The Effects of Thiamine on Breast Cancer Cells.

(1) Background: Thiamine is an important cofactor for multiple metabolic processes. Its role in cancer has been debated for years. Our aim is to determine if thiamine can convert the cellular metabolic state of breast cancer cells from anaerobic to aerobic, thus reducing their growth. (2) Methods: Breast cancer (MCF7) and non-tumorigenic (MCF10A) cell lines were treated with various doses of thiamine and assessed for changes in cell growth. The mechanism of this relationship was identified through the measurement of enzymatic activity and metabolic changes. (3) Results: A high dose of thiamine reduced cell proliferation in MCF7 (63% decrease, p < 0.0001), but didn't affect apoptosis and the cell-cycle profile. Thiamine had a number of effects in MCF7; it (1) reduced extracellular lactate levels in growth media, (2) increased cellular pyruvate dehydrogenase (PDH) activities and the baseline and maximum cellular oxygen consumption rates, and (3) decreased non-glycolytic acidification, glycolysis, and glycolytic capacity. MCF10A cells preferred mitochondrial respiration instead of glycolysis. In contrast, MCF7 cells were more resistant to mitochondrial respiration, which may explain the inhibitory effect of thiamine on their proliferation. (4) Conclusions: The treatment of MCF7 breast cancer cells with 1 µg/mL and 2 µg/mL of thiamine for 24 h significantly reduced their proliferation. This reduction is associated with a reduction in glycolysis and activation of the PDH complex in breast cancer cells.

Cytochrome c in patients undergoing coronary artery bypass grafting: A post hoc analysis of a randomized trial.

PURPOSE: To establish whether plasma cytochrome c is detectable in patients undergoing cardiac surgery, whether cytochrome c levels are associated with lactate/inflammatory markers/cellular oxygen consumption, and whether cytochrome c levels are associated with clinical outcomes. MATERIALS AND METHODS: This was an observational sub-study of a randomized trial comparing thiamine to placebo in patients undergoing coronary artery bypass grafting. Patients had blood drawn before, after, and again 6h after surgery. Cytochrome c, inflammatory markers, and cellular oxygen consumption were measured. RESULTS: 64 patients were included. Cytochrome c was detectable in 63 (98%) patients at baseline with a median cytochrome c level of 0.18ng/mL (quartiles: 0.13, 0.55). There was no difference from baseline level to post-surgical level (0.19ng/mL [0.09, 0.51], p=0.36) or between post-surgical level and 6-hour post-surgical level (0.17ng/mL [0.10, 0.57], p=0.61). There was no difference between the thiamine and placebo groups' change in cytochrome c levels from baseline to after surgery (p=0.22). Cytochrome c levels were not associated with lactate, inflammatory markers, cellular oxygen consumption, or clinical outcomes. CONCLUSIONS: Cytochrome c levels did not increase after cardiac surgery and was not associated with the degree of inflammation or clinical outcomes.

Thiamine as an adjunctive therapy in cardiac surgery: a randomized, double-blind, placebo-controlled, phase II trial.

BACKGROUND: Thiamine is a vitamin that is essential for adequate aerobic metabolism. The objective of this study was to determine if thiamine administration prior to coronary artery bypass grafting would decrease post-operative lactate levels as a measure of increased aerobic metabolism. METHODS: We performed a randomized, double-blind, placebo-controlled trial of patients undergoing coronary artery bypass grafting. Patients were randomized to receive either intravenous thiamine (200 mg) or placebo both immediately before and again after the surgery. Our primary endpoint was post-operative lactate levels. Additional endpoints included pyruvate dehydrogenase activity, global and cellular oxygen consumption, post-operative complications, and hospital and intensive care unit length of stay. RESULTS: Sixty-four patients were included. Thiamine levels were significantly higher in the thiamine group as compared to the placebo group immediately after surgery (1200 [683, 1200] nmol/L vs. 9 [8, 13] nmol/L, p < 0.001). There was no difference between the groups in the primary endpoint of lactate levels immediately after the surgery (2.0 [1.5, 2.6] mmol/L vs. 2.0 [1.7, 2.4], p = 0.75). Relative pyruvate dehydrogenase activity was lower immediately after the surgery in the thiamine group as compared to the placebo group (15% [11, 37] vs. 28% [15, 84], p = 0.02). Patients receiving thiamine had higher post-operative global oxygen consumption 1 hour after the surgery (difference: 0.37 mL/min/kg [95% CI: 0.03, 0.71], p = 0.03) as well as cellular oxygen consumption. We found no differences in clinical outcomes. CONCLUSIONS: There were no differences in post-operative lactate levels or clinical outcomes between patients receiving thiamine or placebo. Post-operative oxygen consumption was significantly increased among patients receiving thiamine. TRIAL REGISTRATION: clinicaltrials.gov NCT02322892, December 14, 2014.

Pyruvate Dehydrogenase Activity Is Decreased in Emergency Department Patients With Diabetic Ketoacidosis.

OBJECTIVES: The pyruvate dehydrogenase complex (PDH) is an essential enzyme in aerobic metabolism. Ketones are known to inhibit PDH activity, but the extent of this inhibition is unknown in patients with diabetic ketoacidosis (DKA). METHODS: We enrolled adult patients presenting to the emergency department in hyperglycemic crisis. Patients were classified as DKA or hyperglycemia without ketoacidosis based on laboratory criteria. Healthy controls were also enrolled. PDH activity and quantity were measured in isolated peripheral blood mononuclear cells. We compared PDH values between groups and measured the relationship of PDH values to measures of acid-base status. RESULTS: Twenty-seven patients (17 with DKA) and 31 controls were enrolled. Patients with DKA had lower PDH activity and quantity compared to the two other groups. PDH activity was significantly correlated with serum bicarbonate and pH and inversely correlated with the anion gap. CONCLUSIONS: DKA is associated with greater suppression of PDH activity than hyperglycemia without ketoacidosis, and this is correlated with measures of acid-base status. Future studies may determine whether PDH depression plays a role in the pathophysiology of DKA and whether modification of PDH could decrease time to DKA resolution.

Cytochrome C in Patients with Septic Shock.

PURPOSE: Cytochrome c is an essential component of the electron transport chain, and circulating cytochrome c might be an indicator of mitochondrial injury. The objective of this study was to determine whether cytochrome c levels are elevated in septic patients, whether there is an association between cytochrome c levels and lactate/inflammatory markers, and whether elevated levels of cytochrome c are associated with poor outcomes. METHODS: This was a single-center, prospective, observational, pilot study within a randomized, placebo-controlled trial. We enrolled adult patients in septic shock and with an elevated lactate (>3 mmol/L). Blood was collected at enrollment and at 12 and 24  h thereafter. Cytochrome c was measured in plasma using an electrochemiluminescence immunoassay. RESULTS: We included 77 patients. Plasma cytochrome c levels were significantly higher in septic patients than in healthy controls (0.70  ng/mL [quartiles: 0.06, 1.99] vs. 0.19  ng/mL [quartiles: 0.03, 1.32], P = 0.008). Cytochrome c levels at enrollment were positively correlated with lactate levels (r(s) = 0.40, P < 0.001) but not with inflammatory markers. Patients who died before hospital discharge had significantly higher cytochrome c levels than survivors (0.99  ng/mL [quartiles: 0.36, 4.09] vs. 0.58  ng/mL [quartiles: 0.03, 1.64], P = 0.01). When analyzed over time, the difference between survivors and nonsurvivors remained significant (P < 0.001). CONCLUSIONS: Cytochrome c levels are higher in septic patients than in controls. In unadjusted analysis, septic nonsurvivors had higher cytochrome c levels than survivors.

Pyruvate Dehydrogenase Activity Is Decreased in the Peripheral Blood Mononuclear Cells of Patients with Sepsis. A Prospective Observational Trial.

RATIONALE: Rodent studies have shown that pyruvate dehydrogenase (PDH) levels are low in sepsis. This may cause cells to shift to anaerobic metabolism, resulting in increased lactate production. Alterations in PDH during sepsis have never been studied in humans. OBJECTIVES: The objective of this pilot study was to measure PDH activity and quantity in patients with severe sepsis. METHODS: We conducted a pilot case-control study at a single urban tertiary care center. We compared PDH activity and quantity between patients with severe sepsis admitted to the intensive care unit and healthy control subjects. PDH activity and quantity were measured in isolated peripheral blood mononuclear cells. We measured PDH activity and quantity in control subjects at baseline and in patients with sepsis at 0 (baseline), 24, 48, and 72 hours. MEASUREMENTS AND MAIN RESULTS: We enrolled 56 patients with sepsis and 20 control subjects with at least one blood sample being drawn from each patient. PDH activity and quantity in the sepsis group were significantly lower than the control group (P < 0.001). In multivariable linear regression adjusting for age, race, sex, and assay plate, the difference remained significant. Patients with sepsis who died had significantly lower PDH activity compared with those who survived (P = 0.03). CONCLUSIONS: PDH activity and quantity is decreased in peripheral blood mononuclear cells of humans with severe sepsis when compared with healthy control subjects, and may be associated with mortality. Whether decreased PDH activity plays a role in lactate metabolism or whether pharmacologic modification of PDH activity may improve outcomes remains unknown.

Immunocapture and microplate-based activity and quantity measurement of pyruvate dehydrogenase in human peripheral blood mononuclear cells.

BACKGROUND: Pyruvate dehydrogenase (PDH) activity is altered in many human disorders. Current methods require tissue samples and yield inconsistent results. We describe a modified method for measuring PDH activity from isolated human peripheral blood mononuclear cells (PBMCs). RESULTS/METHODOLOGY: We found that PDH activity and quantity can be successfully measured in human PBMCs. Freeze-thaw cycles cannot efficiently disrupt the mitochondrial membrane. Processing time of up to 20 h does not affect PDH activity with proteinase inhibitor addition and a detergent concentration of 3.3% showed maximum yield. Sample protein concentration is correlated to PDH activity and quantity in human PBMCs from healthy subjects. CONCLUSION: Measuring PDH activity from PBMCs is a novel, easy and less invasive way to further understand the role of PDH in human disease.