Objective To explore the characteristics of dipyridamole 201 Tl myocardial perfusion imaging (MPI) SPECT in patients with dilated cardiomyopathy. Methods Thirty patients with dilated cardiomyopathy underwent pharmacological stress 201Tl MPI SPECT after intravenous infusion of dipyridamole (0. 56 mg/kg) for 4 min. The early and delayed SPECT images were acquired respectively at 10 and 240 min after 201Tl injection. The images were analyzed and reported by two or three experienced nuclear medicine physicians. Results All patients were found to have abnormal perfusion patterns at delay imaging, however 90.00% (27/30) were also abnormal at early images. Six patients had reverse redistribution. Conclusion Dipyridamole 201Tl MPI SPECT imaging may be of some value for the assessment of patients with dilated cardiomyopathy.

OBJECTIVETo construct a morphine tolerance model in primarily cultured striatal neurons, and screen the differentially expressed genes in this model using suppression subtractive hybridization (SSH).

METHODSSbtracted cDNA libraries were constructed using SSH from normal primarily cultured striatal neurons and long-term morphine treated striatal neurons (10-5 mol/L for 72 hours). To check reliability of the cell culture model, RT-PCR was performed to detect the cAMP-responsive element-binding protein (CREB) mRNA expression. The subtracted clones were prescreened by PCR. The clones containing inserted fragments from forward libraries were sequenced and submitted to GenBank for homology analysis. And the expression levels of genes of interest were confirmed by RT-PCR. Results CREB mRNA expression showed a significant increase in morphine treated striatal neurons (62.85 ± 1.98) compared with normal striatal neurons (28.43 ± 1.46, P < 0.01). Thirty-six clones containing inserted fragments were randomly chosen for sequence analysis. And the 36 clones showed homology with 19 known genes and 2 novel genes. The expression of 2 novel genes, mitochondrial carrier homolog 1 (Mtch1; 96.81 ± 2.04 vs. 44.20 ± 1.31, P < 0.01) and thymoma viral proto-oncogene 1 (Akt1; 122.10 ± 2.17 vs. 50.11 ± 2.01, P < 0.01), showed a significant increase in morphine-treated striatal neurons compared with normal striatal neurons.

CONCLUSIONSA reliable differential cDNA library of striatal neurons treated with long-term morphine is constructed. Mtch1 and Akt1 might be the candidate genes for the development of morphine tolerance.

Analgesics, Opioid ; pharmacology ; Animals ; Cells, Cultured ; Corpus Striatum ; cytology ; Drug Tolerance ; physiology ; Gene Expression Profiling ; Gene Library ; Molecular Sequence Data ; Morphine ; pharmacology ; Neurons ; cytology ; drug effects ; Nucleic Acid Hybridization ; methods ; Rats ; Rats, Wistar ; Reverse Transcriptase Polymerase Chain Reaction ; methods

To study the protective mechanism of Danhong injection on brain microvascular endothelial cells (rBMECs) injured by hypoxic. In the experiment, primary suckling mouse's rBMECs cells were collected and identified with factor VIII to establish the 4 h injury model. Meanwhile, rBMECs were given Danhong injection (25, 50, 100 mL . L-1), and the superoxide dismutase (SOD) activity and the malonyldialdehyde (MDA) level were detected by the biochemical method. Cell MMP-9, ICAM-1 and P53 mRNA expression levels were detected by RT-PCR method. Changes in cells' microscopic structure were observed by transmission electron microscope. According to the results, primary rBMECs were notably injured by hypoxia. Compared with model group, Danhong injection (50, 100 mL . L-1) could remarkably resist the injury induced by hypoxic, increase intracellular SOD activity, decrease MDA level and significantly down-regulate ICAM-1, MMP-9 and P53 mRNA expressions. Danhong injection (100 mL . L-1) could protect the cells' normal morphology and microscopic structure, maintain the close intercellular junction, and inhibit the hypoxia-induced cell apoptosis. The results showed that Danhong injection plays a significant role in protecting rBMECs injured by hypoxia. Its mechanism may be related to the enhancement of cells' antioxidant capacity, the inhibition of inflammatory response and the cell apoptosis.
Animals ; Antioxidants ; metabolism ; Apoptosis ; drug effects ; Brain ; metabolism ; Cell Hypoxia ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Endothelial Cells ; ultrastructure ; Female ; Gene Expression Regulation ; drug effects ; Humans ; Injections ; Intercellular Adhesion Molecule-1 ; metabolism ; Male ; Malondialdehyde ; metabolism ; Matrix Metalloproteinase 9 ; genetics ; Protective Agents ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase ; metabolism ; Tumor Suppressor Protein p53 ; genetics

To study the protective effect of combined administration of active ingredients of Danhong on cultured primary mice's brain microvascular endothelial cells (rBMECs) injured by hypoxia. Primary mice's brain micro-vascular endothelial cells were cultured to establish the 4 h hypoxia model. Meanwhile, active ingredients (protocatechuic aldehyde, salvianolic acid B, hydroxysafflor yellow A and tanshinol) of Danhong were administered in rBMECs. The non-toxic dosage was determined by MTT. The leakage of lactate dehydrogenase(LDH), cell superoxide dismutase (SOD) activity and MDA level were detected by the colorimetric method. The expressions of ICAM-1, MMP-9, P53 mRNA were detected by RT-PCR method. Changes in rBMECs cell cycle and early apoptosis were detected by flow cytometry. Danhong's active ingredients and prescriptions 1, 2, 3, 7, 8, 9 could be combined to significantly restrain LDH in hypoxic cells supernatant. Prescriptions 1, 2, 3, 7, 8, 9 could significantly enhance SOD activity in anoxic cells; Prescriptions 1, 2, 3, 8, 9 could significantly decrease the MDA level; Prescriptions 1, 2, 6, 7, 9 could significantly inhibit the early rB-MECs apoptosis induced by hypoxia. After hypoxia, the up-regulated P53 mRNA expression could cause retardation in G, phase and promote cell apoptosis. This proved that the regulatory function of P53 gene lay in monitoring of calibration points in G, phase. Prescriptions 1, 2, 5, 6, 7, 8, 9 could significantly down-regulate the P53 mRNA expression; Prescriptions 1, 4, 7, 8, 9 could significantly down-regulate the ICAM-1 mRNA expression; Prescriptions 1, 3, 6, 9 could significantly down-regulate the MMP-9 mRNA expression. The combined administration of Danhong's active ingredients showed a significant protective effect on primary cultured rBMECs injury induced by hypoxia Its mechanism may be related to the enhancement of cellular antioxidant capacity and the inhibition of inflammatory response and cell apoptosis. This study could provide ideas for researching prescription compatibility, and guide the clinical medication.
Animals ; Apoptosis ; drug effects ; Brain ; drug effects ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Endothelial Cells ; drug effects ; Hypoxia ; drug therapy ; Microvessels ; drug effects ; Rats ; Rats, Sprague-Dawley

To investigate the cardioprotective effect of formononetin (FMN) on no-reflow (NR) after myocardial ischemia-reperfusion and its molecular mechanism based on integrated pharmacology and experimental verification, firstly, human breast cancer MCF-7 cells and myocardial NR rats were used to confirm the estrogenic activity and the effect of alleviating NR of FMN, respectively. Male SD rats were divided into Sham, NR, FMN (20 mg·kg^-1) and sodium nitroprusside (SNP, 5.0 mg·kg^-1) groups, which were administered once a day for one week, the experiment was approved by the Ethics Committee of Tianjin University of Traditional Chinese Medicine (TCM-LAEC2019095). The pharmacological analysis and in vivo study of NR rats were integrated to reveal the mechanism of FMN improving NR. The results showed that FMN had estrogenic effect and reduced NR by improving cardiac structure and function, reducing NR, ischemic myocardial area and pathological injury of cardiomyocytes. Integrated pharmacology predicts that the mechanism of FMN improving NR is mainly related to phosphatidyinositol-3-kinase-protein kinase B (PI3K-Akt) signal pathway. Phytoestrogens play a role in cardiovascular protection mainly by activating G protein-coupled estrogen receptor (GPER). GPER is also an important regulator in the upstream of PI3K-Akt signaling pathway. This study found that FMN can significantly activate GPER, p-PI3K, p-Akt and phospho-endothelial nitric oxide synthase (p-eNOS). It has good binding ability with GPER and eNOS protein. In this study, through the integration of pharmacology and experimental evaluation, it is revealed that FMN activates PI3K/Akt/eNOS signal pathway by activating GPER, thus significantly improving NR.

italic>Helicobacter pylori (H. pylori) can cause a variety of digestive tract diseases, the serious may develop into gastric cancer. Nowadays, H. pylori infection rate exceeds 50%, and its eradication rate is declining due to the continuous increase of drug resistance, leading to the occurrence of plenty of stubborn infections, which seriously threaten human health. At present, it is difficult to achieve satisfactory curative effect by increasing the types of antibiotics combination or increasing their dose. In this review, the clinical treatments of H. pylori were introduced. Proceed from the characteristics and pathological background of H. pylori infection that makes H. pylori difficult to eradicate, the research advances of drug delivery strategies for improving H. pylori eradication rate were reviewed, such as strategies that could increase drug concentration in stomach (e.g. drug delivery systems with gastric acid-stabilized ability), increase drug concentration in H. pylori colonization sites (e.g. drug delivery systems with gastric retention or H. pylori targeted abilities), overcome H. pylori resistance (metal nanoparticles, anti-biofilm delivery systems), enhance host immune response (vaccine preparation) and so on. Novel drug delivery systems, such as cell membrane coating technology and phage therapy, are comparatively rare in the field of anti-H. pylori, but have broad application prospects. This review would provide reference for the development and application of therapeutic strategies to improve H. pylori eradication rate.

Objective To assess the protection against cisplatin-induced ototoxicity by adenovirus-mediated overexpression of the bcl-2 gene in cultured spiral ganglion cells(SGC).Methods SGC from P3 rats were cultured in vitro and exposed to adenovirus vector carrying green fluorescent protein gene(Ad-GFP),followed by immunocytochemical analysis for expression of the neuron-specific marker Neurofilament200(NF200)and detection under laser scanning confocal fluorescence microscope. Then, SGC were transduced by Ad-bel-2 and the expression of human bcl-2 protein was evaluated by Western Blot. Finally, the cultures of SGC were divided into 4 groups: the group of Ad-bcl-2 transfection followed by cisplatin treatment, the group of Ad-GFP transfection followed by cisplatin treatment, the group of cisplatin treatmentonly and the untreated group. Cisplatin worked for 48 hours at a concentration of 2μg/ml. Outcome measures included survival humber of SGC and longest neurite length by using ImageJ software. Results SGC were cultured successfully in vitro and transfected by adenovirus vector safely and efficiently. By Western Blot, human bcl-2 protein was expressed in the group after exposure to Ad-bcl-2,but not in the Ad-GFP transfected SGC. Cisplatin exposure resulted in shrinking of neuritis and pyknosis of cell body, even cell death. Expression of bel-2 in the SGC provided a significant level of protection against cisplatin-induced SGC degeneration. Conclusions Our results suggest that SGC can be transduced by adenovirus vector safely and efficiently in vitro. Adenovirus-mediated delivery of the bcl-2 gene attenuates cisplatin-induced SGC degeneration.

BACKGROUNDSeveral candidate genes of ossification of the posterior longitudinal ligament (OPLL) susceptibility have been identified, but their polymorphisms account for only a small percent of the total variance. Bone morphogenetic protein-4 (BMP4) is a potent ectopic ossification inducing factor. BMP4 protein and mRNA are present in cells from OPLL patients, but not non-OPLL controls. A single nucleotide polymorphism of 6007C>T(rs17563) of BMP4 has been reported to affect bone density in postmenopausal women. Thus, BMP4 may function in OPLL development. Appropriately, the relationship between BMP4 polymorphisms and OPLL was investigated.

METHODSA case-control association study investigated the genetic etiology in 179 OPLL patients and 298 non-OPLL controls. Extent of OPLL was analyzed by radiologic examinations. Whether single nucleotide polymorphism (SNP) of -5826G>A(rs1957860) 5' of the transcription start site and 6007C>T(rs17563) in exon 4 of the BMP4 gene were statistically associated with genetic susceptibility to OPLL in Chinese Han subjects was assessed.

RESULTSA significant statistical difference in genotype of 6007C>T polymorphism between male OPLL patients and male controls was evident, and the frequency of "TT" genotype in male OPLL patients was significantly higher than in male controls (P = 0.039). The frequency of the "T" allele was also significantly higher in male OPLL subjects than in male controls (P = 0.014, OR = 1.57). A significant difference was also observed between the 6007C>T polymorphism and the number of ossified cervical vertebrae in OPLL patients, while no statistical difference was apparent between the -5826G>A polymorphism and OPLL occurrence.

CONCLUSIONSThe T allele in the 6007C>T polymorphism may be a risk factor for male Han Chinese with ossification of the posterior longitudinal ligament in the cervical spine. Chinese Han male patients with CT and TT 6007C>T genotypes have a genetic susceptibility to OPLL and more extensive OPLL in the cervical spine.

Alleles ; Asian Continental Ancestry Group ; genetics ; Bone Morphogenetic Protein 4 ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Male ; Middle Aged ; Ossification of Posterior Longitudinal Ligament ; genetics ; Polymorphism, Single Nucleotide ; genetics

OBJECTIVETo explore the prediction of the site for microsurgical vasoepididymostomy (VE) in the treatment of epididymal obstructive azoospermia (OA).

METHODSThis study involved 56 infertile men with confirmed OA whose obstruction was suspected to be in the epididymis. Based on their medical history and results of preoperative physical examination and ultrasonography, we predicted the sites for VE. We performed surgical scrotal exploration for the status of epididymal obstruction, conducted palpation and microscopic observation for the epididymal tubules to be anastomosed, and finally decided on the sites for VE by making sure of the presence of motile sperm in the epididymal fluid of the patients. After surgery, we followed up the patients for the rate of pregnancy.

RESULTSAll the patients received bilateral scrotal ultrasonography and surgical scrotal exploration, totaling 112 procedures, including 98 VE procedures. The accuracy rate of the predicted sites for VE was 80.5% (153/190) by medical history and physical examination, 80.3% (90/112) based on the results of ultrasonography, and 87.4% (90/103) according to the first selected epididymal tubules. Of the 28 patients followed up for more than 12 months, motile sperm were found in 19 (67.9% ) at 2 to 12 months and spontaneous pregnancies were achieved in 10 (35.7%), all with the anastomotic sites in the corpus or cauda.

CONCLUSIONMedical history and physical examination contribute to the selection of anastomotic sites and non-invasive scrotal ultrasonography is effective and practical for positioning epididymal obstruction. The epididymal tubules with motile sperm for anastomosis could be easily obtained from the most dilated ones in indurated epididymides.

Azoospermia ; surgery ; Body Fluids ; Epididymis ; diagnostic imaging ; surgery ; Female ; Humans ; Male ; Microsurgery ; methods ; Pregnancy ; Pregnancy Rate ; Scrotum ; diagnostic imaging ; Ultrasonography ; Vas Deferens ; diagnostic imaging ; surgery

OBJECTIVETo analyze the transcription activation and possible regulation mechanism of human X-box binding protein 1(XBP1)gene 5'upstream DNA sequence in different cell lines.

METHODSSix kinds of XBP1 promoter deletion mutants were cloned into pGEM-Teasy vector, which included XBP1 gene 5' upstream -1039 to 66 bp,-859 to 66 bp,-623 to 66 bp,-351 to 66 bp,-227 to 66 bp,-227 to -45 bp respectively. Every deletion mutant sequence was cut from Teasy-XBP1p by KpnI and Xho I, and subcloned into pCAT3-Basic to produce a set of constructs termed as p1-XBP1p, p2-XBP1p, p3-XBP1p, p4-XBP1p, p5-XBP1p, p6-XBP1p, respectively. The transcription activity of each construct was detected after transiently transfecting K562, HepG2,NIH-3T3 and L0(2)cell with FuGENE 6 transfection reagent. Cells transfected by pCAT3-Basic or pCAT3-Promoter were used as negative and positive controls. The activity of chloramphenicol acetyltransferase(CAT), which reflects the transcription activation of the XBP1 gene promoter, was detected by ELISA after 48 hours of transfection.

RESULTSThe reporter vectors of six kinds of XBP1 promoter deletion mutants were successfully constructed, as confirmed by restriction enzyme digestion and sequencing. The activities of p4-XBP1p and p5-XBP1p were higher than the other deletion mutants in K562 and HepG2. And the activity of p5-XBP1p was the highest in HepG2. There was no activity detected from any transfected NIH-3T3.

CONCLUSIONThe XBP1 gene promoter can transactivate its downstream gene to transcription. The core sequence of XBP1 promoter was implied between -227 bp and 66 bp. This sequence was connected with the transcriptional activity of XBP1 promoter closely. Its transcription activity varies with different cell lines. XBP1 promoter might drive gene expression with cell-type specificity.

3T3 Cells ; 5' Flanking Region ; genetics ; Animals ; Base Sequence ; Cell Line ; Chloramphenicol O-Acetyltransferase ; metabolism ; DNA ; analysis ; DNA-Binding Proteins ; genetics ; Gene Deletion ; Gene Expression Regulation ; physiology ; Genes, Reporter ; Humans ; K562 Cells ; Mice ; Molecular Sequence Data ; Nuclear Proteins ; genetics ; Promoter Regions, Genetic ; genetics ; Regulatory Factor X Transcription Factors ; Transcription Factors ; Transcription, Genetic ; physiology ; Transcriptional Activation ; Transfection ; Tumor Cells, Cultured ; X-Box Binding Protein 1