\u3csup\u3e99m\u3c/sup\u3eTc-Labeled C2A Domain of Synaptotagmin I as a Target-Specific Molecular Probe for Noninvasive Imaging of Acute Myocardial Infarction

Abstract: The exposure of phosphatidylserine (PtdS) is a common molecular marker for both apoptosis and necrosis and enables the simultaneous detection of these distinct modes of cell death. Our aim was to develop a radiotracer based on the PtdS-binding activity of the C2A domain of synaptotagmin I and assess 99mTc-C2A-GST (GST is glutathione S-transferase) using a reperfused acute myocardial infarction (AMI) rat model. Methods: The binding of C2A-GST toward apoptosis and necrosis was validated in vitro. After labeling with 99mTc via 2-iminothiolane thiolation, radiochemical purity and radiostability were tested. Pharmacokinetics and biodistribution were studied in healthy rats. The uptake of 99mTc-C2A-GST within the area at risk was quantified by direct γ-counting, whereas nonspecific accumulation was estimated using inactivated 99mTc-C2A-GST. In vivo planar imaging of AMI in rats was performed on a γ-camera using a parallel-hole collimator. Radioactivity uptake was investigated by region-of-interest analysis, and postmortem tetrazolium staining versus autoradiography. Results: Fluorescently labeled and radiolabeled C2A-GST bound both apoptotic and necrotic cells. 99mTc-C2A-GST had a radiochemical purity of \u3e98% and remained stable. After intravenous injection, the uptake in the liver and kidneys was significant. For 99mTc-C2A-GST, radioactivity uptake in the area at risk reached between 2.40 and 2.63 %ID/g (%ID/g is percentage injected dose per gram) within 30 min and remained plateaued for at least 3 h. In comparison, with the inactivated tracer the radioactivity reached 1.06 ± 0.49 %ID/g at 30 min, followed by washout to 0.52 ± 0.23 %ID/g. In 7 of 7 rats, the infarct was clearly identifiable as focal uptake in planar images. At 3 h after injection, the infarct-to-lung ratios were 2.48 ± 0.27, 1.29 ± 0.09, and 1.46 ± 0.04 for acute-infarct rats with 99mTc-C2A-GST, sham-operated rats with 99mTc-C2A-GST, and acute-infarct rats with 99mTc-C2A-GST-NHS (NHS is N-hydroxy succinimide), respectively. The distribution of radioactivity was confirmed by autoradiography and histology. Conclusion: The C2A domain of synaptotagmin I labeled with fluorochromes or a radioisotope binds to both apoptotic and necrotic cells. Ex vivo and in vivo data indicate that, because of elevated vascular permeability, both specific binding and passive leakage contribute to the accumulation of the radiotracer in the area at risk. However, the latter component alone is insufficient to achieve detectable target-to-background ratios with in vivo planar imaging

Let-7b expression determines response to chemotherapy through the regulation of Cyclin D1 in Glioblastoma

BACKGROUND: Glioblastoma is the most common type of primary brain tumors. Cisplatin is a commonly used chemotherapeutic agent for Glioblastoma patients. Despite a consistent rate of initial responses, cisplatin treatment often develops chemoresistance, leading to therapeutic failure. Cellular resistance to cisplatin is of great concern and understanding the molecular mechanisms is an utter need. METHODS: Glioblastoma cell line U251 cells were exposed to increasing doses of cisplatin for 6 months to establish cisplatin-resistant cell line U251R. The differential miRNA expression profiles in U251 and U251R cell lines were identified by microarray analysis and confirmed by Q-PCR. MiRNA mimics were transfected into U251R cells, and cellular response to cisplatin-induced apoptosis and cell cycle distribution were examined by FACS analysis. RESULTS: U251R cells showed 3.1-fold increase in cisplatin resistance compared to its parental U251 cells. Microarray analysis identified Let-7b and other miRNAs significantly down-regulated in U251R cells compared to U251 cells. Transfection of Let-7b mimics greatly re-sensitized U251R cells to cisplatin, while transfection of other miRNAs has no effect or slightly effect. Cyclin D1 is predicted as a target of Let-7b through bioinformatics analysis. Over-expression of Let-7b mimics suppressed cyclin D1 protein expression and inhibited cyclin D1-3’-UTR luciferase activity. Knockdown of cyclin D1 expression significantly increased cisplatin-induced G1 arrest and apoptosis. CONCLUSIONS: Collectively, our results indicated that cisplatin treatment leads to Let-7b suppression, which in turn up-regulates cyclin D1 expression. Let-7b may serve as a marker of cisplatin resistance, and can enhance the therapeutic benefit of cisplatin in glioblastoma cells

126例肠道病毒71型和柯萨奇A组共感的临床特征分析*

Objective: To explore the attack, prevalence, clinical features, diagnosis, therapy and prognosis of enterovirus 71 and coxsackie A16. Methods: Analyzing the general situation, main symptoms and signs, laboratory examination, virus effect analysis and the prognosis of 126 cases of hospital medical records. Results: Among the 126 cases, CA16 is in the majority, and EV71 virus is coexisting. Besides, the ordinary cases have mild symptoms without serious complications, and attention should be paid attention to the fever, blood glucose and myocardial enzyme spectrum and complications, etc. Severe cases need to be discovered early. Conclusion: Timely detection and treatment are the effective ways to reduce the deaths of seriously ill patients lie in full attention to the severe trend.目的  探讨肠道病毒71型（EV71)和柯萨奇A组16型（CoxA16)共感的发病、流行及临床特征、诊断、治疗及预后。方法  分析126例住院病历的一般情况，主要症状及体征，实验室检查，病毒分析情况及治疗预后效果。结果  126例病例中，以CA16为主，有EV71病毒同时存在，普通病例症状轻，无严重并发症，要注意发热、血糖、心肌酶谱及合并症等。尽早发现重症病例。结论  充分重视重症趋势，及时发现，及时治疗，是目前减少重症患者死亡的有效方法