Open Access Dissertation
Nitrate, (NOR 3RP - P) is an oxidized form of nitrogen and it is an integral part of the nitrogen cycle in the natural environment. Nitrates are extensively used in agricultural practices as fertilizers, applied as either animal manures or derived slurries or as chemical fertilizers in the form of ammonium nitrate. Most of the toxicologically relevant nitrate enters the human body through consumption of vegetables, fruits, processed food products, and drinking water. In the ground water system, nitrates are a problematic and widespread contaminant, especially in agricultural areas such as the Midwest, South, and Northeastern U.S. Low-levels of nitrate exposure in drinking water below the EPA level of 10 ppm are not currently considered acutely lethal to humans, but long-term exposure has been increasingly called into question due to concerns about some forms of cancer risk. Nitrate itself is relatively non-toxic, but it transforms to different metabolites once it enters the human body such as nitrite, nitric oxide (NO), and nitrosamine. These metabolites are more harmful to human health than nitrate and cause various kinds of acute human illness such as acute methemoglobinemia (in infants), low blood pressure, increased heart rate, headache, abdominal cramps, vomiting, and, possible chronic disease concerns such as cancer of the digestive tract, thyroid, and ovary. Another chronic disease concern that may be impacted by nitrite/nitrate exposure is Osteoarthritis (OA). OA, the subject of this study, is the most common type of arthritis that wears down the articular cartilage between bones and thus weakens the whole joint. NO production is considered as a mediator for proinflammatory cytokine production that elevates that pain among OA patients. Nitrate and nitrite are responsible for the production of NO endogenously. Therefore, this study is important to explore various concentrations of nitrite exposure and its impact on people having OA and lymphocyte response, stress, and DNA damage. This in-vitro case control study explores whether different levels of nitrite exposure in human mononuclear cells (lymphocytes) has an impact on their proliferation, NO stress activity, and DNA damage. This will increase understanding of the impacts of nitrite/nitrate exposure on human lymphocytes and clarify innate, immunotoxicological impacts at various low doses of nitrite concentration. A total of 32 participants took part in the study, 16 were cases and 16 were controls. Cases were chosen on the criteria of a medically diagnosed OA in any joint, and the controls were participants without OA. Standard cell culture techniques were used to gather lymphocytes from whole blood, culture and expose those lymphocytes to increasing concentrations of sodium nitrite (0, 1, 5, 10, 15 ppm) and challenge the exposed cells (via mitogen) to monitor lymphocyte proliferation, NO activity, and DNA damage via immunoassay and flow cytometry. Additionally, a brief questionnaire about demographics, health history, smoking status, and herbs/supplement, medications was completed by each participant to explore any relationships between the self-report factors and lymphocyte proliferation, NO activity and DNA damage at various nitrite concentrations. Findings indicated that increasing nitrite concentrations suppressed lymphocyte proliferation among all participants (p<0.05). Among case and control participants, the study found increased lymphocyte proliferation, following mitogen stimulation, among the participants not having OA compared to having OA at the exposure levels of 5 ppm. Overall there was no increased NO stress activity observed among the cells challenged with various nitrite concentrations. The DNA damage test showed some interesting results indicating light DNA damage occurred when exposed to 5ppm or more nitrite concentration (p<0.05) for pooled cases and controls. Additionally, questionnaire data indicated some associations between lymphocyte proliferation, NO activity, and DNA Damage under the circumstances stressful life events, multiple prescription drugs, recent immunization and a diagnosis of fibromyalgia. Keywords: Nitrate, Nitrite, Nitric oxide, Lymphocyte Proliferation, NO activity, DNA Damage, Mononuclear cells.
Year of Submission
Doctor of Education
School of Kinesiology, Allied Health, and Human Services
Catherine Zeman, Committee Chair
1 PDF file (xx, 248 pages)
©2018 Junu Shrestha
Shrestha, Junu, "Study of low dose nitrite stress on human mononuclear cells: An in-vitro case-control study on osteoarthritis" (2018). Electronic Theses and Dissertations. 533.