Durk Pearson & Sandy Shaw’s®
Life Extension NewsTM
Volume 16 No. 5 • May 2013


Atherosclerosis

Protection Against Vascular Aging in NOX-2-knockout Mice

At the start of a new paper,1 the authors note that patients with advanced atherosclerosis have severe deficiencies in the ability to generate new blood vessels to adapt to inadequate blood flow. Moreover, advanced age is a major risk factor for coronary and peripheral arterial diseases and is associated with impaired ability to generate new blood vessel formation to increase blood flow around areas of arterial occlusion. In addition, “the number and/or the functional activities of EPCs [endothelial progenitor cells] have been shown to be impaired by aging in both animals and in humans.”1

The researchers, searching for a mechanism to explain the defective neovascularization in the context of aging, have discovered that mice lacking NOX-2 (a subunit of the enzyme NADPH oxidase) were protected against age-associated impairment in reparative neovascularization following ischemia, as well as protecting EPCs against functional impairments associated with aging.

NADPH oxidase is a major source of ROS (reactive oxygen species) critical for neutrophil antimicrobial function.2

As part of the experimental protocol,1 the scientists studied the effect of aging on the expression of NADPH oxidase subunit NOX-2 in mouse ischemic hindlimb muscles. They found NOX-2 expression to be significantly increased in old as compared to young wild type animals. (NOX-2 knockout animals, of course, do not express the NOX-2 subunit in hindlimb muscles.) Hindlimb ischemia (impaired blood flow) was established in experimental animals with and without NOX-2 by surgery. In wild type mice, aging was associated with a significant impairment of blood flow recuperation [restoration by new blood vessel growth] at day 7, whereas, by contrast, NOX-2-/- [knockout animals] were protected against age-dependent impairment of blood flow recuperation. Likewise, wild type aging animals exhibited a significant increase in oxidative stress in ischemic muscles while NOX-2-deficient mice were protected against the age-associated increase in oxidative stress.

The scientists1 report that in previous studies (as cited), NOX-2-deficient mice were protected against the effects of ischemic strokes, some forms of hypertension, and aortic atherosclerosis induced by high blood cholesterol. Moreover, reduced neovascularization is associated with increased oxidative stress and some antioxidants (including red wine) have been reported to restore at least in part the ability to regenerate new blood vessels.

In one study,3 apocynin, thought to be an indirect NADPH oxidase inhibitor, increased collateral growth capacity, whether administered prior to, or 7 days following, arterial ligation. We were particularly interested, however, in the reported efficacy of annatto extract and beta-carotene in inhibiting NADPH oxidase and increasing expression/activity of antioxidant enzymes.4 Both annatto and beta carotene were able to increase the mRNA levels of SOD (superoxide dismutase) and CAT (catalase) in neutrophils from diabetic rats. As NOX-2 has been reported to be required for the microbiocidal function of neutrophils, it is possible that the annatto/beta-carotene treatments inhibited NOX-2.2 Previous papers published by the same group that published this paper3 were reported to show a reduction in ROS (reactive oxygen species) production in neutrophils of diabetic rats treated for 30 days with beta carotene and annatto extract. The researchers note that this study3 is the first (to their knowledge) to report an increased expression of CAT (catalase, an important antioxidant enzyme that protects against hydrogen peroxide) in neutrophils from diabetic rats treated with beta carotene and annatto extract.

The control group and diabetic group of rats3 received a modified standard diet, whereas experimental groups received the diet + annatto (control-annatto), and diet + annatto (diabetic-annatto) or diet + beta carotene (control-beta) and diet + beta carotene (diabetic-beta). The concentration of bixin (a potent carotenoid antioxidant) in the annatto extract was 2.17 g/100 g. As noted by the authors of the annatto/beta carotene study, “bixin, which is present in the extract of annatto, is one of the most effective biological singlet molecular-oxygen quenchers.” Moreover, they note that annatto, which is a natural yellow-red color, is widely used as a food colorant and has a relatively low cost of production and low toxicity, making it a very attractive pigment for the food industry.

The Terpenoid Bixin May Activate PPARgamma, Enhance Insulin Sensitivity and Increase Adiponectin Secretion from Adipocytes

A 2010 paper5 evaluated the effects of various terpenoids derived from herbal and dietary plants on the activity of PPARgamma, an important regulator of insulin sensitivity. (The synthetic glitazones that activate PPARgamma are drugs used in the treatment of diabetes). In a graph showing the effects of 24 hours of incubation of a number of terpenoids at 50 or 100 μM as compared to a vehicle control set at 100%, bixin showed the greatest activation of PPARgamma ligand at five-fold induction (compared to the vehicle control).

Another paper6 reported that, in mice, bixin also activated PPARalpha, a transcription factor that regulates the expression of genes involved in fatty acid oxidation. Activation of fatty acid oxidation in the liver by PPARalpha improved carbohydrate and lipid metabolism in the subject obese mice. Bixin treatment also improved obesity-induced dysfunctions including hyperglycemia, hyperinsulinemia, and adiponectin deficiency.

One of the effects of PPARgamma is to induce differentiation of preadipocytes to adipocytes. The new adipocytes, being small, have various properties that differ from mature adipocytes, such as being more insulin sensitive and secreting more adiponectin, an insulin-sensitizing adipokine.7

References

  1. Turgeon et al. Protection against vascular aging in Nox2-deficient mice: impact on endothelial progenitor cells and reparative neovascularization. Atherosclerosis. 223:122-9 (2012).
  2. Lamb et al. Endotoxin priming of neutrophils requires endocytosis and NADPH oxidase-dependent endosomal reactive oxygen species. J Biol Chem. 287(15):12395-404 (2012).
  3. Miller et al. Antioxidants reverse age-related collateral growth impairment. J Vasc Res. 47:108-14 (2010).
  4. Rossoni, Jr. et al. Annatto extract and beta carotene enhances antioxidant status and regulate gene expression in neutrophils of diabetic rats. Free Radic Res. 46(3):329-338 (2012).
  5. Goto et al. Various terpenoids derived from herbal and dietary plants function as PPAR modulators and regulate carbohydrate and lipid metabolism. PPAR Res. Volume 2010, Article ID 483958, 9 pp. (this is an open access article).
  6. Goto et al. Bixin activates PPARalpha and improves obesity-induced abnormalities of carbohydrate and lipid metabolism in mice. J Agric Food Chem. 60:11952-8 (2012).
  7. Takahashi et al. Bixin regulates mRNA expression involved in adipogenesis and enhances insulin sensitivity in 3T3-L1 adipocytes through PPARgamma activation. Biochem Biophys Res Commun. 390:1372-6 (2009).

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