As a classic fluorescent detect technique, fluorescence resonance energy transfer (FRET) has been widely used in biological researches. Researchers have developed a series of fluorescence detect probes which were based on FRET. Caspase family plays an important role in apoptosis pathway, especially Caspase-8 which located, at the initial of death receptor mediated apoptosis pathway, whose its activation can trigger subsequent precaspases' activation and lead to apoptosis. So it is of great significance to detect the activation of Caspase-8 in apoptosis assay. In this study, a fluorescent probe based on FRET has been designed which can detect the activity change of Caspase-8 in cells. To identify the effectiveness and specificity of the probe, we measure the Caspase-8 activity under the Caspase-8 specifically activated apoptosis inducer RGD-TRAIL with the flow cytometry FRET detection platform. The results show that the probe can respond to the activity change of Caspase-8 in apoptotic cells, and the change can be quantified rapidly by flow cytometry. The study provides a more efficient and convenient detection method of Caspase-8 activity in living cells.
Apoptosis ; Caspase 8 ; metabolism ; Flow Cytometry ; Fluorescence Resonance Energy Transfer ; Fluorescent Dyes ; Humans

In order to clarify material basis of effective parts of liangxue tongyu prescription, blood-heat and blood-stasis rat model induced by dry yeast was established. The changes of rectal temperature, blood viscosity and plasma viscosity were used to evaluate the cooling-blood and activating-blood effects of liangxue tongyu prescription and its parts. Compared with the model group, the extract from liangxue tongyu prescription, its volatile oil and n-butanol part could significantly reduce rectal temperature (P<0.01), and also reduce blood viscosity and plasma viscosity to various degrees (P<0.01 or P<0.05). So volatile oil and n-butanol part were primarily identified as effective parts of liangxue tongyu prescription. By using GC-MS with normalization method of area to analyze volatile oil of liangxue tongyu prescription, 70 compounds were identified, accounting for about 92.54%, mainly as β-asarone, paeonol, α-asarone and shyobunone. 42 compounds such as peony glycosides, tannins, and iridoid glycosides were identified by HPLC-MS techniques and standard comparison. The study determined the effective parts of liangxue tongyu prescription and clarified the chemical composition providing the foundation for further studies on material basis of liangxue tongyu prescription.
Acetophenones ; chemistry ; Animals ; Anisoles ; chemistry ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; Gas Chromatography-Mass Spectrometry ; Oils, Volatile ; chemistry ; Rats ; Tannins ; chemistry

Objective To evaluate the clinical application and safety of CT-guided pereutaneous transthoracic biopsy in the diagnosis of mediastinal masses.Methods Thirty three cases were undertaken CT- guided percutaneous transthoraeie biopsy with automatic biopsy gun and then the sampling specimens were undergone histological examination.The accuracy of puncture,diagnostic correctness and complications were analyzed.Results The operations were performed successfully in all 33 cases(100%),the definite pathologic diagnosis were made in 28 out of 33 cases(85%)and no complications occurred.Conclusion As for midiastinal masses,CT-guided percutaneous transthoracic biopsy is a feasible,successful,efficient interventional diagnostic method with high accuracy in localization,puncture,diagnosis and few complications, which should he recommended in clinical use more widely.(J Intervent Radiol,2007,16:852-854)

Objective To explore ability of the vpr gene of human immunodeficiency virus type 1 ( HIV-1 vpr) to induce cell G_2 arrest and apoptosis, and the influence when it mutated, the relationship between Vpr-induced G_2 arrest and apoptosis inductions. Methods Fourteen mutant vpr fragments selected from Chinese patients with HIV. Both eukaryotic expression vector pcDNA3.1( + ) and PCR products purified, double-cut by Hind Ⅲ and BamH Ⅰ and the cut products legated and transformed into competent cells JM109. The 14 reconstructed plasmids electronically transfected into Jurkat-cells, and established cells with pcDNA3. 1-vpr , pcDNA3. 1-vpr-Fs and pcDNA3. 1 blank cells, and without pcDNA3. 1 cell. Cells were harvested after 24 h. mRNA expression was detected by RT-PCR, the DNA content and percentage of apoptosis were monitored by flow cytometry. Results Transfected with 14 mutant HIV-1 Vpr protein, cells display different G_2 percentage and apoptosis ratio. HIV-1 vpr induce cell cycle G_2 arrest and apoptosis, wherase Vpr Fs with a C-terminal end truncation, vector pcDNA3.1( + ) and the blank cells can not. The G_2 percentage and apoptosis ratio reduced when transfected with vpr expressing mutating of 70V, 85P, 86G, 94G compared to the wild type. Subtype AE has a weaker potential to induce cell cycle G_2 arrest and apoptosis. Preliminary, we find that the higher G_2 percentage followed the higher ratio of apoptosis. Conclusion HIV-1 vpr can induce cell cycle G_2 arrest and apoptosis, wherase Vpr Fs with a C-terminal end truncation can not. We firstly found that mutated sites of 70V, 85P, 86G, 94G may reduce the ability of Vpr to induce cell cycle G_2 arrest and apoptosis, subtype AE of vpr in Chinese HIV-1 patients has a weaker potential to induce cell cycle G_2 arrest and apoptosis. Analysis of various mutations in the vpr gene revealed that the extent of Vpr-induced G_2 arrest correlated with the levels of apoptosis. And investigate the pathegenesis of HIV vpr. This can also make a good foundation for further study on gene therapy.

Adult ; Female ; Humans ; Ovarian Diseases ; diagnosis ; Ovarian Neoplasms ; diagnosis ; Pregnancy

OBJECTIVETo investigate the dynamic changes of expression of matrix metalloproteinases-9 in myocardium of mice with viral myocarditis (VMC) and its significance in the pathogenesis of viral myocarditis.

METHODSVMC model was prepared by an injection of CVB3 in BALB/C mice. The mice receiving an injection of culture solution without virus were used as the control group. Cardiac tissues were obtained 7, 14, 21 and 28 days after injection and made into paraffin sections. Myocardial histopathologic changes were observed by hematoxylin-eosin staining and Masson staining. The expression of MMP-9, type I collagen and type III collagen in cardiac tissues were quantified by SABC immunohistochemical method.

RESULTSThe expression of MMP-9 in the VMC model group was observed on the 7th day, reached a peak on the 14th day, and was significantly higher than that in the control group at all time points (P<0.05). Compared with the control group, the expression of type I collagen in the VMC model group was up-regulated on the 21st day and reached a peak on the 28th day (P<0.05). The expression of type III collagen in the VMC model group was significantly higher than that in the control group on the 28th day (P<0.05). The expression of MMP-9 was positively correlated with myocardial histopathologic scores (r=0.832, P<0.05) and negatively correlated with type I collagen expression (r=-0.791, P<0.05).

CONCLUSIONSMMP-9 is over-expressed at the early stage in VMC mice, and participates in the pathological process of VMC through mediating the degradation metabolism of type I collagen. It may be an important factor that leads to myocardial collagen remodeling and myocardial fibrosis.

Animals ; Collagen Type I ; analysis ; Collagen Type III ; analysis ; Coxsackievirus Infections ; enzymology ; Enterovirus B, Human ; Immunohistochemistry ; Male ; Matrix Metalloproteinase 9 ; analysis ; Mice ; Mice, Inbred BALB C ; Myocarditis ; enzymology ; pathology ; Myocardium ; enzymology ; pathology

OBJECTIVETo study the efficacy of bedside treatment by laser photocoagulation for retinopathy of prematurity (ROP) in preterm infants hospitalized in the Neonatal Intensive Care Unit (NICU).

METHODSThe clinical data of 30 cases of ROP who underwent peripheral laser ablation on bedside in the NICU from March to August 2009 were studied retrospectively.

RESULTSA total of 59 eyes from 30 patients received the laser therapy, with a total cure rate of 95%. According to the International Classification of ROP, 26 eyes of 13 infants had zone 1 disease, and 33 eyes of 17 infants had zone 2 disease. The birth gestational age and birth weight as well as corrected gestational age and corrected weight at operation in the zone 1 disease group were significantly lower than those in the zone 2 disease group. The number of laser spots in the zone 1 disease group was significantly higher than that in the zone 2 disease group. The cure rate in the zone 2 disease group (100%) was significantly higher than that in the zone 1 disease group (88%).

CONCLUSIONSLaser retinal photocoagulation on bedside in the NICU is effective for both zone 1 and zone 2 ROP. As compared with the infants with zone 2 disease, the infants with zone 1 disease may have a poor outcome.

Birth Weight ; Female ; Gestational Age ; Humans ; Infant, Newborn ; Laser Coagulation ; Male ; Retinopathy of Prematurity ; surgery

Aim To explore the role of endoplasmic reticulum stress(ERS)signaling pathway in high fat-induced cell apoptosis in human umbilical vein endo-thelial cells(HUVECs). Methods HUVECs were ex-posed to different concentrations of palmitic acid(0. 1, 0. 2,0. 4,0. 8 mmol·L - 1 )for 24 h and different time points of 0. 4 mmol·L - 1 palmitic acid(0,12, 24,48 h). Cell viability was measured by cell count-ing kit(CCK-8),and the protein expressions of ERS signaling pathway protein such as GRP78,CHOP, PERK,IRE1,ATF6 were determined by Western blot. The level of intracellular apoptosis was detected by immunofluorescence. Results HUVECs exposed to palmitic acid at 0. 4 mmol·L - 1 for 24 h showed a de-crease in their viability and an increase in the expres-sion of ERS signaling pathway proteins (GRP78, CHOP,PERK,IRE1,ATF6)(P < 0. 05);cell ap-optotic levels significantly increased(P < 0. 05). The intracellular apoptosis levels in the vascular endothelial cells of ERS signaling pathway inhibitor 4-phenylbutyr-ic acid (4-PBA,10 mmol · L - 1 )were significantly lower than those of the PA group(P < 0. 05). Conclu-sion Activated ERS signaling pathway might play an important role in the treatment of high fat-induced cell apoptosis in vascular endothelial cells.

OBJECTIVETo investigate the effect of compound qingqin liquid (CQL) on Toll-like receptor 2 (TLR2) and toll-like receptor 4 (TLR4) in rats with urate nephropathy, and to explore its renal protection mechanism.

METHODSTotally 55 SD rats were randomly divided into 5 groups, i.e., the normal control group (n =5), the model group (n =10), the positive drug group (n=10), and the high-, medium-, low-dose CQL groups (n=10) respectively. The urate nephropathy model was induced by intragastrically administering adenine and feeding yeast. Distilled water was intragastrically administered at the daily dose of 10 mL/kg to rats in the normal control group and the model group. Allopurinol was intragastrically administered at the daily dose of 9.33 mg/kg to rats in the positive control group. CQL was intragastrically administered at the daily dose of 3.77, 1.89, 0.94 g/kg to rats in the high-, medium-, and low-dose CQL groups. Rats of each group were executed in batches at the 4th and 6th week respectively. Their kidney tissues were taken out to determine the mRNA transcription level of TLR2 and TLR4 by reverse transcription-polymerase chain reaction (RT-PCR). The protein expression level of TLR2 and TLR4 were determined by Western blot. The protein expression level of TLR4 was also detected by immunohistochemical assay.

RESULTSAt week 4 and 6, the protein expression of TLR2 and TLR4 as well as the mRNA transcription of TLR4 increased in the model group, when compared with the control group (P < 0.05, P < 0.01). Compared with the model group, there was no statistical difference in the transcription level of TLR2 mRNA or TLR4 mRNA among the 3 CQL groups (P > 0.05) at week 4 and 6. Additionally, at week 6, the protein expression of TLR4 and TLR2 could be reduced by CQL (P < 0.05, P < 0.01).

CONCLUSIONCQL might protect kidney tissue against inflammatory injury by inhibiting the protein expression levels of TLR2 and TLR4.

Animals ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Kidney ; drug effects ; metabolism ; Kidney Diseases ; drug therapy ; metabolism ; Male ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Toll-Like Receptor 2 ; genetics ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism ; Uric Acid

OBJECTIVETo describe the pitfalls in positron emission tomography/computed tomography (PET/CT) imaging and classify them according to the principles of their generation.

METHODSWe summarized retrospectively the 18F-fluorodeoxyglucose (FDP) PET/CT imaging pitfalls through reviewing the PET/CT images of 872 patients. The pitfalls were divided into artifacts and infrequent physiological uptake, and the artifacts were further classified according to their causes. Meanwhile, we calculated the incidences of various pitfalls. Whether the PET/CT pitfalls influenced the diagnostic decision was analyzed. The appearances of pitfalls in PET were also described.

RESULTSPitfalls could be found in PET/CT images of 684 (78.4%) patients. Artifacts were found in 664 (76.15%) patients, and could be classified into self-factor artifacts and equipment- or technology-related artifacts. Among self-factor artifacts, respiratory motion (57.5%), postprandial or hyperglycemia artifacts (2.41%), and metal or high density matter artifacts (1.38%) were frequent. As for equipment- or technology-related factors, injection point outleakage or radiotracer contamination (13.88%) and truncation artifacts (1.83%) were most common ones. Infrequent physiological FDG uptakes, including fatty uptake, endometrial uptake, and bilateral breast feeding period uptake, were found in 20 (2.29%) patients. Among all pitfalls, the artifacts in 92 (13.4%) patients and infrequent physiological uptakes in 6 (0.88%) patients affected the diagnostic results. Artifact images in PET could be described as hot or cold area and the images of infrequent physiological uptake were always shown as hot area.

CONCLUSIONSThe incidence of pitfall in PET/CT imaging was high and the causes of pitfalls are various. Among all causes that artifacts generated, respiratory motion is the most common. Some pitfalls may disturb clinical physicians' decision, so it is important to recognize artifacts and physiological uptake, and distinguish them from pathological uptakes.

Adolescent ; Adult ; Aged ; Aged, 80 and over ; Artifacts ; Breast Feeding ; Child ; Child, Preschool ; Diagnostic Errors ; statistics & numerical data ; Drug Contamination ; Endometrium ; metabolism ; Fatty Acids ; metabolism ; Female ; Fluorodeoxyglucose F18 ; Humans ; Hyperglycemia ; Image Interpretation, Computer-Assisted ; Infant ; Injections, Intravenous ; Male ; Middle Aged ; Movement ; Positron-Emission Tomography ; Respiration ; Young Adult