Dino Demirovic

Establishing cellular stress response profiles as biomarkers of homeodynamics, health and hormesis

Aging is the progressive shrinkage of the homeodynamic space. A crucial component of the homeodynamic space is the stress response (SR), by virtue of which a living system senses disturbance and initiates a series of events for maintenance, repair, adaptation, remodeling and survival. Here we discuss the main intracellular SR pathways in human cells, and argue for the need to define and establish the immediate and delayed stress response profiles (SRP) during aging. Such SRP are required to be established at several age-points, which can be the molecular biomarkers of homeodynamic space and the health status of cells and organisms. SRP can also be useful for testing potential protectors and stimulators of homeodynamics, and can be a standard for monitoring the efficacy of potential pro-survival, health-promoting and aging-modulating conditions, food components and other compounds. An effective strategy, which makes use of SRP for achieving healthy aging and extending the healthspan, is that of strengthening the homeodynamics through repeated mild stress-induced hormesis by physical, biological and nutritional hormetins. Furthermore, SRP can also be the basis for defining health as a state of having adequate physical and mental independence of activities of daily living, by identifying a set of measurable parameters at the most fundamental level of biological organization.

Heat Stress and Hormetin-Induced Hormesis in Human Cells: Effects on Aging, Wound Healing, Angiogenesis, and Differentiation:

Accumulation of molecular damage and increased molecular heterogeneity are hallmarks of cellular aging. Mild stress-induced hormesis can be an effective way for reducing the accumulation of molecular damage, and thus slowing down aging from within. We have shown that repeated mild heat stress (RMHS) has anti-aging effects on growth and various other cellular and biochemical characteristics of normal human skin fibroblasts and keratinocytes undergoing aging in vitro. RMHS given to human cells increased the basal levels of various chaperones, reduced the accumulation of damaged proteins, stimulated proteasomal activities, increased the cellular resistance to other stresses, enhanced the levels of various antioxidant enzymes, enhanced the activity and amounts of sodium-potassium pump, and increased the phosphorylation-mediated activities of various stress kinases. We have now observed novel hormetic effects of mild heat stress on improving the wound healing capacity of skin fibroblasts and on enhancing the angiogenic ability of endothelial cells. We have also tested potential hormetins, such as curcumin and rosmarinic acid in bringing about their beneficial effects in human cells by inducing stress response pathways involving heat shock proteins and heme-oxygenase HO-1. These data further support the view that mild stress-induced hormesis can be applied for the modulation, intervention and prevention of aging and age-related impairments.

Hormesis can and Does Work in Humans

If we accept the validity of the general concept of physiological hormesis as being the phenomenon of achieving health beneficial effects by exposure to mild stress, then hormesis is being applied already and successfully to humans. The evidence for this is the well-demonstrated health benefits of regular and moderate exercise. Mild stress-induced activation of one or more intracellular pathways of stress response are central to this. Experimental studies performed on human cells in culture exposed to mild heat shock and other stresses provide biochemical and molecular evidence in support of the application of hormesis to human systems. Although several issues remain to be resolved by more research with respect to the extent and duration of hormetic exposure, making use of the cellular stress response pathways can facilitate discovering novel hormetins for human applications.

Hormesis and Aging

Mild stress-induced hormetic stimulation of protective mechanisms in cells and organisms can result in potential antiaging effects. Detailed molecular mechanisms that bring about the hormetic effects are being increasingly understood and comprise a cascade of stress response and other pathways of maintenance and repair. Although the extent of immediate hormetic effects after exposure to a particular stress may only be moderate, the chain of events following initial hormesis leads to biologically amplified effects that are much larger, synergistic, and pleiotropic. A consequence of hormetic amplification is an increase in the homeodynamic space of a living system in terms of increased defense capacity and reduced load of damaged macromolecules. Hormetic strengthening of the homeodynamic space provides wider margins for metabolic fluctuation, stress tolerance, adaptation, and survival. Hormesis thus counterbalances the progressive shrinkage of the homeodynamic space that is the ultimate cause of aging, diseases, and death. Healthy aging may be achieved by hormesis through mild and periodic but not severe or chronic physical and mental challenges and by the use of nutritional hormesis incorporating mild stress-inducing molecules called hormetins.

Hormesis as a Mechanism for the Anti-Aging Effects of Calorie Restriction

Hormesis in aging is represented by stress-induced stimulation of protective mechanisms and strengthened homeostasis/homeodynamics. Since calorie restriction (CR) generally stimulates various maintenance and repair systems, hormesis has been considered as one of the possible mechanisms. The evidence in favour of the hormetic explanation for the beneficial effects of CR includes the induction of various stress responses including heat shock response, unfolded protein response, autophagy, DNA repair response, sirtuin response, and antioxidant responses. In addition, chronically increased levels of corticosterones and an accompanying decrease in body weight in CR animals is indicative of CR being a stressor and a hormetic agent. Although the exact nature of the low level damage-inducing aspect of CR is yet to be elucidated, a metabolic shift from glycolytic pathways to mitochondrial pathways, which results in enhanced oxidative stress and increased damage to macromolecules and organelles, appears to be the initiator of stress-induced hormetic effects of CR. Periodic modulation of dietary input as a hormetic intervention may be applicable to human beings for slowing down aging.

Basal Level of Autophagy Is Increased in Aging Human Skin Fibroblasts In Vitro, but Not in Old Skin

Intracellular autophagy (AP) is a stress response that is enhanced under conditions of limitation of amino acids, growth factors and other nutrients, and also when macromolecules become damaged, aggregated and fibrillated. Aging is generally accompanied by an increase in intracellular stress due to all the above factors. Therefore, we have compared the basal levels of AP in serially passaged human facial skin fibroblasts undergoing aging and replicative senescence in vitro, and ex vivo in the skin biopsies from the photo-protected and photo-exposed area of the arms of 20 healthy persons of young and old ages. Immunofluorescence microscopy, employing antibodies against a specific intracellular microtubule-associated protein-1 light chain-3 (LC3) as a well established marker of AP, showed a 5-fold increase in the basal level of LC3 in near senescent human skin fibroblasts. However, no such age-related increase in LC3 fluorescence and AP could be detected in full thickness skin sections from the biopsies obtained from 10 healthy young (age 25 to 30 yr) and 10 old (age 60 to 65 yr) donors. Furthermore, there was no difference in the basal level of LC3 in the skin sections from photo-protected and photo-exposed areas of the arm. Thus, in normal conditions, the aging phenotype of the skin cells in culture and in the body appears to be different in the case of AP.

A preliminary attempt to establish multiple stress response profiles of human skin fibroblasts exposed to mild or severe stress during ageing in vitro

Optimal stress response (SR) is an essential aspect of the property of dynamic homeostasis of all biological systems, including cells in culture. Whereas severe stress can induce the so-called stress-induced premature senescence (SIPS), a model developed by Olivier Toussaint, mild stress can strengthen homeodynamics and can postpone senescence through the phenomenon of hormesis. We have attempted to establish multiple stress response profiles (SRP) of early passage young and late passage senescent human facial skin fibroblasts, FSF-1, exposed to either mild (41°C) and severe (43°C) heat shock for 1h, or to mild (2%) and severe (0%) serum deprivation for up to 48h. The results obtained show that FSF-1 cells exposed to two different intensities of stress from two different stressors separately have differential SRP to mild and severe stress, which also vary significantly between young and senescent cells. Establishing multiple and differential SRP to mild and severe stress may facilitate distinguishing between the mild stress-induced beneficial hormetic effects and the harmful effects of severe stress.