To obtain the tervalent fusion toxin gene (named FT),three toxin gene fragments from three species of Vibrio parahaemolyticus,Vibrio vulnificus and Vibrio mimicus were connected with the flexible linker (GGGGS) using overla Pextension PCR. The three toxin gene fragments respectively encode the mature proteins of the thermostable direct hemolysin (TDH) of V. parahaemolyticus,the cytotoxin (VVC) of V. vulnificus and the heat-labile hemolysin (VMH) of V. mimicus. The identity of FT nucleic acid sequence was 99.6% with the corresponding toxin gene fragments. The open reading frame of FT was 3225 bp,encoding 1074 amino acid residues with the predicted molecular weight (MW) of 120.4 kDa. Then,FT was subcloned into the expression vector pET-22b(+). The construction of recombinant expression vector pET-22b-FT was followed by transforming into E. coli BL21(DE3) for expression. The SDS-PAGE electrophoresis results indicated that the MW of the fusion toxin protein was matched to the predicted MW. After induction by 1 mmol/L IPTG at 37℃,the fusion toxin protein was effectively expressed in E. coli BL21(DE3) with the amount of 11.49% through thin layer chromatography scanning (TLCS) analysis. Cavia cobaya was immunized using the purified cytorrhyctes to produce the anti-serum. Through the determination of the optimum working conditions,the sensitivity test,the specificity test,repeatability test and sample simulation test,the indirect ELISA method was established,which is a broad-spectrum,rapid and specific to detect various of food-poisoning Vibrio simultaneously.

AIMTo investigate the role of ICAM-1 on the surface of vascular endothelial cell (VEC) in freezing/thawing injury of VEC, in order to elucidate the pathogenesis of freezing/thawing injury.

METHODSVEC separated and cultured from rat aorta and PMN separated from rat peripheral blood were selected as experiment materials. The frozen/thawed VEC model was founded by freezing VEC with the type WKL-V rate cooling instrument and then rewarming them in a water bath. ICAM-1 expression on the surface of frozen/thawed VEC was detected at 4, 12 and 24 h after freezing/thawing with immunohistochemical method. After coincubating frozen/thawed VEC with normal PMN, the adhesion of VEC to PMN was monitored with rose bengal staining assay and the injury level of VEC was indicated by measuring LDH activity in nutrient solution.

RESULTSThe ICAM-1 expression on the surface of VEC increased from 13.2% +/- 3.6% before freezing/thawing of VEC to 22.3% +/- 4.4% at 4 hour after freezing/thawing, and reached the peak (37.9% +/- 2.5%) at 12 hour after freezing/thawing of VEC. After coincubation of frozen/thawed VEC with normal PMN, the adherence of frozen/thawed VEC to PMN increased from group control 0.204 +/- 0.025 to 0.363 +/- 0.022 (P < 0.01), LDH activity in nutrient solution increased from group control 104.64 +/- 20.14 U/L to 162.33 +/- 27.88 U/L (P < 0.01), monoclonal antibody against ICAM-1 (ICAM-1 Mab) could partially block the adherence of frozen/thawed VEC to PMN (0.270 +/- 0.021, P < 0.01), and diminish LDH activity in nutrient solution (125.39 +/- 22.26 U/L, P < 0.05).

CONCLUSIONThe freezing/thawing of VEC can elicit an increase in ICAM-1 expression on the surface of VEC, and then proceed to VEC-PMN adherence and lead to VEC injury.

Animals ; Cells, Cultured ; Endothelial Cells ; metabolism ; Endothelium, Vascular ; cytology ; Freezing ; Intercellular Adhesion Molecule-1 ; metabolism ; Neutrophils ; cytology ; Rats

AIMTo investigate the effects of shivering on airway rewarming.

METHODSThe hypothermic dog model without shivering was established by immersing an anesthetized dog in cold water and administering atracurium to inhibit the dog shivering. The model dog respired warm fully humidified (40-45 degrees C, RH 99.9%) air and room temperature air(19 +/- 1 degrees C, RH 30% - 75%) to rewarm each for 2 hours, the priority of different temperature air respired was arranged randomly. After rewarming for 4 hours, the relaxed dog breathed warm humidified air by positive pressure ventilation in order to restore its spontaneous respiratory. Then the dog continued to respire warm humidified air spontaneously until the esophageal (Te) and rectal temperature (Tr) of the dog achieved the same degrees as the dog was immersed in the water. The metabolic heat production was detected by indirect calorimetry during the experiment.

RESULTS(1) When the shivering was inhibited, inhaling warm humidified air for 2 hours made the Tr and Te of the dogs increase 0.26-0.39 degrees C and 0.44-1.11 degrees C per hour respectively, inhaling air at room temperature for 2 hours made Tr and Te of the dogs decrease 0.24-0.51 degrees C and 0.58-0.67 degrees C per hour, respectively. And the changes in Tr and Te of the dogs were unrelated to the priority of inhaling air at different temperature. (2) When the dog with shivering respired spontaneously warm humidified air, the rewarming rates of Tr and Te were 2.26-2.33 degrees C/h and 1.96-2.38 degrees C/h respectively, quicker than those of the dogs whose shivering was inhibited. (3) Compared with metabolic heat production of the unshivering dog respiring warm humidified air by positive pressure ventilation, that of the shivering dog respiring warm humidified air spontaneously increased outstandingly, shivering thermogenesis made the rewarming rates increased obviously.

CONCLUSIONAirway rewarming is a method conducive to rewarming of hypothermia. When the body is shivering, the metabolic heat production increases obviously, that makes the rewarming rate increase markedly. So the shivering must be inhibited in order to eliminate the interference of shivering thermogenesis when the effects of airway rewarming are detected.

Animals ; Body Temperature Regulation ; Cold Temperature ; Dogs ; Hypothermia ; physiopathology ; therapy ; Hypothermia, Induced ; Male ; Respiratory Physiological Phenomena ; Shivering

OBJECTIVETo study the clinicopathologic features, differential diagnosis and pathogenesis of sclerosing angiomatoid nodular transformation of spleen.

METHODSTen cases of sclerosing angiomatoid nodular transformation of spleen were retrieved from the archival file. Histochemical and immunohistochemical (EnVision method) studies were performed. Ultrastructural findings were also available in one of them.

RESULTSSclerosing angiomatoid nodular transformation was characterized by micronodular appearance of vascular spaces lined by plump endothelial cells with interspersed ovoid spindle cells. Immunohistochemical study showed that the endothelial cells of vessels in the angiomatoid nodules had various expressions of immunologic phenotypes and could be mainly classified into 3 types: CD34(+)/CD31(+)/CD8⁻ endothelial cells of the capillaries, CD8(+)/CD31(+)/CD34⁻ lining cells of the sinusoids and CD31(+)/CD8⁻/CD34⁻ endothelial cells of the small veins. Collagen network and dilated lymphatic sinuses were evident under transmission electron microscope.

CONCLUSIONSSclerosing angiomatoid nodular transformation of spleen is a rare benign entity. It may represent a reactive condition and bears some relationship with splenic angioma. It needs to be distinguished from borderline or malignant vascular tumors of spleen.

Adult ; Antigens, CD34 ; metabolism ; CD8 Antigens ; metabolism ; Diagnosis, Differential ; Female ; Hemangioendothelioma ; metabolism ; pathology ; Hemangiosarcoma ; metabolism ; pathology ; Histiocytoma, Benign Fibrous ; metabolism ; pathology ; surgery ; ultrastructure ; Humans ; Male ; Microscopy, Electron ; Middle Aged ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Splenic Neoplasms ; metabolism ; pathology ; surgery ; ultrastructure

Colorectal cancer (CRC) is a common malignancy burdening people globally, with increasing morbidity and mortality nowadays, due to the alternation in the diet type and lifestyle in modern society. Berberine, a type of benzylisoquinoline alkaloid, is widely present in numerous medicinal plants, particularly including Coptidis Rhizoma. Mounting evidence reveals that berberine possesses an array of pharmacological effects, such as anti-inflammation, anti-bacterium, anti-cancer, anti-diabetes mellitus and so on. In particular, berberine exhibits substantial inhibition on various types of cancers including CRC. Hereby, we sought to systematically review the suppressive effect of berberine on CRC through the diminishment of the proliferation and metastasis, induction of apoptosis, arrest of cell cycle, regulation of inflammatory reaction, the reverse of chemotherapeutic resistance and restoration of gut microbiota in CRC, so as to shed light on the in-depth mechanisms underlying the treatment of CRC with berberine in the clinical setting.

AIMIn order to study airway rewarming method and rewarming devices for hypothermia, hypothermic dog model was established.

METHODSThe anesthetized dog was immersed in cold water at 16.7 degrees C until the esophageal temperature (Te) of the dog decreased to 34.0 degrees C, the core temperature and skin temperature were monitored by using a 12-channel scanning thermometers. Atracurium besylate, a skeletal muscle relaxant, was injected intravenously when the core temperature of the dog was basically steady after the dog was out of the cold water, the hypothermic dog model was established.

RESULTSRectal and esophageal temperature could stand for the core temperature of the hypothermic dog model, but mixing with each other was prohibited because of leading to mistakes. Administering of atracurium besylate could eliminate the effect of shivering on airway rewarming alone, hypothermic dog model in which shivering was inhibited could be used in determination of airway rewarming technique and rewarming devices for hypothermia.

CONCLUSIONHypothermic dog model in which shivering was inhibited can abolish the interference of shivering, experimental repeatability is good, experimental method quite simple, and the model appropriate for application and dissemination.

Animals ; Body Temperature ; Cold Temperature ; Disease Models, Animal ; Dogs ; Hypothermia ; therapy ; Hypothermia, Induced ; Male ; Respiratory System ; Rewarming ; methods ; Shivering

BACKGROUNDSevere acute respiratory syndrome coronavirus (SARS-CoV) is a newly emerging virus that gives rise to SARS patients with high rates of infectivity and fatality. To study the humoral immune responses to SARS-CoV, the authors evaluated IgG and IgM specific antibodies in patients' sera.

METHODSTwo methods, enzyme-linked immunosorbent assay (ELISA) and indirect immunofluorescent assay (IFA), were used to detect specific serum IgG and IgM against SARS-CoV in 98 SARS patients and 250 controls consisting of patients with pneumonia, health-care professionals and healthy subjects. The serum antibody profiles were investigated at different times over one and a half years in 18 of the SARS patients.

RESULTSThe sensitivity and specificity of ELISA for detecting IgG against SARS-CoV were 100.0% and 97.2% and for IgM 89.8% and 97.6% respectively; the figures using IFA for IgG were 100.0% and 100.0% and for IgM 81.8% and 100.0% respectively. During the first seven days of the antibodies trace test, no IgG and IgM were detected, but on day 15, IgG response increased dramatically, reaching a peak on day 60, remaining high up to day 180 and decreasing gradually until day 540. On day 15, IgM was detected, rapidly reached a peak, then declined gradually until day 180 when IgM was undetectable.

CONCLUSIONThe detection of antibodies against SARS virus is helpful in the clinical diagnosis of SARS.

Adult ; Aged ; Antibodies, Viral ; blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Fluorescent Antibody Technique, Indirect ; Humans ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Male ; Middle Aged ; SARS Virus ; immunology ; Severe Acute Respiratory Syndrome ; immunology ; T-Lymphocytes, Cytotoxic ; immunology

OBJECTIVETo explore the origin and differentiation of gastrointestinal stromal tumors (GISTs).

METHODSImmunohistochemistry staining and electron microscopy were adopted.

RESULTSIn 212 cases of primary GISTs, the positive rates of CD117, CD34, alpha-SMA, MSA, desmin, S-100, PGP9.5 were 96.7%, 77.3%, 19.3%, 15.6%, 1.9%, 16.3%, and 12.3% respectively. Among them, GISTs showed a diffuse and strong positivity for CD117. Electron microscopy of tumor cells demonstrated numerous mitochondria, prominent perinuclear Golgi complex, smooth and rough endoplasmical reticulum and intermediate filaments. Irregular caveolae, dense plaque, incontinuous basal lamina were observed occasionally. Cytoplasmic processes were often observed accompanying with local adhesion present between the processes or between the processes and the cell membrane.

CONCLUSIONSData from both immunophenotype and electron microscopy suggest that GIST might originate from the mesenchymal cells, differentiating to be ICC afterwards, and possessing myoid characteristics in various extent.

Cell Differentiation ; Gastrointestinal Stromal Tumors ; chemistry ; ultrastructure ; Golgi Apparatus ; ultrastructure ; Humans ; Immunohistochemistry ; Microscopy, Electron ; Proto-Oncogene Proteins c-kit ; analysis ; S100 Proteins ; analysis ; Stromal Cells ; chemistry ; ultrastructure ; Ubiquitin Thiolesterase ; analysis

OBJECTIVETo study the antagonistic effect of 3'-daidzein sulfonate sodium (DSS) on benign prostatic hyperplasia (BPH) and its possible mechanism.

METHODSForty healthy mice were randomly divided into five groups: a normal control group without any treatment, a model group of BPH treated by subcutaneous injection of testosterone propionate, a positive control group with the BPH procedure treated by Qianliekang, a 20 mg/(kg x d) DSS group with the BPH procedure and a 40 mg/(kg x d) DSS group with the BPH procedure. After 12 days of the above treatments, the mice were sacrificed for measurement of the prostate glandular wet weight, the index of prostate gland (PI), the morphological changes of prostate gland by light microscopy and the contents of testosterone and estradiol in the serum.

RESULTSThe prostate wet weight and PI decreased dose-dependently after DSS treatment for 12 days compared with the BPH model group (P < 0.05 or P < 0.01). The hyperplastic epithelioglandular papilla waned and even disappeared in the DSS treated groups under the light microscope, the epithelial cells became cubical or flat. The effect of DSS at 40 mg/(kg x d) was similar to that of the positive anti-BPH drug Qianliekang. DSS reduced the serum testosterone, estradiol contents and the T/E2 ratio (P < 0.05 or P < 0.01).

CONCLUSIONDSS has significant antagonistic effect on BPH induced by testosterone propionate in mice, which may involve its regulatory action on the sex hormone balance.

Animals ; Estradiol ; blood ; Isoflavones ; therapeutic use ; Male ; Mice ; Mice, Inbred Strains ; Phytotherapy ; Prostatic Hyperplasia ; drug therapy ; metabolism ; Testosterone ; blood

OBJECTIVETo summarize the experience in surgical treatment of residual shunt after repair of ventricular septal defect and investigate the position of the residual shunts.

METHODSBetween January 1979 and May 2003, re-operations on residual shunt after repair of ventricular septal defect were performed in 37 patients with congenital heart disease including ventricular septal defect, tetralogy of Fallot, double outlet right ventricle in 19, 17 and 1 patients, respectively. It accounted for 0.21% (37/18000) of open heart operations performed during these years. The patients included 26 males and 11 females with age from 3 months to 53 years (mean 16 +/- 12 years). The residual shunt was diagnosed by postoperative murmur and echocardiography. Twenty-six cases were repaired with patch and 11 cases were closed directly with mattresses sutures.

RESULTSTwo patients (2/37, 5%) died within 48 hrs postoperatively. The results in other 35 patients followed up after surgery from 3 months to 15 years were satisfactory.

CONCLUSIONSMost of the residual shunts occurred in base of septal leaflet of tricuspid valve, the second and the first transfer suture respectively. Effects of reoperations on residual shunts were satisfactory.

Adolescent ; Adult ; Cardiac Surgical Procedures ; methods ; Child ; Child, Preschool ; Double Outlet Right Ventricle ; surgery ; Female ; Heart Septal Defects, Ventricular ; surgery ; Humans ; Infant ; Male ; Middle Aged ; Postoperative Complications ; surgery ; Reoperation ; Retrospective Studies ; Tetralogy of Fallot ; surgery