Attenuation of Aging-Related Oxidative Stress Pathways by Phytonutrients: A Computational Systems Biology Analysis.

Aging results from gradual accumulation of damage to the cellular functions caused by biochemical processes such as oxidative stress, inflammation-driven prolonged cellular senescence state, immune system malfunction, psychological stress, and epigenetic changes due to exposure to environmental toxins. Plant-derived bioactive molecules have been shown to ameliorate the damage from oxidative stress. This research seeks to uncover the mechanisms of action of how phytochemicals from fruit/berry/vegetable (FBV) juice powder mitigate oxidative stress. The study uses a computational systems biology approach to (1) identify biomolecular pathways of oxidative stress; (2) identify phytochemicals from FBV juice powder and their specific action on oxidative stress mechanisms; and (3) quantitatively estimate the effects of FBV juice powder bioactive compounds on oxidative stress. The compounds in FBV affected two oxidative stress molecular pathways: (1) reactive oxygen species (ROS) production and (2) antioxidant enzyme production. Six bioactive compounds including cyanidin, delphinidin, ellagic acid, kaempherol, malvidin, and rutin in FBV significantly lowered production of ROS and increased the production of antioxidant enzymes such as catalase, heme oxygenase-1, superoxide dismutase, and glutathione peroxidase. FBV juice powder provides a combination of bioactive compounds that attenuate aging by affecting multiple pathways of oxidative stress.

Mechanistic Understanding of D-Glucaric Acid to Support Liver Detoxification Essential to Muscle Health Using a Computational Systems Biology Approach.

Liver and muscle health are intimately connected. Nutritional strategies that support liver detoxification are beneficial to muscle recovery. Computational-in silico-molecular systems' biology analysis of supplementation of calcium and potassium glucarate salts and their metabolite D-glucaric acid (GA) reveals their positive effect on mitigation of liver detoxification via four specific molecular pathways: (1) ROS production, (2) deconjugation, (3) apoptosis of hepatocytes, and (4) ß-glucuronidase synthesis. GA improves liver detoxification by downregulating hepatocyte apoptosis, reducing glucuronide deconjugates levels, reducing ROS production, and inhibiting ß-Glucuronidase enzyme that reduces re-absorption of toxins in hepatocytes. Results from this in silico study provide an integrative molecular mechanistic systems explanation for the mitigation of liver toxicity by GA.

Attenuation of low-grade chronic inflammation by phytonutrients: A computational systems biology analysis.

BACKGROUND: Low-grade chronic inflammation (LGCI) is a strong and independent risk factor for many chronic diseases, like cardiovascular, musculoskeletal, metabolic, and neurological conditions. Dietary intervention studies have reported evidence for the role of plant-derived flavonoids in modulation of LGCI. This research explores the efficacy of Fruit/Berry/Vegetable (FBV) juice powder on LGCI. METHODS: The study employs computational systems biology: 1) to identify biomolecular mechanisms of LGCI; 2) to identify the bioactive compounds of FBV juice powder and their specific effects on mechanisms of LGCI; and, 3) to predict the quantitative effects of those bioactive compounds on LGCI. RESULTS: Four molecular pathways that are affected by the compounds of FBV include: 1) TNF-α production; 2) CCL2 production; 3) IL-1ß production; and 4) reactive oxygen species production. The bioactive compounds including luteolin, epicatechin, epigallocatechin gallate, lycopene, quercetin, vitamin A, vitamin C and vitamin E in FBV significantly lowered TNF-α production, CCL2 production, IL-1ß production, and reactive oxygen species production. CONCLUSION: FBV provides a combination of active ingredients that synergistically affect multiple modalities of low grade chronic inflammation to help improve blood circulation and energy levels, and lower muscle soreness.

Molecular Systems Architecture of Interactome in the Acute Myeloid Leukemia Microenvironment.

A molecular systems architecture is presented for acute myeloid leukemia (AML) to provide a framework for organizing the complexity of biomolecular interactions. AML is a multifactorial disease resulting from impaired differentiation and increased proliferation of hematopoietic precursor cells involving genetic mutations, signaling pathways related to the cancer cell genetics, and molecular interactions between the cancer cell and the tumor microenvironment, including endothelial cells, fibroblasts, myeloid-derived suppressor cells, bone marrow stromal cells, and immune cells (e.g., T-regs, T-helper 1 cells, T-helper 17 cells, T-effector cells, natural killer cells, and dendritic cells). This molecular systems architecture provides a layered understanding of intra- and inter-cellular interactions in the AML cancer cell and the cells in the stromal microenvironment. The molecular systems architecture may be utilized for target identification and the discovery of single and combination therapeutics and strategies to treat AML.

Bioactive compounds in green tea may improve transplant tolerance: A computational systems biology analysis.

BACKGROUND: Green tea (Camellia sinensis) has bioactive compounds that have been shown to possess nutritive effects on various biomolecular processes such as immunomodulation. This research explores the immunomodulatory effects of green tea in reducing transplant rejection. METHOD: The study employs computational systems biology: 1) to identify biomolecular mechanisms of immunomodulation in transplant rejection; 2) to identify the bioactive compounds of green tea and their specific effects on mechanisms of immunomodulation in transplant rejection; and, 3) to predict the quantitative effects of those bioactive compounds on immunomodulation in transplant rejection. RESULTS: Three bioactive compounds of green tea - epicatechin (EC), gallic acid (GA), and epigallocatechin gallate (EGCG), were identified for their potential effects on immunomodulation of transplant rejection. Of the three, EGCG was the only one determined to enhance anti-inflammatory activity by: 1) upregulating synthesis of HO-1 that is known to promote Treg and Th2 phenotypes associated with enabling transplant tolerance; and, 2) downregulating pro-inflammatory cytokines IL-2, IL-17, IFN-γ, TNF-α, NO, IL-6, and IL-1ß that are known to promote Th1 and Th17 phenotypes associated with transplant rejection. CONCLUSIONS: To the best of our knowledge, this study provides the first molecular mechanistic understanding the clinical nutritive value of green tea, specifically the bioactive compound EGCG, in enabling transplant tolerance.

Computational analysis of nitric oxide biotransport to red blood cell in the presence of free hemoglobin and NO donor.

Red blood cells (RBCs) modulate nitric oxide (NO) bioavailability in the vasculature. Extracellular free hemoglobin (Hb) in the vascular lumen can cause NO bioavailability related complications seen in pathological conditions such as pancreatitis, sickle cell disease and malaria. In addition, the role of extracellular free Hb has been critical to estimate kinetic and transport properties of NO-RBCs interactions in 'competition experiments'. We recently reported a strong dependence of NO transport on RBC membrane permeability and hematocrit. NO donors combined with anti-inflammatory drugs are an emergent treatment for diseases like cancer, cardiovascular complications and wound healing. However, the role of RBCs in transport NO from NO donors is not clearly understood. To understand the significance of extracellular free Hb in pathophysiology on NO availability and estimation of the NO-RBC interactions, we developed a computational model to simulate NO biotransport to the RBC in the presence of extracellular free Hb. Using this model, we studied the effect of hematocrit, RBC membrane permeability and NO donors on NO-RBC interactions in the presence and absence of extracellular free Hb. The plasma NO concentration gradients and average plasma NO concentrations changed minimally with increase in extracellular free Hb concentrations at the higher hematocrit as compared to those at the lower hematocrit irrespective of the NO delivery method, indicating that the presence of extracellular free Hb affects the NO transport only at a low hematocrit. We also observed that NO concentrations increased with NO donor concentrations in the absence as well as in the presence of extracellular free Hb. In addition, NO donor supplementation may increase NO availability in the plasma in the event of loss of endothelium-derived NO activity.

Contribution of membrane permeability and unstirred layer diffusion to nitric oxide-red blood cell interaction.

Nitric oxide (NO) consumption by red blood cell (RBC) hemoglobin (Hb) in vasculature is critical in regulating the vascular tone. The paradox of NO production at endothelium in close proximity of an effective NO scavenger Hb in RBCs is mitigated by lower NO consumption by RBCs compared to that of free Hb due to transport resistances including membrane resistance, extra- and intra-cellular resistances for NO biotransport to the RBC. Relative contribution of each transport resistance on NO-RBC interactions is still not clear. We developed a mathematical model of NO transport to a single RBC to quantify the contributions from individual transport barriers by analyzing the effect of RBC membrane permeability (P(m)), hematocrit (Hct) and NO-Hb reaction rate constants on NO-RBC interactions. Our results indicated that intracellular diffusion of NO was not a rate limiting step for NO-RBC interactions. The extracellular diffusion contributed 70-90% of total transport resistance for P(m)>1 cm s(-1) whereas membrane resistance accounts for 50-75% of total transport resistance for P(m)<0.1 cm s(-1). We propose a narrow P(m) range of 0.21-0.44 cm s(-1) for 10-45% Hct, respectively, below which membrane resistance is more significant and above which extracellular diffusion is a dominating transport resistance for NO-RBC interactions.

Low micromolar intravascular cell-free hemoglobin concentration affects vascular NO bioavailability in sickle cell disease: a computational analysis.

In sickle cell disease, the changes in RBC morphology destabilize the red blood cell (RBC) membrane and lead to hemolysis. Several experimental and clinical studies have associated intravascular hemolysis with pulmonary hypertension in sickle cell disease. Cell-free hemoglobin (Hb) from intravascular hemolysis has high affinity for nitrixc oxide (NO) and can affect the NO bioavailability in the sickle cell disease, which may eventually lead to pulmonary hypertension. To study the effects of intravascular hemolysis related cell-free Hb concentrations on NO bioavailability, we developed a two-dimensional mathematical model of NO biotransport in 50-µm arteriole under steady-state sickle cell disease conditions. We analyzed the effects of flow-dependent NO production and axial and radial transport of NO, a recently reported much lower NO-RBC reaction rate constant, and cell-free layer thickness on NO biotransport. Our results show that the presence of cell-free Hb concentrations as low as 0.5 µM results in an approximately three- to sevenfold reduction in the predicted smooth muscle cell NO concentrations compared with those under physiological conditions. In addition, increasing the diffusional resistance for NO in vascular lumen from cell-free layer or reducing NO-RBC reaction rate did not improve the NO bioavailability at the smooth muscle cell layer significantly for cell-free Hb concentrations ≥1 µM. These results suggest that lower NO bioavailability due to low micromolar cell-free Hb can disturb NO homeostasis and cause insufficient bioavailability at the smooth muscle cell layer. Our results supports the hypothesis that hemolysis-associated reduction in NO bioavailability may play a role in the development of pathophysiological complications like pulmonary hypertension in sickle cell disease that are observed in several clinical and experimental studies.

A computational model for nitric oxide, nitrite and nitrate biotransport in the microcirculation: effect of reduced nitric oxide consumption by red blood cells and blood velocity.

Bioavailability of vasoactive endothelium-derived nitric oxide (NO) in vasculature is a critical factor in regulation of many physiological processes. Consumption of NO by RBC plays a crucial role in maintaining NO bioavailability. Recently, Deonikar and Kavdia (2009b) reported an effective NO-RBC reaction rate constant of 0.2×10(5)M(-1)s(-1) that is ~7 times lower than the commonly used NO-RBC reaction rate constant of 1.4×10(5)M(-1)s(-1). To study the effect of lower NO-RBC reaction rate constant and nitrite and nitrate formation (products of NO metabolism in blood), we developed a 2D mathematical model of NO biotransport in 50 and 200µm ID arterioles to calculate NO concentration in radial and axial directions in the vascular lumen and vascular wall of the arterioles. We also simulated the effect of blood velocity on NO distribution in the arterioles to determine whether NO can be transported to downstream locations in the arteriolar lumen. The results indicate that lowering the NO-RBC reaction rate constant increased the NO concentration in the vascular lumen as well as the vascular wall. Increasing the velocity also led to increase in NO concentration. We predict increased NO concentration gradient along the axial direction with an increase in the velocity. The predicted NO concentration was 281-1163nM in the smooth muscle cell layer for 50µm arteriole over the blood velocity range of 0.5-4cms(-1) for k(NO-RBC) of 0.2×10(5)M(-1)s(-1), which is much higher than the reported values from earlier mathematical modeling studies. The NO concentrations are similar to the experimentally measured vascular wall NO concentration range of 300-1000nM in several different vascular beds. The results are significant from the perspective that the downstream transport of NO is possible under the right circumstances.

An integrated computational and experimental model of nitric oxide-red blood cell interactions.

In blood vessels, nitric oxide homeostasis is maintained by its formation by endothelial nitric oxide synthase and its consumption in smooth muscle cells and in vascular lumen by red blood cell (RBC) encapsulated hemoglobin (Hb). Free hemoglobin has a very high reaction rate (k(Hb-NO) approximately 10(7) M(-1) s(-1)) with NO as compared to RBC-Hb. Mechanisms of reduced NO uptake by RBC-Hb has been extensively studied in recent years. A critical factor in the investigation of NO-RBC interactions is delivery of NO. Common NO delivery methods include use of NO donors and bolus saturated NO solutions, which delivers NO homogeneously and only in the vicinity of bolus, respectively. In this study, we developed a flow system that uses gaseous delivery of NO through a polymeric semi-permeable membrane to obtain precise and uniform NO concentrations for NO-RBC interactions. We conducted experiments using the flow system to study the effect of NO concentrations, hematocrit and RBC suspension flow rates on NO-RBC interactions. We developed a computational model to simulate NO transport and to estimate the reaction rate constant for NO-RBC interaction in the flow system. Our results showed that NO consumption of RBCs (i) increased linearly with an increase in available NO, and (ii) decreased with increase in RBCs suspension flow rate. We estimated the reaction rate constant for NO-RBC interactions to be 0.2 x 10(5) M(-1) s(-1) which is approximately 1250-fold lower than NO consumption by free hemoglobin and approximately 2.5-20 fold slower than reported NO-RBC reaction rate.

Extracellular diffusion and permeability effects on NO-RBCs interactions using an experimental and theoretical model.

Nitric oxide (NO) is a potent vasodilator and its homeostasis depends on interaction with RBCs. A key factor in understanding NO-RBC interactions in vascular lumen is a comprehensive analysis of product identification and quantification. In this context, administration of NO during in vitro NO-RBC interactions becomes a crucial variable. In this study, we designed a bioreactor that maintains a precise NO concentration in the headspace that diffuses to RBCs suspension to study the quantitative effect of NO concentration and hematocrit (Hct) on NO-RBC interactions. The products of NO-RBC reaction (nitrite and total nitrogen species (total NOx)) were measured by chemiluminescence assay. A mathematical model simulating NO biotransport to a single RBC was developed to (1) estimate NO-RBC reaction rate constant; (2) predict the NO concentrations in the bulk RBC suspension and at the RBC membrane for RBC membrane NO permeability (P(m)) values of 0.0415-40 cm/s. Measured nitrite and total NOx concentrations increased with increase in headspace NO concentration whereas nitrite concentrations decreased with hematocrit and total NOx concentrations increased with hematocrit. This indicates that the extracellular resistance is a controlling factor for RBC uptake of NO. Modeling results showed that the effective reaction rate constant (k(eff)) for NO-RBC interactions was 2.32 x 10(4)-1.08 x 10(6) M(-1) s(-1). Results also predict that the membrane permeability in the range of 0.0415-0.4 cm/s is required to maintain physiologically relevant levels of NO at the smooth muscle cell layer. The effective reaction rate constant increased with increase in P(m) and magnitude of increase was higher at 45% Hct. For all P(m) values, the k(hb)/k(eff) ratios were lower for 45% Hct as compared to 5% Hct indicating extracellular resistance is important for RBC NO uptake. Our experimental and mathematical analyses of NO-RBC interactions indicate that both unstirred layer and RBC membrane have a significant effect on NO transport to RBCs. In addition, the membrane permeability in the range of 0.0415-0.4 cm/s is required to maintain sufficient NO concentrations at the smooth muscle cell layer.

Modulation of ADP-induced platelet activation by aspirin and pravastatin: role of lectin-like oxidized low-density lipoprotein receptor-1, nitric oxide, oxidative stress, and inside-out integrin signaling.

Lectin-like oxidized low-density lipoprotein (LDL) receptor-1 (LOX-1), a receptor for oxidized-LDL, is up-regulated in activated endothelial cells, and it plays a role in atherothrombosis. However, its role in platelet aggregation is unclear. Both aspirin and HMG CoA reductase inhibitors (statins) reduce LOX-1 expression in endothelial cells. In this study, we investigated the effect of aspirin and pravastatin on LOX-1 expression on plate-lets. After ADP stimulation, mean fluorescence intensity of LOX-1 expression on platelets increased 1.5- to 2.0-fold. Blocking LOX-1 inhibited ADP-induced platelet aggregation in a concentration- and time-dependent manner. We also established that LOX-1 is important for ADP-stimulated inside-out activation of platelet alpha(IIb)beta(3) and alpha(2)beta(1) integrins (fibrinogen receptors). The specificity of this interaction was determined by arginine-glycine-aspartate-peptide inhibition. Furthermore, we found that LOX-1 inhibition of integrin activation is mediated by inhibition of protein kinase C activity. In other experiments, treatment with aspirin (1-10 mM) and pravastatin (1-5 microM) reduced platelet LOX-1 expression, with a synergistic effect of the combination of aspirin and pravastatin. Aspirin and pravastatin both reduced reactive oxygen species (ROS) released by activated platelets measured as malonyldialdehyde (MDA) release and nitrate/nitrite ratio. Aspirin and pravastatin also enhanced nitric oxide (NO) release measured as nitrite/nitrite + nitrate (NOx) ratio in platelet supernates. Small concentrations of aspirin and pravastatin had a synergistic effect on the inhibition of MDA release and enhancement of nitrite/NOx. Thus, LOX-1 is important for ADP-mediated platelet integrin activation, possibly through protein kinase C activation. Furthermore, aspirin and pravastatin inhibit LOX-1 expression on platelets in part by favorably affecting ROS and NO release from activated platelets.