Objective To investigate children′s myelodysplastic hematopoietic cells reaction to interleukin (IL)-15.Methods CD 34 + cells in bone marrow from 18 myelodysplast syndrome(MDS) patients were purified by an immunomagnetic beads sorting system. Apoptosis of hematopoietic precursors was assayed by propidium iodine staining and flow cytometric analysis.Results On 8th cultured day,when IL-15 concentration was between 0-100 ng/ml,it could suppress apoptosis of hematopoietic cells in MDS patients in a dose-and- time dependent manner. IL-15 in study group significanthy lower than that of control group.Conclusion IL-15 may partly suppress apoptosis of hematopoietic cells in MDS patients.

Objective To investigate the expression of immunohistochemistry of gastrin(GAS),somatostatin(SS),proliferating cell nuclear antigen(PCNA) and Fas-ligand(Fas-L) in the sinus ventriculi of children with pediatric gastritis and to explore the significance of their expression in the pathogenesis of pediatric chronic gastritis.Methods Fifty cases of the sinus ventriculi mucosa samples were enrolled in 3 groups:chronic gastritis,helicobacter pylori(Hp) positive(group A,n=20);chronic gastritis,Hp negative(group B,n=19);control group,normal sinus ventriculi mucosa,Hp negative(group C,n=11).Immunohistochemistry En Vision were carried out including GAS,SS,PCNA and Fas-L.Results In the expression of GAS and SS,the values of group A and B were comparatively higher than those of group C,but there was no significant difference among them in statistics.In the expression of PCNA,the value of group A was comparatively higher and that of group B.The value difference between 2 groups was significant(P=0.019);in the expression of Fas-L,no significant difference was found among these 3 groups.Conclusions Expressions of GAS and SS both increase in children with chronic gastritis and maybe the increase of GAS and SS play a role in the pathogenesis of pediatric chronic gastritis;Hp infection promotes the multiplication of the sinus ventriculi membrana mucosa epithelium cell in pediatric chronic gastritis.

OBJECTIVETo observe the effects of co-transfection of platelet derived growth factor antisense oligodeoxynucleotides (PDGF-AODN) and tissue-type plasminogen activator (tPA) gene on inhibition of intimal proliferation of auto-transplantion artery.

METHODSOne hundred male New Zealand rabbits were randomly divided into four groups (25 in each): Group A (control group), Group B (PDGF-AODN transfection group), Group C (tPA gene transfection group) and Group D (PDGF-AODN and tPA co-transfection group). The left and right external iliac arteries were transplanted reciprocally. The transplanted arteries were respectively soaked in PDGF-AODN, pBudCE4.1/tPA and PDGF-AODN plus pBudCE4.1/tPA solution about 15 minute before transplantation. The rabbits were sacrificed at 3d, 1w, 2w, 4w and 8w after operation. The specimens were harvested for pathologic examination, electron microscopy, chromogenic substrate test, 3H-TdR incorporation test and immunohistochemical staining.

RESULTThe scan electron microscopy showed that there were a few thrombocytes on vas-wall of Group C and D without thrombus, whereas there were abundant thrombocytes and thrombus forming on vas-wall of Group A and B. The intimal area, stenosis ratio of transplanted artery, 3H-TdR incorporation,the number of PDGF positive cell in Group D were significantly less than those in Group A (P<0.01),Group B and Group C (both P<0.05). The activity of tPA gene products in transplanted vas-wall of Group D was significantly higher than that of Group A (P<0.01).

CONCLUSIONLocal co-transfection of PDGF-AODN and tPA gene can effectively inhibit the proliferation of vascular smooth muscle cells, hyperplasia of intima and restenosis of transplanted artery.

Animals ; Endothelium, Vascular ; pathology ; Graft Occlusion, Vascular ; prevention & control ; Hyperplasia ; prevention & control ; Iliac Artery ; transplantation ; Male ; Oligodeoxyribonucleotides, Antisense ; biosynthesis ; genetics ; Platelet-Derived Growth Factor ; biosynthesis ; genetics ; Rabbits ; Random Allocation ; Tissue Plasminogen Activator ; biosynthesis ; genetics ; Transfection ; Tunica Intima ; pathology

OBJECTIVETo investigate the effect of PD98059 on the proliferation and cell cycle of rat hepatic stellate cells (HSCs) stimulated by acetaldehyde and explore its mechanism.

METHODSRat HSCs stimulated by acetaldehyde were incubated with different concentrations of PD98059. Cell proliferation was assessed by MTT colorimetric assay. Cell cycle was analysed by flow cytometry. The mRNA of cyclin D1 and CDK4 were examined by RT-PCR.

RESULTS20, 50, 100 micromol/L PD98059 could significantly inhibit the proliferation of HSCs stimulated by acetaldehyde in a does-dependent manner (0.109+/-0.020, 0.081+/-0.010 and 0.056+/-0.020 vs 0.146+/-0.030, F=31.385, P<0.05) and provoke G0/G1 phase arrest of HSCs stimulated by acetaldehyde in a does-dependent manner (61.9%+/-6.3%, 64.1%+/-3.3% and 70.9%+/-4.8% vs 55.2%+/-4.4%, F=16.402, P<0.05). 50, 100 micromol/L PD98059 could markedly inhibit cyclin D1 mRNA expression of HSC stimulated by acetaldehyde (0.56+/-0.04 and 0.46+/-0.03 vs 0.65+/-0.07, F=68.758, P<0.05) and CDK4 mRNA expression (0.39+/-0.07 and 0.33+/-0.05 vs 0.50+/-0.06, F=29.406, P<0.05).

CONCLUSIONThe Erk signal transduction pathway plays an important role in regulating the proliferation and cell cycle of rat hepatic stellate cells stimulated by acetaldehyde, which may be partly related to its regulative effect on the expression of cyclin D1 gene and CDK4 gene

Acetaldehyde ; pharmacology ; Animals ; Cells, Cultured ; Cyclin D1 ; metabolism ; Cyclin-Dependent Kinase 4 ; Cyclin-Dependent Kinases ; metabolism ; Enzyme Inhibitors ; pharmacology ; Flavonoids ; pharmacology ; Hepatocytes ; drug effects ; Proto-Oncogene Proteins ; Rats

OBJECTIVETo study the effect of platelet-derived growth factor (PDGF) and proliferating cell nuclear antigen (PCNA) antisense oligodeoxynucleotides (AODN) together on inhibiting the proliferation of the stenosis of transplanted vascular.

METHODSThe left and right external iliac arteries (length 1.0 cm) of rabbits were transplanted reciprocally. The transplanted vascular were respectively soaked in liposomes, PDGF-AODN, PCNA-AODN and PDGF-AODN adding PCNA-AODN solution about 20 minute, the vascular anastomotic were sutured by 8/0 suture of soaked in AODN solution. Four weeks later, the specimens were harvested for microscopy. The pathological morphology of transplanted vascular were observed under microscope (HE). The intimal thickness and area, stenosis ratio(%) of transplanted vascular were calculate and analysed statistically among group by computer system. The number of positive cells of PDGF's mRNA in transplanted vascular wall were counted with in situ hybridization histo-cytochemistry and the number of positive cells of PCNA's protein in transplanted vascular wall were counted by S-P immunochemistry.

RESULTSThe intimal thickness and area, stenosis ratio of transplanted vascular, the number of PDGF and PCNA positive cell in PDGF-AODN adding PCNA-AODN group were significantly lower than those in other group (P < 0.01), and that were lower evidently than PDGF-AODN group and PCNA-AODN group.

CONCLUSIONPDGF and PCNA antisense oligodeoxynucleotides together could significantly inhibit the proliferation of vascular smooth muscle cell and stenosis of transplanted vascular.

Animals ; Graft Occlusion, Vascular ; pathology ; prevention & control ; Iliac Artery ; pathology ; transplantation ; Male ; Oligonucleotides, Antisense ; genetics ; Platelet-Derived Growth Factor ; biosynthesis ; genetics ; Proliferating Cell Nuclear Antigen ; biosynthesis ; genetics ; Rabbits ; Transfection ; Transplantation, Homologous

OBJECTIVETo observe the effects of local co-transfection vascular endothelial growth factor165 (VEGF165) and tissue-type plasminogen activator genes on inhibiting intimal hyperplasia and restenosis in rabbits artery after operation injury and possible mechanisms.

METHODSMicrology operation injury was used to establish the model of intimal injury of right external iliac artery in rabbits. To select 120 male New Zealand rabbits and were randomly divided into 3 groups (n = 40, in each group): Group A (physiological brine control group), Group B (pBudCE4.1 group), Group C (pBudCE4.1/VEGF165-tPA group). The vas-wall of micrology operation injury were infused respectively physiological brine, pBudCE4.1 and pBudCE4.1/VEGF165-tPA transfection solution by micro-injector. Each group were divided into 5 subgroups (n = 8, in each subgroup) randomly according to the sacrifice times (2 d, 1 week, 2 week, 4 week and 8 week after operation). The injured vascular specimen were harvested for pathology test, electric microscopy study, reverse transcription-PCR examining and immunochemistry detecting.

RESULTSThe intimal area and narrow ratio of vases in Group C at every time point after operation were significantly lessened than that in Group A and Group B (P < 0.01). The narrow ratio of vases in Group C at 8 week after operation were decreased respectively by 57.9% and 59.0% than that in Group A and B. The expression of VEGF165 mRNA in Group C were increased significantly than that in Group A and B at every time point after operation (P < 0.01), the expression reached the peak at 1 week and continued to 4 week after operation. Immunohistochemical identified that tPA positive cell in Group C were significantly increased than that in Group A and B (P < 0.01) at every time point after operation.

CONCLUSIONLocal co-transfection VEGF165 and tPA genes could restrain intimal hyperplasia and restenosis of vas, which lay a foundation for future multi-gene therapy of vascular intimal hyperplasia.

Animals ; Arteries ; pathology ; Endothelial Cells ; cytology ; Hyperplasia ; prevention & control ; In Vitro Techniques ; Male ; Myocytes, Smooth Muscle ; cytology ; Plasmids ; Rabbits ; Random Allocation ; Tissue Plasminogen Activator ; biosynthesis ; genetics ; Transfection ; Tunica Intima ; pathology ; Vascular Endothelial Growth Factor A ; biosynthesis ; genetics

OBJECTIVETo study the anticoagulant constituents in dried leech (Whitmania pigra Whitman).

METHODThe plasma recalcification time (PRT) as index, the constituents with anticoagulant activity were isolated and purified by anion-exchange chromatography on Sephadex DEAE A-50, gel permeation chromatography on Sephadex G-25 and Sephadex LH-20 columns, and then reversed phase high-performance liquid chromatography successively.

RESULTThree anticoagulant polypeptides were isolated and purified. Compounds 1 and 2 can be translated each other in natural conditions, and their molecular weights are 7100 and 5531, respectively. Compound 3 was identified as a pure polypeptide by HPLC and SDS-PAGE, and its molecular weight was determined as 8 608 by MALDI-TOF-MS. The amino acid composition of compound 3 was also determined.

CONCLUSIONCompound 3 was inferred to be a novel anticoagulant, and named whitmanin.

Animals ; Anticoagulants ; analysis ; chemistry ; isolation & purification ; Leeches ; chemistry ; Molecular Weight

Background Haze formation is a key factor of vision reduce following corneal refractive surgery.Transforming growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA) are documented to participate in haze formation.Laboratory study showed that bevacizumab can not only inhibit corneal neovascularization,but also promote the healing of corneal epithelial basement membrane.However,the impact of bevacizumab on corneal healing after Off-flap epipolis laser in situkeratomileusis (Off-flap Epi-LASIK) is unclear.Objective The present study was to investigate the inhibition effect of bevacizumab on corneal haze after off-flap Epi-LASIK and its active mechanism.Methods Off-flap Epi-LASIK was performed in 24 adult pigmented rabbits and these rabbits were randomized into three groups.Bevacizumab of 0.1 ml (2.5 mg) was subconjunctivally injected 10 minutes after surgery in 16 rabbits and the same amount of bevacizumab was repeatedly injected 4 days after the initial injection in 8 eyes of 16 eyes.In addition,equivalent amount of normal saline solution was used in the same way in the other 8 rabbits.Another 2 health rabbits were used as the blank controls.Operative eyes were examined by slit lamp biomicroscope daily after surgery and haze was scored based on SundarRayde criteria.Corneas were obtained 4 weeks after operation for hematoxylin & eosin and periodic acid Schiff staining.Expressions of TGF-β1 and α-SMA in corneal tissue were detected by immunochemistry.Results Corneal epithelium healed completely in all eyes 4-5 days after operation.The haze scores were lower in the bevacizumab single injection group and repeat injection group than those in the normal saline solution group (P＜0.05) in 1 week and 4 weeks after operation.However,no significant difference was seen in the haze scores between the bevacizumab single injection group and repeat injection group (P ＞ 0.05).The hostopathological examination showed that the fibrosis response of cornea tissue was slight in the bevacizumab single injection group and repeat injection group comparison with the normal saline solution group.At 1 week after operation,the expression levels of TGF-β1 were (49.8 ± 2.1) PU and (38.6 ±4.4) PU in the bevacizumab single injection group and repeat injection group,and those of 4 weeks were (37.7 ±4.8) PU and (28.3 ± 3.5) PU,indicating significant decrease in the TGF-β1 expression compared with (65.1 ±5.3) PU and (51.6±2.2) PU of the normal saline solution group in both 1 week and 4 weeks (P＜0.01).The expression levels of α-SMA in corneas were (67.2±10.0) PU and (32.7±3.1) PU at 1 week,and (34.2±5.7) PU and (22.8±3.0) PU at4 weeks after operation in the bevacizumab single injection group and repeat injection group,which were significantly lower than (87.8±7.7) PU and (59.4±5.6) PU in the normal saline solution group in both 1 week and 4 weeks (P＜0.05).Meanwhile,the expression levels of TGF-β1 and α-SMA were declined in the bevacizumab repeat injection group compared with single injection group (P＜0.01).Periodic acid Schiff staining exhibited that the basement membrane of cornea was intact and continued in bevacizumab injection group.But corneal basement membrane was discontinuous in the normal saline solution group.Conclusions Subconjunctival injection of bevacizumab downregulates the expressions of TGF-β1and α-SMA in cornea after Off-flap Epi-LASIK and thus prevents haze formation.

Objective To investigate the change of the basic fibroblast growth factor(bFGF) leve in human detrusor muscle(DM)in bladder outlet obstruction(BOO)due to benign prostatic hyperplasia(BPH)and its implication.Methods Fifty-four patients with BPH were divided into two groups:the obstructive DM stability and instability groups;and 15 men with bladder tumor who underwent operation in the same period were enrolled in the control group.The bFGF mRNA level in DM was measured by reverse transcription and polymerase chain reaction(RT-PCR)and the bFGF protein level was measured by immunohistochemical staining method.Results The bFGF-mRNA expression level of bladder smooth muscle cells was significantly lower in the control group than that in the obstructive DM stability and instability groups(all P＜0.05),but there was no significant difference between the obstructive DM stability and instability groups(P＞0.05). Conclusions The expression level of bFGF mRNA in bladder DM is elevated in BOO due to BPH,but there is little or no correlation between the increased expression of bFGF mRNA and detrusor muscle instability.

OBJECTIVETo determine the mechanism underlying the therapeutic activities of glycogen synthase kinase 3b (GSK3b) against hepatic ischemia-reperfusion (H-IR) injury by investigating the inhibitive effects of GSK3b on inflammation mediated by Toll-like receptor 4 (TLR4).

METHODSC57BL/6 male mice were subjected to 90 min of warm liver cephalad lobe ischemia, followed by reperfusion for various lengths of time. The mice were divided into three groups: the H-IR untreated model (control group), and the H-IR inflammation-induced models that received an intraperitoneal injection of purified lipopolysaccharide (LPS) endotoxin alone (inflammation group) or with pretreatment of the SB216763 GSK3b-specific inhibitor (intervention group). To create a parallel isolated cell system for detailed investigations of macrophages, marrow-derived stem cells were isolated from femurs of the H-IR control group of mice and used to derive primary macrophages. The cells were then divided into the same three groups as the whole mouse system: control, LPS-induced inflammation model, and inflammation model with SB216763 intervention. Differential expressions of inflammation-related proteins and genes were detected by Western blotting and real-time quantitative PCR, respectively.

RESULTSThe phosphorylation levels of ERK, JNK and p38 MAPK were induced in liver at 1 h after reperfusion, but then steadily decreased and returned to baseline levels by 4 h after reperfusion. In addition, the phosphorylation levels of ERK and JNK were induced in macrophages at 15 min after LPS stimulation, while the phosphorylation level of p38 MAPK was induced at 1 h; SB216763 pretreatment suppressed the LPS-stimulated ERK, JNK and p38 phosphorylation in macrophages. In the mouse model, GSK3b activity was found to promote the gene expression of anti-inflammatory cytokine IL-10 (control: 0.21 ± 0.08, inflammation: 0.83 ± 0.21, intervention: 1.76 ± 0.67; F = 3.16, P = 0.027) but to significantly inhibit the gene expression of pro-inflammatory cytokines IL-12 (control: 0.11 ± 0.05, inflammation: 0.85 ± 0.11, intervention: 0.43 ± 0.10; F = 2.67, P = 0.038), TNF-a, (control: 0.052 ± 0.012, inflammation: 8.11 ± 0.98, intervention: 3.9 ± 0.82; F = 4.13, P = 0.016), IL-6 (control: 0.22 ± 0.08, inflammation: 6.37 ± 0.81, intervention: 2.11 ± 0.63; F = 3.21, P = 0.024), and IL-1b (control: 0.12 ± 0.07, inflammation: 2.51 ± 0.62, and intervention: 1.28 ± 0.33; F = 2.22, P = 0.030).

CONCLUSIONInhibition of GSK3b selectively regulates the expression of anti-inflammatory and pro-inflammatory cytokines in liver Kupffer cells (liver macrophages). This process may be one of the mechanisms underlying the ability of GSK3b to ameliorate hepatic ischemia-reperfusion injury, possibly because inhibition of pro-inflammatory cytokines may indirectly mediate liver cell apoptosis.

Animals ; Cells, Cultured ; Cytokines ; metabolism ; Glycogen Synthase Kinase 3 ; metabolism ; Glycogen Synthase Kinase 3 beta ; Inflammation ; metabolism ; pathology ; Lipopolysaccharides ; adverse effects ; Liver ; pathology ; Macrophages ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Reperfusion Injury ; Toll-Like Receptor 4 ; metabolism