The SRY gene from buffalo (Bubalus bubalis) genome was amplified by the polymerase chain reaction ( PCR) with primers based on the sequence of Hostein SRY gene. The amplified fragment was 2005 bp include 5UTR ( 1 - 504bp) and 3'UTR(1196 - 2005bp). And the amplified fragment was cloned and sequenced. Sequence analysis showed that the coding region of SRY gene (505 - 1195bp) from buffalo was highly homologous with those of other bovine counterpart genes (96% homology) , especially in the HMG box region (99%homology). It was found that there were only signal on male buffalo genome on Southern blot,which indicate SRY gene are highly conservative on evolves.

Objective To investigate the effect of telmisartan on the oxidative stress induced by high glucose in human umbilical vein endothelial cell (HUVEC) in vitro.Methods HUVEC were cocultured with telmisartan (1?10~(-6) mol/L) and various concentration of glucose(5,30 mmol/L) for 0,12,24,36,48 h respectively. The level of MDA in the supernatants of cultured endothelial cells was measured by thiobarbituric acid test,SOD was determined by xanthine oxidase test.The protein expression of peroxisome proliferator activated receptors ? (PPAR-?) in HUVEC 24 h was assessed by Western blot after treatments.Results High glucose significantly increase the levels of MDA (before:1.2?0.06 vs after:1.6?0.1 mmol/mL,P

AIMTo investigate the effect of 8-opioid receptors in the cardioprotection elicited by ischemic postconditioning and the underlying mechanism.

METHODSThe isolated perfused hearts of male Sprague-Dawley rats were subjected to 30 min of global ischemia followed by 120 min of reperfusion. Formazan content of myocardium was measured spectrophotometrically, and the activity of lactate dehydrogenase (LDH) in the coronary effluent was measured. In isolated ventricular myocytes hypoxic postconditioning was achieved by 3 cycles of 5 min reoxygenation/5 min hypoxia starting at the beginning of reoxygenation, and cell viability was measured.

RESULTSIn the Langendorff perfused rat heart model, ischemic postconditioning (6 cycles of 10 s reperfusion/10 s global ischemia starting at the beginning of reperfusion) increased formazan content, reduced LDH release, improved the recovery of the left ventricular developed pressure, maximal rise/fall rate of left ventricular pressure and rate pressure product (left ventricular developed pressure multiplied by heart rate), attenuated the decrease of coronary flow during reperfusion and increased the isolated cell viability. Pretreatment with naltrindole, an antagonist of delta-opioid receptors and calcium-activated potassium channel (KCa) blocker paxilline attenuated the effect of ischemic/hypoxic postconditioning.

CONCLUSIONThe findings indicate that ischemic postconditioning protects myocardium against ischemia/reperfusion injury via activating delta-opioid receptors and opening KCa.

Animals ; Cell Survival ; In Vitro Techniques ; Ischemic Postconditioning ; Male ; Myocardium ; metabolism ; Myocytes, Cardiac ; cytology ; metabolism ; Potassium Channels, Calcium-Activated ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, delta ; antagonists & inhibitors

Child ; Child, Preschool ; Female ; Flow Cytometry ; methods ; Humans ; Infant ; Male ; Thrombasthenia ; classification ; diagnosis

OBJECTIVETo explore the relationship between germ cell apoptosis and the expression as well as the distribution of Sertoli cell vimentin induced by local exposure to heat.

METHODSLocal short-term exposure of prepubertal male rats testis to heat (43 degrees C for 15 min). Histochemical method was used to observe morphological characteristics of seminiferous tubule. The distribution and expression of Sertoli cell cytoskeletons were analyzed by immunohistochemistry and germ cell apoptosis was evaluated by TUNEL technique at different hour-intervals.

RESULTSAfter 2 h and 4 h heat exposure, the disattachment phenomenon between Sertoli cell and spermatogonia occurred. Spermatogonia arranged in disorder and displaced away from the basement membrane of seminiferous tubules. Immunohistochemical staining showed that vimentin positive staining was seen radiating from the Sertoli cell perinuclear region with apical "spoke-like" pattern in controls. There was an intense vimentin immunoreactivity surrounding Sertoli cell nuclei along with the collapse of the apical extensions in 2 h group, but no significant difference compared with the controls. The expressions of vimentin in 12 h and 24 h groups were higher than those of the controls (P <0.01), respectively. TUNEL showed that incidence of apoptosis was observed to increases markedly in 12 h and 24 h groups, but it was found that the incidences of apoptotic events were decreased in these two groups compared with the controls.

CONCLUSIONThe changes of expression and distribution of Sertoli cell vimentin filaments correlate with the increased germ cell apoptosis. Local heat may disrupt spermatogenesis by injuring Sertoli cell directly.

Animals ; Apoptosis ; Hot Temperature ; In Situ Nick-End Labeling ; Male ; Rats ; Rats, Sprague-Dawley ; Sertoli Cells ; metabolism ; Spermatozoa ; pathology ; Vimentin ; biosynthesis

Objective The immunotoxins generated by conjugating monoclonal antibody (mAb) and a certain toxin play an important and promising role in treating hematopoietic malignancies. However, most of the toxins used for the conjugation are toxic proteins, which are immunogenic in the patients. Norcantharidin (NCTD) is a small molecule toxin without immunogenicity, and thus has become a potential new drug for hematopoietic cancers. In this study, we prepared immunotoxin 2E8-NCTD by using the ZCH-4-2E8 cells produced in the laboratory of our hospital, and then detected its targeting effect against CD+19 lymphoid malignant Nalm-6 cells in vitro.Methods 2E8 mAb was obtained from mouse ascites and purified by gel chromatography. After its purity was checked by SDS-PAGE, immunotoxin 2E8-NCTD was generated by conjugating CD19 mAb with NCTD using activated ester method. The binding activity of the immunoconjugate to CD19 antigens on cell surface, and the expression levels of the CD19 antigens on Nalm-6 and K562 cells were determined by flow cytometry. The inhibitory effects of PBS, purified 2E8 mAb, NCTD, and immunotoxin 2E8-NCTD on the cell growth of either Nalm-6 or K562 cells were then compared.Results The purity of the 2E8 mAb was higher than 99% demonstrated by SDS-PAGE assay. 2E8 mAb was detected on the surface of 99.34% of the Nalm-6 cells, while on only 0.98% of the K562. The newly generated immunotoxin had a positive rate of 99.90% on the Nalm-6 with slightly reduced binding activity. Both 2E8-NCTD and NCTD significantly inhibited the growth of CD+19 Nalm-6 cells (P < 0. 001 ), while the purified 2E8 mAb did not show any significant influences on the growth of the same cell line ( P > 0.05 ). Meanwhile, no significant inhibitory effects on the CD-19 K562 cells were identified in the 2E8-NCTD, 2E8 mAb, or control groups, indicating a significant targeting effect of 2E8-NCTD against Nalm-6 cells.Conclusions The immunotoxin 2E8-NCTD can be synthesized by activated ester method. It has target killing effects on CD+19 Nalm-6 leukemia cells in vitro.

OBJECTIVETo investigate the early diagnosis and treatment for trauma around the knee with popliteal vascular injury.

METHODSA retrospective analysis was employed to analyze the clinical data from 15 patients (9 males and 6 females were with a mean age of 39.2 years old,ranging from 26 to 62 years old) with fracture or dislocation around the knee with popliteal vascular injury from January 2007 to January 2013. Combined with clinical symptoms and signs, oxygen saturation monitors, color ultrasound, DSA angiography and interventional surgery were used to determine the vascular injury. The knee fracture and dislocation were fixed with hybrid external fixation and plate-screw fixation, respectively. Then, the blood circulation was reconstructed by thrombectomy, repair and autologous vein graft for individual injured vascular. The average total operation time, average hospitalization days, predictive salvage index (PSI), average blood transfusion amount, average medical expenses and infection cases were recorded to determine the effect of early diagnosis and treatment.

RESULTSThere was one patient with death, 8 patients with amputation, and 6 patients with successful repair surgery for popliteal artery, anterior tibial and posterior tibial arteries. These six patients with surviving limbs were followed up for an average of 28.3 months (ranged, 12 to 60 months). Among the 6 successful patients, the joint function of 4 patients was good and excellent.

CONCLUSIONThe trauma around the knee with popliteal vascular injury is characterized by complex and serious injury, easy misdiagnosis and loss diagnosis, poor prognosis and high risk of amputation. The early diagnosis of trauma around the knee with popliteal vascular injury should depend on the mechanism of trauma, local anatomical characteristics of injury site, clinical presentations and appropriate auxiliary examinations. The appropriate indications for limb salvage and amputation should be used to achieve more effective clinical results.

Adult ; Early Diagnosis ; Female ; Humans ; Knee Injuries ; diagnosis ; surgery ; Male ; Middle Aged ; Popliteal Artery ; injuries ; surgery ; Retrospective Studies

The electrospray ionization-ion mobility spectrometric (ESI-IMS) technique has the potential as an analytical separation tool in analyzing polypeptides and amino acids for fast screening unknown samples in anti-chemical and biological terror attacks. A method for detecting several polypeptides and amino acids was developed based on ESI-IMS using air as drift gas at room temperature. The ion mobility of four amino acids and two polypeptides dissolved in methanol was determined on the system at elution rate of 2 mL/ min. The spectra of these compounds had characteristics of finger-printing maps. The limit of detection of this instrument for Substance P could reach 855 ng / mL in 1 min. The results showed that a small, self-contained ESI-IMS instrument with reservoirs of air could be used to quickly detect and accurately identify polypeptides and amino acids.

OBJECTIVETo study the effect on synthesis of nitric oxide in myocardium by local cryoablation and to investigate its mechanism.

METHODSMyocardium was cryoablated locally by a probe cooled to -60 degrees C and rewarmed by normal salt solution, nitric oxide and its synthesis enzyme were measured before and after cryoablation. L-arginine or methylene blue was added before and during cryoablation and the effect of these drugs on synthesis of nitric oxide was studied.

RESULTSNitric oxide and its synthesis enzyme decreased after cryoablation; L-arginine preserved the synthesis of nitric oxide and methylene blue inhibited the synthesis of nitric oxide. However, nitric oxide in serum did not change.

CONCLUSIONNitric oxide and its synthesis enzyme in myocardium decrease after cryoablation.

Animals ; Cryosurgery ; Myocardium ; metabolism ; pathology ; Nitric Oxide ; biosynthesis ; Nitric Oxide Synthase ; metabolism ; Rabbits

BmNPV bacmid constructed recently and Red recombinant system were used to rapidly disrupted Bombyx monri nucleopolyhedrovirus (BmNPV) orf60 in Escherichia coli (E. coli) BW25113. BmNPV bacmid isolated from E. coli BmDH10Bac was electroporated into E. coli BW25113, which harbors plasmid pKD46 encoding lamda Red recombinase,to produce E. coli BW25113-Bac, which could be used for gene deletion of BmNPV. A linear fragment was amplified by PCR from plasmid pKD3 (containing a chloramphenicol acetyltransferase gene cat) using a pair of primers with length of 63bp,which had 45 bp homologous to the orf60 gene and 18bp homologous to cat sequences. The linear fragment was electroporated into E. coli BW25113-Bac and homologous recombination occurred between the linear fragment and orf60 with the help of lamda Red recombinase. Three specific primer pairs were used to confirm the replacement of orf60 by cat gene. Western blot analysis showed that orf60 was not expressed in BmN cells infected with knockout bacmid.
Animals ; Bacteriophage lambda ; enzymology ; genetics ; Bombyx ; virology ; Electroporation ; Escherichia coli ; genetics ; metabolism ; Gene Expression ; Gene Knockout Techniques ; Genes, Viral ; genetics ; Nucleopolyhedrovirus ; enzymology ; genetics ; Open Reading Frames ; genetics ; physiology ; Recombinases ; genetics ; metabolism ; Viral Proteins ; genetics ; metabolism