Science References

In this day & age, when it comes to your health & medical advice … who do you trust?

I certainly appreciate YOUR trust in me … so who DO I TRUST ?

Before I answer that, let me spell out my biases:

  • safety … first & always … “First, Do No Harm”
  • easy, feasible … before complicated
  • time-honored with modern scientific validation

For Centuries, wise physicians have stated: “follow the time-honored principles, but let your medicine change with the times.”

Interestingly, this principle is the foundation of Dr Tom’s 2020 HealthCare M-A-P, your Most Affordable Protection / Prevention / Prosperity … because your health is your greatest wealth.

In our current age, we are facing uncertainties which have never before been answered. The answers will only solidify over time. When swimming in an ocean of uncertainties, it is prudent to ground on principles, test our hypotheses, and have our findings and practices validated by others.

The CDC predicts each of us will be eventually exposed, and more than 56% are expected to become infected. 

When it comes to COVID, it’s about limiting the severity of infection: mild/moderate is better than severe/critical illness.

How do we do that?

Vaccine theory teaches safe, repetitive immune enhancement prepares an individual for an increasing tide of exposure to pathogens within the community. 

If we are going to be eventually exposed, let’s ALL have a MILD CASE and get on with life.

To wit, below are my current recommendations and the science supporting them:

  • foundational nutrition, using an orthomolecular approach, to optimize self-healing
    • daily ascorbic acid, aka Vitamin C, orally or Intra-Venously
    • oral or injectable Vitamin D3
  • home-based or easy in-office immune enhancement before infection, and immuno-modulation if infected
    • Vitamin D3 as a preventive agent
    • herbal formulations and home-based moxa-bustion or cupping, as safe home-based support of ‘most stressed’ or ‘most vulnerable organs’
    • Immune peptide Thymosin alpha-1 for highly vulnerable individuals to increase baseline immunity
  • use of ‘bio-physics’ – based treatments in addition to bio-chemicals
    • modern adaptations of ancient healing practices, like cupping or moxabustion, which make a difference for those who are house-bound or quarantined
    • photo-dynamic therapy to activate natural or synthetic remedies for their anti-viral and immune enhancing properties

COVID-19 Antibody Seroprevalence in Santa Clara County, California

April 11, 2020

Eran Bendavid1, Bianca Mulaney2, Neeraj Sood3, Soleil Shah2, Emilia Ling2, Rebecca Bromley-Dulfano2, Cara Lai2, Zoe Weissberg2, Rodrigo Saavedra-Walker4, Jim Tedrow5, Dona Tversky6, Andrew Bogan7, Thomas Kupiec8, Daniel Eichner9, Ribhav Gupta10, John P.A. Ioannidis1,10, Jay Bhattacharya1

1 Department of Medicine, Stanford University School of Medicine, Stanford CA
2 Stanford University School of Medicine, Stanford CA
3 Sol Price School of Public Policy, University of Southern California, Los Angeles CA
4 Health Education is Power, Inc., Palo Alto CA
5 The Compliance Resource Group, Inc., Oklahoma City OK
6 Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Stanford CA
7 Bogan Associates, LLC, Palo Alto CA
8 ARL BioPharma, Inc. , Oklahoma City OK
9 Sports Medicine Research and Testing Laboratory, Salt Lake City UT
10 Department of Epidemiology and Population Health, Stanford University School of Medicine, Stanford CA

COVID-19 Antibody Seroprevalence in Santa Clara County, California

Discussion

After adjusting for population and test performance characteristics, we estimate that the seroprevalence of antibodies to SARS-CoV-2 in Santa Clara County is between 2.49% and 4.16%, with uncertainty bounds ranging from 1.80% (lower uncertainty bound of the lowest estimate), up to 5.70% (upper uncertainty bound of the highest estimate). Test performance characteristics are the most critical driver of this range, with lower estimates associated with data suggesting the test has a high sensitivity for identifying SARS- CoV-2, and higher estimates resulting from data suggesting over 30% of positive cases are missed by the test.

These results represent the first large-scale community-based prevalence study in a major US county completed during a rapidly changing pandemic, and with newly available test kits. We consider our estimate to represent the best available current evidence, but recognize that new information, especially about the test kit performance, could result in updated estimates. For example, if new estimates indicate test specificity to be less than 97.9%, our SARS-CoV-2 prevalence estimate would change from 2.8% to less than 1%, and the lower uncertainty bound of our estimate would include zero. On the other hand, lower sensitivity, which has been raised as a concern with point-of-care test kits, would imply that the population prevalence would be even higher. New information on test kit performance and population should be incorporated as more testing is done and we plan to revise our estimates accordingly.

The most important implication of these findings is that the number of infections is much greater than the reported number of cases. Our data imply that, by April 1 (three days prior to the end of our survey) between 48,000 and 81,000 people had been infected in Santa Clara County. The reported number of confirmed positive cases in the county on April 1 was 956, 50-85-fold lower than the number of infections predicted by this study.17 The infection to case ratio, also referred to as an under-ascertainment rate, of at least 50, is meaningfully higher than current estimates.10,18 This ascertainment rate is a fundamental parameter of many projection and epidemiologic models, and is used as a calibration target for understanding epidemic stage and calculating fatality rates.19,20 The under-ascertainment for COVID- 19 is likely a function of reliance on PCR for case identification which misses convalescent cases, early spread in the absence of systematic testing, and asymptomatic or lightly symptomatic infections that go undetected.

medRxiv preprint doi: https://doi.org/10.1101/2020.04.14.20062463. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.
It is made available under a CC-BY-NC-ND 4.0 International license .

The under-ascertainment of infections is central for better estimation of the fatality rate from COVID-19. Many estimates of fatality rate use a ratio of deaths to lagged cases (because of duration from case confirmation to death), with an infections-to-cases ratio in the 1-5-fold range as an estimate of under- ascertainment.3,4,21 Our study suggests that adjustments for under-ascertainment may need to be much higher.

We can use our prevalence estimates to approximate the infection fatality rate from COVID-19 in Santa Clara County. As of April 10, 2020, 50 people have died of COVID-19 in the County, with an average increase of 6% daily in the number of deaths. If our estimates of 48,000-81,000 infections represent the cumulative total on April 1, and we project deaths to April 22 (a 3 week lag from time of infection to death22), we estimate about 100 deaths in the county. A hundred deaths out of 48,000-81,000 infections corresponds to an infection fatality rate of 0.12-0.2%. If antibodies take longer than 3 days to appear, if the average duration from case identification to death is less than 3 weeks, or if the epidemic wave has peaked and growth in deaths is less than 6% daily, then the infection fatality rate would be lower. These straightforward estimations of infection fatality rate fail to account for age structure and changing treatment approaches to COVID-19. Nevertheless, our prevalence estimates can be used to update existing fatality rates given the large upwards revision of under-ascertainment.

While our prevalence estimates of 2.49% to 4.16% are representative of the situation in Santa Clara County as of April 4, other areas are likely to have different seroprevalence estimates based on effective contact rates in the community, social distancing policies to date, and relative disease progression. Our prevalence estimate also suggests that, at this time, a large fraction of the population remains unexposed in Santa Clara County. Repeated serologic testing in different geographies, spaced a few weeks apart, could establish extent of infection over time.

This study had several limitations. First, our sampling strategy selected for members of Santa Clara County with access to Facebook and a car to attend drive-through testing sites. This resulted in an over- representation of white women between the ages of 19 and 64, and an under-representation of Hispanic and Asian populations, relative to our community. Those imbalances were partly addressed by weighting our sample population by zip code, race, and sex to match the county. We did not account for age imbalance in our sample, and could not ascertain representativeness of SARS-CoV-2 antibodies in homeless populations. Other biases, such as bias favoring individuals in good health capable of attending our testing sites, or bias favoring those with prior COVID-like illnesses seeking antibody confirmation are also possible. The overall effect of such biases is hard to ascertain.

The Premier Biotech serology test used in this study has not been approved by the FDA by the time of the study, and validation studies for this assay are ongoing. We used existing test performance data to establish a range of sensitivity and specificity, including reliable but small-size data sourced at Stanford. Test sensitivity varied between the manufacturer’s data and the local data. It is possible that asymptomatic or mildly symptomatic individuals may generate only low-titer antibodies, and that sensitivity may be even lower if there are many such cases.23 Additional validation of the assays used could improve our estimates and those of ongoing serosurveys.

Several teams worldwide have started testing population samples for SARS CoV-2 antibodies, with preliminary findings consistent with a large under-ascertainment of SARS CoV-2 infections. Reports from the town of Robbio, Italy, where the entire population was tested, suggest at least 10% seropositivity;24 and data from Gangelt, a highly affected area in Germany,25 point to 14% seropositivity.

medRxiv preprint doi: https://doi.org/10.1101/2020.04.14.20062463. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity.
It is made available under a CC-BY-NC-ND 4.0 International license .

A recent effort to test the town of Telluride, Colorado is underway, and interim results suggest a prevalence just under 2%.26 Our data from Santa Clara county suggest higher spread of the infection than Telluride but lower than some areas in Europe.

We conclude that based on seroprevalence sampling of a large regional population, the prevalence of SARS-CoV-2 antibodies in Santa Clara County was between 2.49% and 4.16% by early April. While this prevalence may be far smaller than the theoretical final size of the epidemic,27 it suggests that the number of infections is 50-85-fold larger than the number of cases currently detected in Santa Clara County. These new data should allow for better modeling of this pandemic and its progression under various scenarios of non-pharmaceutical interventions. While our study was limited to Santa Clara County, it demonstrates the feasibility of seroprevalence surveys of population samples now, and in the future, to inform our understanding of this pandemic’s progression, project estimates of community vulnerability, and monitor infection fatality rates in different populations over time. It is also an important tool for reducing uncertainty about the state of the epidemic, which may have important public benefits.

Orthomolecular Medicine News Service, Mar 18, 2020

Successful High-Dose Vitamin C Treatment of Patients with Serious and Critical COVID-19 Infection

by Richard Cheng, MD, PhD

(OMNS Mar 18, 2020) A group of medical doctors, healthcare providers and scientists met online March 17, 2020, to discuss the use of high dose intravenous vitamin C (IVC) in the treatment of moderate to severe cases of Covid-19 patients. The key guest was Dr. Enqian Mao, chief of emergency medicine department at Ruijin Hospital, a major hospital in Shanghai, affiliated with the Joatong University College of Medicine. Dr. Mao is also a member of the Senior Expert Team at the Shanghai Public Health Center, where all Covid-19 patients have been treated. In addition, Dr. Mao co-authored the Shanghhai Guidelines for the Treatment of Covid-19 Infection, an official document endorsed by the Shanghai Medical Association and the Shanghai city government. [1]

Dr. Mao has been using high-dose dose IVC to treat patients with acute pancreatitis, sepsis, surgical wound healing and other medical conditions for over 10 years. When Covid-19 broke out, he and other experts thought of vitamin C and recommended IVC for the treatment of moderate to severe cases of Covid-19 patients. The recommendation was accepted early in the epidemic by the Shanghai Expert Team. All serious or critically ill Covid-19 patients in the Shanghai area were treated in Shanghai Public Health Center, for a total of 358 Covid-19 patients as of March 17th, 2020.

Dr. Mao stated that his group treated ~50 cases of moderate to severe cases of Covid-19 infection with high dose IVC. The IVC dosing was in the range of 10,000 mg – 20,000 mg a day for 7-10 days, with 10,000 mg for moderate cases and 20,000 for more severe cases, determined by pulmonary status (mostly the oxygenation index) and coagulation status. All patients who received IVC improved and there was no mortality. Compared to the average of a 30-day hospital stay for all Covid-19 patients, those patients who received high dose IVC had a hospital stay about 3-5 days shorter than the overall patients. Dr. Mao discussed one severe case in particular who was deteriorating rapidly. He gave a bolus of 50,000 mg IVC over a period of 4 hours. The patient’s pulmonary (oxygenation index) status stabilized and improved as the critical care team watched in real time. There were no side effects reported from any of the cases treated with high dose IVC.

Among the international experts who attended today’s video conference were: Dr. Atsuo Yanagisawa, formerly professor of medicine at the Kyorin University, Tokyo, Japan, and the president of the International Society for Orthomolecular Medicine; Dr. Jun Matsuyama of Japan; Dr. Michael J Gonzalez, professor at University of Puerto Rico Medical Sciences, Dr. Jean Drisko, professor of medicine, and Dr. Qi Chen, professor of pharmacology, both at the Kansas University Medical School, Dr. Alpha “Berry” Fowler, professor of pulmonary and critical care medicine, Virginia Commonwealth University, Dr. Maurice Beer and Asa Kitfield, both from NutriDrip and Integrative Medical NY, New York City; Dr. Hong Zhang of Beijing; William T. Penberthy, PhD of CME Scribe, Florida; Ilyes Baghli, MD, president of the Algerian Society of Nutrition and Orthomolecular Medicine (SANMO); Drs. Mignonne Mary and Charles Mary Jr, of the Remedy Room, New Orleans; Dr. Selvam Rengasamy, president of SAHAMM, Malaysia. I, Richard Cheng, MD, PhD of Cheng Integrative Health Center of South Carolina, and Senior Advisor to ShenZhen Medical Association and Shenzhen BaoAn Central Hospital, coordinated this conference.

Albeit a brief meeting of less than 45 minutes due to Dr. Mao’s limited time availability, the audience thanked Dr. Mao for his time and sharing and wished to keep the communication channel open and also able to talk to other clinicians working at the front line against Covid-19.

In a separate meeting, I had the honor to talk to Sheng Wang, M.D., Ph.D., Professor of Critical Care Medicine of Shanghai 10th Hospital, Tongji University College of Medicine at Shanghai China, who also served at the Senior Clinical Expert Team of the Shanghai Covid-19 Control and Prevention Team. There are three lessons that we learned about this Covid-19 infection, Dr. Wang said:

1. Early and high-dose IVC is quite helpful in helping Covid-19 patients. The data is still being finalized and the formal papers will be submitted for publication as soon as they are complete.

2. Covid-19 patients appear to have a high rate of hyper-coagulability. Among the severe cases, ~40% severe cases showed hyper-coagulability, whereas the number among the mild to moderate cases were 15-20%. Heparin was used among those with coagulation issues.

3. The third important lesson learned is the importance for the healthcare team of gearing up to wear protective clothing at the earliest opportunity for intubation and other emergency rescue measures. We found that if we waited until a patient developed the full-blown signs for intubation, then got ready to intubate, we would lose the precious minutes. So the treatment team should lower the threshold for intubation, to allow proper time (~15 minutes or so) for the team to gear up. This critical 15-30 minutes could make a difference in the outcome.

Also, both Drs. Mao and Wang confirmed that there are other medical teams in other parts of the country who have been using high dose IVC treating Covid-19 patients.

For additional reporting and information on China’s successful use of intravenous vitamin C against COVID-19:

Mar 5, 2020Vitamin C Saves Wuhan Family from COVID-19
Mar 3, 2020Shanghai Government Officially Recommends Vitamin C for COVID-19
Mar 1, 2020News Media Attacks Vitamin C Treatment of COVID-19 Coronavirus
Feb 28, 2020Vitamin C and COVID-19 Coronavirus
Feb 23, 2020TONS OF VITAMIN C TO WUHAN: China Using Vitamin C against COVID
Feb 21, 2020Three Intravenous Vitamin C Research Studies Approved for Treating COVID-19
Feb 16, 2020Early Large Dose Intravenous Vitamin C is the Treatment of Choice for 2019-nCov Pneumonia
Feb 13, 2020Coronavirus Patients in China to be Treated with High-Dose Vitamin C
Feb 10, 2020VITAMIN C AND ITS APPLICATION TO THE TREATMENT OF nCoV CORONAVIRUS: How Vitamin C Reduces Severity and Deaths from Serious Viral Respiratory Diseases
Feb 2, 2020Hospital-based Intravenous Vitamin C Treatment for Coronavirus and Related Illnesses
Jan 30, 2020Nutritional Treatment of Coronavirus
Jan 26, 2020Vitamin C Protects Against Coronavirus

References:

1. Expert consensus on comprehensive treatment of coronavirus disease in Shanghai 2019. Chinese Journal of Infectious Diseases, 2020, 38: Pre-published online. DOI: 10.3760/cma.j.issn.1000-6680.2020.0016 and http://rs.yiigle.com/yufabiao/1183266.htm (in Chinese).

Vitam Horm. 2016;102:151-78. doi: 10.1016/bs.vh.2016.04.003. Epub 2016 May 24.

Immune Modulation with Thymosin Alpha 1 Treatment.

King R1Tuthill C2.

Author information

Abstract

Thymosin alpha 1 (Ta1) is a peptide originally isolated from thymic tissue as the compound responsible for restoring immune function to thymectomized mice. Ta1 has a pleiotropic mechanism of action, affecting multiple immune cell subsets that are involved in immune suppression. Ta1 acts through Toll-like receptors in both myeloid and plasmacytoid dendritic cells, leading to activation and stimulation of signaling pathways and initiation of production of immune-related cytokines. Due to the immune stimulating effects of Ta1, the compound would be expected to show utility for treatment of immune suppression, whether related to aging or to diseases such as infection or cancer (italics and bolding mine). Extensive studies in both the preclinical and clinical setting will be summarized in the subsequent sections. These studies have demonstrated improvements in immune system cell subsets and the potential of Ta1 for the treatment of a range of diseases.

© 2016 Elsevier Inc. All rights reserved.

KEYWORDS:

Cancer treatment; Immune modulation; Infectious disease treatment; Thymalfasin; Thymosin alpha 1; Vaccine enhancementPMID: 27450734 DOI: 10.1016/bs.vh.2016.04.003

J Gastroenterol Hepatol. 2004 Dec;19(12):S69-72.

Thymalfasin: an immune system enhancer for the treatment of liver disease.

Sjogren MH1.

Author information

Abstract

Thymalfasin (thymosin-alpha 1) is an immunomodulating agent able to enhance the Thl immune response. It has been evaluated for its immunomodulatory activities and related therapeutic potential in several diseases, including chronic hepatitis B and C, AIDS, primary immunodeficiency diseases, depressed response to vaccination and cancer. The basis for effectiveness in these conditions is primarily through modulation of immunological responsiveness, as thymalfasin has been shown to have beneficial effects on numerous immune system parameters and to increase T-cell differentiation and maturation. Thymalfasin is responsible for reconstitution of immune function when thymic tissue is given back to thymectomized animals. In addition, thymalfasin has been shown to have efficacy in multiple experimental models of immune dysfunction, mainly, infectious diseases such as hepatitis (woodchuck) and influenza (mouse), and cancer such as melanoma (mouse) and colorectal carcinoma (rat) where thymalfasin has shown antitumor effects.

Treatment of 5 Critically Ill Patients With COVID-19 With Convalescent Plasma

Chenguang Shen, PhD1Zhaoqin Wang, PhD1Fang Zhao, PhD1et alYang Yang, MD1Jinxiu Li, MD1Jing Yuan, MD1Fuxiang Wang, MD1Delin Li, PhD1,2Minghui Yang, PhD1Li Xing, MM1Jinli Wei, MM1Haixia Xiao, PhD1,2Yan Yang, MM1Jiuxin Qu, MD1Ling Qing, MM1Li Chen, MD1Zhixiang Xu, MM1Ling Peng, MM1Yanjie Li, MM1Haixia Zheng, MM1Feng Chen, MM1Kun Huang, MM1Yujing Jiang, MM1Dongjing Liu, MD1Zheng Zhang, MD1Yingxia Liu, MD1Lei Liu, MD1Author AffiliationsArticle InformationJAMA. Published online March 27, 2020. doi:10.1001/jama.2020.4783editorial comment icon EditorialCommentrelated articles icon RelatedArticles

Key Points

Question  Could administration of convalescent plasma transfusion be beneficial in the treatment of critically ill patients with coronavirus disease 2019 (COVID-19)?

Findings  In this uncontrolled case series of 5 critically ill patients with COVID-19 and acute respiratory distress syndrome (ARDS), administration of convalescent plasma containing neutralizing antibody was followed by an improvement in clinical status.

Meaning  These preliminary findings raise the possibility that convalescent plasma transfusion may be helpful in the treatment of critically ill patients with COVID-19 and ARDS, but this approach requires evaluation in randomized clinical trials.Abstract

Importance  Coronavirus disease 2019 (COVID-19) is a pandemic with no specific therapeutic agents and substantial mortality. It is critical to find new treatments.

Objective  To determine whether convalescent plasma transfusion may be beneficial in the treatment of critically ill patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.

Design, Setting, and Participants  Case series of 5 critically ill patients with laboratory-confirmed COVID-19 and acute respiratory distress syndrome (ARDS) who met the following criteria: severe pneumonia with rapid progression and continuously high viral load despite antiviral treatment; Pao2/Fio2 <300; and mechanical ventilation. All 5 were treated with convalescent plasma transfusion. The study was conducted at the infectious disease department, Shenzhen Third People’s Hospital in Shenzhen, China, from January 20, 2020, to March 25, 2020; final date of follow-up was March 25, 2020. Clinical outcomes were compared before and after convalescent plasma transfusion.

Exposures  Patients received transfusion with convalescent plasma with a SARS-CoV-2–specific antibody (IgG) binding titer greater than 1:1000 (end point dilution titer, by enzyme-linked immunosorbent assay [ELISA]) and a neutralization titer greater than 40 (end point dilution titer) that had been obtained from 5 patients who recovered from COVID-19. Convalescent plasma was administered between 10 and 22 days after admission.

Main Outcomes and Measures  Changes of body temperature, Sequential Organ Failure Assessment (SOFA) score (range 0-24, with higher scores indicating more severe illness), Pao2/Fio2, viral load, serum antibody titer, routine blood biochemical index, ARDS, and ventilatory and extracorporeal membrane oxygenation (ECMO) supports before and after convalescent plasma transfusion.

Results  All 5 patients (age range, 36-65 years; 2 women) were receiving mechanical ventilation at the time of treatment and all had received antiviral agents and methylprednisolone. Following plasma transfusion, body temperature normalized within 3 days in 4 of 5 patients, the SOFA score decreased, and Pao2/Fio2 increased within 12 days (range, 172-276 before and 284-366 after). Viral loads also decreased and became negative within 12 days after the transfusion, and SARS-CoV-2–specific ELISA and neutralizing antibody titers increased following the transfusion (range, 40-60 before and 80-320 on day 7). ARDS resolved in 4 patients at 12 days after transfusion, and 3 patients were weaned from mechanical ventilation within 2 weeks of treatment. Of the 5 patients, 3 have been discharged from the hospital (length of stay: 53, 51, and 55 days), and 2 are in stable condition at 37 days after transfusion.

Conclusions and Relevance  In this preliminary uncontrolled case series of 5 critically ill patients with COVID-19 and ARDS, administration of convalescent plasma containing neutralizing antibody was followed by improvement in their clinical status. The limited sample size and study design preclude a definitive statement about the potential effectiveness of this treatment, and these observations require evaluation in clinical trials.

Regenerative Medicine for Knee Pain & Arthritis

Review Article

Achieving the Balance between ROS and Antioxidants: When to Use the Synthetic Antioxidants

Borut Poljsak,1 Dušan Šuput,2 and Irina Milisav1,2

1 University of Ljubljana, Laboratory of Oxidative Stress Research, Faculty of Health Sciences, Zdravstvena Pot 5, SI-1000 Ljubljana, Slovenia

2 University of Ljubljana, Faculty of Medicine, Institute of Pathophysiology, Zaloska 4, SI-1000 Ljubljana, Slovenia

Correspondence should be addressed to Irina Milisav; irina.milisav@mf.uni-lj.si

Received 4 February 2013; Accepted 7 April 2013

Academic Editor: Oren Tirosh

Copyright © 2013 Borut Poljsak et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Free radical damage is linked to formation of many degenerative diseases, including cancer, cardiovascular disease, cataracts, and aging. Excessive reactive oxygen species (ROS) formation can induce oxidative stress, leading to cell damage that can culminate in cell death. Therefore, cells have antioxidant networks to scavenge excessively produced ROS. The balance between the production and scavenging of ROS leads to homeostasis in general; however, the balance is somehow shifted towards the formation of free radicals, which results in accumulated cell damage in time. Antioxidants can attenuate the damaging effects of ROS in vitro and delay many events that contribute to cellular aging. The use of multivitamin/mineral supplements (MVMs) has grown rapidly over the past decades. Some recent studies demonstrated no effect of antioxidant therapy; sometimes the intake of antioxidants even increased mortality. Oxidative stress is damaging and beneficial for the organism, as some ROS are signaling molecules in cellular signaling pathways. Lowering the levels of oxidative stress by antioxidant supplements is not beneficial in such cases. The balance between ROS and antioxidants is optimal, as both extremes, oxidative and antioxidative stress, are damaging. Therefore, there is a need for accurate determination of individual’s oxidative stress levels before prescribing the supplement antioxidants.

1. Introduction

Free radicals are reactive chemicals with an unpaired electron

in an outer orbit [1]. Reactive oxygen species (ROS) comprise

(O ), singlet oxygen (1/2O ), and the hydroxyl radical

( OH). There are also reactive nitrogen, iron, copper, and sul- fur species [1, 2] which could attribute to increased ROS for- mation and oxidative stress and impair the redox balance. No matter how careful we are, we cannot avoid endogenous and exogenous free radical formation due to normal metabolism and exposure to environmental oxidants [3]. Free radicals are produced when our cells create energy from food and oxygen and when we are exposed to microbial infections, extensive exercise, or pollutants/toxins such as cigarette smoke, alcohol, ionizing and UV radiations, pesticides, and ozone. The most important endogenous sources of oxidizing agents contribut- ing to aging are mitochondrial: electron transport chain and

nitric oxide synthase reaction. Nonmitochondrial sources of

burst of phagocytic cells [4]. It has been shown that oxidative stress is involved in over 100 diseases, as their cause or consequence [2, 5]. Oxidative stress was first defined by Sies [6] as “a disturbance in the prooxidant to antioxidant balance in favor of the former, leading to potential damage” (Figures 1 and 2). Oxidative stress can be defined as an excessive amount of ROS, which is the net result of an imbalance between production and destruction of ROS (the latter is regulated by antioxidant defences). Oxidative stress is a consequence of an increased generation of free radicals and/or reduced physiological activity of antioxidant defenses against free radicals. All forms of life maintain a reducing environment within their cells. This reducing environment is preserved by enzymes that maintain the reduced state through a constant input of metabolic energy. Disturbances in this normal redox state can cause toxic effects through the production of

both free radical and nonfree radical oxygen containing

2∙− 2

m∙olecules such as hydrogen peroxide (H O ), superoxide

22

450

free radicals are Fenton’s reaction, microsomal cytochrome

P enzymes, peroxisomal beta-oxidation, and respiratory

2

Oxidative Medicine and Cellular Longevity

Antioxidative defenses

Slight prooxidative balance is necessary for optimal cell signaling processes

Th e excessive ROS formation should be corrected only to prevent the accumulation of oxidative damage
(shown by the arrow between straight to dashed line ratio)

Preventive measures

Decrease the formation of ROS by optimal functioning of oxygen metabolism and avoidance of environmental pollutants

Increase the neutralization of ROS by appropriate antioxidant intake

Figure 1: Model antioxidative/oxidative balance of an adult person—the balance is slightly moved towards the increased ROS production (dashed line). The physiological balance is represented by the dashed line and not the dotted line (geometrical balance), since slight pro- oxidative balance is necessary for optimal immune system and cell signaling processes.

The physiological balance Severe oxidative stress

Figure 2: Oxidative stress due to severe disturbance between ROS formation and antioxidative defenses (the physiological balance is represented by the dashed line).

peroxides and free radicals that damage all components of the cell. Severe oxidative stress can cause cell death. The degree of oxidative stress experienced by the cell will be a function of the activity of ROS generating reactions and the activity of the ROS scavenging system. In physiological conditions, the balance between prooxidant and antioxidant substances is kept slightly in favor of prooxidant products, thus favoring a mild oxidative stress (Figure 1) [7].

2. Beneficial Use of Antioxidants

A biological antioxidant has been defined as any sub- stance that is present at low concentrations compared to an oxidizable substrate and significantly delays or prevents the oxidation of that substrate [8]. An ideal antioxidant should be readily absorbed by body and should prevent or quench free radical formation or chelate redox metals at physiologically relevant levels. It should work in aqueous and/or membrane domains and effect gene expression in a positive way [9]. Cellular redox homeostasis is carefully maintained by an elaborate endogenous antioxidant defense system, which includes endogenous antioxidant enzymes such as superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), glutathione (GSH), proteins, and low- molecular-weight scavengers, like uric acid, coenzyme Q, and

lipoic acid. The human antioxidant defense is complex and must minimize the levels of ROS while allowing useful roles of ROS to perform cell signaling and redox regulation [10].

Generation of ROS and the activity of antioxidant defense appear more or less balanced in vivo. In fact, as already mentioned, the balance may be slightly tipped in favor of the ROS so that there is continuous ROS formation and low- level oxidative damage in the human body (Figure 1). This creates a need for a second category of endogenous antiox- idant defense system, which removes or repairs damaged biomolecules before they accumulate and result in altered cell metabolism and permanent damage [11]. Oxidatively damaged nucleic acids are repaired by specific enzymes, oxidized proteins are removed by proteolytic systems, and oxidized lipids are repaired by phospholipases, peroxidases, and acyl transferases [11]. It seems that failures of some or all of the repair systems contribute more to aging and age-related diseases than moderate oscillations in antioxidants and ROS formation [12–14]. Many of the essential maintenance repair systems become deficient in senescent cells, cell damage accu- mulates, for example, lysosomal accumulation of lipofuscin [15, 16]. Age-related oxidative changes are most prominent in nonproliferating cells, such as neurons and cardiac myocytes because there is no “dilution” of damaged structures through celldivision[17].Besides,Dro ̈geandSchipper[18]and Bokov et al. [19] proposed that general signalling failure with aging could be due to insufficient reactive species or wrong reactive species production although it is known that oxidative damage increases with age in a variety of tissues and animal models [19].

The effects of increased fruit and vegetable intake are associated with lowered parameters of cell damage in vitro, for example, lower oxidative stress, DNA damage, malignant transformation rate, and so forth; epidemiologically they seem to result in lowered incidence of certain types of cancer and degenerative diseases, such as ischemic heart

Antioxidative defenses

ROS

ROS

Oxidative Medicine and Cellular Longevity

3

disease and cataract [20–25]. On the other hand, increased or prolonged free radical action can overwhelm ROS defense mechanisms, contributing to development of diseases and aging. Since oxidative damage of our cells increases with age, the increased intake of exogenous antioxidants from fruit and vegetables may support the endogenous antioxidative defense. The antioxidants, like vitamin C and E, carotenoids, and polyphenols (e.g., flavonoids), are presently considered to be the main exogenous antioxidants. Clinical studies imply that eating a diet rich in fruits, vegetables, whole grains, legumes, and omega-3 fatty acids can help humans in disease prevention [26].

3. Use of Synthetic Antioxidants: Their Control and Safety

A dietary supplement, also known as a food or nutritional supplement, is a preparation intended to provide nutrients such as vitamins, minerals, fibres, fatty, or amino acids that are either missing or not consumed in sufficient amounts in person’s diet. Surveys indicate that more than half of the US adult population uses food supplements, many of which con- tain antioxidants, such as vitamin A (retinoids, carotenes), vitamins C and E (tocopherols), lycopene, lutein, ubiquinone, glutathione, polyphenols (flavonoids), resveratrol, and N- acetylcysteine. In the USA, food supplements represent a market of over $7 billion/year [27] and exceed $30 billion worldwide [28].

In the United States of America (and in many other countries), the dietary supplement or dietary ingredient manufacturer is responsible for ensuring that a dietary supplement or ingredient is safe before it is marketed under the Dietary Supplement Health and Education Act of 1994 [29]. Generally, manufacturers neither need to register their products with Food and Drug Administration (FDA) nor get FDA approval before producing or selling dietary supplements. FDA is responsible for taking action against any unsafe dietary supplement product after it reaches the market. Manufacturers must make sure that product label information is truthful and not misleading and are required to submit to FDA all serious adverse event reports associated with use of the dietary supplement in the United States. In contrast, the substances used as drugs must undergo clini- cal studies to determine their effectiveness, safety, possible interactions with other substances, and appropriate dosages before entering the market [30]. FDA independently reviews company’s data and proposed labeling and, if health benefits outweigh its known risks, it approves the drug for sale.

The inappropriate use of dietary supplements may lead to “antioxidative stress.” This term was used for the first time by Dundar and Aslan [31] for description of the negative effects of antioxidants; it is discussed also by recent publication by Poljsak and Milisav [32]. Both “antioxidative” and oxida- tive stresses leading to the antioxidative imbalance can be damaging for the organism and can result in cancerogenesis [33] and aging (Figures 2 and 3). There are a growing number of clinical trials in which individuals received one or more synthetic antioxidants that fail to demonstrate

ROS

The physiological balance

Antioxidative defenses

Severe “antioxidative stress”

Figure 3: Severe disturbance between antioxidative defenses and ROS leads to a state of increased “antioxidative stress.”

beneficial effects of antioxidant supplementation. Some even implied that antioxidant therapy had no effect and even could increase mortality [34–46]. Ristow et al. [47] reported that nutritive antioxidants abolished the life extension by inhibiting a process called mitohormesis. Results of clinical trials on exogenous antioxidants intake are thus conflicting and contradictory. There seem to be homeostatic mechanisms in cells that govern the total antioxidant activity. Modifying the levels of one antioxidant causes compensatory changes in the levels of others, while the overall antioxidant capacity remains unaffected. The intake of only one antioxidant may thus alter the complex system of endogenous antioxidative defence of cells or alter the cell apoptosis pathways [48]. Dosing cells with exogenous antioxidants may decrease the rate of synthesis or uptake of endogenous antioxidants, so that the total “cell antioxidant potential” remains unaltered. Cutler [49, 50] introduced “The oxidative stress compensation model” to explain why dietary supplements of antioxidants have minimal effect on longevity. He explains that most humans are able to maintain their set point of oxidative stress even if they consume additional antioxidant supplements; in other words, there is no further decrease in oxidative stress [49, 50].

4. Importance of the Balance

The production of free radicals increases with age [51], while some of the endogenous defense mechanisms decrease [52]. This imbalance leads to progressive damage of cellular structures, presumably resulting in the aging phenotype [53,

54].The antioxidant defense system must thus minimize the levels of most harmful ROS on one side while still permit enough ROS to remain for their useful purposes (e.g., cell signaling and redox regulation). Cells usually tolerate such mild oxidative stress; this stress can even upregulate cellular repair processes and other protective systems (e.g., chaperones).

5. ‘‘Antioxidative Stress’’ Influences Cell Signaling and Redox Regulation

The beneficial physiological cellular use of ROS is now being demonstrated in different fields, including intracellular signaling and redox regulation. It is well documented that low levels of ROS are signaling molecules, modulating cell proliferation [55], apoptosis [56, 57], and gene expression through activation of transcription factors [58], like NF- kappa-B and hypoxia-inducible-factor-1 (HIF) [59]. The

4

inducers of NF-kappa-B include also tumor necrosis factor alpha (TNF) and interleukin 1-beta (IL-1) [60, 61]. ROS can act as signaling intermediates for cytokines, including IL-1 and TNF [62–64]. These proinflammatory cytokines, tumor necrosis factor (TNF)-, interleukin-1 (IL-1), and interferon- (IFN-), can additionally increase oxidative stress in humans [65], inducing production of ROS [64, 66]. ROS also have a role in vascular cell signaling processes including activation of eNOS [67] and stimulation of cell growth and migration [68] through modulation of intra- cellular calcium [69] and activation of transcription factors such as NF-kappa-B [70] and protein kinases including ERK, p38MAPK, and Akt [71, 72]. ROS signaling is thus integrated into many cellular pathways, including but not limited to (1) proliferation and survival pathways through MAP kinases, PI3 kinase, PTEN, and protein tyrosine phosphatases; (2) ROS homeostasis and antioxidant gene regulation through Ref-1, Nrf-2, thioredoxin, and so forth; (3) aging through p66Shc; (4) DNA damage response through ATM kinase; this may lead to inhibition of mTORC1 resulting in suppression of protein synthesis and activation of autophagy; (5) iron homeostasis through iron-regulatory proteins (IRP) and iron-responsive elements (IRE), and so forth [73].

Oxidative Medicine and Cellular Longevity

2∙− 22

Individuals who overdose antioxidant supplements could enter the status of “antioxidative” stress (Figure 3). If admin- istration of antioxidant supplements decreases the level of free radicals, it may interfere with the immune system to fight bacteria and essential defensive mechanisms for removal of damaged cells, including those that are precancerous and cancerous [84]. Thus, antioxidant supplement overtake may cause harm [35, 36, 56, 85, 86]. When large amounts of antioxidant nutrients are taken, they can also act as prooxidants by increasing oxidative stress [87, 88]. Pro- and antioxidant effects of antioxidants (e.g., vitamin C) are dose dependent, and thus, more is not necessarily better. Our diets typically contain safe levels of vitamins; therefore, high-level antioxidant supplements may upset this important physiolog- ical balance between the ROS formation and neutralization.

The amount of oxidized macromolecules in the cell is the sum of the rate of their formation subtracted by the rate of repair processes. The imposed oxidative damage potential is opposed by the antioxidant defense capacity of the system (Figure 4). In reality, the oxidative damage potential is greater, and thus there is a constant small amount of toxic free radical formation, which escapes the defense of the cell. A certain amount of oxidized proteins and nucleic acids exists in cells at all times; this reflects the oxidative events. Decreased com- pensation of oxidative stress and insufficient repair, in turn, accelerate aging, which consequently leads to further decline of cellular energy levels [89, 90]. Mechanisms that protect cells from oxidative stress (e.g., endogenous antioxidants, DNA repair processes) are consuming significant amounts of energy when being activated in all compartments of the cell for prolonged time. It may require too much energy to pre- vent all oxidative damage throughout the life of an organism. Kowald and Kirkwood predicted that virtual immortality might be achieved if 55% of the total energy of the simulated cell were devoted to repair and/or prevention of free radical and oxidative damage on the quantitative MARS model (mitochondria, aberrant proteins, radicals, and scavengers) [91, 92]. It is the compromise to allocate suboptimal amounts of energy to cell repair systems, with a consequence of gradual deterioration of the body structures with age [93]. Paradox- ically, the efficiency of defense and repair may be enhanced also after the exposure to ROS, since the expression of many DNA repair enzymes is upregulated during the oxidative stress [94–96]. Finkel and Holbrook [97] stated that the best strategy to enhance endogenous antioxidant levels may actually be oxidative stress itself, based on the classical phys- iological concept of hormesis [98]. This is in agreement with a recent Halliwell’s proposal that stimulating the increase in levels of endogenous antioxidants by some prooxidants may be more effective than consuming additional dietary antioxi- dants [10]. Many well-established components of the heart- healthy lifestyle are prooxidant, including the polyunsatu- rated fat, exercise, and moderate alcohol consumption [99].

7. The Importance of Determination of the Oxidative/Antioxidative Status In Vivo

In order to determine the oxidative stress, both, the ROS formation as well as the antioxidative defense potential

The production of O and H O by activated phago-

cytes is the classic example of the deliberate metabolic gen- eration of ROS for useful purposes [74]. H O is recognized

as an ubiquitous intracellular messenger [75–78]. Moderate

amounts of mitochondrial superoxide and hydrogen per-

oxide have important roles in a range of cellular signaling

processes and can activate signaling pathways that promote

2∙− 22

cell survival and disease resistance due to hormesis [79–

81]. Generation2 of O , HOCl, and H O by phagocytes is

∙−

important for defense against various bacterial and fungal

strains [82]. O is produced also by several cell types other than phagocytes, including lymphocytes and fibroblasts [82]. As ROS are important in signal transduction, there seem to be no great reserve of antioxidant defenses in mammals [83].

6. Imbalance between ROS and Antioxidants

6.1. Increased2 Oxidative Stress. The causes of increased ROS production include endogenous reasons (inflammation, ele- vation in O concentration, and increased mitochondrial leakage) and exogenous (environmental pollution, strenuous exercise, smoking, nutrition, chronic inflammation, psycho- logical and emotional stress, and others) [3, 32, 79]. Causes of decreased antioxidant defenses include reduced activity of endogenous antioxidative enzymes and reduced intake or absorption of antioxidants from food.

6.2. Increased “Antioxidative Stress”. Inappropriate antiox- idative intake may cause increased “antioxidative stress.” Antioxidants can neutralize ROS and decrease oxidative stress; however, this is not always beneficial with respect to the development of a disease and its progression (e.g., cancer) or for delaying aging [32] since antioxidants cannot distinguish among the radicals with a beneficial physiological role and those that cause oxidative damage to biomolecules.

22

Oxidative Medicine and Cellular Longevity

5

Antioxidative defenses

Figure 4: The optimal situation: the physiological balance between the ROS production and antioxidative defenses prevents the accu- mulation of damage by ROS and enables enough ROS for signaling. Enzymatic and nonenzymatic antioxidants can neutralize ROS and RNS and decrease oxidative stress and restore the balance.

should be measured; for example, low antioxidant amount is not problematic when the ROS levels are low.

7.1. Determination of ROS. Free radicals have a very short

half-life, which makes them hard to measure in the lab-

oratory. Nevertheless, multiple methods of oxidative stress

measurement are available today, each with their own advan-

tages and disadvantages (see review [101]). Many approaches

are possible: identification of free radicals, either directly by

paramagnetic electron resonance (electron spin resonance,

ESR), or indirectly by identifying some more stable interme-

diates: evaluation of the traces of radical attack on biological

molecules by high performance liquid chromatography, gas-

liquid chromatography, colorimetric tests. The measurement

of antioxidant status can be estimated by colorim∙etric,

These are very reactive species and their quantitation is difficult. In vivo ESR is relatively insensitive and requires steady-state concentrations of free radicals in the micromolar range, which limits its use for measuring ROS in patients. ESR can be applied only through the technique of spin trapping for in vivo samples. Although it seems that toxicity is not a serious problem for most traps, there are no effective spin traps to be administered to humans. Indirect methods are used in order to overcome these problems. Indirect methods usually measure the changes in endogenous antioxidant defense systems or measure the ROS-induced damage of cellular components [101]. Measuring the damage caused by ROS instead of direct measuring of ROS seems logical, since it is the damage caused by ROS that is important rather than the total amount of generated ROS. Methods have been developed to detect and quantify oxidative damage to pro- teins, lipids, and DNA. The principle behind fingerprinting methods is to measure products of damage by ROS, that is, to measure not the species themselves but the damage that they cause. Of course, the end-products must be specific markers of oxidative damage [8]. According to Miwa et al. [102], a good marker of oxidative damage must increase by oxidative stress (i.e., upon the treatment with, e.g., paraquat, diquat, ionizing radiation, hyperoxia), and it must remain unchanged in the absence of the oxidative event.

7.2. Determination of Antioxidant Status. There is a growing interest to measure antioxidant status for clinical assessment [103]. Cellular protection against unwanted oxidation is achieved mainly by enzymes, such as superoxide dismutase (SOD), catalase, and glutathione peroxidase, whereas the

nonenzymatic antioxidants are playing the major role in the plasma. Radical-scavenging antioxidants are consumed during the reactions with ROS, and antioxidant status could be used indirectly to assess the free radical activity. One approach is to measure individual antioxidants (e.g., ascorbate, -tocopherol, urate) in blood, plasma, or tissue homogenates. All of the individual molecules that are cur- rently recognized as antioxidants should be measured [103]. However, this approach has several shortcomings: (1) it is time consuming, expensive, and technically demanding, (2) it may not detect the synergistic effects between the antioxi- dants, and (3) it may not account for the influence of presently unknown antioxidant substances. The other approach is to measure the total antioxidant capacity or activity by sub- jecting the samples to controlled oxidative stress conditions and measuring either the rate of oxidation or how long it takes for oxidation to occur. Determination of antioxidative potential per se is not sufficient, since it is difficult to establish how the individual antioxidants work: by preventing the formation of ROS, by scavenging free radicals, by inducing the signaling pathways, or by repairing the oxidative damage. Additionally, antioxidative status differs significantly between the individuals and between the laboratory methods used in humans [101]. Typical oxidative stress status of an individual is not established so far [104]. There are no reference values on the optimal levels of antioxidants in urine, blood, or even intracellularly. Additionally, several free radicals cannot cross cell membranes due to their charge, or they are so short-lived that their diffusion is negligible. As such they cannot enter the blood from an affected region or organ. As there is no direct correlation between the oxidative stress markers in blood and their levels within the cells measuring the blood samples may be misleading. Besides, unknown are the amounts and combinations of antioxidants needed for the beneficial effect in vivo. The improved methodology for determining the oxidative stress levels in humans may overcome at least some of these drawbacks.

Long-term effects of oxidative stress will occur if antiox- idant status is low and levels of free radicals are high. No specific clinical symptoms or clinical signs are associated with oxidative stress during the early stages of imbalance. There- fore, the oxidative stress is not diagnosed until there is an unavoidable damage and the consequences manifest as a sign of a disease that could last for decades. Thus, oxidative stress should be recognized and the oxidative imbalance should be ameliorated in order to prevent or postpone the free radical- related disease development and premature aging [32]. In practice, it is difficult to determine all types of ROS within the cells or cellular compartments, as well as the overall antioxidative protection and repair of cells and organs at any specific time. Increased oxidative stress could result from increased ROS production or from decreased antioxidative defences. Thus, increased ROS damage could be the result of (a) increased ROS formation, (b) decreased antioxidative defence, or/and (c) altered damage repair (Scheme 2). Since none of the biomarkers can predict the disease development as the consequence of the prolonged oxidative stress [105], it is important to use many methods for detection and quantification of oxidative stress whenever possible in order

immune, or enzymatic methods [100] (Scheme 1). The direct

ROS detection methods measure superoxide, H O , OH.

ROS

Geometrical balance

Physiological balance

22

6

Oxidative Medicine and Cellular Longevity

Direct methods

Methods of free radicals and other reactive species detection

EPR (ESR) and spin trapping

Fluorescent probes

Indirect methods

Measurement of total antioxidant status

Lipid peroxidation

Superoxide dismutase

Catalase

Glutathione peroxidase

Glutathione reductase

Glutathione

4-HNE

8-isoprostaglandin F2𝛼 (8-isoprostane)

MDA

Protein damage

PCC

3-Nitrotyrosine

AGE AOPP

Glycoxidation adducts

Cross-links,

aggregates, fragments

Lipid peroxidation and amino acid oxidation adducts

Others

Measurement of endogenous enzymatic and nonenzymatic antioxidant defense systems

Fingerprinting methods of:

Oxidative DNA damage

8-OHdG

Abasic (AP) sites

Double-strand DNA

breaks

Comet assay (general DNA damage)

Others

Others

Scheme 1: Methods of oxidative stress determination. (8-OHG) = 8-hydroxyguanosine; 4-HNE = 4-Hydroxynonenal; MDA = malondialde- hyde; PCC = protein carbonyl content; AGE = advanced glycation end products; AOPP = advanced oxidation protein products.

to enhance their validity, as each method measures different parameters and has inherent limitations. No single method can measure the oxidative stress or its subsequent damage in vivo accurately at present. Additionally, the living organisms are complex and ever changing systems and therefore any

determination of the oxidative stress levels reflect the tem- porary state that may change considerably over time.

Presently, the use of supplemental antioxidants could be advised only in cases of well-known conditions, where the depletion of antioxidants is known and can be predicted.

Other antioxidative

defense systems and repair processes

Oxidative Medicine and Cellular Longevity

7

Consequence
Defective immune and apoptosis systems

Feedback loops between ROS, antioxidants, and repair processes

Antioxidative stress

Decreased ROS formation Increased antioxidant defense

Cause

Approach

Early detection of (anti)oxidative

stress and/or damage and inplementation of strategies for balance restoration

↓ Oxidative stress
↑ Free radical scavenging ↑ Damage repair

Balance antioxidant level

Feedback loops between ROS, antioxidants, and repair processes

Consequence

Damage

Proteins Lipids DNA

Oxidative stress

Increased ROS formation Decreased antioxidant defense Ineffective repair systems

Aging

Disease

Damage to cells

Scheme 2: Oxidative and “antioxidative” stress: causes, consequences and methods for its control.

Daily use of synthetic supplements has not been proven as beneficial, and excessive use may be harmful. Balanced food still seems to be the best option.

8. Discussion

A complex mix of substances in fruits and vegetables may contribute to improved cardiovascular health and decreased incidence of cancer in individuals who consume more of these foods [22, 23]. Even in elderly subjects a higher daily intake of fruits and vegetables is associated with an improved antioxidant status compared to subjects consuming diets poor in fruits and vegetables [106]. Contrary, many clinical trials in which individuals received one or more synthetic antioxidants failed to prove their benefits. None of the

major clinical trials using mortality or morbidity as the end point has found positive effects of supplementation with antioxidants such as vitamin C, vitamin E, or -carotene. Some recent studies showed that antioxidant therapy had no effect and even increased the mortality [34–38, 107]. The intake of only one antioxidant could alter the endogenous antioxidative defense of cells, modify cell death rates, or decrease the synthesis of endogenous antioxidants. We have to realize that the use of synthetic vitamin supplements is not the alternative to the regular consumption of fruits and vegetables. Cutler explains that most humans are able to maintain their set points of oxidative stress regardless of additional antioxidant supplementation through diet [49, 50]. In contrast, antioxidant supplements do appear to be effective in lowering an individual’s oxidative stress if his/her initial oxidative stress is above normal or above

8

Oxidative Medicine and Cellular Longevity

his/her set point of regulation [49, 50]. Thus, the antioxidant supplements may help the organism to correct the elevated levels of oxidative stress that cannot be controlled by the endogenous antioxidants. There is also a problem of dosing the synthetic antioxidants; for example, there are claims that RDA (recommended daily allowance) levels of vitamin C and E are too low to prevent the oxidative stress. On the other hand, many consumers ingest high amounts of supplements with the antioxidant potential, which may lead to prooxidant effects or to “antioxidative stress” [32]. Therefore, there is a need to determine the individual’s oxidative stress level before administering the supplement therapy. However, the reference values for typical oxidative stress status of an individual are not established so far and oxidative stress is difficult and expensive to measure [104].

9. Conclusion

In vitro and in vivo studies imply that antioxidant nutrients and related bioactive compounds from fruits and vegetables can protect us from oxidative stress. Synthetic antioxidants as dietary supplements may prevent some ROS-induced dam- age in conditions of elevated oxidative stress during elevated environmental oxidant exposure or at weaken endogenous oxidative stress responses of an aged organism. On the other hand, the presented evidence implies that synthetic antioxidant supplements cannot offer appropriate or total protection against oxidative stress and damage in “normal” situations and that the use of antioxidants to prevent disease or aging is controversial in situations of “normal” oxidative stress.

At the moment, it is difficult to evaluate the oxidative stress of the organism also because different criteria of oxidative stress do not correlate with each other. Since there is no universal “scale” of oxidative stress, the future challenge(s) are in determination of total antioxidants and oxidative stress levels in different body fluids (urine, saliva, blood, and cytosol). Further, detection of the increased levels of oxidative stress biomarker in the body fluid does not mean necessarily that the cells of the specific organ or tissue are under oxidative stress. Besides, it is not possible for highly reactive free radical produced within a tissue with a lifetime of microseconds to diffuse into the blood to be detected at the distant site. The researcher is thus limited to determination of secondary products in human body fluids distant from the locus of the ROS production [108]. With indirect oxidative stress markers, the person may be considered being under oxidative stress according to a given criterion but not to another. Therefore, there is an urgent need to compare and standardize the various methods for assessing the oxidative state of biological systems, to establish the universal scale of oxidative stress, and to provide age and gender specific tables of “normal values” for each body fluid. Until these are established, it is prudent to estimate the oxidative stress by combining different methods and biomarkers.

The key to the future success of dietary antioxidant supplementation may be in the fine tuning of the suppression of oxidative damage without disruption of the well-integrated

antioxidant defense networks. The selective enhancement of the defense system could be a major strategy for a successful intervention by antioxidant administration [109].

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