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BACKGROUND: Eosinophil cationic protein (ECP) is an eosinophil-derived protein, which has been shown to be present in circulating neutrophils. OBJECTIVE: To establish whether ECP is produced or internalized by peripheral blood neutrophils. METHODS: This was done using microscopy, flow cytometry, fractionation of cells and RT-PCR techniques. RESULTS: No ECP mRNA was detected after extensive cell purification to eliminate all traces of contaminating eosinophils. Examination of immunostained neutrophils by light, confocal, electron microscopy together with cell fraction experiments, established that ECP is present intracellularly and is mostly associated to cell granules. Uptake studies by flow cytometry and by using both cold and radiolabelled ECP showed that it is internalized by neutrophils and stored in some proportion in their primary granules. Upon stimulation with serum-treated Sephadex particles, the internalized ECP was partially released from cells. CONCLUSION: ECP is not produced but can be internalized by circulating neutrophils, which take it from the environment and partially store it in their primary granules.  相似文献   
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While current donor selection with clinical findings is generally effective, the imprecise nature of the assessment forces clinicians to remain on the conservative side. A reliable biological marker would assist donor selection and would improve donor organ utilization. We collected biopsies from 169 donor lungs before implantation. Expression levels of IL-6, IL-8, IL-10, TNF-alpha, IFN-gamma and IL-1beta were measured by quantitative real-time RT-PCR (qRT-PCR). Seventeen cases died within 30 days after transplantation. No donor factor was significantly associated with 30-day mortality. Univariate analysis of the 84 cases for development of the prediction model showed that IL-6, IL-8, TNF-alpha and IL-1beta were risk factors for mortality and IL-10 and IFN-gamma were protective factors. We analyzed the cytokine expression ratios of risk to protective cytokines. A stepwise logistic regression for 30-day mortality demonstrated that a model containing the ratio of IL-6/IL-10 was the most predictive (p = 0.0013). When applied to the remaining 85 cases for validation, the test of model fit was significant (p = 0.039). Using the cytokine ratio, we were able to define three risk groups with striking differences in survival (p = 0.0003). Multi-cytokine analysis of the donor lung graft with qRT-PCR shows significant promise as a strategy to biologically evaluate the donor lung prior to implantation.  相似文献   
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Liver transplantation, which serves as treatment of familial amyloidotic polyneuropathy (FAP), and domino liver transplantation, which utilizes resected livers from patients with FAP for treatment of liver diseases, may induce changes in transthyretin (TTR), a pathogenic FAP-related protein. To evaluate this possibility, we performed a 70% hepatectomy or administered tacrolimus to Dark Agouti (DA) rats for 7 days and then measured changes in liver TTR mRNA levels and changes in serum TTR concentrations. After hepatectomy, TTR mRNA levels decreased by 77%; at day 3, they returned to preoperative levels. Except for slightly elevated serum TTR concentrations 12 h after operation, serum TTR levels remained unchanged. Thus, partial hepatectomy did not influence serum TTR concentrations. After tacrolimus administration, TTR mRNA declined by 56% 12 h after the experiment started; however, after day 3, a rebound phenomenon occurred until day 7. Tacrolimus may facilitate serum TTR degradation, although production of TTR in the liver also increased. This finding -- that TTR, the source of FAP-inducing amyloid, did not increase after transplantation -- may help post-transplantation treatment of patients who have FAP and other liver diseases.  相似文献   
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Expression of mRNAs encoding the erythroid-specific delta-aminolevulinate synthase (ALAS-E) and the nonspecific delta-aminolevulinate synthase (ALAS-N) were examined in murine Friend virus-transformed erythroleukemia (MEL) cells using nonradioactive in situ hybridization. Following dimethyl sulfoxide (DMSO) treatment, ALAS-E mRNA increased markedly, while ALAS-N mRNA did not increase in wild-type MEL cells. In contrast, in a DMSO-resistant clone of MEL cells, ALAS-E was not detectable before and after DMSO treatment. These findings suggest that ALAS-E and ALAS-N mRNAs are under separate controls and that the expression of ALAS-E mRNA is a critical event in erythroid differentiation.  相似文献   
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目的:建立并优化SYBR GreenI实时RT-PCR体系,定量检测人、小鼠成熟精子中的CatSper1 mRNA。方法:用TRIzol分别提取人、小鼠成熟精子中的总RNA,逆转录后用SYBR GreenI实时PCR定量检测CatSper1 mRNA。SYBR GreenI实时PCR采用普通PCR试剂,加入SYBR GreenI染料,优化退火温度、Mg2+浓度及上、下游引物比例,并在PCR循环时采用四步法以消除引物二聚体的影响。优化完成后用不同浓度的精子cDNA为模板做标准曲线,以检测SYBR GreenI实时PCR的扩增效率。结果:定量检测CatSper1 mRNA的SYBR GreenI实时PCR体系适宜退火温度、Mg2+浓度及上、下游引物比例分别为63℃、3.0mmol/L和1∶1,四步法中采集荧光的温度为88℃。优化后用人和小鼠精子cDNA为模板做标准曲线分别为Y=-3.402log(X)+25.99和Y=-3.409log(X)+24.09,扩增效率分别为96.8%和96.5%,可定量检测人、小鼠成熟精子中的CatSper1 mRNA。结论:用普通的逆转录及PCR系统和试剂,建立了一种方便、廉价、可靠的SYBR GreenI实时荧光定量RT-PCR系统,可用于人、小鼠精子中CatSper1 mRNA定量检测。  相似文献   
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Differential Cellular Gene Expression in Ganglioglioma   总被引:1,自引:0,他引:1  
Summary:  Purpose:  Gangliogliomas (GGs) are neuronal-glial tumors highly associated with epilepsy. We hypothesized that the expression of select gene families including neurotransmitter receptor subunits and growth factors would be distinct in neurons and astrocytes within GG compared with adjacent cortex and that these changes would yield insights into seizure onset and lesion formation.
Methods:  Candidate gene expression was defined in single immunohistochemically labeled neurons and astrocytes microdissected from GG specimens compared with neurons and astrocytes microdissected from morphologically intact cortex adjacent to the GG or normal control cortex.
Results:  Differential expression of 16 genes including glutamate transporter (EAAC1) and receptor (NMDA2C, mGluR5), growth factor (hepatocyte growth factor), and receptor (platelet derived growth factor receptor β, fibroblast growth factor receptor 3) mRNAs was detected in GG neurons compared with control neurons. In astrocytes, altered expression of p75NGF, mGluR3, TGFβ3 and Glt-1 mRNAs was detected. Nestin mRNA, a gene that exhibits enhanced expression in balloon cell cortical dysplasia, was increased in GG neurons. Because of the morphological similarities between GG and cortical dysplasia, we show that there is activation of the mTOR cascade in GG as evidenced by enhanced expression of phospho-p70S6kinase and phosphoribosomal S6 proteins.
Conclusion:  We find differential candidate gene expression in neurons and astrocytes in GG compared with adjacent cortex and show that there is activation of the mTOR pathway. These changes highlight pathways that may be pivotal for epileptogenesis and lesion growth.  相似文献   
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