group (ECOG) performance status (PS) were independently associated with DOT; smoking status, ECOG PS, targeted therapy, and EGFR/ALK/ROS1 status were independently associated with OS. NGS in 22 patients with available samples showed genes with high mutation rates were: TP53 (54.5%), EGFR (50.0%), MYC (18.2%), and PIK3CA (13.6%). When grouped based on progression-free survival (PFS) reported in the PARAMOUNT study, the DOT > 6.9 months set was associated with PIK3CA, ALK, BRINP3, CDKN2A, CSMD3, EPHA3, KRAS, and RB1 mutations, while ERBB2 mutation was observed only in the DOT ≤ 6.9 months set. Conclusion: This study shows that initial chemotherapy with pemetrexed is an effective regimen for advanced lung adenocarcinoma in selected Chinese patients. There is no specific genetic profile predicting the benefit of pemetrexed found by NGS. Biomarkers predicting the efficacy of pemetrexed need further exploration.

Chemical modification of Ce(IV)/EDTA-based artificial restriction DNA cutter for versatile manipulation of double-stranded DNA.

A monophosphate group was attached to the terminus of pseudo-complementary peptide nucleic acid (pcPNA), and two of thus modified pcPNAs were combined with Ce(IV)/EDTA for site-selective hydrolysis of double-stranded DNA. The site-selective DNA scission was notably accelerated by this chemical modification of pcPNAs. These second-generation artificial restriction DNA cutters (ARCUTs) differentiated the target sequence so strictly that no scission occurred even when only one DNA base-pair was altered to another. By using two of the activated ARCUTs simultaneously, DNA substrate was selectively cut at two predetermined sites, and the desired fragment was clipped and cloned. The DNA scission by ARCUT was also successful even when the target site was methylated by methyltransferase and protected from the corresponding restriction enzyme. Furthermore, potentiality of ARCUT for manipulation of huge DNA has been substantiated by site-selective scission of genomic DNA of Escherichia coli (composed of 4,600,000 bp) at the target site. All these results indicate promising applications of ARCUTs for versatile purposes.

Observational Study of Chinese Medicine Syndrome Distribution in Patients with Acute Myocardial Infarction and Its Impact on Prognosis.

OBJECTIVE: To investigate the distribution of Chinese medicine (CM) syndrome in patients with acute myocardial infarction (AMI) on admission and its impact on prognosis. METHODS: A total of 525 AMI patients were prospectively recruited and classifified into 4 groups based on their clinical characteristics: excess-heat, excess-cold, deficiency-heat and deficiency-cold syndromes. Major adverse cardiovascular events (MACEs) were followed up. RESULTS: The excess syndrome was more common than deficiency syndrome (72.95% vs. 27.05%; P<0.05). Totally 495 (94.29%) of 525 AMI patients were followed up (median 277 days). There were 59 (11.92%) MACEs. After adjusted with confounding factors in Cox regression models, the hazard ratio (95% confifidence interval) of excess-heat, excess-cold, defificiency-heat and defificiency-cold syndrome groups were 1, 1.25 (0.63, 2.49; P<0.05), 2.37 (1.14, 4.94; P<0.05), 3.76 (1.71, 8.28; P<0.05), respectively. CONCLUSIONS: Excess syndrome was more common in AMI patients and had better prognosis, while defificiency-cold syndrome had the poorest prognosis. CM syndrome was of value in predicting long-term outcomes in AMI patients.

[Effects of tongxinluo capsule on cardiac ventricle remodeling after myocardial infarction: a multicentre clinical research].

OBJECTIVE: To investigate the effects of tongxinluo (TXL) capsule on cardiac ventricle remodeling after acute myocardial infarction (AMI). METHODS: Seventy AMI patients were randomly divided into 2 equal groups: conventional therapy group treated with conventional Western therapy and TXL treatment group treated with TXL capsule for 6 weeks in addition to the conventional treatment. Cardiac color ultrasound was conducted before and 6 weeks after the treatment to examine the changes of ventricular structure, mass and function. Radioimmunoassay was used to detect the levels of angiotensin II (Ang II) and endothelin-1 (ET-1). RESULTS: Six weeks later, the left ventricular posterior wall thickness of end-diastolic (LVPWTD), left ventricular end-systolic dimension (LVESD), and left atrium dimension (LAD) of the TXL group were 0.93 cm +/- 0.09 cm, 3.71 cm +/- 0.19 cm, and 3.21 cm +/- 0.29 cm respectively, all lower than those before treatment (0.93 cm +/- 0.09 cm, 3.71 cm +/- 0.19 cm, and 3.21 cm +/- 0.29 cm respectively) and those of the conventional treatment group (0.95 cm +/- 0.08 cm, 3.62 cm +/- 0.46 cm, and 3.82 cm +/- 0.30 cm) (all P < 0.05), the ejection-fraction (EF) improvement rate of the TXL group was 63% +/- 7%, significantly higher than that before treatment (52% +/- 6%) and that of the conventional treatment group (59% +/- 8%, both P < 0.05). CONCLUSION: TXL capsule in addition to conventional therapy has a positive effect on the prognosis of AMI via reversing the ventricular remodeling, and improving cardiac function.

Impact of inhalational anesthetics on postoperative cognitive function: Study protocol of a systematic review and meta-analysis.

BACKGROUND: Conflict findings of the impact of inhalational anesthetics on postoperative cognitive function are reported. No systematic review has been performed to solve the problem. The aim of the study was to assess the effect of different inhalational anesthetics on postoperative cognitive function in a network meta-analysis. METHODS: We will search MEDLINE, EMBASE, the Central Register of Controlled Trials in the Cochrane library, and CINAHL for randomized controlled trials or cohort studies assessing the short-term or long-term cognitive function of elderly patients (over 60 years) receiving major surgeries and inhalational anesthetics (desflurane, isoflurane, sevoflurane, halothane, and nitrous oxide) during surgery. Two reviewers will independently screen study eligibility, extract information from eligible studies, and appraise study quality. The impact of inhalational anesthetics will be assessed through: incidence of postoperative cognitive dysfunction at 1 week, 3 months, 1 year, and over 1 year after surgery; incidence of post-operative delirium; test of postoperative cognitive function. RESULTS: The results of this systematic review and meta-analysis will be published in a peer-reviewed journal. CONCLUSION: To our knowledge, this systematic review will be the first to evaluate existing research on the incidence of postoperative cognitive function after inhalational anesthetics. Our study will assess the effect of different inhalational anesthetics on postoperative cognitive function. ETHICS AND DISSEMINATION: The review will be finished in December 2017, and the result will be published in a peer-reviewed journal or disseminated through conference posters or abstracts. REVIEW REGISTRATION NUMBER: CRD42017056675 (www.crd.york.ac.uk/PROSPERO).

Effect of Kuanxiong Aerosol () on Patients with Angina Pectoris: A Non-inferiority Multi-center Randomized Controlled Trial.

OBJECTIVE: To evaluate the effect and safety of Kuanxiong Aerosol (, KA) on patients with angina pectoris. METHODS: Block randomization was performed to randomly allocate 750 patients into KA (376 cases) and control groups (374 cases). During an angina attack, the KA group received 3 consecutive sublingual sprays of KA (0.6 mL per spray). The control group received 1 sublingual nitroglycerin tablet (NT, 0.5 mg/tablet). Log-rank tests and Kaplan-Meier estimations were used to estimate the angina remission rates at 6 time-points after treatment (1, 2, 3, 4, 5, and >5 min). Logistic regression analysis was performed to observe the factors inflfluencing the rate of effective angina remission, and the remission rates and incidences of adverse reactions were compared for different Canadian Cardiovascular Society (CCS) classes of angina. RESULTS: The 5-min remission rates in the KA and control groups were not signifificantly different (94.41% vs. 90.64%, P>0.05). The angina CCS class signifificantly inflfluenced the rate of remission (95% confidence interval = 0.483-0.740, P<0.01). In the CCS subgroup analysis, the 3-and 5-min remission rates for KA and NT were similar in the CCSII and III subgroups (P>0.05), while they were signifificantly better for KA in the CCSI and II subgroups (P<0.05 or P<0.01). Furthermore, the incidence of adverse reactions was signifificantly lower in the KA group than in the control group for the CCSII and III subgroups (9.29% vs. 26.22%, 10.13% vs. 20.88%, P<0.05 or P<0.01). CONCLUSIONS: KA is not inferior to NT in the remission of angina. Furthermore, in CCSII and III patients, KA is superior to NT, with a lower incidence of adverse reactions. (Registration No. ChiCTRIPR-15007204).

Ribosome-inactivation display system.

We present a novel strategy for the connection of phenotype and genotype in vitro that can be used for the selection of functional proteins. The strategy involves the generation of a stable complex among a ribosome, an messenger RNA and its translated protein, without removal of the termination codon, as a result of the action of the ricin A chain during translation. The technique requires no transfection, no chemical synthesis, no ligation, and no removal of the termination codon. Thus, our novel ribosome-inactivation display system should provide, without loss of the pool population, a reliable, simple, and robust selection system for the in vitro evolution of the properties of proteins in a predictable direction by a combination of randomization and appropriate selection strategies.

Thromboembolic event rate in patients with persistent or paroxysmal atrial fibrillation post circumferential pulmonary vein isolation: a single center experience in China.

BACKGROUND: Pulmonary-vein isolation (PVI) is currently used for the treatment of chronic and paroxysmal atrial fibrillation and a major risk of PVI is thromboembolism. The purpose of this study was to observe embolic event rate in patients with persistent or paroxysmal atrial fibrillation (AF) undergone PVI. METHODS: Circumferential PVI (CPVI) was performed in 64 consecutive patients with persistent AF (42 men, aged (60.0 +/- 9.1) years) and in 84 consecutive patients with paroxysmal AF (53 men, aged (61.4 +/- 9.3) years). Warfarin was administrated in all patients before ablation for at least 3 weeks ((5.2 +/- 2.6) weeks) and continued for at least 3 months post ablation with international normalized ratio (INR) of 2.0 - 3.0. During CPVI, intravenous heparin was given at a dose of 5000 - 8000 U or 75 - 100 U/kg, followed by 1000 U or 12 U/kg per hour. RESULTS: In patients with persistent AF, 1 patient developed embolic event during ablation and 3 patients developed embolic events after ablation. In contrast, no thromboembolic event was observed in patients with paroxysmal AF (4/64 vs 0/84, P = 0. 033). CONCLUSION: Thromboembolic event rate related to CPVI is significantly higher in patients with persistent AF than that in patients with paroxysmal AF.

Straightforward detection of SNPs in double-stranded DNA by using exonuclease III/nuclease S1/PNA system.

Single-nucleotide polymorphisms (SNPs) in double-stranded DNA (dsDNA) have been straightforwardly genotyped by matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry (MALDI-TOF MS). Peptide nucleic acid (PNA), a DNA analog, was used as a probe molecule. In its presence, genomic dsDNA was first treated with exonuclease III and then with nuclease S1. By these one-pot reactions, single-stranded DNA fragments including the SNP sites were formed in situ. These fragments were directly analyzed by MALDI-TOF MS, and the identity of the DNA base at the SNP site was determined in terms of mass number. By using two or more PNA probes simultaneously, multiplex analysis was also successful. Various genotypes of apolipoprotein E gene (epsilon2/epsilon2, epsilon3/epsilon3, epsilon4/epsilon4, epsilon2/epsilon3 and epsilon3/epsilon4) were identified from dsDNA obtained by PCR from corresponding patients.

Existence of efficient divalent metal ion-catalyzed and inefficient divalent metal ion-independent channels in reactions catalyzed by a hammerhead ribozyme.

The hammerhead ribozyme is generally accepted as a well characterized metalloenzyme. However, the precise nature of the interactions of the RNA with metal ions remains to be fully defined. Examination of metal ion-catalyzed hammerhead reactions at limited concentrations of metal ions is useful for evaluation of the role of metal ions, as demonstrated in this study. At concentrations of Mn2+ ions from 0.3 to 3 mM, addition of the ribozyme to the reaction mixture under single-turnover conditions enhances the reaction with the product reaching a fixed maximum level. Further addition of the ribozyme inhibits the reaction, demonstrating that a certain number of divalent metal ions is required for proper folding and also for catalysis. At extremely high concentrations, monovalent ions, such as Na+ ions, can also serve as cofactors in hammerhead ribozyme-catalyzed reactions. However, the catalytic efficiency of monovalent ions is extremely low and, thus, high concentrations are required. Furthermore, addition of monovalent ions to divalent metal ion-catalyzed hammerhead reactions inhibits the divalent metal ion-catalyzed reactions, suggesting that the more desirable divalent metal ion-ribozyme complexes are converted to less desirable monovalent metal ion-ribozyme complexes via removal of divalent metal ions, which serve as a structural support in the ribozyme complex. Even though two channels appear to exist, namely an efficient divalent metal ion-catalyzed channel and an inefficient monovalent metal ion-catalyzed channel, it is clear that, under physiological conditions, hammerhead ribozymes are metalloenzymes that act via the significantly more efficient divalent metal ion-dependent channel. Moreover, the observed kinetic data are consistent with Lilley's and DeRose's two-phase folding model that was based on ground state structure analyses.

[Circulating endothelial cells in the peripheral blood of advanced NSCLC patients].

OBJECTIVE: To investigate the changes and clinical value of circulating endothelial cells (CEC) in the peripheral blood of advanced NSCLC patient. METHODS: Sixty-seven advanced NSCLC patients were randomly divided into either the treatment group with NP plus endostatin or control group with NP alone. Level of CEC and cytokeratin (CK) in the peripheral blood were measured by flow cytometry. RESULTS: The response rate and benefit rate was 44.4%, 80.0% in the treatment group, and 27.3%, 50.0% in the control group, respectively (P = 0.176 and P = 0.012). Time to tumor progression (TTP) was 146.7 days in the treatment group and 91.1 days in the control group (P = 0.061). However, when the cut-off of TTP was defined as > 170 days, there was a significant difference between two groups (cut-off = 170, P = 0.034; cut-off = 180, P = 0.009). The number of CEC decreased by 0.29 +/- 0.47 in the treatment group and by 0.01 +/- 0.43 in the control group (P = 0.033). The correlation between CEC and CK was found to be positive either before (r = 0.381, P = 0.013) or after the treatment (r = 0.450, P = 0.004). CONCLUSION: Chemotherapy combined with endostatin is superior to chemotherapy alone in the treatment of NSCLC. CEC, as a biomarker, may be useful in predicting the efficacy of the combined treatment.