OBJECTIVETo construct a recombinant bacillus Calmette-Guérin vaccine (rBCG) secreting human interferon-alpha 2b (IFN alpha-2b).

METHODSBCG Ag85B signal sequence and IFN alpha-2b gene were amplified from the genome of BCG and of human peripheral blood by polymerase chain reaction (PCR), respectively. IFN alpha-2b gene was cloned in E. coli-BCG shuttle-vector pMV261 to get pMV261-IFN alpha-2b. A new recombinant plasmid pMV261-IFN alpha-2b was constructed by inserting BCG Ag85B signal sequence into pMV261-Ag85B-IFN alpha-2b. Then, BCG was transformed with this recombinant plasmid by electroporation, and designated as rBCG-IFN alpha-2b. The DNA and protein expressions of IFN alpha-2b gene in rBCG were determined by PCR and Western blot respectively. Also the quantity of IFN alpha-2b protein secreted by rBCG in culture supernatants was determined by enzyme linked immunosorbent assay (ELISA).

RESULTSBy partial nucleotide sequencing, the DNA sequences of human IFN alpha-2b and BCG Ag85B were consistent with that in the Gene Bank, and were correctly inserted into the shuttle expression vector pMV261 to construct recombinant plasmid pMV261-Ag85B-IFN alpha-2b. BCG was successfully transformed with this recombinant plasmid by electroporation and the recombinant BCG (rBCG-IFN alpha-2b) was capable of synthesizing and secreting cytokine IFN alpha-2b. The concentration of IFN alpha-2b in culture supernatants was quantified by ELISA and calculated to be approximately 301.45 pg/ml.

CONCLUSIONSRecombinant BCG secreting human IFN alpha-2b (rBCG-IFN alpha-2b) was constructed successfully and the specific IFN alpha-2b protein can be expressed highly and steadily by rBCG vaccine.

BCG Vaccine ; genetics ; immunology ; metabolism ; Gene Expression ; Genetic Vectors ; Humans ; Interferon-alpha ; genetics ; metabolism ; Plasmids ; genetics ; Recombinant Proteins ; Transformation, Bacterial

OBJECTIVETo examine the chemopreventive effect of selective cyclooxygenase-2 (COX-2) inhibitor celecoxib for Barrett's esophagus in rats.

METHODSFifty 8-week-old male Sprague Dawley rats underwent esophagojejunostomy to induce Barrett's esophagus model. Four weeks after operation the animals were given celecoxib 10 mg/(kg*d(-1))(celecoxib group), or saline 1 ml (control group). Another 10 rats were sham operation group. All animals were sacrificed at 20 week after surgery. The degree of inflammation, Barrett's esophagus, adenocarcinoma, COX-2 expression and PGE(2) of animals were assessed.

RESULTAmong 60 rats, 6 rats died in celecoxib group, 8 rats died in control group, 1 rat died in sham operation group, and 45 (75%) rats completed the study. The incidence of mild, moderate and severe degree esophageal inflammation in celecoxib group and control group was 14/19(73.68%), 4/19(21.05%), 1/19(5.26%); 4/17(23.53%), 5/17(29.41%), 8/17(47.06%)(P<0.05), respectively. The incidence of Barrett's esophagus was 7/19(36.84%), 13/17(76.47%) in two group respectively(P<0.05); The incidence of Barrett's esophagus with dysplasia was 2/19(10.53%), 8/17(47.06%)(P<0.05), respectively. The expression of COX-2 was 1/7(14.29%), 10/13(76.92%)(P<0.05) in two groups. PGE2 content was significantly lower in the celecoxib group than that in control group(P<0.001). No esophageal pathological changes were found in sham operation group.

CONCLUSIONSelective COX-2 inhibitors celecoxib can inhibit inflammations, development of Barrett's esophagus and esophagus adenocarcinoma.

Animals ; Barrett Esophagus ; metabolism ; prevention & control ; Celecoxib ; Cyclooxygenase 2 ; metabolism ; Cyclooxygenase 2 Inhibitors ; therapeutic use ; Dinoprostone ; metabolism ; Male ; Pyrazoles ; therapeutic use ; Rats ; Rats, Sprague-Dawley ; Sulfonamides ; therapeutic use

OBJECTIVETo establish animal models of reflux esophagitis in rats.

METHODSSeventy male Sprague Dawley rats aged 8 weeks were randomly divided into 4 groups: in Group A (n=20) esophagojejunostomy was performed to induce a gastro-jejuno-esophageal reflux; in Group B (n=20) esophagoduodenostomy was performed to induce a gastro-duodeno-esophageal reflux; in Group C (n=20) total gastrectomy plus esophagojejunostomy was performed to induce a jejuno-esophageal reflux; in Group D (n=10) only was performed sham operation (control).

RESULTAmong 70 rats, 6 died in Group A, 7 died in Group B, 6 died in Group C, and 72.9 %(51/70) animals were completed in the study. After 12 weeks the incidence of esophageal inflammation was 100.0%; in Groups A, B and C erosion occurred in 11/14 (78.6%), 10/13 (76.9%), 3/14 (21.4%) of animals, respectively; squamous dysplasia was in 10/14 (71.4%), 10/13 (76.9%), 5/14 (35.7%) of rats, respectively; Barrett's esophagus was in 6/14 (42.9%), 5/13 (38.5%), 1/14 (7.1%), respectively. One esophageal adenocarcinoma was found in Group A; no histological changes were observed in Group D.

CONCLUSIONThe animal models of reflux esophagitis can be induced by esophagojejunostomy, esophagoduodenostomy or total gastrectomy plus esophago-jejunostomy in rats; and the former two surgical modalities are better than the later.

Animals ; Barrett Esophagus ; Disease Models, Animal ; Esophagitis, Peptic ; classification ; Esophagus ; surgery ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley

A 17-year-old young man with a history of swollen leg and intermittent jaundice was presented to Peking Union Medical College Hospital with acute fever and mental disturbance.He developed deep venous thrombosis,acute myocardial infarction and plantar skin necrosis during the past four years,and was presented with an acute episode of fever,thrombocytopenia,acute kidney injury,acute myocardial infarction,mental disturbance,and obstructive jaundice.Laboratory tests showed schistocytes on peripheral blood smear.High titer of antiphospholipid antibodies was detected.Strikingly,the activity of a disintegrin and metalloprotease with a thrombospondin type 1 motif,member 13 (ADAMTS13)was significantly decreased without the production of inhibitors.Images indicated stenosis of the common bile duct,common hepatic duct,and cystic duct,which caused dilation of bile ducts and the gall bladder.Corticosteroids and anticoagulation therapy were effective at first,but the disease relapsedonce the corticosteroids tapered down.Plasma exchange was administrated for 17 times,which was effective temporarily during this episode.Methylprednisolone pulse therapy,intravenous immunoglobulin,rituximab,anticoagulation therapy,and bile drainage,were all tried but still could not control the disease.The patient's family agreed to withdraw treatment after he developed septic shock.

OBJECTIVETo investigate the role of B7-H1 expression in IL-10 production, the B7-H1 and IL-10 expression levels in pancreatic carcinoma tissues and to analyze the correlation between B7-H1 expression and IL-10 level.

METHODSThe mRNA and protein levels expressions of B7-H1 and IL-10 in 35 cases of pancreatic cancer and corresponding paracarcinoma tissues and 5 cases of normal pancreas tissues were detected by RT-PCR, Western blot and immunohistochemistry respectively.

RESULTSThe findings for the first time provided the evidences that there was a clear trend for B7-H1 and IL-10 expressions to be most highly expressed in carcinoma tissue, intermediately expressed in paracarcinoma tissue, and expressed at the lowest level in normal pancreatic tissue at mRNA and protein levels. Moreover, there were statistically significant differences in B7-H1 and IL-10 expression between pancreatic carcinoma tissues, corresponding paracarcinoma tissues and normal pancreatic tissues at mRNA and protein levels (P < 0.05). Furthermore, the immunohistochemistry indicated that there were high expression levels of B7-H1 (60.5% +/- 12.7%) and IL-10 (65.3% +/- 16.2%) in pancreatic carcinoma tissues while there were no significant expressions in normal pancreatic tissues. Meanwhile, correlation analysis revealed that B7-H1 expression was significant associated with IL-10 level in tumor tissues at mRNA (P = 0.008, r = 0.841) and protein levels (P = 0.007, r = 0.838).

CONCLUSIONSOver-expression of B7-H1 may be responsible for the increasing IL-10 production in pancreatic cancer, which caused reduced immune response to tumor cells and contributed to pancreatic carcinoma escape from immune attack.

Antigens, CD ; immunology ; B7-H1 Antigen ; Humans ; Immune Evasion ; Interleukin-10 ; immunology ; Pancreatic Neoplasms ; immunology

OBJECTIVETo observe degenerative intervertebral disc and to examine innervation of degenerative discs in the rabbit anular-injury model.

METHODSTwo different magnitudes of anular injury at 5 mm depth were performed by 11 blade or 16 gauge needle at the L3-L4 or L5-L6 discs in New Zealand white rabbits (n=48, 2.5-3.0 kg). Disc degeneration was evaluated by radiographic, MRI and histological examination at different time points after surgery. To identify nerve ingrowth into disc, two general markers PGP 9.5 and GAP 43, for nerve fibers were examined by immunohistochemistry.

RESULTSignificant decreases in disc height and signal intensity in magnetic resonance imaging were observed in 11 blade group and 16 G puncture group (P<0.01). 16 G puncture group induced slower and more progressive disc degeneration companed with the stab group and control group. At the 12-week time point, nucleus pulposus tissues were extruded and scar tissues formed outside the disc. In stab discs, nerve ingrowth was scattered on the surface of injury site and in the deeper part of the scar tissues, more than 1 mm from the surface. However, in punctured discs, PGP 9.5 and GAP 43-immunoreative fibers were only observed in the outmost part of the scar tissues and superficial area. More nerve fibers were observed in stab group.

CONCLUSIONInnervation may act as a source of discogenic pain which is associated with intervertebral disc degeneration caused by disc anular injury.

Animals ; GAP-43 Protein ; metabolism ; Intervertebral Disc ; injuries ; innervation ; pathology ; Intervertebral Disc Degeneration ; diagnosis ; diagnostic imaging ; etiology ; Low Back Pain ; etiology ; Lumbar Vertebrae ; Male ; Nerve Fibers ; pathology ; Rabbits ; Radiography ; Random Allocation ; Ubiquitin Thiolesterase ; metabolism

Animals ; Apoptosis ; Blood Pressure ; Disease Models, Animal ; Flow Cytometry ; Fluid Therapy ; In Situ Nick-End Labeling ; Intestinal Mucosa ; pathology ; Lactic Acid ; blood ; Male ; Rats ; Rats, Sprague-Dawley ; Resuscitation ; Shock, Hemorrhagic ; mortality ; pathology ; therapy ; Survival Rate

OBJECTIVETo investigate the effect of intravesical instillation of antifibrinolytic agents with bacillus Calmette-Guerin (BCG) on preventing recurrence of superficial bladder transitional cell carcinoma (BTCC) after surgical management.

METHODSA total of 326 cases of superficial BTCC undergoing transurethral resection of bladder tumor (TURBT) or partial cystectomy were divided into 5 groups. Then the different dosage BCG with or without antifibrinolytic agents was regular instilled into bladders (once a week, then once a month after 6 times). Group A including 66 cases received intravesical instillation of 100-120 mg BCG plus 100 mg para-aminomethyl benzoic acid (PAMBA). Group B including 64 cases: instillation of 50-60 mg BCG plus 100 mg PAMBA; Group C including 65 cases: 100-120 mg BCG plus 2.0 g epsilon-aminocaproic acid (EACA); Group D including 64 cases: 50-60 mg BCG plus 2.0 g EACA; Group E (control group) including 67 cases: 100-120 mg BCG. All the cases had been followed up for 4 to 69 months (mean, 28.5 months). Not only was cystoscopy performed every 3 months, but also biopsy was carried out to identify recurrence when necessary. Side effect was recorded after instillation.

RESULTSThe rate of tumor recurrence of Group A, Group B, Group C and Group D was 12%, 10%, 9%, 9% respectively, which was significantly lower than that of Group E (30%) (chi(2) = 5.699, 6.818, 7.380, 7.867, P = 0.017, 0.009, 0.007, 0.005). And there was no significant difference of tumor recurrence rate between Group A and Group B or between Group C and Group D (Group A and Group C: high dosage BCG plus antifibrinolytic agents, while Group B and Group D: low dosage BCG plus antifibrinolytic agents) (P > 0.05). But the side effects developing in Group B and Group D after BCG instillation were less than those in Group A and Group C.

CONCLUSIONSThe efficacy of BCG on prevention the recurrence of superficial BTCC can be enhanced when combined with antifibrinolytic agents. Even if the dosage of BCG was reduced by half, the efficacy didn't changed. A new approach of low dosage BCG plus antifibrinolytic agents is recommended in the prophylaxis of recurrence of bladder cancer.

4-Aminobenzoic Acid ; administration & dosage ; Adjuvants, Immunologic ; therapeutic use ; Administration, Intravesical ; Adult ; Aged ; Aged, 80 and over ; Aminocaproic Acid ; administration & dosage ; Antifibrinolytic Agents ; therapeutic use ; BCG Vaccine ; therapeutic use ; Carcinoma, Transitional Cell ; drug therapy ; surgery ; Combined Modality Therapy ; Drug Therapy, Combination ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Neoplasm Recurrence, Local ; prevention & control ; Urinary Bladder Neoplasms ; drug therapy ; surgery ; para-Aminobenzoates

OBJECTIVETo measure the plasma levels of transforming growth factor beta1 (TGFbeta1), the protein expression of alpha-SMA in hepatic stellate cells and urokinase plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1), and study on the relationships between the plasma levels of TGFbeta1, the protein expression and the serum hyaluronic acid (HA) in patients with different grades of hepatic fibrosis.

METHODSThirty seven cases with hepatic fibrosis of different grades were classified according to HE and VG staining categories from 0 to 4, in which there were 8 cases in grade 1, 9 cases in grade 2, 7 cases in grade 3, 13 cases in grade 4. The plasma levels of TGFbeta1 and the serum levels of HA were detected by ELISA. The protein expressions of a-SMA, uPA and PAI-1 in fibrotic liver tissue were observed by immunohistochemistry.

RESULTSWith the progression of hepatic fibrosis, the plasma levels of TGFbeta1 and the protein expression of a-SMA, uPA and PAI-1 in fibrotic liver tissue were increased. In grade 3 and 4, the plasma levels of TGFbeta and the protein expression of a-SMA and PAI-1 in fibrotic liver tissue were significantly increased, but the protein expression of uPA in cirrhosis liver tissue did not increased.

CONCLUSIONTGFbeta1, a-SMA, uPA and PAI-1 play an important role in the progression of hepatic fibrosis. Inhibiting the early activation of latent TGFbeta1 or increasing uPA and inhibiting PAI-1 over express may contribute to matrix degradation and retard the progression of hepatic fibrosis.

Actins ; blood ; Adult ; Aged ; Female ; Hepatitis B, Chronic ; blood ; complications ; Humans ; Liver Cirrhosis ; blood ; etiology ; Male ; Middle Aged ; Plasminogen Activator Inhibitor 1 ; blood ; Transforming Growth Factor beta ; blood ; Transforming Growth Factor beta1 ; Urokinase-Type Plasminogen Activator ; blood

OBJECTIVESTo measure the plasma levels of urokinase plasminogen activator (uPA), urokinase plasminogen activator receptor (uPAR) and plasminogen activator inhibitor-1 (PAI-1), and study the relationship between the plasma levels of uPA, PAI-1 and the serum albumin (Alb), collagen type IV (CIV), the serum hyaluronic acid (HA), prothrombin time (PT) and prothrombin activity (PTA) in patients with different stages of liver cirrhosis following chronic hepatitis B.

METHODS72 cases with liver cirrhosis of different stages were classified according to child-pugh's categories A, B, C, in which there were 23 cases in child A, 29 cases in child B, and 20 cases in child C. The plasma levels of uPA, uPAR, PAI-1 and the serum levels of HA, CIV were detected by ELISA. The serum PCIII concentration was determined by radioimmunoassay.

RESULTSWith the progression of hepatic fibrosis, the plasma levels of uPA, uPAR and PAI-1 were (1.36+/-0.43) microg/L, (3.03+/-1.48) microg/L and (24.09+/-7.14) microg/L respectively in group A, (1.79+/-0.62) microg/L, (4.80+/-2.22) microg/L and (41.40+/-17.52) microg/L respectively in group B. The highest levels were in child C, whose levels were (1.88+/-0.64) microg/L, (4.82+/-2.02) microg/L and (52.60+/-16.87) microg/L respectively, compared with group A and group B, t value were from 2.81 to 7.38, all of P value were less than 0.01. There was negative correlation between the plasma levels of uPA and the serum PCIII (r=-0.4785, P<0.05) in child A, but, positive correlation between the plasma PAI-1 and the serum HA (r=0.5447, P<0.01) in child C. The value of PAI-1/uPA was significantly decreased in child A, but increased in child B and child C.

CONCLUSIONIn the late of liver cirrhosis, increased PAI-1 together with uPA, uPAR are associated with overall inhibition of matrix degradation. The plasma levels of uPA and PAI-1 were correlation to the progression of liver cirrhosis.

Female ; Hepatitis B, Chronic ; complications ; Humans ; Liver Cirrhosis ; blood ; Male ; Middle Aged ; Plasminogen Activator Inhibitor 1 ; blood ; Receptors, Cell Surface ; blood ; Receptors, Urokinase Plasminogen Activator ; Urokinase-Type Plasminogen Activator ; blood