Objective To evaluate the influential factors of iron acquisition during Francisella tularensis LVS infection of mouse macrophages.Methods F.tularensis LVS expressing green fluorescent protein was used to infect murine macrophage J774A.1 cells.Transferrin receptor 1(Tfr1)was detected with mono-antibody and visualized with a goat-anti mouse IgG conjugated to Alexa 594.The expression profile of 5 iron metabolism related genes of J774A.1 murine macrophages uninfected or infected with F.tularensis LVS was determined with real-time PCR.Immunoblot analysis was used to compare the Tfr1 expression of live Francisella infected macrophage with dead bacteria.Tfr1 knock-off in J774A.1 cells was performed with siRNA.The transfected cells were infected with Francisella for immunoblotting and microscopy and infection assay.Results It was revealed that the live vaccine strain of F.tularensis induced the expression of Tfr1 in host macrophages.Gene expression analysis indicated that F.tularensis LVS drove an active iron acquisition program with induction of Tfr1 and iron regulatory proteins(Irp1 and Irp2).It was shown by Western-blotting that the siRNA-Tfrc-1 could knock off about 75% of Tfr1 in J774A.1 cells.It was determined by infection assay that,Tfr1 was knocked off,the bacteria number at 1h infection with Francisella was not different from that of control(F=1.06,P=0.326 5),while it was decreased significantly after 24h of infection(F=24.12,P=0.000 6).Conclusions It is demonstrated that upregulation of the Tfr1 may be mediated by post-transcriptional regulation during early infection,but sustained later through increased expression of Irp 1 and Irp 2.Increased expression of Tfr1 expands the intracellular iron pool through transferrin-mediated delivery and may thus be readily available for uptaking by Francisella.Knocking off the expression of Tfr1 does not affect bacterial invasion.Francisella,however,may fail to proliferate in macrophages in which the expression of transferrin receptor has been suppressed.

OBJECTIVETo study clinic therapeutic effect about reconstruction of severe orbital and cul-de-sac deformity after the radiotherapy with transcranial orbitotomy advancement combining cascade free flap both dorsum pedis flap and anterior tibial fascial flap.

METHODSFive cases was subjected to orbital and cut-de-sac severe deformities after both operation and radiotherapy because of retinoblastoma. The technique included transcranial orbital advancement by anterior orbital osteotomy and rigid fixed with titanic plate by coronal incision, and meanwhile incising the cul-de-sac which would be extended circumference around the central incision separation, and then designing extent of cascade flap consisted of dorsum pedis flap and anterior tibial fascial flap according to the size of cul-de-sac defect and extent of temporal depression. Then, the aforementioned two parts of cascade flap were transplanted into cul-de-sac and temple respectively. There is either the superficial temporal artery and vein or facial artery and jugular vein to chose vascular anastomosis.

RESULTSAll flaps survived. After 3 to 6 months following up, the results showed satisfactory orbital contour and temporal depression improved significantly in all cases. After the conjunctival sac were fixiformed with prefabricated eye prosthesis mode about 3 months. 3 cases have good appearance with wearing eye prosthesis and the other 2 cases' appearance is poor. One of the poor appearance cases, with depressed eye socket, have orbital implant underlying conjunctival sac in secondary operation. The other one, with swallowed inferior fornix, is transplanted autogenous hard palatal mucosa into inferior fornix in secondary operation. In addition, delayed healing in donor site of dorsum pedis occurred in one of the 4 cases.

CONCLUSIONSIt is a reliable procedure about reconstruction of severe orbital and cul-de-sac deformity after both the operation and radiotherapy with transcranial orbitotomy advancement combining cascade free flap both dorsum pedis flap and anterior tibial fascial flap. All deformity was corrected by one staged procedure which lessen patient's suffering and shorten patient's hospital stay and spare patient's costs.

Adolescent ; Adult ; Female ; Humans ; Male ; Orbital Diseases ; etiology ; surgery ; Osteotomy ; Radiotherapy ; adverse effects ; Reconstructive Surgical Procedures ; methods ; Retinoblastoma ; radiotherapy ; surgery ; Surgical Flaps ; Young Adult

OBJECTIVETo study the feasibility of the modified osteotomy of transcranial orbitotomy in the treatment of intraorbital tumor.

METHODSWe treated 8 patients with intraorbital tumor during six years. By the bicoronary incision, all cases underwent double bone flap osteotomy on the frontal bone: the superior orbital rim bone flap and roof flap instead of single fronto-orbital bone flap in the conventional transcranial orbitotomy. After removal of bone flaps, intracranial and intraorbital operation was performed. Then, two bone flaps were reduced respectively and fixated with titanic micro-plates and nails. At last, the scalp flap was sutured.

RESULTSThe operative field was very well exposed. It was found that the retrobulbar tumor was located at the superolateral, median and superonasal area respectively, which was coincided with the preoperative CT and MRI. The tumor included adenocarcinoma of the lacrimal gland, neurinoma, meningioma, and cavernous hemangioma. Four patients had blood transfusion during the operation. No other postoperative complications happened except 3 cases of diplopia and one case of blindness. After 3 to 6 months follow up, diplopia of the 3 cases gradually disappeared. Of the 8 cases, 6 reached the same visual acuity as the preoperative state. One decreased visual acuity and one lost light perception. There was well-balanced fronto-orbital appearance and eyeball position compared with the healthy side in all cases.

CONCLUSIONSThis surgical method for the treatment of intraorbital tumor is safe with well-exposed operative field. It has advantages not only in simpleness and less trauma, but also in keeping orbital roof and anterior fossa intact and decreasing complications.

Adult ; Craniotomy ; methods ; Exophthalmos ; surgery ; Feasibility Studies ; Female ; Humans ; Male ; Orbit ; surgery ; Orbital Neoplasms ; surgery ; Surgical Flaps ; Young Adult

OBJECTIVETo demonstrate the ablation of segmental glissonian pedicle by ultrasound-guided radiofrequency in precision liver resection in porcine model.

METHODSThe study was carried out on 6 Bama miniature pigs. Coagulative desiccation of the vessels feeding the segment to be resected was induced by introduction of a "cooled-tip" radiofrequency electrode under intraoperative ultrasound guidance at the level of the vessels. Then, an area of marked discoloration on the surface of the liver became obvious. Liver parenchymal transection followed without any form of hepatic inflow occlusion. Three segments (segment 3, 5, and 6) were resected on each pig. The criteria studied the local and general tolerance, the change of blood flow parameters of each segmental arterial and portal branches, and the microscopic appearance of the segmental arterial, portal and biliary tract branches. Using t test as the statistical methods.

RESULTSPostoperative mortality and morbidity were 0 and all serum parameters had normalized by 14 days. Demarcation area was obtained in 15 segments. The flow rate of the resected segmental portal branches was 0 cm/s after radiofrequency and the value of the arterial branches was (28 ± 18) cm/s which was significantly lower than pre-radiofrequency, which were (10.2 ± 3.0) cm/s and (60 ± 17) cm/s (t = 14.439, 5.502; P = 0.000). There was no significant difference in the flow rate of the remnant segmental arterial and portal branches among pre-radiofrequency, post-radiofrequency and 14 days after operation. On microscopic observation, the mean ablated length of segmental portal, arterial and biliary tract branches were (1.6 ± 0.4) cm, (1.2 ± 0.4) cm, and (1.7 ± 0.3) cm respectively.

CONCLUSIONSThe ablation of the segmental glissonian pedicle by ultrasound-guided radiofrequency is quick, safe and effective. The probe needs to be away from vessels feeding segments 1.5 cm at least to avoid thermal injuries.

Animals ; Catheter Ablation ; methods ; Female ; Hepatectomy ; methods ; Male ; Swine ; Swine, Miniature ; Ultrasonography

BACKGROUND: The poor retention and survival of donor cells implanted into the myocardium limit the efficacy of cell therapy for myocardial infarction. Embedding cells in natural or synthetic biomaterials is a strategy to address this issue. OBJECTIVE: To explore the effects of bone marrow mesenchymal stem cells (BMSCs) encapsulated in hyaluronic acid (HA) hydrogel on cardiac function after myocardial infarction. METHODS:BMSCs from male Sprague-Dawley rats were isolated and cultured,and then HA-encapsulated BMSCs were cultured in vitro in the three-dimensional manner. A model of myocardial infarction was made by cutting the anterior descending artery of female Sprague-Dawley rats. After 1 week, the model rats were screened by ultrasonic testing and then eligible ones were randomly divided into four groups: PBS group (n=8), HA group (n=8), BMSCs group (n=29), and HA-encapsulated BMSCs group (n=29). At 1 week after modeling, the model rats underwent the secondary thoracotomy and the implants were injected into the marginal zone and infarcted region in corresponding groups. The survival rate and apoptosis of implanted cells were examined at post-injection day 1, week 1 and week 2 by RT-PCR and TUNEL respectively. At post-injection week 4, changes of cardiac microstructure and function were evaluated by histological examination and echocardiography. RESULTS AND CONCLUSION: Compared with the BMSCs group, HA hydrogel significantly enhanced the survival rate and reduced the apoptotic rate of BMSCs at post-injection day 1 and week 2 (both P < 0.05). At post-injection week 4, the HA+BMSCs combined treatment yielded the best recovery of cardiac function (P < 0.05). To conclude, HA hydrogel can act as a vehicle for BMSCs delivery and improve the beneficial effects of implanted BMSCs in early myocardial repair(within 2 weeks after infarction)via enhancing cell retention and survival.

Aged ; Female ; Humans ; Hypertriglyceridemia ; blood ; epidemiology ; Male ; Middle Aged ; Prevalence ; Uric Acid ; blood

OBJECTIVETo investigate the influence of the amount of portal blood stasis removal on endotoxemia and liver function after liver transplantation.

METHODSForty-seven patients who received liver transplantation from February 2006 to November 2007 were divided into 2 groups according to the amount of portal blood stasis removal during operation: group A (n = 26) 50 ml and group B (n = 21) 200 ml of portal blood stasis removal respectively. The levels of plasma endotoxin, D-lactate, tumor necrosis factor-alpha, interleukin-6, liver function and blood coagulation were examined and analyzed.

RESULTSUnder the condition of no significant difference in sex, age, primary liver diseases and Child-pugh's classification, cold ischemic time, total operation and anhepatic time, operation methods, volume of blood loss and transfusion, and all preoperative observations. Most of observations showed the restoration of the patients in group B was better than that in group A. The plasma levels of endotoxin, D-lactate, tumor necrosis factor-alpha, interleukin-6, alanine aminotransferase, aspartate aminotransferase, prothrombin time and activated partial thromboplastin time in group B were significantly lower than those in group A (P < 0.05). The level of plasma prealbumin in group B was significantly higher than that in group A (P < 0.05).

CONCLUSIONSThe removal of 200 ml portal blood stasis leads to a better results than that of 50 ml, and it can help alleviate endotoxemia and facilitate the restoration of the liver function after liver transplantation.

Adult ; Aged ; Bloodletting ; methods ; Endotoxemia ; prevention & control ; Female ; Humans ; Liver ; physiopathology ; Liver Transplantation ; Male ; Middle Aged ; Portal Vein ; surgery ; Postoperative Complications ; prevention & control ; Reperfusion Injury ; prevention & control

OBJECTIVETo investigate the effect and mechanism of portal blood stasis on lung and renal injury induced by hepatic ischemia reperfusion.

METHODSA rabbit hepatic ischemia reperfusion injury model was established by hepatic portal occlusion and in situ hypothermic irrigation for 30 min. Twenty-four New Zealand white rabbits were employed and randomly divided into 3 groups equally by different dosage of portal blood stasis removal: group A5 (5 ml blood removal), group A10 (10 ml blood removal),and group B (no blood removal). Eight rabbits were served as controls with no hepatic portal occlusion and hypothermic irrigation. After reperfusion 4 h serum endotoxin content, tumor necrosis factor-alpha (TNF-alpha), urea nitrogen (BUN), and creatinine (Cr) were examined respectively, meantime lung and kidney tissues were sampled to determine the content of malondialdehyde (MDA), superoxide dismutase (SOD), the pathology, and wet to dry weight ratio, broncho-alveolar lavage fluid protein content in lung tissues.

RESULTSRemoving portal blood stasis ameliorated lung and renal injury as shown by decreasing the level of serum endotoxin, TNF-alpha, BUN, Cr, wet to dry weight ratio, broncho-alveolar lavage fluid protein content, MDA, SOD. TNF-alpha, Cr, broncho-alveolar lavage fluid protein content in lung tissues and MDA in kidney tissue in group A5 were significantly reduced compared with those in group B (P < 0.05), while in lung tissue in group A10 were also markedly reduced (P < 0.05). The activation of SOD in group A5 were significantly increased (P < 0.05).

CONCLUSIONSRemoval of portal blood stasis before the resume of splanchnic circulation may ameliorate the lung and renal injury induced by hepatic ischemia reperfusion. The possible mechanism may be that portal blood stasis removal reduces endotoxin absorption, and further decreases production of serum TNF-alpha.

Animals ; Disease Models, Animal ; Female ; Ischemia ; metabolism ; pathology ; Kidney ; metabolism ; pathology ; Liver ; blood supply ; Lung ; metabolism ; pathology ; Male ; Portal Vein ; pathology ; Rabbits ; Random Allocation ; Reperfusion Injury ; metabolism ; pathology

UNLABELLEDTo reduce the huge labor-cost in the screening in traditional monoclonal antibody generation, We established a new system for monoclonal antibody generation integrating with protein array. BALB/c mice were immunized by eight recombinant proteins respectively, and the positive hybridoma cells were obtained by cell fusion and ELISA screening. All the eight kinds of positive hybridoma cells were mixed, cloned, screened by protein array, and definite dilution cloned.

RESULTS175 single cell clones were obtained by complex cloning, and 119 of those were positive clones. Then 8 positive cell lines were generated by the following 2 rounds definite dilution cloning. By comparing with the traditional method, we got 8 monoclonal antibodies using the combined protein array screening and multiplex cloning method in 1 cycle, and fewer amounts of antigens were used. As a result, the combined protein array and multiplex cloning method could be used as an economical, rapid and simple tool applying in high throughput monoclonal antibody generation.

Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; isolation & purification ; Female ; Humans ; Hybridomas ; metabolism ; Mice ; Mice, Inbred BALB C ; Protein Array Analysis

OBJECTIVETo observe the effect of glycoprotein receptor blockade tirofiban in acute anterior myocardial infarction patients without ST segment resolution after primary percutaneous coronary intervention (PCI).

METHODSFrom April 2006 to April 2008, 157 acute anterior myocardial infarction patients without ST segment resolution after PCI were randomly allocated to tirofiban (intravenous bolus 10 microg/kg followed by intravenous infusion of 0.15 microgxkg(-1)xmin(-1) for 48 h, n = 80) or equal volume saline (control group, n = 77). Baseline characteristics, PCI features and clinical outcomes during hospitalization, left ventricular ejection fractions (LVEF) and major adverse cardiac events (MACE, including death, re-infarction and target vessel revascularization) at 30 and 180 days after discharge were compared between the two groups.

RESULTSThe baseline clinical characteristics were comparable between the two groups. Compared to control group, the MACE rates and re-infarction rates at 30 days (6.3% vs.18.2%, P < 0.05; 1.3% vs.9.1%, P < 0.05, respectively) and 180 days (10.0% vs.23.4%, P < 0.05; 2.5% vs.10.4%, P < 0.05, respectively) were significantly reduced in tirofiban group. LVEF value was significantly higher in tirofiban group at 30 days and 180 days compared with those in control group [(51 +/- 6)% vs. (46 +/- 8)%, P < 0.05; (57 +/- 7)% vs. (50 +/- 9)%, P < 0.05]. Hemorrhagic complications were similar between the two groups.

CONCLUSIONUse of tirofiban for acute anterior myocardial infarction patients without ST segment resolution after PCI is safe and can significantly improve 30 and 180 days clinical outcomes after discharge.

Aged ; Angioplasty, Balloon, Coronary ; Anterior Wall Myocardial Infarction ; diagnosis ; drug therapy ; therapy ; Electrocardiography ; Female ; Humans ; Male ; Middle Aged ; Platelet Aggregation Inhibitors ; therapeutic use ; Prognosis ; Treatment Outcome ; Tyrosine ; analogs & derivatives ; therapeutic use