Bacteremia induced by periodontal infection is an important factor for periodontitis to threaten general health. P. gingivalis DNA/virulence factors have been found in the brain tissues from patients with Alzheimer's disease (AD). The blood-brain barrier (BBB) is essential for keeping toxic substances from entering brain tissues. However, the effect of P. gingivalis bacteremia on BBB permeability and its underlying mechanism remains unclear. In the present study, rats were injected by tail vein with P. gingivalis three times a week for eight weeks to induce bacteremia. An in vitro BBB model infected with P. gingivalis was also established. We found that the infiltration of Evans blue dye and Albumin protein deposition in the rat brain tissues were increased in the rat brain tissues with P. gingivalis bacteremia and P. gingivalis could pass through the in vitro BBB model. Caveolae were detected after P. gingivalis infection in BMECs both in vivo and in vitro. Caveolin-1 (Cav-1) expression was enhanced after P. gingivalis infection. Downregulation of Cav-1 rescued P. gingivalis-enhanced BMECs permeability. We further found P. gingivalis-gingipain could be colocalized with Cav-1 and the strong hydrogen bonding between Cav-1 and arg-specific-gingipain (RgpA) were detected. Moreover, P. gingivalis significantly inhibited the major facilitator superfamily domain containing 2a (Mfsd2a) expression. Mfsd2a overexpression reversed P. gingivalis-increased BMECs permeability and Cav-1 expression. These results revealed that Mfsd2a/Cav-1 mediated transcytosis is a key pathway governing BBB BMECs permeability induced by P. gingivalis, which may contribute to P. gingivalis/virulence factors entrance and the subsequent neurological impairments.
Animals ; Rats ; Bacteremia/metabolism* ; Blood-Brain Barrier/microbiology* ; Caveolin 1/metabolism* ; Gingipain Cysteine Endopeptidases/metabolism* ; Permeability ; Porphyromonas gingivalis/pathogenicity* ; Transcytosis ; Virulence Factors/metabolism*

Aim To explore the anti-apoptotic function of cardiac progenitor cells(CPCs)-derived exosome in vitro.Method CPCs were isolated from mouse heart using Magnetic Cell Sorting(MACS)system.Flow Cy-tometry(FC)determine the purity of stem cell surface antigen-1 positive(Sca-1 +)CPCs.Exosome was puri-fied from conditional medium,and confirmed by West-ern blot using CD63 as a marker,Nanoparticle Traffic-king Analysis(NTA)was used to detect the diameters and concentration of exosome.Then the cells were di-vided into control groups and CPC-exosome pre-protec-tion groups.H2 O2 was added into H9c2 cells to induce oxidative stress.Western blot was adopted to determine the expression of cleaved caspase-3.Results ① Im-munofluorescence showed that CPCs isolated by MACS were positively expressing Sca-1 protein;FC analysis showed that typical purity of Sca-1 +CPCs from the first
preparations was more than 95%.② WB demonstrated that CD63 of exosome isolated from CCMwas positively expressed,and NTA results showed that the diameters of exosome were (82.33 ±3.06)nm(n =3).Micro-scope detected PKH-26 labeled exosome appeared in the cytoplasma of H9c2 cells.③ Western blot showed the CPC-exosome pre-protection groups significantly down-regulated the levels of cleaved caspase-3 com-pared to the control groups(P <0.05).Conclusion CPC can secrete exosome which carries many important cargos,which can effectively gather in H9c2 cells. CPC-exosome can protect H9c2 cells from the oxidative stress induced by H2 O2 .Our results highlight a new perspective strategy for cardiac disease.

In this study ,the objective is to establish a nested‐PCR assay for the detection of H .bilis with high sensitivity and specificity .The nested primers were designed based on sequences of 16S rRNA gene of seventeen subtypes of H .bilis .Af‐ter optimizing reaction condition ,the sensitivity and specificity of the assay were examined via the detection of feces simulated samples ,mice infection model samples and clinic patients’ samples .The detection sensitivity of H .bilis strain for feces simu‐lated samples was 10 CFU/100 μL .H .bilis was successfully detected in the liver ,caecum and feces of experimentally infected mice .Moreover ,H .bilis was successfully detected in the bile ,cholecyst mucous membrane and feces samples from two of ten patients with cholelithiasis .Due to the PCR assay’s high sensitivity and specificity ,the method may be used to detect the infec‐tion of H .bilis .

Objective To establish ultra-high performance liquid chromatography-evaporative light scattering detector (UPLC-ELSD) method to determine the content of astragaloside in Guiqi Yiqi Yangxue Mixture and optimize the best water decoction process of the mixture.Methods The UPLC-ELSD conditions to determine the content of astragaloside in the mixture were as follow.The method was carried out on a Waters Acquity UPLC BEH C18 column (2.1 mm ×50 mm,1.7 μm) by using acetonitrile-water (35∶65) as mobile phase, and the flow rate was 0.3 mL/min and the column temperature 30℃.The parameter of ELSD:the drift tube temperature was set at 60℃, and the gas flow rate of N2 2.0 L/min.The content of paeoniflorin in the mixture was detected by previously established HPLC method.The orthogonal test was designed to optimize the water decoction process of the amount of water (A), decoction time (B) and decoction times (C), and the extract yield and the contents of astragaloside and paeoniflorin were evaluated as the comprehensive markers for the purpose.Results The UPLC-ELSD determination of astragaloside had a good linear relationship (r =0.9999), its mean recovery was 103.0%, and it had a good precision, specificity and repeatability.The orthogonal experimental results showed that decoction times (C) >the amount of water (A) >decoction time (B) were major factors affecting the decoction process in order and the ANOVA result showed that decoction times has significant effect on the decoction process (P<0.05).The optimal process of Guiqi Yiqi Yangxue Mixture was adding eight-fold water each time to decoct twice for 1.5 h totally.Conclusion This optimized water decoction process is feasible and stable, and suitable for industrialized production.

Objective To investigate the diagnosis and treatment of ectopic para-aortic pheochromocytoma. Methods Clinical data of 2 cases with ectopic para-aortic pheochromocytoma in the Third Affiliated Hospital of Sun Yat-sen University from August 2012 to December 2013 were analyzed retrospectively. The informed consents of both patients were obtained and local ethical committee approval had been received. Both patients were female with the age of 54, 57 years old. The occurrence of disease, diagnosis, treatments and efficacy were observed. Results Case 1 was admitted in hospital for complaint of 1-month distending pain in upper abdominal and ifnding of retroperitoneal space-occupying lesion for 8 d. Solid space-occupying lesion at the right side of mid-upper abdomen was found by abdominal ultrasound and computer tomography (CT) examination. The preoperative diagnosis was abdominal mesenchymoma. Case 2 was admitted in hospital for complaint of palpitation, chest pain and ifnding of hepatic space-occupying lesion for 10 d and had a history of 5-year hypertension. Cystic solid space-occupying lesion in the hepatic caudate lobe was found by abdominal ultrasound, CT and magnetic resonance imaging (MRI) examinations and cystadenoma was suspected. The 2 patients underwent resection of ectopic para-aortic pheochromocytoma under general anesthesia by tracheal intubation. The intraoperative exploration found that touching the tumor had great impacts on blood pressure and heart rate and ectopic para-aortic pheochromocytoma was diagnosed. The patients were transferred to surgical intensive care unit (ICU) for monitoring and treatment. Pheochromocytoma was confirmed by the pathological examination. The patients were recovered and discharged from hospital and remained well till the paper submission date. Conclusions Ectopic para-aortic pheochromocytoma is a rare disease without characteristic clinical features. It is dififcult for preoperative diagnosis. The conifrmation is mainly depends on the pathological ifndings. Surgical resection is the priority of treatment. Fully assessment of operative risk before operation, avoiding touching tumor tissues during operation, complete excision of tumor capsule, close monitoring on the blood pressure after operation can achieve a good clinical treatment outcome.

Human cytomegalovirus glycoprotein complex Ⅱ (gC Ⅱ ) consists of two glycoproteins, gM and gN. Although gC Ⅱ specific IgG purified from HCMV positive patient sera can neutralize HCMV, there has been no report on the generation of virus-neutralizing antibodies by immunizing with one epitope of gM. The epitope, termed MAD, was screened from random phage peptide library by subtractive strategy. The peptide sequence of MAD was highly homologous with 32~38 amino acids of HCMV gM. Mice immunized with MAD coupled with keyhole limpet hemocyanin (KLH) could produce specific antibodies against MAD, and the antibodies obtained could bind not only native HCMV particles, but also the recombinant gM30～78 peptide. ELISA analysis results showed that MAD could specifically bind HCMV-positive human serum samples. Virus-neutralizing assay results demonstrated that the antibodies against MAD could inhibit HCMV strain AD169 entering the human embryonic lung cells. The results suggested that MAD could be used as a new potential protective antigen in the development of HCMV vaccine.

OBJECTIVETo search for the dystrophin gene mutations of Duchenne muscular dystrophy (DMD) patients without gross deletions, in order to offer accurate genetic counseling and prenatal diagnosis for those families.

METHODSAll 79 exons of the dystrophin gene as well as its 5'-UTR and 3'-UTR of 14 Chinese DMD/Becker muscular dystrphy (BMD) patients without detectable gross deletions were screened by denaturing high performance liquid chromatography (DHPLC) and heteroduplex fragments were identified by subsequent sequencing.

RESULTSSeven causative point mutations, including two novel ones, were detected in 7 patients. Fourteen known polymorphisms and 7 unknown intronic variations were also detected. Five mothers of the patients were obligate carriers.

CONCLUSIONDHPLC is an efficient way of identifying point mutations and the female carriers in DMD families.

Adolescent ; Asian Continental Ancestry Group ; genetics ; Base Sequence ; Child ; Child, Preschool ; DNA Mutational Analysis ; Dystrophin ; genetics ; Exons ; genetics ; Female ; Genetic Counseling ; Genetic Testing ; methods ; Humans ; Introns ; genetics ; Male ; Muscular Dystrophy, Duchenne ; diagnosis ; genetics ; Point Mutation ; Polymorphism, Genetic ; Pregnancy ; Prenatal Diagnosis ; Sequence Deletion ; genetics

Objective To evaluate a thrombolytic system of portal vein port-catheter kit (PC) in the treatment of portal vein thrombosis (PVT). Methods In this study, 42 PVT patients with liver cirrhosis and portal hypertension after splenectomy from 2005 to 2007 were divided into two groups. In group A (20 eases) thrombolysis was administered through the PC device. Urokinase at the dosage of 1000 U?kg-1?h-1 was given for a consecutive 3 -6 days through the PC, and then the therapy was converted to 100 AxaIU/kg of low molecular heparin twice a day for 7 days subcutaneously. In group B, the thrombolysis was performed on 22 patients through peripheral veins. The therapy was same as in group A except for that the urokinase dosage was doubled. The complete thrombolysis rate, the effective thrombolysis rate, the time of thrombolysis, the long-term recurrence rate and the incidence of complication were compared between the two groups. Results The complete thrombolysis rate and the effective thrombolysis rate in group A were 75%, 90% respectively, compared with that of 41%, 59% respectively in group B. The significant differences in the complete thrombolysis rate, the effective thrombolysis rate, the time of thrombolysis and the incidence of complication were found between the two groups, while the thrombolysis recurrence rate had no significant difference between the two groups. Conclusion PC regime is an effective and safe method for the treatment of portal vein thrombosis.

Objective To isolate and identify the potential binding partners of LRRK2,a gene linked to both dominant familial form and sporadic form of Parkinson's disease,thus to further our knowledge of its function.Methods We used a sequence containing full-length of COR domain and part of ROC and MAPKKK domain as bait.The bait amplified by polymerase chain reaction(PCR) was then cloned into a yeast expression plasmid pGBKT7.After being sequenced and analyzed,pGBKT7-bait was transformed into the yeast strain AH109.Western blot was performed to confirm the expression of pGBKT7-bait in AH109 yeast strain.Then human fetal brain cDNA library was trarnsformed into that yeast strain.which could express pGBKT7-bait fusion protein.The yeast strain which contained pGBKT7-bait and human fetal brain cDNA library was plated on quadruple dropout medium (SD/-Trp/-Leu/-His/-Ade)containing X-a-gal.We retested these positive colonies using 2 independent yeast strains AH109 contained pGBKT7-bait or pGBKT7,respectively.At last,these plasmids isolated from these true positive colonies were analyzed by bioinformatics.Results We obtained 9 true positive colonies,these colonies were sequenced, and we performed sequence Blast in GenBank.Three colonies of the 9 positive colonies were not in open reading-frames.Among other 6 colonies,there were known proteins including spermatid perinuclear RNA-binding protein(STRBP)and BCL2-associated athanogene 5 isoform b(BAG5),as well as unknown proteins including tyrosine phosphatase non-receptor type(PTPN23),1(3)mbt-like 3 isoform b(L3 MBTL3),RALY RNA binding protein-like isoform 1(RALYL),and Homo sapiens mRNA for KIAA1783 protein,partial cds(KIAA 1783).Conclusion True positive colonies of LRRK2 are successfully obtained by the yeast 2-hybrid.Our screened proteins may provide a new research clue for revealing biological functions of LRRK2,pathogenesis of Parkinson's disease,and other neurodegerations.

Objective To investigate the status and influencing factors of knowledge,attitude and practice(KAP) about AIDS among the migrant miners in a nonferrous metal company of Anhui Province and to provide evidence for constituting strategies and measures for AIDS health education and behavioural intervention.Methods 303 migrant miners in a nonferrous metal company were surveyed.Univariate analysis,Logistic regression and correlation analysis were used to explore their influencing factors.Results The general rate of awareness of AIDS was 87.9%,and the main factor affecting miners knowledge level of AIDS was their educational level.Only 55.1% of migrant miners thought people with AIDS should not be isolated;50.8% of them would like to contact people with HIV;38.0% did not object to extramarital sex.There were 32(10.6%) migrant miners admitting having had extramarital sex in the recent three months,but 11(34.4%) admitted never having used condoms,only 7(21.9%) having used each time.Multivariate analysis identified the influencing factors for condom utilization including age and attitude to people with HIV/AIDS.Conclusions ①The migrant miners have sufficient knowledge about AIDS.②The traditional sexual beliefs are changing,and an increasing number of people are tolerant to extramarital sex.Consequently,it is likely that extramarital sex might rise.③The promotion of condom use should be strengthened.