Effects of dizocipine maleate on mitochondrial ultramicrostructure in neurons following traumatic brain injury in neonatal rats A quantitative time-course analysis

BACKGROUND： The effects of N-methyl-D-aspartic acid （NMDA） receptor antagonist on neurodegeneration in the immature brain following traumatic brain injury （TBI） are still widely unknown. OBJECTIVE： To study the effects of dizocipine maleate （MK-801）, a non-competitive NMDA receptor antagonist, on mitochondrial ultramicrostructure of neurons in the ipsilateral cingulate cortex and hippocampus after TBI in neonatal rats, and to analyze the optimal time interval of MK-801 administration （1 mg/kg）. DESIGN： Completely randomized controlled study. SETTING： Shanghai Jiao Tong University. MATERIALS： Eight 7-day-old neonatal SD rats, irrespective of gender, were provided by Experimental Animal Center, Medical College of Fudan University. The experiment was approved by a local ethics committee. MK-801 was provided by Sigma. A CM-120 transmission electron microscope （Philips, Holland） was used for tissue analysis. METHODS： This study was performed at the Departments of Anatomy, Neuromorphology, and Biophysics, Medical College of Shanghai, Jiaotong University, between October 2006 and January 2007. Focal models of contusion and laceration of brain were established by the free-falling impact method. Eight rats were randomly divided into a normal control group （n = 2 ） and a MK-801 group （n = 6）. Rats in the normal control group did not receive model establishment and administration, and they were only analyzed by an electron microscope. In the MK-801 group, the cingulate cortex was damaged using a contusion device. MK-801 （1 mg/kg） was intraperitoneally injected 30 minutes before lesion, immediately after lesion, and 30 minutes after lesion （n = 2 for each time point）.MAIN OUTCOME MEASURES： The cingulate cortex and hippocampal tissues from the injured side were removed 24 hours after lesion and routinely processed for analysis of neuronal ultramicrostructure using transmission electron microscopy. RESULTS： Differential therapeutic effects of MK-801 （1 mg/kg） at distinct administration time points： thirty minutes before lesion, the shape of cortical and hippocampal neurons was similar to that observed during excitotoxicity-induced cell death. Organelles were enlarged, the nuclear membrane of cortical neurons was complete with gear wheel-like changes, and the nuclear chromatin was irregularly aggregated around the edge. When MK-801 was applied 30 minutes after lesion, the cingulate cortex contained apoptotic neurons in early and late stages. The nuclear membrane of hippocampal neurons displayed incisures. The chromatin shape was not similar to necrosis in an early stage. Immediate administration of MK-801 after lesion slightly altered the neuronal architecture, such that mitochondria were enlarged. The neuronal shape in the control group was normal. Effects of immediate administration of MK-801 on mitochondrial injury following TBI were that the mitochondria in cortical and hippocampal neurons were damaged to a certain degree in the MK-801 group. Mitochondrial injury was reversible, when MK-801 was applied 30 minutes before lesion and immediately after lesion. Application 30 minutes after lesion produced irreversible changes. In addition, mitochondrial injury occurred earlier than other organelle and nuclear changes. CONCLUSION： Mitochondrial injury occurs earlier than other organelle and nuclear changes. Early administration of MK-801 （1 mg/kg） can prevent or reduce necrosis following TBI, decrease the degree of neuronal injury, and protect nerve cells.

Autoantibodies against MMP-7 as a novel diagnostic biomarker in esophageal squamous cell carcinoma

AIM： To evaluate the diagnostic values of serum autoan tibodies against matrix metalloproteinase-7 （MMP-7） in patients with esophageal squamous cell carcinoma （ESCC）. METHODS： The MMP-7 cDNA was cloned from ESCC tissues, and MMP-7 was expressed and purified from a prokaryotic system. MMP-7 autoanUbodies were then measured in sera from 50 patients with primary ESCC and 58 risk-matched controls, using a reverse capture enzyme-linked immunosorbent assay （ELISA） in which autoantibodies to MMP-7 bound to the purified MMP-7 proteins. In addition, MMP-7 autoantibody levels in sera from 38 gastric cancer patients and from control serum samples were also tested. RESULTS： The optimum conditions for recombinant MMP-7 protein expression were determined as 0.04 mmol/L Isopropyl-13-D-Thiogalactopyranoside （IPTG）induction at 37℃ for four hours. The levels of serum autoantibodies against MMP-7 were significantly higher in patients with ESCC than in the matched-control samples （OD450 = 1.69 ±0.08 vs OD450 = 1.55 ± 0.10, P 〈 0.001）. The area under the receiver operating character- istic （ROC） curve was 0.87. The sensitivity and specificity for detection of ESCC were 78.0% and 81.0%, respectively, when the OD450 value was greater than 1.65. Although the levels of autoantibodies against MMP-7 were also significantly higher in patients with gastric cancer compared to control samples （OD450 = 1.62± 0.06 vs OD450 = 1.55±0.10, P 〈 0.001）, the diagnostic accuracy was less significant than in ESCC patients. The area of ROC curve was 0.75, whereas the sensitivity and specificity were 60.5% and 71.7%, respectively, when the cut-off value of OD450 was set at 1.60. CONCLUSION： Serum autoantibody levels of MMP-7 may be a good diagnostic biomarker for esophageal squamous cell carcinoma,

Biological function of a novel gene overexpressed in human hepatocellular carcinoma

Obejctive To clone the full length of a differentially expressed cDNA fragment, LC27, and study its biological function tentatively Methods Northern blot was used to analyze the expression pattern of LC27 in hepatocellular carcinoma, matched nontumor liver tissues, fetal liver and normal adult liver tissues, as well as BEL 7402 hepatocellular carcinoma cell line ESTs splicing and 5' rapid amplification of cDNA ends (5' RACE) were used to clone the full length of LC27 cDNA An antisense oligodeoxynucleotide approach was used to investigate the biological role of the gene in the proliferation of BEL 7402 cells Results A 2186?bp novel cDNA with an open reading frame encoding a 283 amino acid protein was cloned Analysis of the deduced amino acid sequence indicated that it is 38% (88/229) identical to human Golgi 4 transmembrane spanning transporter MTP The gene and the encoded protein was termed hepatocellular carcinoma overexpressed transmembrane protein (hotp) and HOTP, respectively Hotp mRNA was almost undetectable in normal adult liver and fetal liver tissues However, it was significantly up regulated in hepatocellular carcinoma and some matched nontumor liver tissues, as well as BEL 7402 cells The proliferation of BEL 7402 cells was suppressed by an antisense oligodeoxynucleotide against hotp mRNA at a concentration of 50?μg/ml Conclusion HOTP may be an integral membrane transporter protein The overexpression of the gene in hepatocellular carcinoma may play an important role in hepatocarcinogenesis and disease progression

人热休克转录因子1功能反应与其半胱氨酸巯基氧化还原性能的相关性

目的 评价半胱氨酸巯基氧化还原介导剂对人热休克转录因子 1(hHSF1)的氧化还原状态、结构和功能的作用 ,并试图去解释氧化作用在细胞衰老中对hHSF1转录反应的影响。 方法 通过改良的Westernblot方法检测不同氧化还原介导剂对hHSF1构象的影响 ;通过凝胶电泳迁移率变动试验检测hHSF1的DNA的结合活性。 结果 促进二硫键交联的氧化型介导剂二酰胺(DM )、一氧化氮 (NO+ )载体亚硝基谷胱甘肽 (GSNO)与含hHSF1的HeLa细胞S10 0提取液预培育 ,能在体外形成一个致密的、电泳中移动快速、分子内二硫键交联的氧化型hHSF1(ox hHSF1)单体构象 ;在电泳前加入还原型介导剂二硫苏糖醇 (DTT)能完全逆转。由于ox hHSF1形成 ,使hHSF1在体外不能被热诱导激活形成三体 ,失去和DNA结合的活性 ,而且这个作用仅对含半胱氨酸巯基的转录因子有特异性。 结论 hHSF1功能反应与其半胱氨酸巯基的氧化还原化学性能相关 ,氧化作用和hHSF1分子内二硫键交联有可能是衰老细胞热休克转录反应呈渐减性的原因。