Effects of Rhodiola rosea and honeybush extracts on stress-induced defects in bioenergetics: from mitochondria to the synapse

Mitochondria are essential organelles that are responsible mainly for the energy production in the form of adenosine triphosphate (ATP) through oxidative phosphorylation (OXPHOS) but also for various other cellular functions such as calcium and redox homeostasis, synaptic plasticity, neurotransmission and cell death. Cellular and organismal health is highly dependent on mitochondria because of the diverse role of the latter. The role of mitochondria is particularly prominent in the brain since neurons conduct a number of complex processes which require a great amount of energy. Mitochondria “fund” neuronal functions by providing ATP and thus neurons highly depend on these organelles. Mitochondria play pivotal roles not only in health but also in the pathogenesis of several diseases. Mitochondrial dysfunction, oxidative stress, impaired energy metabolism and synaptic dysregulation have been identified as common features in many neurodegenerative diseases but also in stress-related brain disorders. For example, tauopathies such as Alzheimer’s disease (AD) and chronic traumatic encephalopathy are characterized by decreased ATP levels, increased reactive oxygen species (ROS) levels and impaired synaptic transmission leading to neuronal death in the brain. Lately, there is an increasing interest on mitochondria-targeting pharmacological strategies with a particular focus on substances that increase bioenergetics (respiration and ATP levels) and decrease ROS.
Current pharmacological strategies against neurodegenerative and stress-related mental disorders are inadequate in stopping the progression of these diseases. They mainly treat symptoms but often present severe adverse effects. Nature has always been a source of therapeutic plants and phytochemicals which were used by local healers against a variety of diseases and discomforts. Of note, a few of the current AD medications are naturally-derived phytochemicals (e.g. galantamine and rivastigmine). Therefore, plant extracts and phytochemicals derived from plants could be used as medications or supplementations in order to alleviate symptoms as they do not present severe adverse effects while they exert multi-targeted functions. In order to develop novel plant-based strategies against complex diseases, such as neurodegenerative, we need to study and understand the molecular mode of action of phytochemicals.
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Therefore, the aim of this PhD thesis was to identify phytochemicals that enhance mitochondrial functions and test their ability to counteract deficits caused by different types of stress. In this matter, we evaluated in vitro the effects of two extracts: Honeybush extract (Cyclopia species) and Rhodiola rosea extract as well as the Honeybush extract compound mangiferin against oxidative stress, glucocorticoid- or abnormal tau-induced stress respectively.
Honeybush (Cyclopia species) is a South African endemic plant and certain species have a long tradition of use as herbal tea, known as honeybush tea. Increased global demand for honeybush tea led to a scientific interest for the bioactivity of its extracts due to their phenolic composition. Indeed, honeybush extracts have been indicated to possess antioxidant activities. However, their neuroprotective properties have never been tested before. Due to their antioxidant activities we hypothesized that honeybush extracts would exert antioxidant properties as well as beneficial effects on mitochondria in neuronal cells. Therefore, we tested the effects of four honeybush extracts on human neuroblastoma SH-SY5Y cells. The effects of the aqueous extracts of C. subternata, C. genistoides, C. longifolia and a 70% ethanolic extract of C. genistoides in intrinsic health status were firstly evaluated. We showed that treatment with honeybush extracts had a beneficial effect on bioenergetics as it increased ATP production, respiration and mitochondrial membrane potential (MMP). Next, their ability to counteract oxidative stress-induced deficits was tested. The aqueous extracts of C. subternata and C. genistoides, in particular, showed a protective effect by rescuing the bioenergetic and mitochondrial deficits under H2O2-induced oxidative stress conditions. In detail, the extracts partially rescued the ATP levels and respiration and they completely rescued MMP levels under oxidative stress as well as reduced the H2O2-induced ROS. The most promising effects were exerted by the aqueous extract of C. subternata followed by the aqueous extract of C. genistoides which is relevant as these two extracts are largely used for the production of honeybush tea. Our data indicate the potential neuroprotective effect of Honeybush extracts against oxidative stress while we demonstrated for the first time their mitochondria-enhancing properties. Although more research is required, these findings set the basis for the development of a condition-specific nutraceutical.
Rhodiola rosea is a perennial plant growing in mountainous areas of Central Asia and Europe. Extracts of Rhodiola rosea have been characterized as adaptogens as they help organisms to adapt to new or stressful situations and have been used by Vikings and the Russian army to enhance performance during battles. Rhodiola has been shown to alleviate symptoms of stress-related mental disorders such as chronic stress and burnout in clinical studies. These conditions lead to the dysregulation of the stress system, namely the hypothalamic-pituitary-adrenal (HPA) axis, resulting in the overproduction of glucocorticoids such as cortisol in humans as end-products. Overaccumulation of glucocorticoids causes detrimental cellular effects by compromising mitochondrial functions and overall cellular health. Due to its adaptogenic properties, we hypothesized that Rhodiola rosea extract (RRE) would be able to protect neuronal cells against glucocorticoid stress induced by the synthetic glucocorticoid dexamethasone. Therefore, we evaluated its effects against dexamethasone-induced deficits both in neuroblastoma SH-SY5Y cells and in murine hippocampal HT22 cells as well as the ability of RRE itself to enhance mitochondrial functions under non-stressful conditions. We showed that RRE protected both cell lines from dexamethasone-induced cell death and normalized the increase in ROS levels in an inverted U-shape and U-shape manner respectively. Also, we showed that RRE increased ATP production and metabolic activity in intrinsic health status in both cell lines and again in an inverted U-shape manner. These biphasic response curves reflect the activation of the «adaptive cellular stress response» and thus we confirmed that RRE has the typical features of an adaptogen with regard to the modulation of stress response and adaptive homeostasis. In addition, we showed that RRE induced neurite outgrowth in neuroblastoma SH-SY5Y as it increased the number and length of neurites as well as their endpoints and attachment points compared to the untreated control condition with a concomitant upregulation of the brain-derived neurotrophic factor (BDNF) mRNA and protein levels. Therefore, RRE promoted neurite outgrowth possibly via the BDNF pathway. Taken together, our results support the use of RRE as an adaptogen in stress-related mental disorders while we showed that the BDNF pathway might mediate RRE`s beneficial activities.
Mangiferin is a xanthonoid polyphenol present in the fruit, leaves and bark of Mangifera indica, but also in Honeubush extracts, and has been shown to exhibit antioxidant and neuroprotective properties including amyloid beta (Aβ)-related AD models. However, studies showing its benefits in tau-related AD models are missing. Tauopathies are a group of neurodegenerative diseases (e.g. AD) characterized by an abnormal accumulation of the protein tau within neurons, excessive tau hyperphosphorylation and formation into neurofibrillary tangles which are neurotoxic. As many other cases of neurodegenerative diseases, tauopathies are characterized by mitochondrial dysfunction, disturbed bioenergetics and excessive oxidative stress. We hypothesized that mangiferin could have beneficial effect on a cellular tauopathy model by boosting mitochondrial functions and by promoting neurite outgrowth in neuroblastoma cells stably transfected with the mutant tau P301L (P301L cells). Indeed, we demonstrated that mangiferin not only increased ATP levels and MMP in the P301L cells but also promoted neurite outgrowth and brought the P301L cells in a morphological state of resembling the healthy mock neuroblastoma cells (cells transfected with the empty vector). These findings suggest that mangiferin might be a promising candidate against tauopathies but of course more studies are required to establish such use.
Collectively, our data show that indeed plant extracts and phytochemicals can exert beneficial properties against various types of stress by enhancing mitochondrial functions, by scavenging ROS and by promoting neurite outgrowth. These benefits lead to the development of more complex neural networks, enhanced synaptic transmission and strengthened synaptic plasticity. Therefore, they exert benefits at the mitochondrial, cellular and at the synapse level. Overall, the studies performed in this PhD thesis helped to comprehend the molecular mode of action of phytochemicals both in healthy conditions and against different types of stress. We showed the promising effects of Honeybush extracts (Cyclopia species) against oxidative stress, of Rhodiola rosea extract against glucocorticoid stress and of the compound mangiferin against abnormal tau-induced cellular stress. All of those benefits were achieved through the improvement of mitochondrial functions. Our findings could contribute to the development of pharmacological strategies against specific conditions.