No description available.

No description available.

Tuberculosis (TB), a severe disease caused by Mycobacteria tuberculosis (MTb), remains a globally severe health problem. In modern days, emergence of drug resistant TB is a new threat to public health since it can lead to treatment failure, increased transmission of TB to other hosts and further development of drug resistant complications. The traditional diagnostic method for TB by Acid Fast Bacilli (AFB) smear and Löwenstein–Jensen medium (LJ) culture are poor in sensitivity and time consuming respectively. There is a need for rapid diagnosis and identification of MTb. Nucleic acid amplification like PCR and real-time PCR are options for rapid diagnosis. However, such techniques require sophisticated technique and complex equipment. The high cost would constitute a barrier for countries with a high demand but only limited resources. Loop-mediated isothermal amplification (LAMP) assay is a novel technique, proven by many studies as to its high sensitivity and highly specificity to MTb. Most importantly, LAMP assay is economical and affordable by developing countries. The first objective of this study was to evaluate the analytical sensitivity and specificity of LAMP assay. The specificity of LAMP assay was determined by performing LAMP assay on 19 clinical isolates, which had already been identified previously. The clinical isolates included 14 mycobacteria tuberculosis complex (MTb), and five mycobacteria other than tuberculosis (MOTT) strains that were positive for AFB smear and LJ culture but negative for IS6110 single-tube nested real time PCR. The specificity was 100%. The analytical sensitivity as well as the limit of detection (LOD) were determined by testing on a duplicate set of serial DNA dilution, where each duplicate consisted of dilution of 100,000, 10,000, 1000, 300, 100, 10 and 1 colony forming unit/milliliter (CFU/ml). The LOD of LAMP assay was about 3 CFU per reaction. The second objective of this study evaluated the diagnostic performance of LAMP assay against AFB culture and IS6110 single tube nested real time PCR for identification of MTb in 200 respiratory specimens from 123 patients. All the specimens have already been tested for IS6110 single tube nested real time PCR, and culture results and AFB smears results have been obtained for all the specimens. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of three diagnostic methods (AFB smear microscopy, LAMP assay amplification and IS6110 single-tube nested real time PCR) were calculated with 95% confidence interval using LJ culture as gold standard. The LAMP assay had a sensitivity of 80%, specificity of 92.6%, PPV of 60% and NPV of 97.1%. However, MTb might fail to grow on the LJ culture medium for various reasons, for instance, MTb might already be dead after antibiotic treatment of the patient, or there might be poor laboratory practices during the processing of the specimens. Since the LJ culture method could produce false negatives in the situations described above, an alternative to the LJ culture method, ‘Hybrid Method’ was used as the gold standard. Under this method, a specimen was regarded as positive if the LJ culture result was positive. On the other hand, if a specimen generated a negative result using the LJ culture method, the results from the LAMP assay and IS6110 nested real time PCR would be considered, i.e., if both the LAMP assay and IS6110 nested real time PCR gave positive results while the results of LJ culture were negative, the specimen was referred to be positive in this case. In other words, a specimen would be regarded as negative if and only if the LJ culture result was negative and at least one of the LAMP assay or IS6110 was negative at the same time. Along the same line, the sensitivity, specificity, PPV and NPV of the three diagnostic methods (AFB smear microscopy, LAMP assay amplification and IS6110 single-tube nested real time PCR) were calculated with 95% confidence interval against the Hybrid Method. After resolution, the LAMP assay had a sensitivity of 87.0%, specificity of 100%, PPV of 100% and NPV of 97.1%. Our results showed that the LAMP assay has a great potential to be a new TB diagnostic test, especially in developing countries, with its lot of advantages like ease of use, cheap and fast. The LAMP assay in the study showed a high specificity, however, the sensitivity has to be improved before application in clinical use. For comparison of clinical performance, IS6110 single tube nested real time PCR had a higher sensitivity than that of LAMP assay (100% vs 80% using culture as gold standard; 100% vs 87% using ‘Hybrid Method’ as gold standard). However, LAMP assay had a higher specificity than that of IS6110 single tube nested real time PCR (92.6% vs 90.7% using culture as gold standard; 100% vs 98% using ‘Hybrid Method’ as gold standard). LAMP had been proven to be a potential and powerful tool in clinical diagnosis of MTb. Further improvement on its sensitivity is required to enable its extensive use in the clinic in the future. / published_or_final_version / Medical Sciences / Master / Master of Medical Sciences

Diss. (sammanfattning) Uppsala : Univ., 2003. / Härtill 4 uppsatser.

Common amplicons were delineated throughout the NPC genome in a large panel of NPC cell lines, xenografts, and primary tumors by two high-density genomic arrays with ∼1 Mb and 35 kb resolution. Apart from the genetic changes reported in previous studies, a number of novel chromosomal aberrations were discovered, including gains at 7p11, 16p13.3, 19p13, 19q13-q43 and 20q13. Most distinctively, common amplicons at 11q13 and 12p13 were found in this cancer. Two smallest amplification regions with 5.4 Mb and 2.16 Mb were delineated at 11q13.1-q13.3 and 12p13.31 respectively. The high prevalence of these 2 amplified regions have led to the hypothesis that activation of the target oncogenes in these regions are critical events for NPC development. / Expression of candidate genes located within 11q13.3 was examined and consistent overexpression of CCND1 in cell lines and xenografts were identified in the 11q13.3. Frequent concordant gains and overexpression of CCND1 were further confirmed in primary tumors. Knockdown of CCND1 mRNA by siRNA technique was found to inhibit cell growth and lead to cell cycle arrest at G1. Alterations of protein expressions of other cell cycle components were also observed. Moreover, inactivation of p16 and overexpression of cyclin D1 were commonly occurred in NPC. These findings provided evidence that cyclin D1 may have cell cycle-independent functions, which is critical in NPC tumongenesis. / Frequent gains of 12p13.31 region were confirmed by fluorescence in situ hybridization (FISH) analysis. According to expression array and real-time RT-PCR results, LTbetaR, TNFRSF1A and FLJ10665 were the three genes showing concordant amplification and overexpression in NPC xenograft. The LTbetaR protein, which is a lymphotoxin beta receptor, was confirmed to be recurrently overexpressed in NPC primary tumors and its overexpression may be involved in the activation of NF-kappaB in NPC. The findings suggested that it is one of the candidate oncogenes of this cancer. / In summary, three candidate NPC-associated oncogenes locating at 3q26.32, 11q13.3 and 12p13.31 were identified by genome-wide mapping analysis. Molecular and functional characterizations of these genes have provided evidences that they play critical roles in NPC tumorigenesis. / In this study, detailed investigation was carried out on a candidate NPC-associated oncogene, PIK3CA at 38q26.32, an amplicon reported previously. Copy number gains and amplifications of this gene, but not mutation, were demonstrated to be common events in NPC. The findings hence implied the importance of PIK3CA in NPC tumorigenesis. / Nasopharyngeal carcinoma is a common cancer in Southern China. Despite multiple genetic changes have been reported previously, limited information of NPC-associated oncogene is available. Since amplification is one of the major mechanisms in oncogene activation, a comprehensive characterization of common amplicons in human cancers is expected to facilitate the identification of the oncogenes involved in tumorigenesis. The aims of the present study is to define and characterize the common amplicons in NPC genome and then to identify NPC-associated oncogenes. / Or Yan Yan. / "July 2006." / Adviser: Kwok Wai Lo. / Source: Dissertation Abstracts International, Volume: 68-09, Section: B, page: 5715. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2006. / Includes bibliographical references (p. 179-201). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

No description available.

published_or_final_version / Microbiology / Master / Master of Medical Sciences

published_or_final_version / Pathology / Master / Master of Medical Sciences

Cancer is a disease that involves genomic instability, to which c-Myc contributes during its initiation and progression. Over 70% of all human cancers show deregulated levels of c-Myc protein. The term genomic instability refers to genetic and/or epigenetic changes that alter the normal organization and function of genes and chromosomes. Genomic instability is a hallmark of cancer and often is associated with cancer. Deregulated c-Myc expression generates genomic instability by initiating intra- and extrachromosomally locus-specific gene amplification, gene rearrangements and karyotypic instability that includes translocations, fusions, insertions and deletions. Out of the several outlined pathways by which deregulated levels of c-Myc can lead to genomic instability, the work described in this thesis focuses on three with direct relevance to tumorigenesis; gene amplification (increase in gene copy number), remodeling of the chromosomal and telomeric structures in the interphase nucleus and comparing the effect of Myc to that of Epstein Bar virus (EBV) infection in remodeling the nuclear structure that may lead to genomic instability.

Cancer is a disease that involves genomic instability, to which c-Myc contributes during its initiation and progression. Over 70% of all human cancers show deregulated levels of c-Myc protein. The term genomic instability refers to genetic and/or epigenetic changes that alter the normal organization and function of genes and chromosomes. Genomic instability is a hallmark of cancer and often is associated with cancer. Deregulated c-Myc expression generates genomic instability by initiating intra- and extrachromosomally locus-specific gene amplification, gene rearrangements and karyotypic instability that includes translocations, fusions, insertions and deletions. Out of the several outlined pathways by which deregulated levels of c-Myc can lead to genomic instability, the work described in this thesis focuses on three with direct relevance to tumorigenesis; gene amplification (increase in gene copy number), remodeling of the chromosomal and telomeric structures in the interphase nucleus and comparing the effect of Myc to that of Epstein Bar virus (EBV) infection in remodeling the nuclear structure that may lead to genomic instability.