Dissertation
Defining the pathophysiology of C3 glomerulopathy - an ultra-rare complement-mediated renal disease -
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Autumn 2020
DOI: 10.17077/etd.005691
Abstract
C3 glomerulopathy (C3G), is a complex ultra-rare complement-mediated renal disease affecting approximately two to three individuals per 1,000,000 per year, as a result of the dysregulation of the complement alternative pathway (AP) proteins in the fluid-phase breakdown products are deposited in the glomeruli of the kidney. C3G encompasses two major disease sub-types, C3 glomerulonephritis (C3GN) and dense deposit disease (DDD). Many patients with C3G have abnormal levels of proteins of the AP in their bloodstream. In particular complement component C3, an essential protein of the AP has been demonstrated to be decreased in the circulation. The increased breakdown products are indicative of on-going complement activity. In measuring the complement protein levels, and their breakdown products, we can define patient-specific biomarker profiles that are characteristic of C3GN and DDD. These biomarker profiles are unique “signatures” that can be determined for each person. They have demonstrated the importance of evaluating complement proteins in the disease state. C3GN and DDD have commonalities and differences when a broad panel of alternative pathway complement proteins are compared. A pilot study evaluation of AP proteins indicated that C3 is decreased in both C3GN and DDD when compared to normal controls, and that reduction was greater in DDD patients. Additionally, the study also demonstrated greater terminal pathway activity, soluble C5b-9, in C3GN than in DDD. These tests localize the sites of on-going complement dysregulation is C3G.
SOMAscan, a robust, multiplexed proteomic DNA microarray technology targeting >1300 proteins in patients’ plasma samples was used to validate the complement biomarkers currently used clinically for predicting complement dysregulation in C3G. Additionally, SOMAscan provides an opportunity for the discovery of novel biomarkers in association with C3G to further inform our understanding of C3G. In this study, C3G patient plasma samples (C3GN; N=25 and DDD; N=33) were analyzed employing SOMAscan technology compared to normal controls (no renal phenotype; N=34) and Anti-Neutrophil Cytoplasmic Antibodies (ANCA) glomerulonephritis pool (N=10). The C3G samples were retrospectively screened for additional clinical factors: dialysis, transplant, and Eculizumab treatment.
Our SOMAscan study showed that renin was among the top differentially expressed proteins in C3G (p-value = 0.0013, one-way ANOVA) as well as in other diseases (e.g. hypertension) as a result of the treatments received and not the underlying disease. Renin is an aspartyl protease and key protein of the renin-angiotensin-aldosterone system (RAAS) playing a major role in blood pressure regulation. Whether renin may play a contributory role in the pathogenesis of C3G is unknown. Therefore, we sought to test renin cleavage of C3 using a broad range of renin concentrations using silver staining to detect cleavage activity.
In conclusion, clinical data should be considered when analyzing biomarkers since therapies can alter protein expression. Renin’s increased expression in C3G was an incidental finding, secondary to non-disease specific treatments. The two recombinant renin (rRenin) proteins used in this study produced different results. One of which did not cleave C3 (BioVision) and one which cleaved C3 (AbCam). No C3 breakdown products were identified at biologically relevant concentrations of renin. The rRenin that cleaved C3 contained PRSS1, a serine protease, likely the reason for cleavage of C3. This finding supported previous findings of serine proteases cleaving C3. Although, we demonstrated that renin was elevated in patients with C3G our renin studies did not provide evidence to support a direct role for renin in cleaving C3. Uncovering novel biomarkers using SOMAscan may be of value as diagnostic markers, or for the development of disease-specific drug therapies, as well as aid in refining our understanding of C3G.
Details
- Title: Subtitle
- Defining the pathophysiology of C3 glomerulopathy - an ultra-rare complement-mediated renal disease -
- Creators
- Bertha Martin
- Contributors
- Richard J.H. Smith (Advisor)John F. Engelhardt (Committee Member)Carla M. Nester (Committee Member)Todd E. Scheetz (Committee Member)Val C. Sheffield (Committee Member)Tina Tootle (Committee Member)Charles Yeaman (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Anatomy and Cell Biology
- Date degree season
- Autumn 2020
- DOI
- 10.17077/etd.005691
- Publisher
- University of Iowa
- Number of pages
- xx, 143 pages
- Copyright
- Copyright 2020 Bertha Martin
- Language
- English
- Description illustrations
- color illustrations
- Description bibliographic
- Includes bibliographical references (pages 113-143).
- Public Abstract (ETD)
The kidneys are one of the vital organ systems in our body carrying out a crucial role for our well-being, that of filtering our blood. The kidneys filter out the excess fluid, toxins (e.g. drugs), wastes as well as participating in other key roles. Some of these roles include blood pressure control, stimulate the production of red blood cells, and produce key minerals for strong bones. The complement system part of the innate immune system contains over 40 proteins that protect us from invading pathogens.
The complement system activation can be triggered through one of three pathways each containing the necessary proteins to respond. One of these pathways is the alternative pathway (AP), and the focus of this thesis, it serves as a surveillance mechanism for scouring the body for the presence of pathogens as it is always on at low levels ready to attack and protect our bodies. Although these pathways are centuries old, in C3 glomerulopathy (C3G), an ultra-rare complement-mediated disease, there is still more to understand.
Disruptions to the normal functions of the kidneys due to proteinuria also compromise the kidney’s structure. Kidney disease can progress to a more chronic state if processes are not corrected: blood pressure control and urine protein reduction. Chronic progression of kidney disease leads to kidney failure a state in which the normal function of the kidneys is lost.
The research undertaken in this thesis explores the importance of biomarkers in the disease setting concerning chronic kidney disease which affects the normal pathway activation and regulation of the complement system. We used a combination of molecular diagnostic tools: silver stain in the assessment of the presence of proteins in reactions between proteins and a more sensitive proteomics tool for the identification of protein in patients’ plasma samples. We found that protein markers are a great source for following the course of a patient’s disease. Furthermore, proteins may have differing roles, but the innate role seems to predominate.
- Academic Unit
- Anatomy and Cell Biology
- Record Identifier
- 9984035694202771
Metrics
66 File views/ downloads
42 Record Views