The effects of monoterpene perilly alcohol (POH) alone or in combination with STI571 on the proliferation and apoptosis of the cell line K562 positive for Ber/Abl were investigated. By using cell culture, the effect of the drugs on the proliferation of the cells was studied. TUNEL and flow cytometry assay of FITC-Annexin V and PI labeled cells were applied to detect the effects of the drugs on the apoptosis of the cells. The results showed that at 36 h, IC50 of POH on K562 positive for Bcr/Abl and HL-60 negative for Bcr/Abl were 81.0±11.3 μmol/L and 113.6±23.4 μmol/L respectively (P>0.05). POH could inhibit the proliferation of K562 in a time- and dose-dependent manner with the inhibitory rate of 100 μmol/L POH on K562 cells at 36 h being (53.2±3.65)%. K562 cells were more sensitive to STI571 than POH. IC50 of STI571 on K562 cells in 36 h was (0.256±0.054) μmol/L. In a time- and dose-dependent manner, POH induced the apoptosis of K562 cells with the percentage of apoptotic cells by 100 μmol/L POH at 40 h being (21.0±3.3) %. Both 100 μmol/L POH and 0.2 μmol/L STI571 had the same inhibitory effects on the K562 cells at 36 h. But at 12 and 24 h, the inhibitory rate of POH was significantly higher than that of STI571 (P<0.05) and the ability of STI571 inducing apoptosis at 36 h was greater than that of POH. 50 μmol/L, 100 μmol/L and 200 μmol/L POH in combination with 0.2 μmol/L STI571 could obviously increase the inhibitory effects on the cellular proliferation. Combined use of 50 μmol/L, 100 μmol/L, 200 μmol/L with 0.2 μmol/L STI571 could strongly induced apoptosis, especially 200 μmol/L POH in combination with 0.2 μmol/L STI571. It was concluded that the antileukemia effect of POH had no obvious Bcr/Abl positive selectivity. POH can inhibit the proliferation of K562 and induce the apoptosis in a time- and dose-dependent manner. K562 cells were more sensitive to STI571 than POH. POH in combination with STI571 could obviously enhance the abilities of STI571 inhibiting the proliferation and inducing apoptosis of K562 cells.
To investigate TLR2 (Toll-like receptor 2) mRNA expression in ischemic hepatic lobes under the condition of partial hepatic ischemia/reperfusion injury in BALB/c mice and its relationship with liver function impairment. A partial ischemia/reperfusion injury model was established. The portal vein and hepatic artery supply to the median and left lobes of the liver were obstructed by an atraumatic artery micro-clip, with the obstruction lasting for about 60 min. Then reperfusion was fulfilled by removal of the clip. The liver samples were collected at the 4th h after the restoration of blood inflow. Total RNA was extracted from the liver samples and analyzed quantitatively by method of real-time PCR. At the same time, portal vein serum and plasma were taken respectively for further detection of the level of endotoxin, tumor necrosis factor alpha (TNF-alpha) and plasmic alanine aminotransferase (pALT). The results indicated that TLR2 mRNA in ischemic lobe was up-regulated markedly in mice partial liver ischemia/reperfusion injury model compared to that in sham operation group (ΔCt: 1.05±1.02 vs 5.08±1.36,P<0.001). The level of portal vein pALT and TNF-alpha increased significantly (112.32±17.56 pg/ml vs 6.07±5.33 pg/ml,P<0.01; 890±127 μ/L vs 30±5 μ/L,P<0.001). However, the level of portal vein endotoxin remained below the normal line, suggesting a state of non-endotoxemia. TLR2 mRNA expression in ischemic lobe, as well as portal vein pALT and TNF-alpha, was up-regulated in the model of mice partial ischemia/reperfusion injury, suggesting the involvement of TLR2 in ischemia/reperfusion pathological process.
The pathogenesis of aplastic anemia (AA) was explored and the effects of AA serum on the expression of crucial cyclin D isoform (cyclin D3) in umbilical cord blood hematopoietic stem/progenitor cells were observed. The CD34+ cells were isolated from the cord blood with MIDI-MACS Semi-solid methylcellulose culture technique was used to measure the formation of CFU-GM; The expression level of cyclin D3 was assayed by semi-quantitative RT-PCR and Western-blot after the hematopoietic stem/progenitor cells were incubated in AA serum. The results showed that the AA serum could inhibit the formation of CFU-GM and down regulate the expression level of the cyclin D3 at the mRNA and protein level respectively. In conclusion, the AA serum could inhibit the proliferation of hematopoietic stem cells and down regulate level of cyclin D3, which might be one mechanism of hematopoiesis inhibition in AA.
To investigate the role of connective tissue growth factor (CTGF) in transdifferentiation of human renal tubular epithelial cell (HKC),in vitro cultured HKC cells were divided into 3 groups: negtive control, low dose CTGF-treated group (rh CTGF, 2.5 ng/ml) and high dose CTGF-treated (rhCTGF, 5. 0 ng/ml). Then the expression of α-smooth muscle actin (α-SMA) were assessed by indirect immuno-fluorescence, and the percentage of α-SMA positive cells were assessed by flow cytometry. RT-PCR were also performed to examine the mRNA level of α-SMA. Upon the stimulation of different concentrations of rhCTGF, the expression of α-SMA were markedly stronger than that in negative controls. The percentages of α-SMA positive cells were significantly higher in the stimulated groups than that of negative controls (38.9%, 65.5% vs 2.4%,P<0.01). α-SMA mRNA levels were also up-regulated by the stimulation of rhCTGF (P<0.01). These results suggest that CTGF can promote the transdifferentiation of human renal tubular epithelial cells towards myofibroblast (Myo-F).
Heart fatty acid-binding protein (H-FABP) is supposed to be the most sensitive biomarker of early acute myocardial infarction (AMI). To evaluate the diagnostic value of H-FABP for AMI in the early stage, the plasma levels of H-FABP were measured by sandwich ELISA in 93 patients with suspected AMI at admission within 6h after onset of chest pain and 69 normal healthy subjects. The plasma concentrations of cardiac troponin-I (cTnI), creatine kinase-MB (CK-MB) and myoglobin (Mb) were assayed at the same time by using corpuscle chemiluminescence for those patients. The patients were classified as AMI group (n=32) and non-AMI group (n=61) retrospecitively. The diagnostic validity was evaluated in terms of sensitivity, specificity and receiver operating characteristics (ROC) curve analysis. The results showed the cutoff value of H-FABP for AMI was 16.8 ng/ml, and it diagnostic sensitivity for AMI was 64.29% within 3h and 84.38% within 6 h after onset of chest pain, and the diagnostic specificity for non-AMI was 100% within 3 h and 91.8% within 6 h. H-FABP had higher sensitivity than that of cTnI and CK-MB at all time points (P<0.05), whereas there was no significant difference in specificity among the four markers. But the area under the ROC curve of H-FABP was significantly greater than that of cTnI, CK-MB and Mb within 3 h. These results revealed that H-FABP possessed high diagnostic sensitivity and specificity for AMI in early stage, especially within 3 h after onset of persistent angina pectoris. In conclusion, H-FABP can be used as a sensitive marker for AMI in the early stage.