Original articleLonger period of oral administration of aspartame on cytokine response in Wistar albino ratsEfectos de la administración oral crónica de aspartamo sobre la respuesta de citocinas en ratas albinas Wistar
Introduction
Nowadays consumers are increasingly concerned about the quality and safety of many products of industrialized countries, in particular the use of artificial sweeteners, flavorings, colorings, preservatives and dietary supplements. General apprehension also exists regarding the possible long-term health effects of the raw materials and technologies used for the packaging, sterilization and distribution of foods. Many non-nutritive sweeteners have been used in foods and beverages to allow people to enjoy the sweet taste without consuming sugar-associated calories. One of these sweeteners is aspartame. This sweetener is incorporated into a number of foodstuffs (drinks, desserts, sweets, etc.) and in table sweeteners, under different brand names and into some 600 medicines.1 Its sweetening power is 180–200 times greater than that of sucrose.2 Because it contains no calories, aspartame is considered a boon to health-conscious individuals everywhere; a recent observation indicated that aspartame is slowly making its way into ordinary products used every day, which do not carry any indication as being for people on diets or diabetic patients. Aspartame in dry products is fairly stable even at high temperatures. However, in solution, its stability is a function of time, temperature, pH and available moisture. Aspartame is most stable between pH values of 3 and 5, even with increasing temperature.3 However, it breaks down and loses its sweetness in normal cooking or baking. Thus its use is limited to a table-top sweetener (Equal®TM – The NutraSweet Co., Deerfield, IL) and as NutraSweet in dry foods, soft drinks, and frozen foods like ice cream. It is slightly soluble in water (about 1.0% at 25 °C), sparingly soluble in alcohol and insoluble in fats and oils. Being a peptide, it is amphoteric and is metabolized extensively to release its constituent amino acids and methanol.3 Upon ingestion, this artificial sweetener is immediately absorbed from the lumen and metabolized by gut esterases and peptidases to phenylalanine, aspartic acid and methanol. Orally ingested aspartame components are immediately absorbed from the lumen and reach the portal blood in a manner similar to that of amino acids arising from dietary protein or polysaccharides.4, 5 Their concentrations are found increased in the blood stream.6 This sweetener and its metabolic breakdown products (phenylalanine, aspartic acid and methanol) have been a matter of extensive investigation for more than 20 years, including experimental animal studies. Ten per cent of aspartame consists of methanol. Methanol is a toxicant that causes systemic toxicity.7 The primary metabolic fate of methanol is the direct oxidation to formaldehyde and then into formate. The severity of clinical findings in methanol intoxication correlated better with formate levels.8 Methanol is gradually released in the small intestine when the methyl group of aspartame encounters the enzyme chymotrypsin,4 but methanol is more readily generated by the body (thus becoming even more dangerous) when it is heated above 30 °C before being ingested. This occurs when soft drinks are left out in the sun or foods containing aspartame are heated. Methanol breaks down into formaldehyde and formic acid in the body. Formaldehyde is an embalming fluid, as a preservative in vaccines and a deadly neurotoxin. Formic acid causes cells to become too acidic, thereby producing metabolic acidosis. Acidosis damages cellular health by causing enzymes to stop functioning. Oxidative stress arises from the imbalance between pro-oxidants and antioxidants in favor of the former, leading to the generation of oxidative damage.9 Generation of free radicals is an integral feature of normal cellular functions, in contrast, excessive generation and/or inadequate removal of free radical results in destructive and irreversible damage to the cell.10 A stressor is a stimulus that is either internal or external, which activates the hypothalamic pituitary adrenal axis and the sympathetic nervous system resulting in a physiological change11 Corticotropin-releasing hormone is released during stress and stimulates the release of adrenocorticotropic hormone,12 which in turn releases corticosterone from the adrenal cortex. Elevation in the corticosterone level accelerates the generation of free radicals13 and alters the normal homeostasis of organ.14
Cytokines belong to the family of signaling molecules. They are released by specific cells of the immune system. Cytokines are small glycoprotein chemical structures that act in a paracrine and endocrine fashion as soluble signals between cells and play a pivotal role in the immune response. They are the hormonal messengers responsible for most of the biological effects in the immune system, such as cell-mediated immunity and humoral immunity. T lymphocytes are a major source of cytokines. Immune cells produce mediators of inflammatory and immune reactions called cytokines. These low-molecular weight glycoproteins in small concentrations are indispensable for normal functioning of the immune system. Their excessive secretion, however, leads to immune cell dysfunctions. Inflammatory cytokines may be produced by mononuclear cells of the immune system in response to numerous agents, such as microorganisms and their products (e.g., lipopolysaccharide – LPS)15, 16 as well as some xenobiotics.17 It is postulated that xenobiotics can influence the concentrations of inflammatory cytokines through oxidative stress mechanisms.18, 19, 20 The experimental and epidemiological data currently available to evaluate the above toxigenic risks of aspartame are insufficient and often unreliable, due to the inadequate planning and conduct of previous experiments. Recently from previous studies on aspartame and its metabolite, altering the oxidative status of the cells was investigated. Oral aspartame (75 mg/kg body weight/day) consumption causes oxidative stress in brain21 and its (40 mg/kg bw/day) consumption caused oxidative stress in brain,22 liver and kidney,23 and also in immune organs.24 This inadequacy, combined with the general limited knowledge about the safety/potential toxigenic effects of substances widely present in the industrialized diet, motivated the design of the current study. However, little is known about the effects of aspartame on cytokine expression. The detailed mechanisms of the effects of aspartame on cytokines are still unclear; therefore, the present study aimed at clarifying whether longer time of aspartame consumption has any effect on cytokine expression in serum of Wistar albino rats.
Section snippets
Chemicals
Pure aspartame powder and methotrexate were purchased from Sigma Aldrich chemical, (St. Louis, USA) and all other chemicals used were of analytical grade obtained from Sisco research laboratory (Mumbai, India). ELISA kits for cytokine estimations were obtained from Ray biotech system (USA).
Animal model
Animal experiments were carried out after approval from the Institutional Animal Ethical Committee (IAEC No: 01/21/14) and the Committee for the Purpose of Control and Supervision of Experiments on Animals
Statistical analysis
Data are expressed as mean ± standard deviation (SD). All data were analyzed with the SPSS for windows statistical package (version 20.0, SPSS Institute Inc., Cary, North Carolina, USA). Statistical significance between the different groups was determined by one-way analysis of variance (ANOVA). When the groups showed significant difference, then Tukey's multiple comparison tests were followed and the significance level was fixed at p < 0.05.
Effect of aspartame on plasma corticosterone level
The results are summarized in Table 1. The control, as well as the folate-deficient rats, did not show any significant variation among themselves in the plasma corticosteroid levels. However, both control animals as well as folate-deficient animals when treated with aspartame for 90 days, whether they were un-immunized or immunized, showed a marked increase in corticosteroid levels when compared to control as well as the folate-deficient animals. Among the aspartame-treated animals, the
Discussion
A homeostatic balance exists between the formation of free oxygen radicals and their removal by endogenous scavenging antioxidants.44 In this study, the folate-deficient diet-fed animals were used to mimic the human methanol metabolism. However, the folate-deficient diet-fed animals did not show any significant changes in the parameters studied and remained similar to controls animals. The increase in corticosterone level indicates that aspartame may act as a chemical stressor. Changes in
Acknowledgments
The authors gratefully acknowledge the University of Madras for their financial support [UGC No. D.1.(C)/TE/2012/1868]. The authors acknowledge Mr. Sunderaswaran Loganathan for his constant support and help.
References (71)
- et al.
In vitro effect of aspartame in angiogenesis induction
Toxicol In Vitro
(2011) The aspartame story: a model for the clinical testing of a food additive
Am J Clin Nutr
(1987)- et al.
Formaldehyde derived from dietary aspartame binds to tissue components in vivo
Life Sci
(1998) - et al.
Repeated ingestion of aspartame—sweetened beverage effect on plasma amino acid concentrations in normal adults
Metabolism
(1988) - et al.
Glucocorticoids increase the accumulation of reactive oxygen species and enhance adriamycin-induced toxicity in neuronal culture
Exp Neurol
(1996) - et al.
Evidence that shockinduced immune suppression is mediated by adrenal hormones and peripheral-adrenergic receptors
Pharmacol Biochem Behav
(1990) - et al.
Toxic metals stimulate inflammatory cytokines in hepatocytes through oxidative stress mechanisms
Toxicol Appl Pharmacol
(1998) - et al.
A critical involvement of oxidative stress in acute alcohol-induced hepatic TNF-α production
Am J Pathol
(2003) The toxicity of methanol
Life Sci
(1991)- et al.
Review of the genotoxicity of formaldehyde
Mutat Res
(1988)
Role of catalase and hydroxyl radicals in the oxidation of methanol by rat liver microsomes
Biochem Pharmacol
Protein measurement with the Folin phenol reagent
J Biol Chem
Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction
Anal Biochem
Analysis of nitrate, nitrite, and nitrate in biological fluids
Anal Biochem
Colorimetric assay of catalase
Anal Biochem
Muscle damage is linked to cytokine changes following a 160-km race
Brain Behav Immun
The placebo effect and relaxation response: neural processes and their coupling to constitutive nitric oxide.
Brain Res Rev
A role for the extracellular signal-regulated kinase signal pathway in depressive-like behavior
Behav Brain Res
Enzymatic repair of oxidative damage to human apolipoprotein A-I
FEBS Lett
Aspartame and its effects on health
BMJ
Methanol poisoning
Intensive Care Med
Serum formate concentrations in methanol intoxication as a criterion for hemodialysis
Ann Intern Med
Free radicals in biology and medicine
Antioxidants, programmed cell death, and cancer
Nutr Res
Cytokines for psychologists: implications for bidirectional immune-to-brain communication for understanding behavior, mood, and cognition
Psychol Rev
Physiology and pharmacology of corticotropin-releasing factor
Pharmacol Rev
Effect of endotoxins isolated from Desulfovibrio desulfuricans soil and intestinal strain on the secretion of TNF-α by human mononuclear cells
Pol J Environ Stud
Effect of inositol hexsaphosphate on lipopolysaccharide-stimulated release of TNF-α from human mononuclear cells
Pol J Environ Stud
Xenobiotics and inflammation
The role of Kupffer cell oxidant production in early ethanol induced liver disease
Free Radic Biol Med
Effect of chronic exposure to aspartame on oxidative stress in the brain of albino rats
J Bio Sci
Aspartame (a widely used artificial sweetener) and oxidative stress in cerebral cortex
Int J Pharm Biomed Sci
Effect of aspartame on some oxidative stress parameter in liver and kidney of rats
Afr J Pharm Pharmacol
Imbalance of oxidant–antioxidant status by aspartame in the organs of immune system of Wistar albino rats
Afr J Pharm Pharmacol
Cited by (20)
Updated systematic assessment of human, animal and mechanistic evidence demonstrates lack of human carcinogenicity with consumption of aspartame
2023, Food and Chemical ToxicologyCitation Excerpt :Down regulation of genes related to positive acute phase response and inflammation in mice exposed to aspartame in utero was also observed (Collison et al., 2018). In rats, Choudhary and Sheela Devi (2015) reported mixed findings following a 90-day exposure of 40 mg/kg-day (single dose; no positive control); IL-2 and IFNg were decreased, and IL-4 was increased. Schiano et al. (2019) reported no effects of aspartame administered for 8 weeks on the modulation of carrageenan-induced mouse paw edema.
The debate over neurotransmitter interaction in aspartame usage
2018, Journal of Clinical NeuroscienceCitation Excerpt :It is possible that secreted neurotransmitters of microorganisms inside the intestinal lumen may additionally induce epithelial cells to release molecules that in turn modulate neuronal signaling within the ENS, or act directly on primary afferent axons [86]. Hence, as mentioned above, the altered gut microbes [53,54] as well as altered cytokine with immune dysfunction [51,87] after aspartame ingestion may have potential to modulate neuronal signaling and result in neurobiological impairments. At present, we all know that aspartame metabolite specially Phy and its interaction with neurotransmitter and aspartic acid by acting as excitatory neurotransmitter causes this pattern of impairments.
The impact of non-caloric artificial sweetener aspartame on female reproductive system in mice model
2023, Reproductive Biology and EndocrinologyThe Effect of Aspartame and Stevia on the Histological Structure and the Related Biological Markers in the Alveolar Bone of Albino Rats
2023, Egyptian Journal of Histology