Serum biomarkers regulated by TGF-beta to predict skin disease in patients with diffuse cutaneous systemic sclerosis (scleroderma)
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Scleroderma is a chronic connective tissue disease that occurs one in about every 5,000 individuals. It is characterized by skin fibrosis along with other organ involvement, such as interstitial lung disease, gastro-esophageal reflux disease, and renal crisis. SSc is generally preceded by a condition known as Raynaud's Syndrome. Raynaud's presents with the purpling or whitening of the finger tips and toes, especially in cold weather. Raynaud's is caused by vascular dysfunction in the circulation in the fingers and toes. About 10-15% of people over the age of 35 developing Raynaud's will eventually develop some sort of connective tissue disease. Patients diagnosed with SSc can be placed into one of two categories, Limited or Diffuse Cutaneous Disease. Limited Disease, also known as CREST Syndrome, presents with skin disease limited to the forearms and face, typically with limited systemic involvement. Diffuse Disease presents with more proximal skin disease with typically more severe internal organ involvement. To help diagnose patients, physicians can test for autoantibodies in the serum, specifically anti-topoisomerase antibodies associated with Diffuse Disease and anti-centromere antibodies associated with Limited Disease. Clinically physicians assess the severity of skin disease with a technique known as the Modified Rodnan Skin Score, which determines the extent of skin disease throughout the body. While the molecular pathways involved in the pathogenesis of SSc are largely unknown, some researchers believe that SSc is caused primarily by excessive inflammation, leading to vascular injury, and excessive collagen production causing systemic fibrosis. The pathways involved in inflammation and collagen deposition involve numerous different cytokines, including TGF-β, IL-6, and IFN Type I. All three of these cytokines further regulate gene expression of other proteins involved in inflammation and collagen deposition. One previous study has determined a useful 4-gene biomarker of skin disease using skin biopsy samples from diffuse patients. However, skin samples are hard to obtain so physicians are searching a useful set of biomarkers of disease activity in serum. Researchers seek to identify these proteins in serum and determine their involvement in the pathogenesis of SSc. Identification of these proteins might provide insights on the molecular mechanisms involved and might also indicate therapeutic targets. Serum biomarkers might predict disease severity at different points in time and thus allow physicians to track disease progression over time. In this study we examined TGF-β regulated proteins as potential serum biomarkers. These potential biomarkers included cartilage oligomeric protein (COMP), matrix metalloproteinase-9 (MMP-9), osteopontin and periostin. All four of these proteins have been implicated in the pathogenesis of skin fibrosis in previous studies and therefore are classified as potential biomarkers for SSc skin disease. We studied the levels of each of the four biomarkers in serum samples through ELISA analysis. Results for COMP analysis showed a statistically significant increase between control and diffuse SSc serum samples and results for osteopontin showed a moderate correlation between MRSS values. COMP levels did not correlate with the MRSS, and osteopontin levels showed no statistical significance between controls and diffuse samples. In contrast to previous reports, MMP-9 results showed no correlation with MRSS values or statistical significance between control and diffuse samples. Periostin analysis showed a negative correlation with the MRSS but no statistical significance between control and diffuse samples. Periostin's negative cmrelation with MRSS values is of particular interest as it implies a potential protective role of periostin for fibrosis. The results of this current study varied greatly from previous studies. This variation in results between the current study and previous studies is likely due to a different patient population in Boston. The variation could also be due to the use of different ELISA kits and protocols. It is also possible that the correlations found in previous studies are not as robust as researchers once believed as none of the studies included a validation cohort. In an attempt to detetmine the utility of each biomarker in determining disease trajectory, the four potential biomarkers were compared to delta MRSS values over six months. Interestingly, osteopontin showed a high negative correlation with delta MRSS values, while periostin and COMP showed a high positive correlation with delta MRSS values. Further research is needed to determine whether the proteins in question can be useful biomarkers for SSc skin disease. Also, further research including IFN regulated proteins might help develop an efficient multiple gene biomarker for serum in diffuse patients.
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