Objective To observe the effect of astragaloside IV (A) and SB203580 antagonize cadmium (Cd) toxicity on expression of associated protein and blood-testis barrier(BTB) in rats and to study the protective mechanism of A on it.Methods Totally 21 SD male rats were randomly divided into 7 groups, 3 rats per group:Cd [ intraperitoneally injected with 0.1%CdCl2,1mg/(kg?d)],Cd+A [at the above dose of CdCl2,at the same time with A,10mg/(kg?d)], and Cd +SB203580 [at the above dose of CdCl2,at the same time with SB203580,100μg/(kg?d)], each of the above groups was further divided into continuous five and ten days treatment groups .The control group was intraperitoneally injected with equal dosage of normal saline .The testes were studied by light , electron microscopy , immunohistochemistry and Western blotting .Results In the control group ,irregular and lightly stained nuclei of Sertoli cell ( Sc) in seminiferous tubules were observed by HE staining .A continuous electron density line of tight junction ( TJ) and normal ultrastructure of BTB were observed .After Cd treatment ,the vesicular formation in the Sc was observed .The ultrastructural damage of Sc and TJ was observed .Compared with the corresponding time point of Cd group ,these were weakened in morphology of testis and ultrastructure of TJ after Cd +A or Cd +SB203580 treatment .The positive products of zonula occludens-1 ( ZO-1 ) and claudin-11 were localized mainly in the base of the seminiferous tubule .After Cd treatment , the average absorbance (AA) of ZO-1 and Claudin-11 was decreased significantly compared with that of the control group (P<0.05).After Cd +A or Cd +SB203580 treatment,AA of ZO-1 and Claudin-11 were increased significantly compared with that of the Cd group(P<0.05),though lower than that of the control group .The result of Western blotting showed that phosphorylation-p38MAPK in Cd group was increased significantly compared with that of the control group (P<0.05).After Cd +A or Cd+SB203580 treatment, it was decreased significantly compared with that of the Cd group (P<0.05).Conclusion Cd decreases ZO-1 and Claudin-11 expression and damages ultrastructure of TJ in BTB , asⅣhas protective effect on it , and is related to inhibiting activation of p 38 MAPK pathway .

Bacteria isolated from rotten hides,one identified as Pseudomonas aeraginosa later, were shown to decompose fresh hides completely in 48 hours at room temperature. Strongly collagenolytic activity was detected when native collagen from calfskin was used as substrate. The optimal reaction pH value and temperature of the collagenace produced by P.aeraginosa are 7.5 and 32℃ respectively. EDTA and EGTA inhabit the collagenolytic activity severely while PMSF has little effect on it. Not only media components but also fermentation conditions have differents effect on the production of this collagenolyic enzyme.

Objective To investigate the toxic effect of cadmium (Cd) on the ultrastructure ,expression of related protein and the signal molecule phosphrylated P38 mitogen-actived protein kinase(P-P38MAPK) of primary cultured rat sertoli cell(Sc) ,and the protective effect of astragaloside (A ) on it .Methods The primary cultured rat Sc were divided into the control group ,Cd (50 mol/L)group and Cd(50 mol/L) plus A(10 mg/L) group ,they were used for the electron microscope observation and the im-munohistochemistry detection of vimentin ,E-cadherin ,-catenin and P-P38MAPK .Results The Sc ultrastructural changes included that the swelled mitochondria ,abundant lipid droplets and dilated endoplasmic reticulum were found in the Cd group .Further ,apop-tosis occurred in some Sc .However these ultrastructure changes above mentioned were slighter in the Cd plus A group ;the immu-nohistochemistry showed that the positive products of vimentin ,E-cadherin and-catenin were obviously decreased in the Cd group (P<0 .05) ,and those in the Cd plus A group were higher compared with the Cd group(P<0 .05);the expression of P-P38MAPK in cytoplasm was increased in the Cd group ,and showed the trend to move from cytoplasm to nucleus ,meanwhile ,the positive prod-ucts expression in the Cd plus A group was lower than that in the Cd group (P<0 .05) .Conclusion Cadmium can cause the injury of the Sc ultrastructure ,damage of cytoskeletal protein and fibronectin ,and increase of P-P38MAPK level ;astragaloside can antago-nize the toxicity of cadmium on Sc ,the protective effect maybe related with the decrease of P-P38MAPK in Sc .

BACKGROUND:The production and release of a large amount of inflammatory factors caused by immune system inflammatory response mainly contributes to secondary spinal cord injury. OBJECTIVE:To investigate the effects of umbilical cord Wharton’s jely mesenchymal stem cel transplantation on repair of injured neurological function and expression of inflammatory factors monocyte chemoattractant protein 1 and interleukin 10 in rats with acute spinal cord injury. METHODS: Eighty-one healthy adult male Sprague-Dawley rats were randomly and equaly divided into sham operation, model and cel transplantation groups, with 27 rats per group. Rats in the latter two groups were subjected to hemisection of the spinal cord to establish acute spinal cord injury models. Rat models in the cel transplantation group received umbilical cord Wharton’s jely mesenchymal stem cel injection (1×106)via the tail vein. Rat neurological function was evaluated using the BBB score at different time points after spinal cord injury. The expression of monocyte chemoattractant protein 1 and interleukin 10 in injured spinal cord tissue was detected using ELISA assay at different time points after spinal cord injury. Migration and neuronal differentiation of umbilical cord Wharton’s jely mesenchymal stem cels in the injured spinal cord tissue were determined using immunohistochemical staining method. RESULTS AND CONCLUSION:Compared with the sham operation and model groups, rat neurological function was significantly recovered in the cel transplantation group (P < 0.05). Compared to the model group, monocyte chemoattractant protein 1 level in the serum and monocyte chemoattractant protein 1 mRNA and protein expression in the injured spinal cord tissue were significantly lower (P < 0.05), but interleukin 10 mRNA and protein expression in the injured spinal cord tissue was significantly higher (P < 0.05), in the cel transplantation group. In the cel transplantation group, umbilical cord Wharton’s jely mesenchymal stem cels could migrate to the injured region and express glial fibrilary acidic protein. These findings suggest that umbilical cord Wharton’s jely mesenchymal stem cels promote rat neurological function recovery by regulating the inflammatory response in the injured spinal cord tissue, which is likely to be one of mechanisms by which transplantation of umbilical cord Wharton’s jely mesenchymal stem cels treats spinal cord injury.

Based on site-specific transposition of an expression cassette into a baculovirus shuttle vector (Bacmid) which propagated in Escherichia coli, the Bac-to-Bac System provides a rapid and efficient method to generate recombinant baculoviruses and is widely used for high level expression of heterologous proteins. And the efficiency of recombinant baculovirus infecting cells plays an important role on the protein expression. In this study, we introduced an EGFP expression cassette driven by polyhedrin promoter into the p74 locus of Bacmid by homologous recombination. The target Bacmid-egfp was then transformed into E. coli DH10B containing the transposition helper plasmid to gain a new transposition receipt strain E. coli DH10Bac-egfp. Because of the intact attTn7 sites and lacZ', target gene cloned in a pFastBac vector can be transposed into the Bacmid-egfp shutter vector to construct recombinant baculovirus, which would allow the tracing of the target protein expression and the recombinant Bacmid transfection or recombinant baculoviral infection under fluorescence microscopes. Recombinant virus Bac-egfp-DsRed was constructed by transposing DsRed into the Bacmid-egfp in E. coliDHl0Bac-egfp, and the Sf9 cells infected with the recombinant virus expressed DsRed and EGFP efficiently. Another protein IL-6 fused with 6 x his tag was expressed and purified sucessfully from Sf9 cells infected with recombinant virus Bac-egfp-6 x his-IL6 constructed by the improved Bac-to-Bac system.
Animals ; Baculoviridae ; genetics ; Green Fluorescent Proteins ; genetics ; Interleukin-6 ; biosynthesis ; genetics ; Plasmids ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; Spodoptera

OBJECTIVETo investigate the association between the C311S polymorphism of paraoxonase 2 (PON2) gene and ischemic stroke in Chinese type 2 diabetes mellitus (T2DM) patients.

METHODSA case-control study of 279 Chinese subjects (including 162 T2DM with or without ischemic stroke and 117 non-diabetic control) was performed. Genotype frequencies of C311S polymorphism were studied by polymerase chain reaction-restriction fragment length polymorphism(PCR-RFLP) analysis with DdeI digestion.

RESULTSC311S polymorphism of PON2 gene was detected in Chinese with the C/S allele frequencies 0.145 and 0.855. The frequency distribution showed significant difference between Chinese and Asian Indian. Furthermore, the genotype distribution (SS, CS and CC) of the PON2 C311S gene polymorphism exhibited a significant difference between T2DM patients complicated with ischemic stroke and T2DM without ischemic stroke, the former had a significantly higher C allele frequency(P<0.05).

CONCLUSIONThe above data indicate that the polymorphism at codon 311(Cys --> Ser)in the PON2 gene is associated with ischemic morbidity in Chinese T2DM patients and C allele might be a risk factor.

Adult ; Aryldialkylphosphatase ; genetics ; Asian Continental Ancestry Group ; genetics ; Diabetes Mellitus, Type 2 ; complications ; genetics ; Female ; Humans ; Male ; Middle Aged ; Polymorphism, Genetic ; Stroke ; etiology ; genetics

OBJECTIVETo express P1B, P2A, P3AB and P3D cDNA gene fragments in prokaryotic system using thioredoxin fusion expression system; to investigate the antigenicity and application of recombinant protein.

METHODSBy using PCR technique, P1B, P2A, P3AB and P3D gene fragments were cloned. Choosing M47 as the expressive vector, the recombinant plasmid P1B, P2A, P3AB and P3D was constructed and expressed in Escherichia coli after inducing by IPTG. By anion exchange and affinity chromatography, purified recombinant protein was obtained. By using Western Blot analysis and indirect ELISA to detect its antigenic activity.

RESULTSFour recombinant plasmids was proved to be constructed successfully by sequencing and the correct molecular weight of their expression products. Recombinant proteins were obtained in BL21 (DE3) and purified after Ni2+ affinity chromatography. Western Blot analysis and indirect ELISA showed that P2a had specific antigenicity.

CONCLUSIONThe P2a protein expressed in prokaryotic system was proved to have specific antigenicity. The indirect ELISA distinguished 24 positive sera from 24 negative sera. It is very likely that P2a can be an antigen to diagnose acute patients of hepatitis A and differentiate inactivated vaccine-induced immunity from an infection.

Blotting, Western ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; genetics ; Gene Expression ; Hepatitis A virus ; genetics ; immunology ; metabolism ; Humans ; Polymerase Chain Reaction ; Recombinant Proteins ; immunology ; metabolism ; Viral Proteins ; genetics ; immunology ; metabolism

BACKGROUNDMost of the studies on traditional Chinese medicine (TCM) 'spleen' deficiency syndrome in the recent 30 years were conducted only on the basis of single functional index, neglecting the study on the pathophysiologic internal relationship between spleen deficiency syndrome and gastric diseases in modern medicine. But it was at the subcellular molecular biological level that we explored the pathophysiologic basis of classification of spleen deficiency in chronic gastritis by detecting the bioactive substances in gastric mucosa nuclei and mitochondria.

METHODSBy means of optical microscope, scanning electron microscope (SEM), transmission electron microscopy (TEM) and histochemical staining, we conducted histopathological, subcellular ultrastructural analysis and nuclei and mitochondrial ultrastructural analysis of gastric mucosa of 188 spleen deficiency patients and of 42 voluntary blood donors. At the same time, bioactive substances were measured by means of X-ray energy dispersive analysis system (EDAX) image analysis system, radioimmunoassay method and chemiluminescence method.

RESULTSThe content of cAMP, superoxide dismutase (SOD), Zn and Cu in gastric mucosa, and the content of Zn and Cu in mitochondria decreased progressively in order of groups: healthy control (HC), spleen Qi deficiency without organic lesion (F-SQD), spleen Yang deficiency without organic lesion (F-SyangD), disease without symptoms group, spleen Qi deficiency with organic lesion (G-SQD), spleen Yang deficiency with organic lesion (G-SyangD), spleen Yin deficiency (SyinD) and spleen deficiency with Qi stagnation (SDQS), chronic spleen deficiency gastritis (CSG) and chronic atrophic gastritis (CAG); decreased in order of HC, intestinal metaplasia (IM)Ia, IMIb, IMIIa and IMIIb, P < 0.05. The content of DNA, Zn and Cu in nuclei progressively increased in order mentioned above, P < 0.05.

CONCLUSIONSThe quantitative changes of gastric mucosal cAMP, SOD, Zn, Cu, of mitochondrial Zn, Cu and of nuclear DNA, Zn and Cu are not only the substance base on which the lesion of gastric mucosa tissue structure occurs, but also the substance base on which spleen deficiency is classified. G-SQD and G-SyangD were more likely to be found in low-grade or middle-grade CSG and CAG, while SyinD and SDQS in middle-grade or high-grade CSG, CAG and IMIIb.

Adult ; Aged ; Chronic Disease ; Cyclic AMP ; analysis ; Female ; Gastric Mucosa ; pathology ; ultrastructure ; Gastritis ; metabolism ; pathology ; Humans ; Lipid Peroxides ; blood ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Splenic Diseases ; classification ; Superoxide Dismutase ; analysis

OBJECTIVETo investigate the incidence of radiation-induced maxillary malignancy after radiotherapy for head and neck cancer.

METHODSA total of 273 patients who suffered from osteoradionecrosis after radiotherapy for head and neck cancer were evaluated. Among them, 6 patients were presented with carcinoma and sarcoma arising from maxillary area after radiotherapy.

RESULTSRadiation-induced maxillary cancers happened at a rate of 2.2% in the patients with osteoradionecrosis. There were no statistically significant differences in age, sex and the time interval between the radiotherapy and the cancer occurrence.

CONCLUSIONSRadiation-induced malignancy after radiotherapy for head and neck cancer is mainly located in maxilla, presenting as squamous cell carcinoma or sarcoma of the maxillary sinus.

Adolescent ; Adult ; Aged ; Female ; Head and Neck Neoplasms ; radiotherapy ; Humans ; Incidence ; Jaw Neoplasms ; etiology ; Male ; Middle Aged ; Neck ; diagnostic imaging ; Neoplasms, Radiation-Induced ; Osteoradionecrosis ; etiology ; Radiography ; Radiotherapy ; adverse effects ; Retrospective Studies ; Young Adult

BACKGROUNDMagnetic resonance imaging (MRI) is prone to be deformed by artifacts caused by the presence of metallic materials. The aim of this study was to evaluate the artifacts from galvano-ceramic and metal-ceramic crowns in MRI, in order to analyze their influences on diagnostic interpretation of MRI.

METHODSGalvano-ceramic and metal-ceramic crowns (Bio98, Wiron99, SP-78, BioKC97) were fabricated with the same model. All materials were imaged by means of 1.5T MRI apparatus with three different sequences, T1-weighted spin-echo (T1-weighted SE), T2-weighted spin-echo (T2-weighted SE) and Gradient echo (GE). Mean and standard deviation of distilled water signal intensity (SI) around the sample in the region of interest (500 mm2) enclosing the whole artifacts were determined, and compared for evaluation of the homogeneity of signal intensity. Images around the sample were acquired and evaluated.

RESULTSThere were statistically significant differences in the values of signal intensity between acrylic resin control and BioKC97, Wiron99 in the three sequences (P<0.001). The mean values of signal intensity for Bio98, SP-78 were significantly different from that of acrylic resin control (RE) in GE sequence (P<0.001). No difference was showed between acrylic resin control and galvano-ceramic crown (P>0.05). Images showed that the greatest artifact was a 25 mm ring with distortion in Wiron99 in GE sequence.

CONCLUSIONSThis in vitro study suggested that galvano-ceramic crown had no influence on the MRI, while metal-ceramic crowns caused moderate artifacts in the MRI. Therefore, galvano-ceramic restoration is a valuable alternative in dentistry.

Artifacts ; Ceramics ; Crowns ; Dental Alloys ; Humans ; Magnetic Resonance Imaging ; methods ; Metals