published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

Ren-Wang Jiang. / "July 2002." / Thesis (Ph.D.)--Chinese University of Hong Kong, 2002. / Includes bibliographical references. / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Introduction: Numerous studies have found associations between exposure to airborne particulate matter and respiratory morbidity and mortality. However, the varying composition, dependant on the different sources of particulate matter, and its effect on processes of inflammation and oxidative stress in the airways has not been completely elucidated. The use of airway biomarkers, fractional exhaled nitric oxide (FENO) and exhaled breath condensate (EBC), can provide valuable insight into processes of inflammation and oxidative stress in the airways. In these studies we sought to characterize the associations between airborne pollutant exposures and their sources and airway biomarkers. Methods:The study population of interest for Chapters III-V is a subset of children currently enrolled in a birth cohort under the Columbia Children's Center for Environmental Health. Chapter III refers to the validation in this population of a method for partitioning FENO contributions from the proximal and distal airways. Chapters IV and V refer to the implementation of this method in the study of ambient metals and residential proximity to relevant sources of particulate matter (PM) and black carbon (BC). Briefly, the children of African-American and Dominican mothers living in low-income NYC neighborhoods had FENO samples collected offline at constant flow rates of 50, 83 and 100 mL/sec at ages 9 and/or 11. Surrogate measures for bronchial flux NO (JNO) and alveolar (Calv) NO concentrations were estimated using a previously validated mathematical model. Wheeze in the last 12 months was assessed by the International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire. Seroatopy was determined by specific IgE at age 9. For Chapter IV, in order to examine the associations between metal fractions of particulate matter and airway inflammation, ambient measures of Ni, V, Zn and Fe were obtained from a local central monitoring site and averaged over nine days based on three 24 hour measures every third day. Seroatopy was determined by specific IgE at age 7. For Chapter V, residential distance to a major road, truck route and bus stop density, area covered by major roads, stationary point sources (SPS), toxic release inventory sites (TRIS) and commercial buildings, and number of buildings burning residual oil within 250-meters of each child's home were determined. These variables were selected a priori as potential important sources or airborne PM and BC. For Chapter VI, the study population was comprised of seven- and eight-year-old children enrolled in an asthma case-control study in New York City. Seven day averages of domestic levels of particulate matter <2.5 microns (PM2.5), BC and environmental tobacco smoke (ETS) were measured. Urea and 8-isoprostane were measured by liquid chromatography tandem mass spectrometry (LC/MS/MS) in EBC collected during home visits. All data were analyzed with SPSS. Results: In our first study, children with seroatopy had significantly higher median JNO p<0.001) when compared to non-seroatopic children; however, median Calv was not significantly different between these two groups (p=0.644). Children with wheeze in the past year had significantly higher median Calv (p<0.001), but not JNO (295 vs. 165 pL/s, p=0.241) when compared with children without wheeze. In our second study, ambient V and Fe concentrations were associated positively with FENO50 (p=0.018, p=0.027). Ambient Fe was associated positively with JNO (p=0.017). Ambient Ni and V concentrations were associated positively with Calv (p=0.004, p=0.018 respectively). A stronger association of Ni concentrations with Calv was observed among the children with seroatopy. In our third study, there were no significant associations between any of the air pollution indicator variables and FENO parameters in adjusted models. In our final study, PM2.5and BC, but not ETS, were significantly associated with increases in 8-isoprostane (p<0.05 for both) after adjustment for covariates. In a co-pollutant model including PM2.5, BC and ETS, only BC remained a statistically significant predictor of 8-isoprostane. Discussion: Recent exposure to airborne pollutants was associated with increased levels of biomarkers of airway inflammation and oxidative stress measured in exhaled breath. The metal and BC fractions of PM might be more relevant to the understanding adverse respiratory outcomes related to air pollution exposure.

Abstract not available

Background. Obstructive Sleep Apnoea (OSA) is associated with an increase in nocturnal gastro-oesophageal reflux (nocturnalGOR) events and symptoms, however the mechanism for this remains undefined. Treatment of OSA with continuous positive airway pressure (CPAP) has been shown to reduce nocturnalGOR in individuals with OSA however the reasons for this reduction are not clear. The combination of OSA and nocturnalGOR could be particularly problematic for individuals who have had a lung transplant in whom Bronchiolitis Obliterans Syndrome (BOS) limits survival. It is thought that GOR plays a role in the development of BOS in these individuals. Methods and Results. Five interrelated studies were undertaken. The first two studies sought to determine and compare the prevalence and risk factors of nocturnalGOR in OSA patients with the general population. To do this, a GOR questionnaire was completed by 2,042 members of the general community as part of the Busselton Health Survey and by 1,116 patients with polysomnography-diagnosed OSA. Risk of OSA in the general population was determined using a standardised sleep questionnaire. 137 of the OSA patients completed the questionnaire before and after treatment with CPAP. The prevalence of nocturnalGOR symptoms reported more than once a week (frequent symptoms) was greater in OSA patients (10.1%) than the general population (5.8%) (p<0.001), in individuals from the general population at high (11.2%) than low risk of OSA (4.5%) (p<0.001) and in patients with severe (14.7%) than mild OSA (5.2%) (p<0.001). Treatment of OSA with CPAP decreased the prevalence of frequent nocturnalGOR from 9.0% to 3.8% (p=0.04). In the general population, high risk of OSA was independently associated with a 2.4-fold increased risk of frequent ABSTRACT vi nocturnalGOR symptoms than low risk. In the OSA group, disease severity was independently associated with nocturnalGOR symptoms, with an adjusted odds ratio of 1.7 for frequent nocturnalGOR symptoms.

Highly pathogenic avian influenza H5N1, which is panzootic in poultry, continues to spread and becomes endemic in Asia, Africa, and Europe. It causes human disease with high fatality (about 60%) and continues to pose a pandemic threat. The pathological lesions associated with human H5N1 disease is Acute Respiratory Distress Syndrome (ARDS). The biological basis underlying the development of ARDS in human H5N1 disease remains controversial. Clinical, animal and in vitro studies suggested that the differences between H5N1 influenza viruss and low pathogenic influenza viruses in regard to viral replication, tissue tropism and cytokine dysregulation may contribute to disease pathogenesis. We previously found delayed onset of apoptosis in influenza H5N1 virus infected human peripheral blood monocyte-derived macrophages. This may allow a longer survival time for the virus in target cells for prolonged viral replication, which may contribute to the pathogenesis of H5N1 disease. As human bronchial and alveolar epithelial cells are target cells of influenza virus, I explored if influenza H5N1 and H1N1 virus infected human respiratory epithelial cells displayed differential apoptotic response and dissected the apoptotic pathways triggered by influenza virus infection. In this study, the apoptotic response in highly pathogenic influenza H5N1 viruses, A/HK/483/97 and A/Vietnam/1203/04, their precursor avian influenza H9N2 virus, A/Quail/HK/G1/97, and seasonal H1N1 virus, A/HK/54/98 infected primary human alveolar and bronchial epithelial cells was compared by TUNEL. A delayed onset of apoptosis in influenza H5N1 viruses and avian H9N2 virus infected alveolar epithelial cells was observed; the pattern was similar in bronchial epithelial cells. Concomitantly, by Western blotting, a delay in caspase 3 activation in H5N1 virus (A/HK/483/97) infected alveolar epithelial cells compared to H1N1 virus (A/HK/54/98) infected cells was shown. Also, influenza H5N1 and H1N1 virus induced apoptosis through both intrinsic and extrinsic pathways in human alveolar epithelial cells. Chemokine IP-10 was differentially up-regulated in influenza H5N1 virus infected alveolar epithelial cells, but its relationship to the delayed onset of apoptosis requires further studies. TRAIL, an upstream signaling molecule of extrinsic apoptotic pathway, mRNA was up-regulated in influenza H5N1 infected alveolar epithelial cells but not in influenza H1N1 infected cells. Using recombinant viruses, I showed that the 627 amino acid residue on PB2 of H5N1 virus and mutation of amino acids on 253 and 591 residues on PB2 of H9N2 virus contribute to the TRAIL upregulation. Immunohistochemical staining of physiologically relevant ex vivo model of human bronchus showed that influenza H5N1 (A/Vietnam/3046/04) and H9N2 (A/Quail/HK/G1/97) virus did not infect human bronchi as well as human H1N1 (A/HK/54/98) virus. Profiling of apoptosis related genes showed that TRAIL tends to be up-regulated in H5N1 virus infected bronchi ex vivo. This study demonstrated the delayed onset of apoptosis by H5N1 virus infected respiratory epithelial cells may be a mean for influenza virus to have prolonged replication within the human respiratory tract and contribute to disease severity. The results generated provide a robust research agenda, yielding critical information that elucidate molecular mechanisms, such as TRAIL up-regulation, that may contribute to the virulence and pathogenesis in human H5N1 disease. / HKU 3 Minute Thesis Award, 2rd Runner-up (2011) / published_or_final_version / Pathology / Master / Master of Philosophy

Background: Respiratory diseases are common worldwide, which are caused by various respiratory viruses. As symptoms caused by these viruses are similar, laboratory diagnosis is essential to distinguish the virus. Conventionally, respiratory viruses are isolated by cell culture with a panel of cell lines. However, handling of several cell lines is labour intensive, and the turnaround time of conventional culture is long. In previous study, the use of human colon adeno-carcinoma (Caco-2) in conventional culture was investigated. The study has proven that Caco-2 is generally susceptible to the eight common respiratory viruses, i.e. Adenovirus, Influenza A and B, Respiratory Syncytial virus, Parainfluenza virus 1, 2,3 and 4. As turnaround time of conventional culture is long; therefore, in this study, rapid shell vial culture using Caco-2 cells were evaluated. Moreover, the application of Caco-2 shell vial culture on recovering human metapneumovirus (hMPV) was also investigated. Materials and methods: This study consisted of four stages. First, recovery of viruses by conventional culture and shell vial culture of Caco-2 were compared. Specimens were added to conventional culture and shell vial simultaneously. For conventional culture, formation of CPE was examined daily and IF staining was performed when CPE was indicated; meanwhile, shell vial culture were incubated for seven days and stained with IF to detect infected cells. In stage two, the effect of incubating shell vial culture in rolling drum was investigated. Shell vials inoculated with the same specimen in duplicate were incubated in rolling drum and without rolling drum simultaneously. IF staining was performed in day 2, and results were obtained. For those which are IF negative in day 2, second shell vial was further incubated to seven days before harvest. In the next stage, a large batch of samples was used to evaluate on the use of Caco-2 shell vial culture in day 2 and day 7. Lastly, Caco-2 shell vial and conventional culture and LLC-MK2 conventional culture were tested for isolation of hMPV. Results: Compared to Caco-2 conventional culture, recovery rate of shell vial culture was elevated slightly. When experimenting on the effect of incubation in rolling drum, results showed that recovery rate was raised in shell vial with rolling drum in day 2, moreover, the percentage of positive cells were increased significantly (p value < 0.05). Furthermore, in the evaluation of Caco-2 shell vial in day 2 and day 7, 75% of samples were isolated in day 2 while 85% were recovered in day 7. Lastly, in the investigation on recovery of hMPV, 53%, 42% and 17% hMPV positive cases were isolated by Caco-2 shell vial, Caco-2 conventional culture and LLC-MK2 conventional culture respectively. Conclusion: First, although recovery rate by shell vial and conventional culture were similar, turnaround time was reduced from a week to a few days by shell vial culture. Therefore, Caco-2 shell vial culture is a more efficient than Caco-2 conventional culture in isolating respiratory viruses. The study also showed that incubation of shell vial in rolling drum able to increase the number of positive cells. Furthermore, in this study, Caco-2 cells were also shown to be more efficient in isolating hMPV when compare to LLC-MK2 cells. / published_or_final_version / Microbiology / Master / Master of Medical Sciences

No abstract available. / Thesis (M.Med.)-University of KwaZulu-Natal, 2004.

To investigate the possibility that malnutrition is an important factor that prolongs respiratory failure (PRF), I studied the effects of pharmacologic injections of recombinant human growth hormone (rhGH), an important anabolic stimulus, on nutritional and respiratory parameters in patients requiring mechanical ventilation for more than three days. Patients were excluded from consideration if dominating factors known to prolong ventilatory failure had not been stabilized. Over ten months, 106 patients in PRF were evaluated, but only six met the selection criteria. Three patients were randomized to receive standard nutritional support, and three into a group that received the equivalent nutrition plus 5 mg/day of rhGH for 14 days or until withdrawal of mechanical ventilation. Baseline characteristics of the selected patients were divergent as demonstrated by body mass indexes ranging from 14 to 42 (kg/m$ sp2),$ baseline maximal inspiratory pressures (PI$ sb{ max}$ from $-$15 to $-$70 cm H$ sb2$O, and Day 1 N balances from $-$13.5 to 1.2 g N/day. Despite increased plasma insulin-like growth factor-1 concentrations, the mean daily N balances of the rhGH-treated group were no better than the controls (1.3 $ pm$ 5.0 vs. 0.4 $ pm$ 2.6 g N/day; Mean $ pm$ SD), nor were there differences in PI$ sb{ max},$ level of ventilatory assistance required, and days to weaning. The persistence of respiratory failure in the overwhelming majority of patients in PRF appears to be due to factors already known to prevent weaning from mechanical ventilation. Even the carefully selected patients enrolled in the present study were insufficiently homogeneous or stable enough to allow proper testing of the experimental hypothesis.

A cross-sectional epidemiologic survey of the respiratory health status was conducted in the adult (15 years and older) Indian South African population resident in Lenasia, Johannesburg to study the distribution and determinants of respiratory symptoms, disease and lung function level. A slightly modified self-administered version of a standardised respiratory health questionnaire and a wedge spirometer was used. There were a high proportion of current smokers among men. Although women smoked less than men in other communities they nevertheless smoked on average more heavily than other Indian South African women. Indian men and women who smoked had a high prevalence of respiratory symptoms. The women also demonstrated an increased susceptibility to the effects of cigarette smoking when compared with women in other communities. Indians in this study had spirometric lung function levels that were lower than that recorded in recent studies in Blacks and Whites in South Africa. Respiratory symptoms, disease and lung function level were examined in a multiple logistic regression model which contained all the potential determinants recorded in the present study. Voluntary tobacco smoking, recent chest illnesses and any kind of heart trouble was associated with a significant risk for having most of the respiratory symptoms and diseases in men and women. In addition exposure to dust in the work environment, little or no exercise,>Std. 8 education a history of any kind of chest trouble and respiratory trouble before the age of 16 years was associated with an increased risk for having respiratory symptoms in men in this model. An increased risk for respiratory symptoms was demonstrated in women only with age. Age and standing height were the most important determinants of lung function level in men and women in the regression model. Dust exposure in the work environment was associated with a significantly lower lung function level in men. Alcohol consumption and a history of whooping cough was also independently associated with a lower lung function level in men but were of borderline significance. In women involuntary /passive tobacco smoke exposure and respiratory trouble before the age of 16 years were associated with a lower lung function level. Women who spent most of their lives in a rural area and those who had a university education had a higher lung function level. The deleterious effects of smoking on lung function were minimal in this study possibly because lung function was performed only in subjects in the 18-45 year age category. A "healthy smoker" effect was demonstrated in men. Men who ever smoked and were without cardiorespiratory symptoms had significantly higher lung function levels compared to men who never smoked and were without symptoms. / Thesis (M.D.)-University of Natal, Durban, 1992.