Aging-related change of erectile function and the expression of SIRT1 / 中华男科学杂志

<p><b>Objective</b>To explore aging-related changes in erectile function and the expressions of SIRT1 and other relevant factors in rats.</p><p><b>METHODS</b>We divided 40 male SD rats into four age groups of equal number: 2-month-old (2 mo), 8-month-old (8 mo), 14-month-old (14 mo), and 20-month-old (20 mo), measured the intracavernous pressure (ICP), mean arterial pressure (MAP), and ICP/MAP ratio by electrostimulation of the cavernous nerve, evaluated fibrosis in the corpus cavernosum by Masson's trichrome staining, detected the expressions of SIRT1, P53, and FOXO3a by Western blot, and determined the levels of NO and cGMP using the NO/cGMP kit.</p><p><b>RESULTS</b>Both the ICP/MAP ratio and the cGMP level were elevated with aging, reaching the peak at 8 months and then gradually decreased. Masson staining showed an aging-related increase of collagen fibers in the corpus cavernosum.The expression of SIRT1 was reduced while those of P53 and FOXO3a increased with aging.</p><p><b>CONCLUSIONS</b>Aging-related erectile dysfunction may be attributed to the reduced activity of the NO/cGMP pathway, apoptosis and oxidative stress, and SIRT1 may play a role in aging-related erectile dysfunction.</p>

Application of fast-track surgery concept in the perioperative nursing care of andrological patients: A randomized controlled study / 中华男科学杂志

<p><b>Objective</b>To evaluate the application of the fast-track surgery (FTS) concept in the nursing care of andrological patients during the perioperative period.</p><p><b>METHODS</b>A total of 200 males to be treated by andrological surgery were included in a control group and another 200 in an observation group, the former received conventional perioperative nursing care, while the latter underwent an FTS nursing care procedure including a variety of proven effective methods to reduce surgical stress and achieve a quick recovery during the perioperative period. Comparisons were made between the two groups of patients in the postoperative enterokinesia time, anal exhaust time, eating time, off-bed time, defecating time, bowel preparation complications, and degree of comfort and satisfaction.</p><p><b>RESULTS</b>Compared with the controls, the patients in the observation group showed significantly earlier postoperative enterokinesia time (［5.8±0.9］ vs ［4.4±1.4］ h, P<0.01), anal exhaust time (［10.8±1.8］ vs ［7.7±2.0］ h, P<0.01), eating time (［12.9±0.7］ vs ［6.3±0.7］ h, P<0.01), off-bed time ［14.3±2.7］ vs ［8.2±1.4］ h, P<0.01), and defecating time (［49.2±2.6］ vs ［39.6±2.5］ h, P<0.01), a lower incidence of bowel preparation complications (P<0.01), and a higher degree of comfort (P<0.01) and satisfaction (［97.5±0.7］% vs ［99.4±+0.3］ %, P<0.01).</p><p><b>CONCLUSIONS</b>The FTS concept can be safely and effectively applied to the perioperative nursing care of andrological patients to achieve a faster recovery and higher degree of comfort and satisfaction postoperatively.</p>

Combination of acupressure and magnetic sticker improved the quality of life in patients with advanced gastroenteric tumor: a clinical observation / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To explore the clinical effect of combination of acupressure and magnetic sticker on the quality of life (QOL) including appetite, defecation, and sleep in patients with advanced gastroenteric tumor.</p><p><b>METHODS</b>Totally 147 patients with advanced gastroenteric tumor were assigned to 4 groups according to different treatment methods, i.e., the supportive treatment group (A, 20 cases), the acupressure treatment group (B, 41 cases), the magnetic sticker treatment group (C, 40 cases), and a combination of acupressure and magnetic sticker treatment group (D, 46 cases). They were respectively treated with different methods, supportive treatment for group A, acupressure for group B, magnetic sticker for group C, and a combination of acupressure and magnetic sticker for group D. The scores of food intake, defecation frequency, sleep time, Karnofsky, and QOL were compared before treatment and at day 14 after treatment.</p><p><b>RESULTS</b>After treatment, the scores of food intake, defecation frequency, and sleep time were obviously improved in B, C and D groups (P < 0.01). There was statistical difference between group D and group A (P < 0.01). In addition, in comparison with A group, both Karnofsky score and QOL score increased in B, C and D groups (P < 0.01).</p><p><b>CONCLUSION</b>The assisted therapy of the combination of acupressure and magnetic sticker could ameliorate QOL such as the digestive functions and sleep in patients with advanced gastroenteric tumor.</p>

Establishment and preliminary application of dengue virus envelope domain III IgG antibody capture enzyme-linked immuno-absorbent assay / 中华预防医学杂志

<p><b>OBJECTIVE</b>To establish a highly sensitive and specific assay to detect dengue virus (DENV) envelope protein domain III (EDIII) IgG antibody, and to explore its value in the diagnosis and seroepidemiological survey of dengue.</p><p><b>METHODS</b>The DENV EDIII IgG antibody capture ELISA was developed using the recombinant full-length DENV EDIII, which was prepared by Pichia yeast expression system as the capture antigen. The serum samples were collected from the same group of 35 DENV-1 patients of primary infection during disease period in 2006 and their follow-up phase in 2010; and the sensitivity of the assay was compared to that of the commercial Panbio DENV IgG ELISA.</p><p><b>RESULTS</b>The sensitivity of DENV EDIII IgG ELISA in detecting the serum samples from disease period and follow-up phase was 87% (20/23) and 94% (33/35), respectively; whereas the sensitivity of Panbio DENV IgG ELISA was 71% (25/35) and 0, respectively. The sensitivity of DENV EDIII IgG ELISA in detecting the serum samples from both periods was similar, without statistical significance (χ(2) = 0.946, P = 0.331). For serum samples from disease period, the sensitivity of DENV EDIII IgG ELISA was comparable with that of Panbio DENV IgG ELISA (χ(2) = 1.924, P = 0.165). However, DENV EDIII IgG ELISA demonstrated a significantly higher sensitivity than Panbio DENV IgG ELISA in detecting the serum samples from follow-up phase (χ(2) = 62.432, P = 0.000).</p><p><b>CONCLUSION</b>DENV EDIII IgG capture ELISA is highly sensitive in detecting IgG in the serum samples from either disease period or follow-up phase. This method might be a promising alternative for diagnosis and seroepidemiologic survey of dengue.</p>

Gene cloning and immunogenicity analysis of the structural proteins VP1-VP4 of enterovirus 71 / 南方医科大学学报

<p><b>OBJECTIVE</b>To clone the genes encoding the structural proteins VP1-VP4 of enterovirus 71 and investigate the immunogenicity of the expressed recombinant proteins.</p><p><b>METHODS</b>The VP1-VP4 cDNAs were amplified by RT-PCR from the extracted viral RNA and cloned into pMD19-T vector. The cloned VP1-VP4 genes were then inserted into the multi-cloning sites of plasmid pQE30a and expressed in E. coli M15 with IPTG induction. After washing with 8 mol/L urea and purification with Ni-affinity chromatography, the recombinant proteins obtained were tested for immunogenicity by Western blotting and ELISA using rabbit antisera against enterovirus 71 and Coxsackie Virus A16.</p><p><b>RESULTS</b>The recombinant VP1-VP4 proteins were highly expressed in E. coli M15 and the purified proteins could be specifically recognized by the rabbit sera against enterovirus 71.</p><p><b>CONCLUSION</b>The expressed enterovirus 71 structural proteins show good immunogenicity and can be used for developing enterovirus 71 vaccine and detection kits.</p>

Characterization of specific monoclonal antibodies to Aspergillus conidia by flow cytometry / 南方医科大学学报

<p><b>OBJECTIVE</b>To characterize the specific monoclonal antibodies to Aspergillus conidia.</p><p><b>METHODS</b>Flow cytometry was used to examine the reactivity of the specific monoclonal antibodies to Aspergillus conidia.</p><p><b>RESULTS</b>Both the monoclonal antibodies MA3 and Con2 showed specific reactivity to Aspergillus conidia suspensions. MA3 was capable of binding to the conidia of A.fumigatus, A.flavus, A.niger and A.terreus, while Con2 was reactive only to the conidia of A.fumigatus.</p><p><b>CONCLUSION</b>Two specific monoclonal antibodies (MA3 and Con2) to Aspergillus conidia have been obtained.</p>

Characterization and secreted expression of dengue virus type I-IV envelope glycoprotein domain III in Pichia pastoris / 中华预防医学杂志

<p><b>OBJECTIVE</b>To achieve secretory and extracellular production of recombinant dengue virus serotypes I-IV envelope glycoprotein domain III (DENV-1-4 EDIII) in Pichia pastoris.</p><p><b>METHODS</b>EDIII genes of DENVI-IV were amplified and cloned into vector pPIC9K, respectively. These recombinant plasmids were then linearized and transferred into Pichia pastoris strain GS115. Clones highly produced in 4.0 mg/ml G418 were amplified and induced by methanol to achieve the secreted recombinant proteins. Ni-NTA agarose beads were used for purification, while SDS-PAGE and Western blotting were used for identification.</p><p><b>RESULTS</b>The recombinant plasmids pPIC9K-DENV-1-4 EDIII were constructed and successfully transferred into Pichia pastoris strain GS115. The recombinant EDIII proteins were expressed in a secretory way with the molecular weight about 12 × 10(3) and specifically identified by anti-His monoclonal antibody and anti-DENVI-IV mice sera.</p><p><b>CONCLUSION</b>DENVI-IV EDIII proteins are successfully achieved from Pichia pastoris expression system and could be used for development of dengue vaccines, diagnostic reagents and study of biological function of the E protein.</p>

To evaluate the neutralizing abilities of anti-dengue virus antibodies with nonstructural protein 1 antigen capture enzyme-linked immunosorbent assay / 中华预防医学杂志

<p><b>OBJECTIVE</b>To produce neutralizing antibodies against envelope protein domain III (EDIII) of dengue virus serotype I (DENV-1) and evaluate the nonstructural protein 1 (NS1) antigen capture enzyme-linked immunosorbent assay (ELISA) for identification of antibody neutralizing abilities.</p><p><b>METHODS</b>Five BALB/c mice and one New Zealand Rabbit were immunized with recombinant EDIII protein of DENV-1 for the production of hybridomas and hyperimmune sera. Indirect ELISA, immunofluorescence assay (IFA) and Western Blot analyses were applied to identify specificity of antibodies. Comparing to plaque reduction neutralization test (PRNT), the new established DENV-1 specific NS1 antigen capture ELISA was used for detecting the neutralizing abilities of these antibody.</p><p><b>RESULTS</b>Four strains of monoclonal antibodies (mAbs) named 1A1, 1B3, 3D3 and 9D6 and one hyperimmune serum of rabbit were obtained, all of which were approved to have neutralizing abilities to DENV-1 with the PRNT titer of 1:1024, 1:512, 1:256, 1:4096 and 1:4096. MAb 3D3 with the lowest neutralization titer in PRNT had not shown neutralizing ability to DENV-1 in NS1 antigen capture ELISA, while MAbs 1A1, 1B3 and 9D6 and the rabbit hyperimmune serum could protect the C6/36 from being infected by DENV-1 with the neutralization titer of 1:32, 1:32, 1:128 and 1:128 in this assay.</p><p><b>CONCLUSION</b>NS1 antigen capture ELISA could be used to identify antibody neutralizing abilities to DENV, it was a faster and more convenient way to screen antibodies with high neutralization titer and might also be used as one of the methods to evaluate the effects of vaccines.</p>

Expression and identification of H5 subtype hemagglutinin of avian influenza A virus in insect cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To clone and express avian influenza A virus [A/Hong Kong/482/97(H5N1)] H5 subtype hemagglutinin in baculovirus-insect cell expression system and investigate the antigenicity and bioactivity of the recombinant protein.</p><p><b>METHODS</b>H5 gene of influenza A virus was amplified by PCR. The recombinant bacmid was obtained by cloning the gene to the donor plasmid of pFastBacHTB and transformed into DH10Bac competent cells. The recombinant baculovirus stock was prepared by transfecting the recombinant bacmid DNA into the insect cell line for protein expression after amplification. Immunofluorescene assay (IFA) and Western blotting were performed to identify the antigenicity of the recombinant protein, and hemagglutination assay was used to identify its bioactivity.</p><p><b>RESULTS</b>The recombinant his-H5 protein was expressed in the insect cells with a relative molecular mass of 64,000, which showed erythrocyte-agglutinating activities with the red blood cells of guinea pig. Western blotting and IFA demonstrated that the recombinant his-H5 could be recognized and bound by standard anti-H5 sera.</p><p><b>CONCLUSION</b>The recombinant his-H5 with a post-translation modification is successfully obtained in insect cells, which may provide a potential source for further study of the antigen's biological function and for production of the subunit vaccine or monoclonal antibodies.</p>

Antigenicity analysis of nucleocapsid proteins of 3 human coronaviruses SARS-CoV, 229E and OC43 with their monoclonal antibodies / 南方医科大学学报

<p><b>OBJECTIVE</b>To prepare and characterize monoclonal antibodies (mAbs) against the recombinant nucleocapsid (N) protein of 3 human coronaviruses SARS-CoV, 229E and OC43 and study the antigenic relationship between the 3 N proteins.</p><p><b>METHODS</b>BALB/c mice were immunized with the recombinant N proteins of SARS-CoV, 229E and OC43 to obtain the mAbs by means of hybridoma. Screening and identification of the mAbs were performed using indirect enzyme-linked immunosorbent assay (ELISA), Western blotting and indirect immunofluorescence assay. Cross-reactivity between the N proteins of the 3 coronaviruses was analyzed with the prepared mAbs.</p><p><b>RESULTS</b>The mAbs against the recombinant N proteins of SARS-CoV, 229E and OC43 were obtained, which reacted specifically with the corresponding viral N protein as shown by indirect ELISA, Western blotting and indirect immunofluorescence assay. No cross-reactivity was found between the 3 N proteins.</p><p><b>CONCLUSION</b>The prepared mAbs against the recombinant N proteins may provide valuable assistance in studying antigenic relationships of N proteins between the 3 human coronaviruses.</p>

Development and application of triple antibodies-based sandwich ELISA for detecting nucleocapsid protein of SARS-associated coronavirus / 中华流行病学杂志

<p><b>OBJECTIVE</b>To prepare and characterize monoclonal antibodies (mAb) and polyclonal antibodies against nucleocapsid (N) protein of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) and to establish antibodies-based sandwich ELISA for detecting N protein of SARS-CoV, which might apply to early diagnosis of patients with SARS-CoV infection.</p><p><b>METHODS</b>BALB/c mice were immunized with purified recombinant N protein of SARS-CoV for producing mAbs, and New Zealand white rabbits were immunized for producing polyclonal antibodies. The identification of antibodies was performed using indirect enzyme-linked immunosorbent assay (ELISA), indirect fluorescent-antibody assay (IFA), and Western immunoblotting. Capturing and detecting antibodies were selected by pairing the mAbs and polyclonal antibodies one by one and an antibodies-based sandwich antigen capture ELISA was used for detecting N antigen of SARS-CoV.</p><p><b>RESULTS</b>Nine mAbs and hyperimmune rabbit polyclonal antibodies, specifically against SARS-CoV nucleocapsid protein were obtained. Using paired ELISA assay, three mAbs N1E8, N8E1 and N10E4 were selected as capturing antibody and rabbit polyclonal antibodies as detecting antibody then triple antibodies-based sandwich ELISA was established following horseradish peroxidase (HRP)-conjugated goat anti-rabbit immunoglobulin G. The recombinant N protein was used as a standard to establish a detection sensitivity of approximated 50 pg/ml with this assay. When tested with 420 serum specimens from serologically confirmed SARS patients, the positive rates of serum N protein were 90.1%, 23% and 0%, in which sera collected from 1 to 10 days, 11 to 20 days and beyond 21 days respectively after the onset of symptoms. The specificity of the assay was 99.86% in 715 control serum specimens. There was no cross-reaction with other respiratory viruses and coronaviruses.</p><p><b>CONCLUSION</b>Specific and high affinity mAbs and rabbit polyclonal antibodies were obtained. By paired and optimized sandwich ELISA, a sensitive and specific antigen capture ELISA was established for detecting N antigen of SARS-CoV, which might apply to early diagnosis, source tracing and epidemiological studies of SARS.</p>

Cloning, purification, and antigenic characterization of three recombinant fragments derived from SARS-CoV S1 domain / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>The present study aimed to clone and express three fragments of genomic RNA derived from SARS associated coronavirus (SARS-CoV) S1 domain and to study its immunogenicity.</p><p><b>METHODS</b>The S1 domain gene was amplified by PCR with specific primers and was inserted into the prokaryotic expression vector pQE-30. Three fragments (40-751, 746-1344 and 746-2001 bp) derived from S1 domain produced after the recombinant plasmid (pQE-30/S1) was digested by restriction endonucleases. The three fragments were cloned into pQE-30 and expressed in M15 strains of Escherichia coli. The expression products, designated S1a, S1b and S1c respectively, were purified by Ni affinity chromatography. The immunogenicity was analyzed by Western Blot and ELISA using serologically confirmed sera from SARS patients and the sera from healthy donors was used as control at the same assay.</p><p><b>RESULTS</b>Three recombinant plasmids (pQE-30/S1a, pQE-30/S1b, pQE-30/S1c) were constructed.Fusion proteins with relative molecular mass of 26,700, 22,500 and 46,000 dalton were successfully expressed with amounts of 35%, 35% and 30% of total cell protein and purified by Ni affinity chromatography, respectively. Western Blot and ELISA analysis showed that the S1c protein could be specifically recognized by the sera from SARS patients.</p><p><b>CONCLUSION</b>The recombinant S1c protein was a good immunogen and has the potential to be used as a vaccine against SARS-CoV infection.</p>