OBJECTIVETo study telomerase activity (TA) and its variation in bone marrow mononuclear cells from patients with myelodysplastic syndrome (MDS) at different stages in comparison with normal bone marrow cells and leukemic cells.

METHODSThe TA was semi-quantitatively determined in mononuclear cells from 20 normal bone marrow samples, 21 patients with MDS at different stages and 32 cases of acute leukemia by using a polymerase chain reaction-enzyme linked immuno-sorben assay (PCR-ELISA) kit.

RESULTSThe TA in normal bone marrow cells was in the range of 0 to 0.3 units (U) with a mean of 0.11 +/- 0.08 U. Among them, 3 samples were considered positive in accordance with the standard recommended by the kit's pamphlet. In bone marrow cells from patients with acute leukemia, the TA was ranging from 0 to 0.96 U with a mean value of 0.42 +/- 0.26 U. The positive rate was 78.1% which was significantly different from that in normal bone marrow (BM) (P < 0.01). In case of myelodysplastic syndrome, the average level of TA was 0.27 +/- 0.19 U (ranging from 0 to 0.97 U) with a positive rate of 66.7%. In comparison with normal BM cells, the difference was significant (P < 0.05). Particularly, the MDS high-risk subgroup exhibited a significantly higher activity of telomerase (P < 0.05). In comparison with INT-1 and INT-2 subgroups in MDS patients based on international prognostic scoring system (IPPS), the difference in TA was also significant (P < 0.05). The abnormality in cell karyotype was not correlated with TA.

CONCLUSIONThe normal bone marrow cells demonstrate TA at a marginal level while a remarkably increasing level may be seen in acute leukemia patients. The BM cells from MDS patients display a moderate TA among which the high risk MDS subgroup with a poor prognostic IPPS score exhibited markedly higher TA.

Adolescent ; Adult ; Bone Marrow Cells ; enzymology ; Child ; Female ; Humans ; Leukocytes, Mononuclear ; enzymology ; Male ; Middle Aged ; Myelodysplastic Syndromes ; enzymology ; Telomerase ; metabolism

BACKGROUNDSevere sepsis and septic shock are the leading causes of morbidity and mortality in hospitalized patients. This study aimed to investigate the association of poly (ADP-ribose) polymerase-1 (PARP-1) activity in circulating mononuclear cells with myocardial dysfunction in patients with septic shock.

METHODSA total of 64 patients with septic shock were divided into the survival group (n = 41) and the nonsurvival group (n = 23) according to mortality at 28 days after enrollments. PARP-1 activity in circulating mononuclear cells, brain natriuretic peptide, Acute Physiology and Chronic Health Evaluation II score, the cardiac index (CI), the cardiac function index (CFI), global ejection fraction (GEF), and the left ventricular contractility index (dp/dt max) were measured after admission to the intensive care unit.

RESULTSPARP-1 activity in circulating mononuclear cells of nonsurvival patients with septic shock was significantly higher than that in survival patients. PARP-1 activity in circulating mononuclear cells was strongly, negatively correlated with the CI, the CFI, GEF, and dp/dt max. Multiple Logistic regression analysis showed that PARP-1 activity in circulating mononuclear cells was an independent risk factor of myocardial dysfunction. The optimal cutoff point of PARP-1 activity for predicting 28-day mortality was 942 nmol/L with a sensibility of 78.2% and specificity of 65.1%.

CONCLUSIONPARP-1 activity in circulating mononuclear cells is significantly associated with myocardial dysfunction and may have prognostic value in patients with septic shock.

Aged ; Aged, 80 and over ; Female ; Humans ; Leukocytes, Mononuclear ; enzymology ; Male ; Middle Aged ; Poly(ADP-ribose) Polymerases ; metabolism ; Shock, Septic ; enzymology

Telomerase play a key role in the maintenance of telomere length and chromosome integrity. We have evaluated the association between telomerase activity and the risk of lung cancer in peripheral blood. Telomerase activity in peripheral blood mononuclear cells was measured by a PCR-designed telomeric repeat amplification protocol in 63 lung cancer patients and 190 healthy controls that were matched for age, gender, and smoking status. Telomerase activity was significantly lower in the lung cancer patients than in controls (mean +/- standard deviation; 1.32 +/- 1.65 vs 2.60 +/- 3.09, P < 1 x 10(-4)). When telomerase activity was categorized into quartiles based on telomerase activity in the controls, the risk of lung cancer increased as telomerase activity reduced (Ptrend = 1 x 10(-4)). Moreover, when the subjects were categorized based on the median value of telomerase activity, subjects with low telomerase activity were at a significantly increased risk of lung cancer compared to subjects with high telomerase activity (adjusted odds ratio = 3.05, 95% confidence interval = 1.60-5.82, P = 7 x 10-4). These findings suggest that telomerase activity may affect telomere maintenance, thereby contributing to susceptibility to lung cancer.
Age Factors ; Aged ; Case-Control Studies ; Female ; Humans ; Leukocytes, Mononuclear/enzymology/immunology ; Lung Neoplasms/*enzymology/*etiology ; Male ; Middle Aged ; Odds Ratio ; Risk Factors ; Sex Factors ; Smoking ; Telomerase/*blood

To explore the expression of heme oxygenase-1 (HO-1) in the peripheral blood mononuclear cells (PBMCs) and its relationship with pulmonary ventilation function in asthmatic patients, 18 asthmatic patients and 18 healthy subjects were selected. HO-1 protein and mRNA levels in PBMCs were measured by immunohistochemical staining and reverse transcription-polymerase chain reaction (RT-PCR), respectively. Blood carbon monoxide Hb (COHb), serum total IgE and pulmonary ventilatory function were observed. Our results showed that the percentage of cells positive for immunohistochemical staining of HO-1 were significantly higher in asthmatic patients (41.72 +/- 7.44) % than that in with healthy subjects (10.45 +/- 4.36) % (P < 0.001) and the optical density of PBMC HO-1 mRNA was higher in asthmatic patients (26.05 +/- 4.14) than that in healthy subjects (10. 82 +/- 4.26) (P < 0.001). The relation analysis showed that PBMC HO-1 protein and mRNA levels had significantly negative relation with FEV1%, PEFR, MEFR50%, respectively (r = -0.51-0.89, P < 0.05-0.001, respectively) and a positive relation with COHb and serum total IgE (r = 0.48-0. 85, 0.05-0.001, respectively). It is concluded that the expression of PBMC HO-1 protein and mRNA increased significantly in asthmatic patients, and HO-1 may play a significant role in the pathogenesis of asthma. The expression of HO-1 may bear a relation with severity of asthma.
Asthma/blood ; Asthma/*enzymology ; Carbon Monoxide/blood ; Heme Oxygenase-1/*biosynthesis ; Heme Oxygenase-1/blood ; Immunoglobulin E/*blood ; Leukocytes, Mononuclear/*enzymology ; RNA, Messenger/blood

To investigate the effect of total panax notoginseng saponins (tPNS) to induce the differentiation of mononuclear cells (MNC) in cord blood into endothelial cells, the DMEM culture media containing tPNS were used to induce the MNC of cord blood. Then, the morphology of the adherent cells was observed by the light microscopy and the fluorescence microscopy, the changes of cell surface markers (UEA-1), function marker (vWF) and CD31 were detected by flow cytometry. The results showed that the number of adherent cells produced by 250 mg/L tPNS and the positive rate of cells expressing CD31 and UEA-1 were higher than those in the groups of other concentrations (P < 0.05). There was no significant difference in the number of adherent cells expressing CD31 and UEA-1 between 50 ng/ml VEGF + 250 mg/L tPNS and 50 ng/ml VEGF. It is concluded that the traditional Chinese drug tPNS can induce partial MNC in the cord blood to differentiate into endothelial cells. No synergistic effect has been found between tPNS and VEGF.
Cell Differentiation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; enzymology ; Fetal Blood ; cytology ; Humans ; Leukocytes, Mononuclear ; cytology ; Panax notoginseng ; chemistry ; Saponins ; isolation & purification ; pharmacology

OBJECTIVETo study the effects of rare earth exposure on human telomerase and apoptosis of human peripheral mononuclear cells (PBMNs).

METHODSRare earth mine lot in Xunwu county, the biggest ion absorptive rare earth mine lot of China, was selected as the study site. Another village of Xunwu county, with comparable geological structure and social environment was selected as the control site. Thirty healthy adults were randomly selected from the study site as exposure group and another 30 healthy adults randomly selected from the control site as control group. The blood content of 15 rare earth elements, including La, Ce, Pr, Nd, Sm, Eu, Gd, Tb, Dy, Ho, Er, Tm, Yb, Lu and Y, were determined by inductive coupled plasma-source mass spectrometry (ICP-MS). The total contents of rare earth elements in the blood were calculated. The TRAP and FCM assays were carried out to analyse the telomerase and apoptosis of human PBMNCs respectively.

RESULTSIn the exposure group, the concentration of La, Ce, Dy and Y were significantly higher (P<0.001), and Pr, Nd, Sm, Gd and Yb were higher than those in the control group (P<0.05). The total content of rare earth in the blood of exposure group showed significant difference compared with control group (P<0.001). Telomerase activity in PBMNs of the exposure group was higher than that in the control group (P<0.05); there were 11 adults in the exposure group (30 adults) and 5 adults in control group (30 adults) showed positive telomerase activity. The average age of the exposure group was (38.69 +/- 8.02) years-old, while the control group was (40.45 +/- 9.02) years-old (P >0.05). It was found that there was a significant relationship between telomerase activity and the total content of rare earth elements (P <0.01). 3. The proportion of apoptosis was not different between the two groups (P >0.05), but the cells in the S-phase and G2-M phase were increased (P <0.01) in the exposed group.

CONCLUSIONThe telomerase activity of PBMNs in the rare earth elements exposed group was higher than that of the control group, and there is no effect on apoptotic rate of PBMNs, but may promote the diploid DNA replication, and increase the percentage of G2/M and S phase cells.

Apoptosis ; drug effects ; Environmental Exposure ; adverse effects ; Female ; Humans ; Leukocytes, Mononuclear ; cytology ; enzymology ; Male ; Metals, Rare Earth ; adverse effects ; analysis ; Telomerase ; metabolism

OBJECTIVEThe beta-D-endoglycosidase heparanase (Hpa) is HS-specific which leads to the degradation of heparan sulfate (HS). An increased permeability of the glomerular basement membrane (GBM) for proteinuria was suggested to relate to a decrease of HS side chains in the GBM. However, whether an up-regulated expression of Hpa exists in steroid responsive nephrotic syndrome (SRSNS) remains unknown. The purpose of this study was to evaluate the role of Hpa in the development of SRSNS and the correlation with the proteinuria.

METHODSForty-three children with SRSNS were selected and included the active stage group (n = 23), the restoration stage group (n = 10) and the remission stage group (n = 10). There were 23 nephritic nephrosis children, 15 purpura nephritis children and 15 healthy children as controls. By using the method of reverse transcriptase-PCR (RT-PCR), Hpa gene expression in the peripheral blood leukocytes (PBLs) was assayed.

RESULTS(1) All patients with nephrotic syndrome exhibited higher levels of Hpa mRNA than those of the healthy group (P < 0.05). The highest expression was in the active stage group of SRSNS (1.27 +/- 0.36, P < 0.001), while there was no difference between the patients of nephritic nephrosis group (0.62 +/- 0.15) and purpura nephritis group (0.55 +/- 0.17) (P > 0.05). (2) In contrast with the healthy group, there was a significant difference in the active stage group of SRSNS (P < 0.001). So was the restoration stage group (P < 0.05), but there was no difference to the remission stage group (P > 0.05). (3) There was a positive correlation between the expression level of Hpa mRNA and the quantity of urinary protein (r(s) = 0.751, P < 0.001).

CONCLUSIONUp-regulated expression of Hpa mRNA may be important to the loss of glomerular negative charge in GBM and lead to proteinuria in SRSNS.

Adolescent ; Child ; Child, Preschool ; Female ; Gene Expression Regulation, Enzymologic ; Glucuronidase ; genetics ; Humans ; Leukocytes, Mononuclear ; metabolism ; Male ; Nephrotic Syndrome ; enzymology ; genetics ; Proteinuria ; enzymology ; genetics ; RNA, Messenger ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

To establish a quantitative assay for telomerase activity and analyze the telomerase activity in peripheral blood mononuclear cells (PBMNC) from patients with acute leukemia, a fluorescent dye, PicoGreen, was added to the products after telomere repeat amplification protocol. The samples were excited at 480 nm and the fluorescence emission intensity was measured at 520 nm using a spectrofluorometer. Telomerase activity was detected in PBMNCs from 20 cases of normal individuals and 25 patients with acute leukemia. The results showed that the fluorescence of PicoGreen binding to double-stranded DNA specifically was enhanced with increase of DNA quantities. In conclusion, the met hod is rapid, simple and quantitative, the telomerase activities of PBMNCs from acute leukemia patients are significantly higher than that of the normal controls.
Acute Disease ; Adolescent ; Adult ; Aged ; Cell Line ; DNA, Neoplasm ; genetics ; metabolism ; Female ; Fluorescent Dyes ; chemistry ; Humans ; Leukemia ; blood ; enzymology ; genetics ; Leukocytes, Mononuclear ; enzymology ; Male ; Middle Aged ; Organic Chemicals ; Telomerase ; chemistry ; genetics ; metabolism

OBJECTIVETo observe in vitro the effect of Sinomenine, a pure alkaloid extracted from the chinese medical plant Sinomenium acutum on the activity of cyclooxygenase (COX-1 and COX-2) and the expression of COX-1 and COX-2 mRNA.

METHODMononuclear leukocytes were obtained from healthy adults. Isolated mononuclear leucocytes from human peripheral blood (PBMC) were incubated (1 x 10(6).mL-1) with or without sinomenine (or indomethacin), after incubated for 24 hours at 37 degrees C with 5% CO2; the media were assayed for the PGE2 by radioimmunoassay (RIA). LPS was used to stimulate the monocytes at a concentration of 5 micrograms.mL-1. And by RT-PCR, both COX-1 and COX-2 mRNAs were detected in Mononuclear leukocytes after incubation for different hours with drug (sinomenine or indomethacin) or not.

RESULTLPS (stimulated) induced the production of PGE2 in PBMC increasing with high expression of COX-2 mRNA; sinomenine reduced PGE2 production in LPS stimulated human monocytes more than in non-stimulated human monocytes. In comparative experiments, indomethacin, a non selective COX inhibitor, reduced the production of PGE2 equally in both states. Meanwhile, neither sinomenine(0.1-1 mmol.L-1) nor indomethacin(0.5-10 mumol.L-1) inhibited the expression of both COX-1 and COX-2 mRNAs by RT-PCR with beta-actin as reference.

CONCLUSIONIn contrast with indomethacin, Sinomenine shows a preferential inhibitory effect on COX-2 over COX-1, These results suggest that Sinomenine is a selective COX-2 inhibitor, which may be directly related to suppressing cyclooxygenase activity.

Adult ; Cyclooxygenase 1 ; Cyclooxygenase 2 ; Dinoprostone ; blood ; Humans ; Isoenzymes ; biosynthesis ; Leukocytes, Mononuclear ; enzymology ; Membrane Proteins ; Morphinans ; isolation & purification ; pharmacology ; Plants, Medicinal ; chemistry ; Prostaglandin-Endoperoxide Synthases ; biosynthesis ; metabolism ; RNA, Messenger ; biosynthesis ; Sinomenium ; chemistry

OBJECTIVETo study the clinical and enzymological characteristics of the children with mitochondrial respiratory chain complex III deficiency.

METHODThe clinical manifestations of five patients (3 males, 2 females) were summarized. Spectrophotometric assay was used for the analysis of respiratory chain complex I to V enzyme activity in peripheral blood leukocytes, after obtaining venous blood.

RESULT(1) Five patients were hospitalized at the age of 1 month to 15 years. Three patients had Leigh syndrome with progressive motor developmental delay or regression and weakness. One had severe liver damage and intrahepatic cholestasis. One presented muscle weakness. (2) Deficient complex I + III activity was identified in five patients. Their complex I + III activities in peripheral blood leukocytes were 3.0 to 14.2 nmol/min per mg mitochondrial protein (control: 84.4 ± 28.5 nmol/min per mg mitochondrial protein). The ratio of complex I + III to citrate synthase decreased to 3.5 to 22.9% (normal control 66.1 ± 14.7%). The activities of complex III decreased to 10.4 to 49.3% of the lowest control value, while complex I, II, IV and V activities were normal. The results supported the diagnosis of isolated respiratory chain complex III deficiency.

CONCLUSIONComplex III deficiency is a kind of disorder of energy metabolism with various manifestations. The complex I + III activities and the ratio of complex I + III to citrate synthase were lower than those of the control. The activities of complex I, II, IV and V were normal.

Adolescent ; Child ; Child, Preschool ; Electron Transport Complex I ; metabolism ; Electron Transport Complex II ; metabolism ; Electron Transport Complex III ; metabolism ; Female ; Humans ; Infant ; Leigh Disease ; Leukocytes, Mononuclear ; enzymology ; Male ; Mitochondrial Diseases ; diagnosis ; metabolism ; physiopathology