OBJECTIVETo investigate the pro-apoptotic effect of Her-2 targeted recombinant caspase-6 fusion protein on osteosarcoma SOSP-9607 cells.

METHODSRecombinant immunocasp-6 was generated by sequential fusion of the genes of a signal peptide, a single-chain Her-2 antibody (e23sFv), a PEA translocation domain (PEA aa253-364) and an active caspase-6. The immunocasp-6 gene was cloned into pCMV plasmid to construct a kind of eukaryotic expression vector, i.e. pCMV-e23sfv-PE II-caspase-6 (abbr. pCMV-6) and transfected into SOSP-9607 cells. Murine xenograft models were randomly divided into two groups that received i.m. injections of liposome encapsulated pCMV-6 or pCMV alone. The tumor volume and weight of the nude mice and the tumor weight of the cured mice were observed and statistically analyzed. The morphological changes of the tumors were examined with HE staining, apoptotic morphology of the tumor was observed by TUNEL staining and the gene expression was analyzed by immunohistochemical staining.

RESULTSThe tumor growth of the mice in the treatment group was significantly slower than that of the control group (P = 0.001). The weight of the nude mice in the treatment group was significantly higher than that of the control group (P = 0.0002). The tumor weight of the mice in the treatment group was significantly lower than that of the control group (P = 0.0006). HE and TUNEL staining of the tumor of nude mice in the treatment groups showed typical characteristics of apoptosis, while normal structure was found in the control group. Furthermore, caspase-6 was not found in the tumor and muscle tissues in the control group, but only in the treatment group by immunohistochemistry.

CONCLUSIONImmunocasp-6 can selectively recognize and bind to and kill HER-2 positive osteosarcoma cells, therefore, to offer some foundation for the clinical treatment of osteosarcoma.

ADP Ribose Transferases ; genetics ; Animals ; Apoptosis ; Bacterial Toxins ; genetics ; Bone Neoplasms ; metabolism ; pathology ; Caspase 6 ; genetics ; metabolism ; Cell Line, Tumor ; Exotoxins ; genetics ; Humans ; Mice ; Mice, Inbred BALB C ; Mice, Nude ; Neoplasm Transplantation ; Osteosarcoma ; metabolism ; pathology ; Plasmids ; Random Allocation ; Receptor, ErbB-2 ; genetics ; Recombinant Fusion Proteins ; genetics ; metabolism ; Transfection ; Tumor Burden ; Virulence Factors ; genetics

OBJECTIVETo determine the expressions of ryanodine receptor type 1 (RyR1) and Cav1.3 L-type calcium channel (Cav1.3) in the vaginal smooth muscle cells of castrated rats and investigate the correlation of RyR1 and Cav1.3 with estrogen in female sexual dysfunction.

METHODSForty female SD rats of 8 weeks were randomly divided into Groups A (2-week sham operation), B (4-week sham operation), C (2-week castration) and D (4-week castration). Two and 4 weeks after surgery, the serum estradiol level was determined with the automated immunochemiluminescence system and the expressions of RyR1 and Cav1.3 in the vaginal smooth muscle were detected by immunohistochemistry and RT-PCR. Gray scale ratio was used to represent the mRNA expression levels of RyR1 and Car1.3, and the optical density value to denote their protein expression levels.

RESULTSSerum estradiol was significantly decreased in Group C ([0.210 +/- 0.026] nmol/L) as compared with A ([0.505 +/- 0.053] nmol/L) (P < 0.01), and so was it in Group D ([0.130 +/- 0.031] nmol/L) in comparison with B ([0.476 +/- 0.058] nmol/L) (P < 0.01). RyR1 and Cav1.3 were expressed in all groups. The mRNA expressions of RyR1 and Cav1.3 were significantly reduced in Group C (0. 680 +/- 0.073 and 0.580 +/- 0.043) as compared with A (0.950 +/- 0.064 and 0.870 +/- 0.019) (P < 0.01), as well as in Group D (0.220 +/- 0.032 and 0.190 +/- 0.020) in comparison with B (0.890 +/- 0.072 and 0.820 +/- 0.021) (P < 0.01). The protein expressions of RyR1 and Cav1.3 were significantly down-regulated in Group C (96.67 +/- 7.75 and 87.97 +/- 6.96) as compared with A (123.69 +/- 10.66 and 106.46 +/- 8.04) (P < 0.01), and so were they in D (86.45 +/- 8.16 and 69.43 +/- 8.30) in comparison with B (109.31 +/- 9.87 and 97.38 +/- 7.56) (P < 0.01).

CONCLUSIONBoth RyR1 and Cav1.3 were expressed in the vaginal smooth muscle cells of the rats, and estrogen might be involved in the regulation of female sexual reaction by acting on the expressions of RyR1 and Cav1.3.

Animals ; Calcium Channels ; metabolism ; Estrogens ; blood ; Female ; Myocytes, Smooth Muscle ; metabolism ; Ovariectomy ; Rats ; Rats, Sprague-Dawley ; Ryanodine Receptor Calcium Release Channel ; metabolism ; Vagina ; cytology

OBJECTIVETo examine the expression of vascular endothelial growth factor (VEGF) and its receptors (VEGFR1, VEGFR2) in transdifferentiated human proximal tubular epithelial (HK-2) cell induced by transforming growth factor beta1 (TGFbeta1).

METHODSThe transdifferentiation of HK-2 cells was detected by evaluation of expression of alpha-SMA by cytoimmunochemistry and RT-PCR. The VEGF mRNA was evaluated with RT-PCR. The secreted VEGF in the culture media was measured with ELISA. The cellular VEGF, VEGFR1, and VEGFR2 were measured with Western blot.

RESULTSThe immunostain of alpha-SMA were positive in HK-2 cell induced by TGFbeta1 at the concentration of 5 and 8 ng/ml for 72 h. The expression of alpha-SMA mRNA was induced by TGFbeta1 in concentration- and time-dependent manners. The expressions of mRNA and protein of VEGF were upregulated by TGFbeta1 at the concentration of 0.1 and 1 ng/ml for 72 h and at the concentration of 8 ng/ml for 12 h and 24 h when compared with the control. But expressions of mRNA and protein of VEGF were downregulated by TGFbeta1 at the concentration of 3, 5, and 8 ng/ml for 72 h and at the concentration of 8 ng/ml for 36, 48, and 72 h, respectively. Meanwhile, Protein levels of VEGFR1 and VEGFR2 were upregulated by TGFbeta1 in concentration- and time- dependent manners.

CONCLUSIONSIncreased expression of VEGFR1 and VEGFR2 and two-phase change in VEGF expression occurred in the process of tubular epithelial transdifferentiation induced by TGFbeta1. Reduced expression of VEGF may contribute to tubular epithelial transdifferentiation in a vicious circle.

Cell Differentiation ; Epithelial Cells ; cytology ; Humans ; Kidney Tubules, Proximal ; cytology ; RNA, Messenger ; metabolism ; Receptors, Vascular Endothelial Growth Factor ; metabolism ; Transforming Growth Factor beta ; pharmacology ; Transforming Growth Factor beta1 ; Vascular Endothelial Growth Factor A ; metabolism ; Vascular Endothelial Growth Factor Receptor-1 ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; metabolism

OBJECTIVEIn order to explore the role of toll-like receptors 2 (TLR2) in initiating inflammatory response, the expression of TLR2 of the liver and IL-18, TNF-alpha and IFN-gamma of plasma in fulminant hepatic failure was analysed.

METHODSD-galactosamine (D-Gal, 900 mg/kg) and lipopolysaccharide (LPS, 10 microg/kg) were administered intraperitoneally into the BALB/C mice. To evaluate the hepatic injury, serum transaminase (ALT and AST) and plasma IL-18, TNF-alpha and IFN-gamma were determined and the mortality was observed at various time points following the intraperitoneal injection. The level of TLR2 mRNA was measured by semiquantitative RT-PCR. The protein expression of TLR2 in the liver was detected by immunohistochemistry. The data was analyzed by SAS software.

RESULTSAfter 4 hours of intraperitoneal injection of D-Gal/LPS, the serum transaminase and plasma IL-18, TNF-alpha and IFN-gamma levels were elevated. The treated mice began to die at 7 hours. The mortality reached up to 80% at 10 h. TLR2 mRNA was expressed at a low level in liver tissues of normal mice, while it was significantly increased and maintained at a higher level following intraperitoneal injection with D-Gal/LPS. The expression of TLR2 protein was similar to that of the TLR2 mRNA, and the expression of TLR2 mRNA was positively correlated with the concentration of plasma IL-18, TNF-alpha and IFN-gamma (r=0.36, P=0.02; r = 0.48, P 0.003; r = 0.72, P<0.001) at different time points.

CONCLUSIONSOur results showed that TLR2 was involved in initiating and inducing the expression of proinflammation cytokines in this model of fulminant hepatic failure. The results suggest that adjusting the expression of TLR2 might be a new strategy in preventing the development of infectious diseases

Animals ; Galactosamine ; Interferon-gamma ; blood ; Interleukin-18 ; blood ; Lipopolysaccharides ; Liver ; metabolism ; Liver Failure, Acute ; chemically induced ; metabolism ; Male ; Mice ; Mice, Inbred BALB C ; RNA, Messenger ; biosynthesis ; genetics ; Toll-Like Receptor 2 ; biosynthesis ; genetics ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo observe the effect of bone morphogenetic protein-7 (BMP-7) on the transdifferentiation of cultured human tubular epithelial cell (HKC) induced by TGF-beta1 and to elucidate its possible mechanism.

METHODSThe cultured HKC cells were divided into 5 groups: serum-free group (negative control); single TGF-beta1 treated group (positive control); single BMP-7 treated group; combined TGF-beta1 and BMP-7 treated group; and BMP-7 pre-treated group. Expression of keratin of HKC cells was assessed by indirect enzyme immunohistochemistry (IEI), expression of alpha-smooth muscle actin (alpha-SMA) and E-cadherin by immunohistological method, percentage of alpha-SMA positive HKC cells by flow cytometry, and mRNA expression of alpha-SMA, TGF-beta1, and TGF-beta type II receptor by reverse transcription PCR.

RESULTSThe expression of alpha-SMA and the percentage of alpha-SMA positive HKC cells markedly increased after having been treated by TGF-beta1 while the expression of E-cadherin and keratin decreased. In the group pre-treated with BMP-7 (50 ng/ml) and then added with TGF-beta1 (8 ng/ml), expression of alpha-SMA was significantly lower than in the positive control group, while expression of E-cadherin and keratin significantly higher than in the positive control group. Measurement of the percentage of alpha-SMA positive HKC found significant deference between the combined TGF-beta1 and BMP-7 treated group and the positive control group (9.7% vs 19.8%; 5.8% vs 19.8%; P < 0.05). Significant difference existed between the BMP-7 (50 ng/ml) pre-treated group and the positive control group (8.7% vs 19.8%, P < 0.05). mRNA expression of alpha-SMA was measured by RT-PCR and the results showed that it significantly decreased in the group treated or pre-treated with BMP-7 (50 ng/ml) (15% and 12% of the results in the positive control group, respectively). The mRNA expression levels of both TGF-beta1 and its type II receptor significantly decreased (28% and 19%; 47% and 36%, compared with the positive control group, respectively).

CONCLUSIONTransdifferentiation of cultured renal epithelial cell induced by TGF-beta1 can be inhibittd by certain levels of BMP-7, cultured together with TGF-beta1 or pretreated. BMP-7 can prevent and inhibit the mRNA expression of TGF-beta1 and its type II receptor, which may be an important mechanism by which BMP-7 inhibit the transdifferentiation of renal tubular epithelial cell.

Actins ; biosynthesis ; genetics ; Bone Morphogenetic Protein 7 ; Bone Morphogenetic Proteins ; pharmacology ; Cell Differentiation ; drug effects ; Cells, Cultured ; Epithelial Cells ; cytology ; Humans ; Kidney Tubules ; cytology ; metabolism ; Polymerase Chain Reaction ; RNA, Messenger ; biosynthesis ; genetics ; Transforming Growth Factor beta ; pharmacology ; Transforming Growth Factor beta1

OBJECTIVETo investigate the antimicrobial resistance and penicillin resistance-associated genes (TEM and pbp2B) of Streptococcus pneumoniae (S. pneumoniae) isolated from sputum specimens of Guangzhou children with respiratory tract infection.

METHODSE-test and Kirby-Bauer methods were applied to detect the antibiotic susceptibility of 44 strains of S. pneumoniae. PCR was used to detect resistance genes pbp2B and TEM, followed by DNA sequence analysis of pbp2B gene. The sequence results were compared to those of penicillin-susceptible S. pneumoniae R6.

RESULTSOf the 44 isolates of S. pneumoniae, only 5 (11.4%) were susceptible to penicillin. All strains were resistant to erythromycin but susceptible to ofloxacin and vancomycin. The resistance rate of the isolates to clindamycin and trimoxazole was more than 90%. The S. pneumoniae isolates showed a high susceptibility to amoxicillin, imipenem and ceftriaxone, with a resistance rate of 0, 2.6% and 3.9%, respectively. The sequence analysis showed that more than 99% nucleotide sequence of pbp2B gene of five penicillin-susceptible isolates was the same as penicillin-susceptible S. pneumoniae R6, without any amino acid replacement. Site mutation was found in the remaining 39 penicillin-nonsusceptible isolates with a nucleotide mutation rate ranging from 13.2% to 23.1% and amino acid replacement rate from 6.5% to 10.9%. The 39 penicillin-nonsusceptible isolates were classified into 4 types according to the mutation site between Ser391 and Thr492 of pbp2B: type I (n=30), type II (n=7), type III (n=1) and type IV (n=1). No TEM gene was detected in all the 44 S. pneumoniae isolates.

CONCLUSIONSThe S.pneumoniae isolates from Guangzhou children with respiratory tract infection are resistant to penicillin and erythromycin. Amoxicillin and the third generation cephalosporin may be recommended for treating S. pneumoniae infection. The mutation of pbp2B gene plays an important role in the development of S. pneumoniae resistance to penicillin.

Aminoacyltransferases ; genetics ; Child, Preschool ; Drug Resistance, Bacterial ; genetics ; Female ; Humans ; Infant ; Male ; Microbial Sensitivity Tests ; Penicillin Resistance ; genetics ; Penicillin-Binding Proteins ; genetics ; Respiratory Tract Infections ; microbiology ; Streptococcus pneumoniae ; drug effects ; genetics ; beta-Lactamases ; genetics

The risk factors of high trait anger of juvenile offenders were explored through question naire study in a youth correctional facility of Hubei province,China.A total of 1090 juvenile offenders in Hubei province were investigated by self-compiled social-demographic questionnaire,Childhood Trauma Questionnaire (CTQ),and State-Trait Anger Expression Inventory-Ⅱ (STAXI-Ⅱ).The risk factors were analyzed by chi-square tests,correlation analysis,and binary logistic regression analysis with SPSS 19.0.A total of 1082 copies of valid questionnaires were collected.High trait anger group (n=316) was defined as those who scored in the upper 27th percentile of STAXI-Ⅱ trait anger scale (TAS),and the rest were defined as low trait anger group (n=766).The risk factors associated with high level of trait anger included:childhood emotional abuse,childhood sexual abuse,step family,frequent drug abuse,and frequent internet using (P＜0.05 or P＜0.01).Birth sequence,number of sibling,ranking in the family,identity of the main care-taker,the education level of care-taker,educational style of care-taker,family income,relationship between parents,social atmosphere of local area,frequent drinking,and frequent smoking did not predict to high level of trait anger (P＞0.05).It was suggested that traumatic experience in childhood and unhealthy life style may significantly increase the level of trait anger in adulthood.The risk factors of high trait anger and their effects should be taken into consideration seriously.

BACKGROUNDRelapses occur frequently in patients with lupus nephritis. Renal biopsy is the gold standard for assessing renal activity and hence guiding the treatment. Whether repeat renal biopsy is helpful during flares of lupus nephritis remains inconclusive. In the present study, we retrospectively reviewed the patients with lupus nephritis who had more than one renal biopsy with the hope to find the clinical value of repeat biopsy.

METHODSPatients who had a diagnosis of lupus nephritis and two or more renal biopsies were selected from the database of the patient pathology registration at this renal division. Renal biopsy was evaluated according to the International Society of Nephrology/Renal Pathology Society (ISN/RPS) classification of lupus nephritis. The pathological patterns and treatment regimens were analyzed after a repeat biopsy.

RESULTSWe identified 44 systemic lupus erythematosus patients with serial renal biopsies. In total, there were 94 renal biopsies. Overall, the pathological transition occurred in 64% instances according to the ISN/RPS class. When the transition was analyzed according to proliferative, membranous or mix lesions, it showed different profile: 35% in patients with proliferative lesion, 23.5% patients with mix lesions, 100% in patients with pure membranous lesion. The pathological transition could not be predicted by any clinical characteristics. After the repeat renal biopsy, 34% of patients had a change in their treatment regimens.

CONCLUSIONSThe pathological conversion was very prevalent in patients with lupus nephritis. However, the transitions became less prevalent when they were analyzed according to pure membranous, proliferative, and mix lesion. Repeat biopsy might be helpful to avoid unnecessary increased immunosuppression therapy.

Adult ; Aged ; Biopsy ; Female ; Humans ; Kidney ; pathology ; Lupus Nephritis ; diagnosis ; pathology ; Male

OBJECTIVETo investigate the findings of contrast-enhanced multislice computed tomography (MSCT) that characterize intraductal papillary neoplasms of bile ducts (IPNB).

METHODSThe MSCT findings and clinical data of 16 cases of IPNB proven by surgical pathology were reviewed retrospectively.

RESULTSAmong the 16 cases, nine were adenoma (multi-lesions, n = 5; single lesions, n = 4) and seven were adenocarcinoma (multi-lesions, n = 4; single lesions, n = 3). Among the nine adenoma cases, seven showed nodules or masses in the expanding intrahepatic bile ducts with asymmetrical low density on plain scan, and two showed obvious expansion of biliary ducts and the inner wall of bile ducts was rough. All seven of the adenocarcinoma cases showed nodules or masses in the expanding intrahepatic bile ducts with asymmetrical low density-like adenoma. When contrast enhancement was applied, the nine adenoma cases manifested slight-to-moderate degrees of asymmetrical enhancement. For the seven adenocarcinoma cases, two showed asymmetrical enhancement similar to that of the adenoma cases and five showed continued enhancement; one case showed malignant infiltration of the bile duct and evident damage in the adjacent hepatic tissue. The CT plain scan findings for the two groups (adenoma and adenocarcinoma) were not significantly different (t = -1.17, P = 0.2632). Significantly different findings were obtained with the MSCT imaging analysis for the arterial phase (t = 6.53, P less than 0.01) and the portal vein phase (t = 5.63, P less than 0.01). All cases showed asymmetrical expansion of intrahepatic biliary ducts, diffuse or local, and four cases showed moderate expansion of the common bile duct. One adenocarcinoma case showed intumescence in the celiac lymph node by moderate asymmetrical enhancement.

CONCLUSIONMSCT is helpful for the differential diagnosis of IPNB from other hepatic lesions.

Adult ; Aged ; Bile Duct Neoplasms ; diagnostic imaging ; Bile Ducts, Intrahepatic ; diagnostic imaging ; Female ; Humans ; Male ; Middle Aged ; Papilloma, Intraductal ; diagnostic imaging ; Retrospective Studies ; Tomography, X-Ray Computed ; methods

OBJECTIVETo investigate the clinical effect of acellular dermal matrix (ADM) combined with auto-skin grafting on deep burn wound ,and the result of long-term follow-up and histological examination.

METHODSOne hundred and fifty-two patients with deep burn hospitalized from February 2000 to July 2003 were repaired with porcine ADM and auto split-thickness graft. Wound healing rate was assessed 1 week after operation. Degree of cicatricial hyperplasia was examined 1, 3, 6, 12 months after operation. Wound samples from 5 patients were harvested for histological examination 72 months after operation, for which transmission electron microscopy were employed in 2 cases.

RESULTSGrafts completely survived was seen in 116 patients (accounting for 76.3% of cases), survival rate over 95% were observed in 23.7% of cases. One hundred and twenty-seven patients were followed up 1 month after operation, in whom mild local contraction, cord like scar was seen along its junction with skin, its texture was soft ,and there was no pruritus or blister formation. One hundred and one patients were followed up 3 months after operation, and the graft showed mild contraction less marked when compared with that of the site where auto split-thickness skin grafting was used. Articular function was good. Eighty-two patients were followed up 6 months after operation,color and texture of grafts were similar to normal skin with no obvious cicatricial hyperplasia. Fifty-eight patients were followed up 12 months after operation, the texture of grafts was similar to normal skin without obvious reject reaction. Sixteen patients were followed up over 72 months after operation, the grafts appeared dry compared with normal skin. Histological examination showed: tissue structure of grafts was similar to normal skin, intact small sweat gland and sweat gland cells were not found in dermal layer.

CONCLUSIONHeterologous ADM combined with auto split-thickness graft can survive in human body without obvious immune rejection reaction for a long time. No intact small sweat gland or sweat gland cells in dermis is a problem worth of study in regeneration of skin function.

Adolescent ; Adult ; Aged ; Burns ; surgery ; Dermis ; transplantation ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Skin Transplantation ; Skin, Artificial ; Transplantation, Autologous ; Young Adult