激光多普勒血流仪测定兔后肢电损伤局部血流变化

目的:应用激光多谱勒血流仪监测局部血流变化以判断高压电损伤程度。方法:实验于2003-10/2005-01在深圳市第二人民医院烧伤整形科实验室完成。选择健康新西兰兔32只,采用自制电击设备建立电损伤重度损伤模型,将电击区域均分为A,B,C,D,E5个区,依次为肢体远端到近端。在A,B,E区于伤前、伤后0,2,4,24,48,72h,4,5,6,7d测定局部皮肤表面及肌肉中血流值变化情况。结果:①损伤前A、B、E区皮肤表面平均血流值分别为75.74,78.86,84.21PU,表明肢体近端至远端血流值逐渐减少,各点之间血流值差异无显著性意义(P>0.05)。②A区皮肤表面伤后2h血流值明显低于伤后0,4,24,48h,差异有显著性意义[分别为(27.22±22.80),(91.60±74.37),(63.97±44.41),(58.80±52.50),(58.72±39.02)PU,t=2.357,2.856,2.364,2.998,P<0.05],此时组织学表现为细胞水肿明显,有少量中性粒细胞浸润。A区皮肤表面伤后4h血流值增加,但至伤后4d才逐渐恢复到伤前水平。A区肌肉伤后2h血流值低于伤后24h,4d,差异有显著性意义[分别为(83.70±73.30),(135.10±63.07),(160.26±71.32)PU,t=2.383,2.367,P<0.05]。此时组织学表现为肌纤维肿胀,部分横纹消失,肌纤维的胞浆呈红染的疏松水波状和网状结构。③B区皮肤表面伤后4,24h血流值低于损伤前和伤后0h,差异有显著性意义[分别为(50.75±41.69),(47.23±35.36),(78.86±26.47),(83.89±32.77)PU;t=2.324,2.732,2.826,3.189,P<0.05,0.01],组织学表现同A区。B区局部肌肉伤后4h血流值低于伤后0,48,72h,差异有显著性意义[分别为(64.65±34.08),(107.11±36.50),(130.30±75.04),(180.56±72.59)PU,t=2.545,2.712,2.963,P<0.05]。此时组织学表现基本上同A区伤后2h。④E区皮肤表面各时相点之间血流值差异无显著性意义(P>0.05)。组织学表现同A区。E区局部肌肉伤后2h血流值低于伤后0,24,72h,差异有显著性意义[分别为(129.11±65.08),(175.02±71.67),(169.14±71.05),(196.64±67.34)PU,t=2.776,2.521,2.895,P<0.05]。此时组织学表现为肌纤维溶解断裂及中性粒细胞浸润。结论:用激光多谱勒血流仪监测损伤局部血流变化可以判断高压电损伤程度。     

Lithium augments GM-CSA generation in canine cyclic hematopoiesis

Cyclic hematopoiesis in gray collie dogs can be cured by lithium treatment. We examined the mechanism of lithium's effect by developing an assay for the canine equivalent of GM-CSF (called GM-CSA). Phytohemagglutinin (PHA)-stimulated canine blood mononuclear cells produce GM-CSA in a dose-dependent manner; this GM-CSA stimulates more neutrophil-containing colonies than does endotoxin-treated dog serum. Production of GM-CSA by PHA-stimulated normal dog cells was not altered by lithium. However, cells from gray collies during their neutrophilic period increased their GM-CSA when lithium (2 mEq/L) was added to low doses of PHA, whereas neutropenic gray collie cells did not. These data suggest that lithium could modulate cyclic hematopoiesis by increasing intramedullary GM-CSA at the time when marrow neutrophilic progenitor cells are at their nadir.     

Fibronectin in artery subendothelium is important for platelet adhesion

The role of subendothelial fibronectin in platelet interaction with subendothelium was studied. Human umbilical artery subendothelium was exposed to flowing blood containing 111In-labeled platelets in an annular perfusion chamber. Platelet adhesion was determined from the 111In radioactivity on the vessel wall. When perfusions were performed for five minutes at a wall shear rate of 1,800 s-1, platelet adhesion was the same whether normal plasma or fibronectin-free plasma was used. Preincubation of subendothelium with rabbit anti-human fibronectin serum, however, resulted in a marked inhibition of platelet adhesion. Preincubation with normal rabbit serum had no effect. Platelet adhesion was also diminished when the vessel wall was preincubated with anti- fibronectin IgG fraction or F(ab')2 fragment. After the latter preincubations, frozen sections of 4 micron were incubated with fluorescein isothiocyanate-conjugated goat anti-rabbit IgG, F(ab')2 fragment specific. Fluorescence was seen throughout the subendothelium both before and after perfusion. No fluorescence was seen when subendothelium was preincubated with normal rabbit IgG or F(ab')2 or with anti-fibronectin IgG that had been absorbed with purified fibronectin. After absorption of anti-fibronectin IgG with purified fibronectin, the inhibiting effect on platelet adhesion was also no longer present. Preincubation of the vessel wall with anti-fibronectin IgG reduced platelet adhesion significantly at a wall shear rate of 800 s-1. This effect was even greater at 1,800 s-1. At low shear rate (400 s-1), there was no inhibition.     

A randomized controlled phase III trial of recombinant human granulocyte colony-stimulating factor (filgrastim) for treatment of severe chronic neutropenia

Patients with idiopathic, cyclic, and congenital neutropenia have recurrent severe bacterial infections. One hundred twenty-three patients with recurrent infections and severe chronic neutropenia (absolute neutrophil count < 0.5 x 10(9)/L) due to these diseases were enrolled in this multicenter phase III trial. They were randomized to either immediately beginning recombinant human granulocyte colony- stimulating factor (filgrastim) (3.45 to 11.50 micrograms/kg/d, subcutaneously) or entering a 4-month observation period followed by filgrastim administration. Blood neutrophil counts, bone marrow (BM) cell histology, and incidence and duration of infection-related events were monitored. Of the 123 patients enrolled, 120 received filgrastim. On therapy, 108 patients had a median absolute neutrophil count of > or = 1.5 x 10(9)/L. Examination of BM aspirates showed increased proportions of maturing neutrophils. Infection-related events were significantly decreased (P < .05) with approximately 50% reduction in the incidence and duration of infection-related events and almost 70% reduction in duration of antibiotic use. Asymptomatic splenic enlargement occurred frequently; adverse events frequently reported were bone pain, headache, and rash, which were generally mild and easily manageable. These data indicate that treatment of patients with severe chronic neutropenia with filgrastim results in a stimulation of BM production and maturation of neutrophils, an increase in circulating neutrophils, and a reduction in infection-related events.     

Characterization of endogenous cytokine concentrations after high-dose chemotherapy with autologous bone marrow support

Endogenous cytokines are thought to mediate numerous biologic processes and may account for some adverse effects experienced following the administration of recombinant proteins. This study describes the pattern of endogenous cytokine exposure following high-dose chemotherapy. Blood concentrations of interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), macrophage colony-stimulating factor (M-CSF), and erythropoietin (EPO) were measured by enzyme-linked immunosorbent assay (ELISA) methods in 68 patients receiving the same ablative chemotherapy regimen (cyclophosphamide, cisplatin, carmustine). Patients were grouped according to cellular support (autologous bone marrow [BM] CSF-primed peripheral blood progenitor cells [PBPCs]) and prescribed growth factor (recombinant human granulocyte or granulocyte-macrophage colony-stimulating factor [rHuG- CSF or rHuGM-CSF]). Leukocyte reconstitution was most accelerated in the groups treated with PBPCs and rHuG-CSF. IL-6, M-CSF, and TNF-alpha concentrations were higher in the groups treated with rHuGM-CSF and without PBPCs. Maximal endogenous cytokine concentrations occurred approximately 12 days after BM reinfusion. High concentrations of EPO occurred in patients experiencing significant hypotension despite routine transfusions for hematocrit < 42%. High M-CSF and IL-6 levels were associated with increased platelet transfusion requirements. Concentrations of all four cytokines were significantly higher in patients experiencing renal or hepatic toxicity, with elevations occurring in a predictable sequence and M-CSF elevations occurring first. This report shows that endogenous cytokine concentrations may be influenced by either cellular or CSF support and are associated with differences in platelet reconstitution and organ toxicity.     

Advantages of using recombinant measles viruses expressing a fluorescent reporter gene with vibratome slice technology in experimental measles neuropathogenesis

Aims: In this study of experimental measles neuropathogenesis, the utility of enhanced green fluorescent protein (EGFP) as a sensitive indicator of measles virus (MV) cell-to-cell spread in the central nervous system (CNS) has been assessed in vibratome-cut brain slices to demonstrate the degree and mechanism of viral spread in the rodent CNS. Methods: Recombinant MVs expressing EGFP were visualized at different levels in 200-μm vibratome-cut brain sections from infected animals by confocal scanning laser microscopy (CSLM). Comparison was made with 7-μm microtome sections, stained for the N protein of measles by immunocytochemistry (ICC). Results: The recombinant viruses were readily visualized in infected brain tissue, with no loss of neuropathogenicity. No difference was found in the sites of infection when MV infection was detected through EGFP fluorescence or by ICC. MV-infected cells were detected in the cerebral cortex, olfactory bulb and tract, hippocampus, thalamus, hypothalamus, ependyma and subventricular zone. However, the 200-μm vibratome-cut sections and confocal microscopy proved excellent for demonstrating virus distribution in neurites and for in-depth analysis of the extent of tract infection in the white matter of the cerebral hemispheres such as selective infection of the internal capsule and anterior commissure. Conclusions: The use of self-tracing recombinant MVs, viewed in thick vibratome-cut sections by CSLM, demonstrated that in experimental MV neuropathogenesis the infection is selective and spreads predominately by neurites using defined anatomical pathways.     

Serum, plasma and paraffin-embedded tissues as sources of DNA for studying cancer susceptibility genes

The ability to isolate DNA from archived human serum, plasma and paraffin-embedded human tissues enhances opportunities to study breast, lung and other cancer risk factors. We report herein a simple and fast protocol for the extraction of genomic DNA from these sources. Using a phenol-based extraction method, the recovery for DNA is quantitative and reproducible. DNA yields in serum (250 microl) were between 162 and 1060 ng (n = 18 subjects), in plasma (250 microl) were between 165 and 375 ng (n = 5 subjects) and in embedded tissues (5-microm thick sections for ethanol fixed, and between 5- and 20-microm sections for formaldehyde fixation) were between 1 microg and 11.7 microg (n = 32 subjects). The extraction method was combined with newly designed PCR- based assays for cancer susceptibility marker genes such as CYP1A1 (exon 7), CYP2E1 (Dra1, Rsa1), GSTM1 and NAT2 [NAT2*5A (C481T), NAT2*6A (G590A), NAT2*7A (G857A)]. Genotyping results from the serum and paraffin-embedded tissues compared favorably to results from archived freshly frozen tissues, where concordance was 98% for serum, 100% for ethanol-fixed embedded tissues, and 97% for formaldehyde-fixed and paraffin-embedded tissues. This facile method will allow for the use of archived tissue samples of prospective cohort and other studies where intact DNA was not previously available.      

Delayed rupture of renal artery after renal percutaneous transluminal angioplasty

Two cases are reported in which rupture of the renal artery occurred many hours after renal percutaneous transluminal angioplasty. Delayed rupture can be recognized by the angiographic appearance and by the presence of persistent flank pain. The typical angiographic finding is a poorly defined zone of contrast medium at the site of perforation.     

A computerized method for rapid quantification of gallbladder volume from real-time sonograms

A computerized method that requires only 1-2 minutes to quantify gallbladder volume from real-time sonograms is described. This time is considerably shorter than that required using the hand-calculation method. There was a highly significant correlation between gallbladder volumes calculated by computer and hand (r = 0.97; P less than .001).