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1.
Medicine (Baltimore) ; 102(38): e35126, 2023 Sep 22.
Artículo en Inglés | MEDLINE | ID: mdl-37746991

RESUMEN

BACKGROUND: Research on opioid-free anesthesia has increased in recent years; however, it has never been determined whether it is more beneficial than opioid anesthesia. This meta-analysis was primarily used to assess the effect of opioid-free anesthesia compared with opioid anesthesia on the incidence of postoperative nausea and vomiting. METHODS: We searched the electronic databases of PubMed, the Cochrane Library, Web of Science and Embase from 2014 to 2022 to identify relevant articles and extract relevant data. The incidence of postoperative nausea and vomiting, time to extubation, pain score at 24 hours postoperatively, and time to first postoperative rescue analgesia were compared between patients receiving opioid-free anesthesia and those receiving standard opioid anesthesia. Differences in the incidence of postoperative nausea and vomiting were evaluated using risk ratios (95% confidence interval [CI]). The significance of the differences was assessed using mean differences and 95% CI. The heterogeneity of the subject trials was evaluated using the I2 test. Statistical analysis was performed using the RevMan 5.4 software. RESULTS: Fourteen randomized controlled trials, including 1354 participants, were evaluated in the meta-analysis. As seen in the forest plot, the OFA group had a lower risk of postoperative nausea and vomiting than the control group (risk ratios = 0.41, 95% CI: 0.33-0.51, P < .00001; n = 1354), and the meta-analysis also found that the OFA group had lower postoperative analgesia scores at 24 hours (P < .000001), but time to extubation (P = .14) and first postoperative resuscitation analgesia time (P < .54) were not significantly different. CONCLUSIONS: Opioid-free anesthesia reduces the incidence of postoperative nausea and vomiting while providing adequate analgesia without interfering with postoperative awakening.


Asunto(s)
Anestesia , Anestesiología , Humanos , Analgésicos Opioides/efectos adversos , Incidencia , Náusea y Vómito Posoperatorios/epidemiología , Ensayos Clínicos Controlados Aleatorios como Asunto
2.
Int J Surg Case Rep ; 110: 108660, 2023 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-37579630

RESUMEN

INTRODUCTION: Herein, we describe a case of Meigs' syndrome, a complex condition that poses a challenge for anesthesiologists to manage. Good anesthetic management of this syndrome is necessary to preserve the prognosis. PRESENTATION OF CASE: An 80-year-old woman was admitted to the emergency department with complaints of abdominal pain, particularly in the left lower abdomen, with aggravation after activity. The patient was unable to sleep in a supine position. Her serum carbohydrate antigen 125 level was 253.15 U/mL-1, and laboratory examinations were nonspecific. On auscultation, breath sounds were absent from the base of the right lung. Abdominal computed tomography (CT) was performed to screen for a possible tumor consisting of both solid and cystic components, but the findings were inconclusive. Chest CT showed large right pleural effusions and hiatal hernia. DISCUSSION: A multidisciplinary team conducted careful preoperative preparation, while the anesthesiology team prepared detailed peri-anesthesia management strategies to regulate acid-base and electrolyte balance and maintain respiratory and hemodynamic stability. The surgeon resected the tumor successfully. The patient was discharged after 1 week. A postoperative pathology test confirmed fibrothecomas. CONCLUSION: We provided an effective strategy for the anesthetic management of Meigs' syndrome, which remains a complex challenge for anesthesiologists. It is important that anesthesiologists perform adequate preoperative evaluation and prudent peri-anesthesia management to ensure that patients have a good prognosis and discharge healthily. A multidisciplinary team is essential when caring for patients with Meigs' syndrome.

3.
Aging Clin Exp Res ; 35(9): 1873-1879, 2023 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-37479909

RESUMEN

OBJECTIVE: To explore the correlation between intraoperative burst suppression (BS) and postoperative delirium (POD) in elderly patients, and provide more ideas for reducing POD in clinical. METHODS: Ninety patients, aged over 60 years, who underwent lumbar internal fixation surgery in our hospital were selected. General information of patients was obtained and informed consent was signed during preoperative visits. Patients were divided into burst suppression (BS) group and non-burst suppression (NBS) group by intraoperative electroencephalogram monitoring. Intraoperative systolic blood pressure (SBP), diastolic blood pressure (DBP), mean arterial pressure (MAP), and heart rate (HR) were recorded, and the variation and minimum value were obtained by calculating. Hemoglobin (HGB), C-reactive protein (CRP), system immune inflammatory index (SII) at 24 and 72 h after surgery, the incidence of postoperative adverse reactions, postoperative hospital stay, and total cost were recorded after operation. POD assessment was performed using CAM within 7 days after surgery or until discharge. SPSS25.0 was used for statistical analysis. RESULTS: Compared with the NBS group, the number of elderly patients with high frailty level in BS group was more (P = 0.048). There is correlation between BS and POD (OR: 4.954, 95%CI 1.034-23.736, P = 0.045), and most of the POD patients in BS group behave as hyperactive type. CONCLUSION: The occurrence of intraoperative BS is associated with POD, and elderly patients with frailty are more likely to have intraoperative BS. BS can be used as a predictor of POD.


Asunto(s)
Delirio del Despertar , Fragilidad , Anciano , Humanos , Persona de Mediana Edad , Estudios Prospectivos , Proteína C-Reactiva , Electroencefalografía
4.
Front Med (Lausanne) ; 9: 989341, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-36186818

RESUMEN

Background: Controlled hypotension technique was usually used to reduce intraoperative bleeding, and it could improve visualization of the surgical field during total knee arthroplasty (TKA). However, inappropriate controlled hypotension, through reducing cerebral blood flow or cerebral perfusion pressure, may cause postoperative cognitive dysfunction (POCD), so it is important to identify the appropriate level of controlled hypotension. Objective: To investigate the effects of different levels of controlled hypotension on regional cerebral oxygen saturation and postoperative cognitive function in patients undergoing TKA. Methods: Patients meeting inclusion criteria were enrolled through preoperative visits and basic information was obtained. The patients were randomly divided into three groups: Group A, MAP was maintained at 90-100% of the baseline; Group B, MAP was maintained at 80-90% of the baseline; Group C, MAP was maintained at 70-80% of the baseline. The MAP, HR, and rSO2 were observed and recorded during the operation. The C-reactive protein (CRP), hemoglobin (Hb) and MMSE score at 1, 3, and 7 days after operation were recorded. SPSS25.0 was used for data analysis. Result: When the MAP had a decrease among the three groups, rSO2 did not decrease significantly, and none of the patients experienced POCD which was measured by MMSE. And there was no correlation between the decline in rSO2 and that in MAP. Conclusion: No POCD was experienced in the three groups, and we recommend that the controlled hypotensive target indicated by MAP was maintained at 70-80% of the baseline which not only decreases intraoperative bleeding and improve the quality of the surgical field, but also is still within safe levels.

5.
Biomed Res Int ; 2022: 5372603, 2022.
Artículo en Inglés | MEDLINE | ID: mdl-35224095

RESUMEN

BACKGROUND: Cognitive dysfunction after total knee arthroplasty (TKA) is very common in elderly patients. Postoperative cognitive dysfunction (POCD), as a form of cognitive dysfunction, may affect patients' short- and long-term recoveries. The identification of meaningful risk factors may help reduce the occurrence of POCD in the future. OBJECTIVE: Our goal was to retrospectively investigate the risk factors for early POCD in elderly patients undergoing TKA and to further analyze the relationship between the intensity of risk factors and the level of cognitive function. METHODS: The related indicators and the Montreal Cognitive Function Assessment Scale (MOCA) scores of 105 elderly patients were collected by searching the electronic case system. According to the postoperative MOCA score, patients were divided into three groups: normal group (group N), mild POCD group (group M), and severe POCD group (group S). SPSS 25.0 software was used for statistical analyses. RESULTS: At baseline, the preoperative MOCA score was significantly different in patients with POCD (P ≤ 0.001), while other baseline indicators were not significantly different. In terms of changes in hemoglobin levels, statistically significant differences were observed between group M, group S, and group N (P = 0.039). Among inflammatory indicators, only postoperative CRP levels showed a statistically significant difference in patients with POCD (P = 0.041). Postoperative pain was also significantly different among the three groups (P = 0.009). The multivariate regression analysis revealed that a low preoperative MOCA score and severe postoperative pain were independent risk factors for mild and severe cognitive impairment, while a high postoperative CRP level was only an independent risk factor for mild cognitive impairment. CONCLUSIONS: Our study found that the level of preoperative cognitive function, postoperative CRP level, and postoperative pain were independent risk factors for POCD. Moreover, the levels of preoperative cognitive function and postoperative pain were more strongly correlated with severe POCD than postoperative CRP levels.


Asunto(s)
Artroplastia de Reemplazo de Rodilla , Complicaciones Cognitivas Postoperatorias/epidemiología , Anciano , China/epidemiología , Femenino , Humanos , Masculino , Estudios Retrospectivos , Factores de Riesgo
6.
Biochim Biophys Acta Mol Basis Dis ; 1868(1): 166292, 2022 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-34710568

RESUMEN

Tuberculosis (TB) remains a major cause of mortality and morbidity worldwide, and it is instant to discover novel anti-TB drugs due to the rapidly growing drug-resistance TB. Mycobacterium tuberculosis (Mtb) secreted effector ESAT6 plays a critical role in modulation miRNAs to regulate host defense mechanisms during Mtb infection, it can be a possible target for new tuberculosis drugs. The non-tuberculous mycobacteria Mycobacterium smegmatis (M. smegmatis) and Mtb have high gene homology but no pathogenicity. We used ESAT6 to interfere with macrophages or mice infected by M. smegmatis and determined that it enhanced the survival rate of bacteria and regulated miR-222-3p target PTEN. Expression of miR-222-3p reduced and PTEN enhanced with the progression of macrophages infected by M. smegmatis with ESAT6 co-incubation. MiR-222-3p overexpression diminished M. smegmatis survival and upregulated proinflammatory cytokines. VO-Ohpic trihydrate (PTEN inhibitor) reduced M. smegmatis survival and upregulated proinflammatory cytokines in vivo and in vitro, and VO-Ohpic trihydrate reversed the tissue damage of mouse organs caused by ESAT6. These results uncover an ESAT6 dependent role for miR-222-3p and its target PTEN in regulating host immune responses to bacterial infection and may provide a potential site for the development of anti-tuberculosis drugs that specifically antagonize the virulence of ESAT6.


Asunto(s)
Antígenos Bacterianos/genética , Proteínas Bacterianas/genética , MicroARNs/genética , Fosfohidrolasa PTEN/genética , Tuberculosis/genética , Animales , Modelos Animales de Enfermedad , Interacciones Huésped-Patógeno/genética , Humanos , Inmunidad Innata/genética , Ratones , Mycobacterium smegmatis/genética , Mycobacterium smegmatis/patogenicidad , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/patogenicidad , Tuberculosis/inmunología , Tuberculosis/patología
7.
J Orthop Surg Res ; 15(1): 530, 2020 Nov 12.
Artículo en Inglés | MEDLINE | ID: mdl-33183324

RESUMEN

BACKGROUND: There are various techniques to reduce blood loss in total knee arthroplasty (TKA), including the use of a tourniquet and tranexamic acid (TXA). In this study, we studied the combined effect of TXA with a tourniquet on blood loss in the setting of primary TKA. METHODS: Randomized controlled trials (RCTs) of nine treatment methods were included (placebo, intravenous [i.v.] TXA, topical TXA, i.v.-combined topical TXA, oral TXA, placebo + tourniquet, i.v. TXA +tourniquet, topical TXA + tourniquet, and i.v.-combined topical TXA + tourniquet). The patients were divided into eight groups according to the different treatment strategies, with 30 cases per group. The differences in the total blood volume, the number of patients transfused, the hemoglobin before and after the operation, and complications after the operation were compared. RESULTS: Totally 15 RCTs meeting our inclusion criteria were collected in this study. Compared with the placebo + tourniquet group, the i.v. TXA + tourniquet group displayed lower hemoglobin reduction value, pulmonary embolism (PE) incidence, total blood loss, and blood transfusion risk; the topical TXA + tourniquet group showed reduced PE incidence, total blood loss, and blood transfusion risk, and the i.v.-combined topical TXA and i.v.-combined topical TXA + tourniquet groups showed decreased total blood loss and lower blood transfusion risk. Retrospective clinical study results also demonstrated that the efficacy of i.v.-combined topical TXA was the best. CONCLUSIONS: Our meta-analysis indicates that i.v.-combined topical TXA provides a low total blood loss without increasing the blood transfusion risk in patients undergoing total knee replacement surgery.


Asunto(s)
Artroplastia de Reemplazo de Rodilla/efectos adversos , Pérdida de Sangre Quirúrgica/prevención & control , Técnicas Hemostáticas , Torniquetes , Ácido Tranexámico/administración & dosificación , Administración Tópica , Anciano , Estudios de Casos y Controles , Terapia Combinada , Femenino , Estudios de Seguimiento , Humanos , Infusiones Intravenosas , Masculino , Persona de Mediana Edad , Ensayos Clínicos Controlados Aleatorios como Asunto , Estudios Retrospectivos , Resultado del Tratamiento
8.
Anaesth Crit Care Pain Med ; 36(2): 123-124, 2017 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-28012999

RESUMEN

We describe the anaesthetic management for caesarean section in a 32-year-old patient with pituitary dwarfism. In addition to supportive treatment, we offered a postoperative epidural analgesia pump. The patient recovered well without any complications.


Asunto(s)
Anestesia Epidural/métodos , Anestesia Obstétrica/métodos , Cesárea/métodos , Enanismo Hipofisario/complicaciones , Femenino , Humanos , Embarazo , Resultado del Tratamiento
9.
Pak J Pharm Sci ; 28(1 Suppl): 325-8, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25631510

RESUMEN

In order to explore two kinds of nano-liposomes in lidocaine hydrochloride nano-liposomes on in vitro permeability of drug, and conduct comparison and analysis, this paper investigates cumulative infiltration situation of lidocaine hydrochloride flexible nano-liposomes and ordinary nano-liposomes by using modified Franz diffusion pool on mice vitro skin. Cumulative osmotic quantity of lidocaine hydrochloride flexible nano-liposomes for 9h was higher than ordinary nano-liposomes. (t)max(Maximum osmotic quantity time) of lidocaine hydrochloride flexible nano-liposomes and ordinary nano-liposomes in mice skin was 5 and 60min, the former (C)max (maximum dosage time) was 1.2 times of the latter. Drug was not found in mice plasma of ordinary nano-liposomes group, traces of drugs was detected in 0.5 and 1h in flexible nano-liposomes group, but the concentration was lower than the effective concentration. Compared with the classic skin transparent promoter and ordinary liposome, flexible nano-liposomes have more advantages, but its stability is less than ordinary nano-liposomes because of the addition of surface active substance. Flexible nano-liposomes have great development potential as a carrier of transdermal drug delivery field.


Asunto(s)
Anestésicos Locales/administración & dosificación , Lidocaína/administración & dosificación , Piel/metabolismo , Animales , Técnicas In Vitro , Lidocaína/farmacocinética , Liposomas , Ratones , Nanopartículas , Permeabilidad
10.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-271767

RESUMEN

As the dominant antigens, early secreted antigenic target 6 (ESAT6, E6) and culture filtrate protein 10 (CFP10, C10) had once been the focus of tuberculosis (TB) vaccine due to their capability of inducing strong cell immune response in the host. They are also endowed with promising future of prevention against and diagnosis of TB. In this review, we systematically introduce recent research progress of E6 and C10, especially in structure-function, biological characteristics, protein expression and secretion, host immunity and vaccine development, and the prospects of their application are also discussed.


Asunto(s)
Humanos , Antígenos Bacterianos , Química , Genética , Alergia e Inmunología , Proteínas Bacterianas , Química , Genética , Alergia e Inmunología , Epítopos Inmunodominantes , Alergia e Inmunología , Biología Molecular , Fragmentos de Péptidos , Química , Genética , Alergia e Inmunología , Vacunas contra la Tuberculosis , Genética , Alergia e Inmunología , Vacunas de ADN , Alergia e Inmunología
11.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-325114

RESUMEN

<p><b>OBJECTIVE</b>To clone and express the Rv3871 gene related to the virulent protein secretion of Mycobacterium tuberculosis and analyze its molecular structure, function and homology using bioinformatic approach.</p><p><b>METHODS</b>A pair of primers was designed to amplify the Rv3871 gene, which was subcloned into the prokaryotic plasmid pET32a(+). The recombinant plasmid was identified by sequence analysis and the expressed recombinant protein by SDS-PAGE. The structure, function and homology alignment of Rv3871 were analyzed comparatively against other mycobacteria.</p><p><b>RESULTS</b>The restriction fragments through molecular cloning matched perfectly in size with our prediction. The gene sequence was consistent with the corresponding sequence in GenBank. The expression protein was detected by SDS-PAGE with a molecular weight of 84 kD. Two FtsK/SpoE III domains were found by bioinformatic analysis. The homology results showed distinct differences between Rv3871 of the pathogenic M. tuberculosis and its counterparts in non-pathogenic mycobacteria.</p><p><b>CONCLUSION</b>Molecular cloning, expression and sequencing identify the structural and functional characteristics of Rv3871. The structural and functional differences of the gene between pathogenic and non-pathogenic mycobacteria identified by bioinformatics provide some evidence for the pathogenesis and drug targets of tuberculosis.</p>


Asunto(s)
Proteínas Bacterianas , Genética , Metabolismo , Clonación Molecular , Biología Computacional , Mycobacterium tuberculosis , Genética , Metabolismo , Virulencia , Proteínas Recombinantes , Genética , Metabolismo , Virulencia , Genética
12.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-280194

RESUMEN

This study was conducted to construct eukaryotic recombinant vector of LipL32-HlyX fusion gene from Leptospira serovar Lai and express it in mammalian cell. Both of LipL32 gene and HlyX gene were amplified from Leptospira strain O17 genomic DNA by PCR. Then with the two genes as template, LipL32-HlyX fusion gene was obtained by SOE PCR (gene splicing by overlap extension PCR). The fusion gene was then cloned into pcDNA3.1 by restriction nuclease digestion. Having been transformed into E. coli DH5alpha, the recombiant plasmid was identified by restriction nuclease digestion, PCR analysis and sequencing. The recombinant plasmid was then transfected into COS7 cell whose expression was detected by RT-PCR and Western blotting analysis. RT-PCR amplified a fragment about 2000 bp and Western blotting analysis found a specific band about 75 KD which was consistent with the expected fusion protein size. In conclusion, the successful construction of eukaryotic recombinant vector containing LipL32-HlyX fusion gene and the effective expression in mammalian have laid a foundation for the application of Leptospira DNA vaccine.


Asunto(s)
Animales , Proteínas de la Membrana Bacteriana Externa , Genética , Metabolismo , Células COS , Chlorocebus aethiops , Fusión Génica , Vectores Genéticos , Secuencias Hélice-Asa-Hélice , Genética , Leptospira , Genética , Lipoproteínas , Genética , Metabolismo , Proteínas Recombinantes de Fusión , Genética , Metabolismo
13.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-384015

RESUMEN

Objective To study on the expression of the eukaryotic recombinant vector carrying HlyX gene of Leptospira serovar Lai in mammalian cell and explore the humoral immune response in BALB/c mice immunized with the recombinant plasmid. Methods The HlyX gene was amplified from Leptospira serovar Lai genomic DNA by PCR and inserted into pcDNA3.1 vector. After transformed into E. coli DH5α,the recombinant plasmid was assayed for identification by PCR analysis,restriction nuclease enzyme digestion and sequencing. The recombinant plasmid was transfected into COS-7 cells,then RT-PCR and Western blot were performed to test the expression of the target gene. The recombinant plasmid was injected intramuscularly into BALB/c mice for three times at intervals of two weeks,and the antibody titer was measured by ELISA. Results PCR showed the full length HlyX gene was about 1100 bp. PCR analysis,restriction nuclease enzyme digestion and sequencing indicated the recombinant vector was constructed successfully. After the plasmid Was transfected into COS-7 cells,a fragment about 1100 bp was found by RT-PCR and a specific band relative molecular mass(Mr)about 40×103,which was consistent with the expected size of the target proteins was showed by Western blot. ELISA showed the antibody titer in BALB/c mice immunized by the HlyX gene of Leptospira serovar Lai can elicit high-titer antibody in BALB/c mice,which has laid the foundation for the application of the DNA vaccine.

14.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-298255

RESUMEN

<p><b>OBJECTIVE</b>The mechanism by which M.tuberculosis persists and survives in host macrophage is not fully understood, however, the M. tuberculosis chromosome-encoded TA loci perform functions possibly of signaling to these processes. To explore the biological functions of M. tuberculosis chromosome-encoded TA loci, the Rv1494 and Rv1495 genes of M.tuberculosis H37Rv strain were cloned and expressed.</p><p><b>METHODS</b>The hypothetical proteins Rv1494 and Rv1495 were bioinformatically analyzed by means of Bioedit software, Dnaman software and Pfam database. The complete open-reading frame sequences of Rv1494 and Rv1495 genes were amplified by PCR using M.tuberculosis H37Rv genomic DNA as the template, and the PCR products were cloned into prokaryotic expression vector pET32a(+), respectively. After induction of expressions in E.coli host strain BL21 (DE3), the recombinant proteins were purified and detected by Western blotting.</p><p><b>RESULTS</b>According to bioinformatic analysis, the hypothetical proteins of Rv1494 and Rv1495 genes shared some homologies with mazEF family, one of E. coli chromosomal TA loci (homology at 26% and 29.5%). Sequence analysis showed that the inserted target genes and its reading frames were completely correct. The recombinant plasmids were induced with IPTG to effectively express the fusion proteins with relative molecular mass coincident with prediction. The specific positive signals were identified from the immunoblots.</p><p><b>CONCLUSION</b>For the first time, the Rv1494 and Rv1495 genes of M.tuberculosis H37Rv strain were cloned and its prokaryotic expression vectors were constructed successfully in this experiment, which may facilitate further functional study of this mazEF-like gene pair.</p>


Asunto(s)
Proteínas Bacterianas , Genética , Metabolismo , Western Blotting , Clonación Molecular , Electroforesis en Gel de Poliacrilamida , Escherichia coli , Genética , Regulación Bacteriana de la Expresión Génica , Genes Bacterianos , Vectores Genéticos , Genética , Mycobacterium tuberculosis , Genética , Metabolismo , Reacción en Cadena de la Polimerasa , Proteínas Recombinantes de Fusión , Genética , Metabolismo
15.
Journal of Biomedical Engineering ; (6): 1298-1302, 2006.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-331426

RESUMEN

This study was conducted to amplify the cfpl0-esat6 fusion gene by SOE and insert into the integrating shuttle plasmid pMV361 to form the recombinant plasmid. Then another recombinant plasmid was constructed by insertinga-A g signal sequence of BCG. The two recombinant plasmids were introduced into BCG and the induced products from recombinant BCG were analyzed. In conclusion,the successful construction of rBCG expressing the fusion protein CFP10-ESAT6 will be the base of the development of novel Mycobacterium tuberclosis vaccines.


Asunto(s)
Antígenos Bacterianos , Genética , Proteínas Bacterianas , Genética , Mycobacterium bovis , Genética , Metabolismo , Mycobacterium tuberculosis , Genética , Plásmidos , Proteínas Recombinantes de Fusión , Genética , Alergia e Inmunología , Vacunas contra la Tuberculosis
16.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-282884

RESUMEN

<p><b>OBJECTIVE</b>Objective To construct recombinant Mycobacterium smegmatis expressing ESAT-6 of the human pathogen Mycobacterium tuberculosis.</p><p><b>METHODS</b>ESAT-6 gene was amplified from M. tuberculosis genomic DNA and inserted into an E.coli-mycobacterium shuttle vector under the control of HSP60 promoter. The recombinant vector was transformed into M. smegmatis by electroporation. To assess the ability of recombinant M. smegmatis to activate macrophage, mouse macrophage ANA-1 was cocultured with recombinant M. smegmatis. The apoptosis of ANA-1 cells was detected by flow cytometry and iNOS mRNA expression of the cells was detected by reverse transcription-polymerase chain reaction (RT-PCR). The survival of M. smegmatis strains in ANA-1 cells was evaluated.</p><p><b>RESULTS</b>The recombinant vector was verified by restriction endonuclease digestion and DNA sequencing. ESAT-6 protein was expressed in M. smegmatis in response to heat shock and the molecular weight of the expression product was identical to the expected value. The growth curve of the new recombinant M. smegmatis was consistent with that of the wild-type strain, suggesting the absence of ESAT-6 protein toxicity against M. smegmatis. The recombinant M. smegmatis did not induce significant changes in mouse macrophage ANA-1 apoptosis. Coculture of the macrophages with recombinant M. smegmatis for 4 to 24 h could induce iNOS expression in the former, and the CFU of recombination M. smegmatis grown in ANA-1 cells was much less than that of the control bacteria.</p><p><b>CONCLUSION</b>The recombinant M. smegmatis expressing M. tuberculosis ESAT-6 gene possess immunogenicity, which provides experimental evidence for the development of novel M. smegmatis-based vaccine against tuberculosis.</p>


Asunto(s)
Animales , Humanos , Ratones , Antígenos Bacterianos , Genética , Alergia e Inmunología , Apoptosis , Alergia e Inmunología , Proteínas Bacterianas , Genética , Alergia e Inmunología , Línea Celular , Electroforesis en Gel de Poliacrilamida , Escherichia coli , Genética , Citometría de Flujo , Vectores Genéticos , Activación de Macrófagos , Alergia e Inmunología , Macrófagos , Biología Celular , Alergia e Inmunología , Metabolismo , Mycobacterium smegmatis , Genética , Metabolismo , Óxido Nítrico Sintasa de Tipo II , Genética , ARN Mensajero , Genética , Metabolismo , Proteínas Recombinantes , Genética , Alergia e Inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transformación Genética
17.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-327088

RESUMEN

This study was conducted to potentiate the expression of outer membrane protein OmpL17 of the strong virulent L. interrogans serovar Lai and investigate its immunogenicity in rabbits. The OmpL17 was cloned into prokaryotic expression vector pGEX-1lambdaT. The recombination expression plasmid pGEX-OmpL17 was transformed into E. Coli JM109. The GST fused protein GST-OmpL17 was expressed after induction by IPTG, then GST-tag was by thrombin and purified using Bulk GST purification Modules. SDS-PAGE and Western blotting analysis indicated that the molecular weight of GST-OmpL17 and OmpL17 was about 54 KDa and 28 KDa respectively. The outer membrane protein OmpL17 was subcutaneously injected into rabbits and high titre anti-OmpL17 antibody was obtained (1:4896) which could conjugate specifical with OmpL17. In conclusion, OmpL17 and specifical anti-OmpL17 antibody were obtained, which provided an experimental basis for researching pathogenic effect and immunity functions of OmpL17.


Asunto(s)
Humanos , Proteínas de la Membrana Bacteriana Externa , Genética , Alergia e Inmunología , Proteínas Bacterianas , Genética , Alergia e Inmunología , Clonación Molecular , Leptospira interrogans , Genética , Alergia e Inmunología , Porinas , Genética , Alergia e Inmunología , Proteínas Recombinantes , Genética , Alergia e Inmunología , Virulencia
18.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-280037

RESUMEN

<p><b>OBJECTIVE</b>To gain new insights into the molecular pathogenesis of the 109(InsG) and 139(C--> T) mutations and their roles in familial oligodontia.</p><p><b>METHODS</b>The region of PAX9 paired domain (PAX9PD) was amplified and the expression plasmids were constructed in pGEXlambda -1T by PCR-based cloning. PAX9PD proteins were prepared on the basis of GST instruction. The binding of wild type and two novely mutant PAX9 paired domain to double-stranded DNA targets were analyzed by gel mobility shift assay.</p><p><b>RESULTS</b>Wild type PAX9PD protein bind to the high affinity paired domain recognition sequences, CD19-2(A-ins) and Pax6CON, the 109(InsG) and 139(C--> T) mutant PAX9PD protein were unable to bind to these cognate DNA-binding sites.</p><p><b>CONCLUSION</b>The functional defects in DNA binding of mutant 109(InsG) PAX9 and 139(C--> T) PAX9, as well as loss-of-function of PAX9 most likely result in its haploinsufficiency during the patterning of dentition and the subsequent loss of posterior teeth.</p>


Asunto(s)
Humanos , Anodoncia , Genética , Secuencia de Bases , Cromatografía Líquida de Alta Presión , Análisis Mutacional de ADN , Ensayo de Cambio de Movilidad Electroforética , Salud de la Familia , Mutación , Factor de Transcripción PAX9 , Genética , Metabolismo , Reacción en Cadena de la Polimerasa
19.
Chinese Journal of Stomatology ; (12): 266-270, 2005.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-273240

RESUMEN

<p><b>OBJECTIVE</b>To investigate the mutational characteristics of PAX9 gene in Chinese patients with congenital oligodontia and thus to provide a molecular basis for studying the pathogenesis of oligodontia.</p><p><b>METHODS</b>Thirteen individuals with oligodontia and 9 healthy individuals, from 4 unrelated autosomal dominant families, and 16 sporadic patients with hypodontia in China, as well as 196 healthy control individuals (without oligodontia or hypodontia) were screened. Congenital absence of teeth was confirmed by panoramic X-ray analysis. Mutations of PAX9 gene were detected using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis. After the finding of abnormal SSCP bands, analysis was carried out with DNA sequencing.</p><p><b>RESULTS</b>PCR-SSCP detected SSCP bands alteration in exon2 of PAX9 gene in two unrelated families. Sequencing of PAX9 gene revealed a novel frameshift mutation (109InsG) and a novel missense mutation (C139T). All the affected members of each family were heterozygous for the mutations. In sporadic patients and the other two families, no similar sequence changes in PAX9 gene were found.</p><p><b>CONCLUSIONS</b>The results extend the spectrum of mutations in PAX9 gene associated with oligodontia. The novel mutations will play an important role in gene diagnosis of oligodontia.</p>


Asunto(s)
Adolescente , Adulto , Niño , Femenino , Humanos , Masculino , Persona de Mediana Edad , Adulto Joven , Pueblo Asiatico , Genética , Mutación del Sistema de Lectura , Mutación Missense , Factor de Transcripción PAX9 , Genética , Linaje , Pérdida de Diente , Genética
20.
Artículo en Chino | WPRIM (Pacífico Occidental) | ID: wpr-238344

RESUMEN

Total RNA was extracted from human LAK cell, and a cDNA encoding mature peptide HMG-17 and its alpha helix domain was amplified by RT-PCR. The recombinant prokaryotic expression vector pGEX-1lambdaT-HMG-17 and pGEX-1lambdaT HMG-17alpha helix was constructed. Using affinity chromatography, thrombin cleaving and AU-PAGE elution, we obtained the purified HMG-17. Analyses of MIC, MEC and MBC indicated that HMG-17 and HMG-17alpha had strong antibacterial activity. MIC of the alpha-helic domain was almost the same as that of HMG17, suggesting that the alpha-helic structure would be essential for the antibacterial activity of HMG-17.


Asunto(s)
Humanos , Antibacterianos , Farmacología , Escherichia coli , Genética , Metabolismo , Proteína HMGN2 , Genética , Farmacología , Células Asesinas Activadas por Linfocinas , Química , Péptidos , Genética , Farmacología , Células Procariotas , Metabolismo , Proteínas Recombinantes , Genética , Farmacología
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