Doctor of Philosophy (PhD) / Healthcare services and health research aim to improve the physical and psychosocial well being of consumers, and to offer responsive services needed and valued by them. Research in chronic kidney disease (CKD) has predominantly focused on investigating biomedical aspects and evaluating technological or pharmacological treatment interventions to improve medical management. While research into assessing patients’ and caregivers’ quality of life, and symptom burden, is growing minimal attention has been given to gaining a broad and in-depth understanding about the experiences, psychosocial issues and needs of patients and their caregivers. These need to be considered when planning and delivering patient-centred care and health research across the whole trajectory of CKD. The studies that form the major part of this thesis explore the perspectives, needs and experiences of CKD patients and their caregivers, within a broad and multidimensional framework encompassing aspects of the nature of the health and illness experiences and consumer perspectives. In Chapter 2, to understand what is known about parental experiences of caring for a child with CKD, the relevant qualitative literature was systematically reviewed and synthesized. Three inter-related clusters were identified: intrapersonal, interpersonal and external experiences. In Chapter 3, to gain a more detailed and broader understanding of this topic, in-depth interviews were conducted with parents of 20 children with CKD and 4 major themes were identified: absorbing the clinical environment, medicalising parenting, disrupting family norms, and coping strategies and support structures. In Chapter 4, to assess the effectiveness of support interventions for caregivers of patients with CKD, a systematic review was conducted which identified only three eligible studies that assessed only the effect of educational material on caregiver knowledge, not other domains. In Chapter 5, to describe and compare the broad range and depth of experiences and perspectives from predialysis, dialysis and transplantation patients, data from patient focus groups were analysed. The 5 themes that emerged from this data were: personal meaning of CKD, managing and monitoring health, lifestyle consequences, family impact, and informal structures. In Chapter 6, the focus groups were also used to elicit research priorities and identify reasons that patients used to develop their research priorities. A patient focused research agenda was elicited for CKD and 5 reasons that patients used to develop their research priorities were identified: normalisation of life, altruism, economic efficiency, personal concerns and clinical outcomes. During the focus groups, participants repeatedly expressed frustration about the poor public profile, and lack of community-based information on CKD prevention. So in Chapter 7, to assess how Australian news media covered prevention and early detection of CKD, I analysed television and newspaper stories that referred to CKD prevention or early detection. Kidney disease in general, and particularly the prevention and early detection of CKD, received virtually no media attention. When mentioned, it was mainly in the context of transplantation and donor stories, and seldom prevention or early detection, which appears largely unnewsworthy in its current form. At best, CKD received peripheral mention as a secondary concern in diabetes and obesity news stories which focused on lifestyle solutions. In Chapter 8, to develop a checklist for explicit and comprehensive reporting of qualitative studies (in-depth interviews and focus groups), I performed a comprehensive search in relevant publications for existing checklists used to assess qualitative studies. Seventy-six items from 22 checklists were compiled into a comprehensive list. All items were grouped into three domains: 1) research team and reflexivity, 2) study design, and 3) data analysis and reporting. The overarching purpose of these studies was to gain a better understanding about the needs, experiences and perspectives of CKD patients and their caregivers. The findings describe the permanent, profound and pervasive impact of CKD on the lives of patients and caregivers across the whole illness trajectory. A more detailed and broader understanding about patient and caregiver perspectives, as presented in this thesis, can support a move towards advancing patient-centred healthcare and research in CKD.

Six different isoforms of Type IV collagen (colIVα1-6) have been identified. The individual isoforms of colIV are termed alpha chains and are translated from six different COLIV genes (COLIVA1-A6). Collagen Type IV gene products compose the structural framework of basement membranes. The glomerular basement membrane (GBM) is a specialized basement membrane involved in the ultrafiltration processes of the kidney. The colIVα1-α5 chains are expressed in the human GBM while the colIV α1-α6 chains are expressed in the canine GBM. Many inherited diseases of the kidney have been reported and mutations in genes regulating kidney function have been identified. Alport syndrome (AS) is the most common form of human hereditary nephritis (HN). AS is defined as an inherited progressive kidney disorder associated with sensoneural deafness and is characterized by extensive thickening and multilamminar splitting of the GBM when examined by electron microscopy. AS has both X-linked (XLAS) and autosomal (ARAS) modes of inheritance. Mutations in the COLIVA5 gene are responsible for XLAS. A form of HN with characteristic splitting of the GBM with X-linked inheritance has been described in Samoyed dogs. A specific mutation in the COLIVA5 gene has been identified in Samoyed dogs affected with HN. Mutations in the COLIVA3 and COLIVA4 genes are responsible for ARAS. A form of HN has been identified in English cocker spaniel dogs (ECS) that has been described as autosomal in inheritance and includes GBM abnormalities including extensive lammination characteristic of ARAS. Both ARAS and ECS-HN show loss of the colIVA3 and colIVA4 chains in the GBM when examined with monoclonal anitibodies. ECS-HN has been hypothesized to have the same molecular basis of disease as ARAS. As such, we have isolated and characterized canine COLIVA3 and COLIVA4 sequences from normal dogs and ECS dogs affected with HN and compared the coding regions of these candidate genes.

In patients with chronic kidney disease (CKD), overhydration, uremic toxins and left ventricular (LV) dyssynchrony are factors that may lead to LV dysfunction and conduction abnormalities and thus contribute to the high cardiac mortality. Colour tissue velocity imaging (TVI) allows a detailed quantitative analysis of cardiac function in CKD patients, opening new possibilities to evaluate longitudinal myocardial motion, rapid isovolumetric events, LV filling pressure and LV synchronicity. Aims: Using TVI technique: 1. To evaluate myocardial function disturbances and their relations to risk factors in CKD patients. 2. To assess LV synchronicity in HD patients, both at baseline and after HD, and 3. To study acute cardiac effects of HD and i.v. furosemide in HD patients. Methods: 40 predialysis CKD (stages I, II, III, IV and V) (Study II) and 59 HD (Studies I, III, IV and V) patients were studied. In both groups of patients LV function was evaluated using TVI, and in HD patients LV synchronicity was also assessed using tissue synchronization imaging (TSI). In HD patients the evaluations were performed before and after HD (Studies III and V) and i.v. furosemide infusion (Study IV). Results: 1. TVI detected: a) LV contraction disturbances in CKD patients with LVH and normal ejection fraction. b) An increase of LV contractility after HD. c) No changes in cardiac function induced by furosemide. 2. TSI detected the presence of LV dyssynchrony and its improvement after HD. 3. In CKD, cardiac dysfunction seemed to be related to high levels of PTH, phosphate and blood pressure. Conclusions: TVI is a sensitive tool for studies on cardiac function in CKD, allowing a detailed and accurate evaluation of disturbances in LV function. TVI also provides the possibility to follow the changes in LV function and synchronicity induced by different therapeutical interventions. The obtained information may contribute to a better management of CKD patients. / QC 20100809

Thromboxane receptor (TPr) activity is elevated in diabetes and contributes to complications of diabetic kidney disease (DKD). TPr blockade appears to have therapeutic potential. Several rodent models of DKD show attenuation of renal damage and proteinuria upon administration of the TPr antagonist, S18886. However, the cellular targets that underlie the injurious effects of TPr activation in DKD remain to be elucidated. A pilot study in our laboratory subjected a conditionally-immortalized mouse podocyte cell line to high glucose (25 mM D-glucose) and equibiaxial mechanical stretch (an in vitro simulator of increased glomerular capillary pressure associated with glomerular hyperfiltration in early diabetes). qRT-PCR revealed that exposure of podocytes to mechanical stretch (10% elongation) and high glucose for 6 hours yielded a 9-fold increase in TPr mRNA levels vs. controls (non-stretch, 5mM D-glucose + 25mM L-glucose) (p<0.05, n=5). We hypothesized that TPr expression and activity are increased in podocytes during the onset of DKD resulting in maladaptive effects on this key glomerular filtration barrier cell type. We showed that enhanced TPr signaling threatens podocytes viablility. Cultured podocytes treated with the TPr agonist, U-46619 (1 μM) for 24 hours are more vulnerable to apoptosis as quantified by Hoescht 33342 (20% cell death p<0.001, n=3) , TUNEL (30-fold increase, ns, n=3) and Annexin-V labeling (3-fold increase, p <0.001, n=3). To further support these in vitro findings, we developed a transgenic mouse with podocyte-specific overexpression of TPr. A construct consisting of a desensitization resistant mutant of the human TPr with both N- and C-terminal HA-epitope tags under the control of an 8.3 kb fragment of the immediate 5’ mouse NPHS1 promoter was cloned, isolated and injected into FVB/n oocytes that were implanted into pseudopregnant CD1 females. Founders were characterized for TPr transgene expression, and TPr transgene mRNA levels were detected by qRT-PCR. Our in vitro results suggest that increased TPr expression in podocytes of diabetic mice may contribute to filtration barrier damage and have important implications in the development and progression of DKD.

No description available.

The impact of habitually consuming a high protein (HP) diet at the upper limit of the acceptable macronutrient distribution range (AMDR) on kidney health is unknown. The current study was designed to test the hypothesis that long-term consumption of a diet providing 35% of energy as protein will have negative consequences on renal health, as assessed in a pig model. Methods: Adult female, non-pregnant, commercial pigs (Genesus) were randomized to receive either NP (15% energy from protein) or HP (35% energy from protein) isocaloric diets for either 4 or 8 months. Diets contained whole protein sources with an animal: plant ratio of 2:1 in the NP diet to mimic the average Canadian diet. The increased protein in the HP diet was achieved by increasing egg and dairy protein sources. Body composition was measured by dual-energy X-ray absorptiometry. Glomerular volume and kidney fibrosis were evaluated on kidney sections by quantitative image analysis. The inflammatory marker monocyte chemoattractant protein-1 (MCP-1) and the growth factor transforming growth factor beta-1(TGFβ1) were assessed in renal tissue using commercial ELISA kits. Results: Pigs given the HP diet had lower body weights and percentage of body fat. Pigs consuming the HP diet had significantly higher glomerular filtration rates (GFR) and larger kidneys. Renal MCP-1 levels and renal fibrosis also were significantly higher in pigs given the HP diet, while proteinuria and renal TGFβ1 expression did not differ. Conclusion: These findings suggest that, despite the potential benefit of the HP diet on body composition, long-term intakes of protein at the upper limit of the AMDR may compromise renal health in healthy female pigs. / October 2008

Thromboxane receptor (TPr) activity is elevated in diabetes and contributes to complications of diabetic kidney disease (DKD). TPr blockade appears to have therapeutic potential. Several rodent models of DKD show attenuation of renal damage and proteinuria upon administration of the TPr antagonist, S18886. However, the cellular targets that underlie the injurious effects of TPr activation in DKD remain to be elucidated. A pilot study in our laboratory subjected a conditionally-immortalized mouse podocyte cell line to high glucose (25 mM D-glucose) and equibiaxial mechanical stretch (an in vitro simulator of increased glomerular capillary pressure associated with glomerular hyperfiltration in early diabetes). qRT-PCR revealed that exposure of podocytes to mechanical stretch (10% elongation) and high glucose for 6 hours yielded a 9-fold increase in TPr mRNA levels vs. controls (non-stretch, 5mM D-glucose + 25mM L-glucose) (p<0.05, n=5). We hypothesized that TPr expression and activity are increased in podocytes during the onset of DKD resulting in maladaptive effects on this key glomerular filtration barrier cell type. We showed that enhanced TPr signaling threatens podocytes viablility. Cultured podocytes treated with the TPr agonist, U-46619 (1 μM) for 24 hours are more vulnerable to apoptosis as quantified by Hoescht 33342 (20% cell death p<0.001, n=3) , TUNEL (30-fold increase, ns, n=3) and Annexin-V labeling (3-fold increase, p <0.001, n=3). To further support these in vitro findings, we developed a transgenic mouse with podocyte-specific overexpression of TPr. A construct consisting of a desensitization resistant mutant of the human TPr with both N- and C-terminal HA-epitope tags under the control of an 8.3 kb fragment of the immediate 5’ mouse NPHS1 promoter was cloned, isolated and injected into FVB/n oocytes that were implanted into pseudopregnant CD1 females. Founders were characterized for TPr transgene expression, and TPr transgene mRNA levels were detected by qRT-PCR. Our in vitro results suggest that increased TPr expression in podocytes of diabetic mice may contribute to filtration barrier damage and have important implications in the development and progression of DKD.

Six different isoforms of Type IV collagen (colIVα1-6) have been identified. The individual isoforms of colIV are termed alpha chains and are translated from six different COLIV genes (COLIVA1-A6). Collagen Type IV gene products compose the structural framework of basement membranes. The glomerular basement membrane (GBM) is a specialized basement membrane involved in the ultrafiltration processes of the kidney. The colIVα1-α5 chains are expressed in the human GBM while the colIV α1-α6 chains are expressed in the canine GBM. Many inherited diseases of the kidney have been reported and mutations in genes regulating kidney function have been identified. Alport syndrome (AS) is the most common form of human hereditary nephritis (HN). AS is defined as an inherited progressive kidney disorder associated with sensoneural deafness and is characterized by extensive thickening and multilamminar splitting of the GBM when examined by electron microscopy. AS has both X-linked (XLAS) and autosomal (ARAS) modes of inheritance. Mutations in the COLIVA5 gene are responsible for XLAS. A form of HN with characteristic splitting of the GBM with X-linked inheritance has been described in Samoyed dogs. A specific mutation in the COLIVA5 gene has been identified in Samoyed dogs affected with HN. Mutations in the COLIVA3 and COLIVA4 genes are responsible for ARAS. A form of HN has been identified in English cocker spaniel dogs (ECS) that has been described as autosomal in inheritance and includes GBM abnormalities including extensive lammination characteristic of ARAS. Both ARAS and ECS-HN show loss of the colIVA3 and colIVA4 chains in the GBM when examined with monoclonal anitibodies. ECS-HN has been hypothesized to have the same molecular basis of disease as ARAS. As such, we have isolated and characterized canine COLIVA3 and COLIVA4 sequences from normal dogs and ECS dogs affected with HN and compared the coding regions of these candidate genes.

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Thesis (M.Sc.)--Memorial University of Newfoundland, 2000. / Restricted until November 2001. Bibliography: leaves 82-97.