Praveen K. Vayalil

Date Fruits (Phoenix dactylifera Linn): An Emerging Medicinal Food

Date palm is one of the oldest trees cultivated by man. In the folk-lore, date fruits have been ascribed to have many medicinal properties when consumed either alone or in combination with other herbs. Although, fruit of the date palm served as the staple food for millions of people around the world for several centuries, studies on the health benefits are inadequate and hardly recognized as a healthy food by the health professionals and the public. In recent years, an explosion of interest in the numerous health benefits of dates had led to many in vitro and animal studies as well as the identification and quantification of various classes of phytochemicals. On the basis of available documentation in the literature on the nutritional and phytochemical composition, it is apparent that the date fruits are highly nutritious and may have several potential health benefits. Although dates are sugar-packed, many date varieties are low GI diet and refutes the dogma that dates are similar to candies and regular consumption would develop chronic diseases. More investigations in these areas would validate its beneficial effects, mechanisms of actions, and fully appreciate as a potential medicinal food for humans all around the world. Therefore, in this review we summarize the phytochemical composition, nutritional significance, and potential health benefits of date fruit consumption and discuss its great potential as a medicinal food for a number of diseases inflicting human beings.

Therapeutic value of small molecule inhibitor to plasminogen activator inhibitor-1 for lung fibrosis.

Fibrosis is a final stage of many lung diseases, with no effective treatment. Plasminogen activator inhibitor-1 (PAI-1), a primary inhibitor of tissue-type and urokinase-type plasminogen activators (tPA and uPA, respectively), plays a critical role in the development of fibrosis. In this study, we explored the therapeutic potential of an orally effective small molecule PAI-1 inhibitor, TM5275, in a model of lung fibrosis induced by transforming growth factor-β1 (TGF-β1), the most potent and ubiquitous profibrogenic cytokine, and in human lung fibroblasts (CCL-210 cells). The results show that an intranasal instillation of AdTGF-β1(223/225), an adenovirus expressing constitutively active TGF-β1, increased the expression of PAI-1 and induced fibrosis in murine lung tissue. On the other hand, treating mice with 40 mg/kg of TM5275 for 10 days, starting 4 days after the instillation of AdTGF-β1(223/225), restored the activities of uPA and tPA and almost completely blocked TGF-β1-induced lung fibrosis, as shown by collagen staining, Western blotting, and the measurement of hydroxyproline. No loss of body weight was evident under these treatment conditions with TM5275. Furthermore, we show that TM5275 induced apoptosis in both myofibroblasts (TGF-β1-treated) and naive (TGF-β1-untreated) human lung fibroblasts, and this apoptosis was associated with the activation of caspase-3/7, the induction of p53, and the inhibition of α-smooth muscle actin, fibronectin, and PAI-1 expression. Such an inhibition of fibrotic responses by TM5275 occurred even in cells pretreated with TGF-β1 for 6 hours. Together, the results suggest that TM5275 is a relatively safe and potent antifibrotic agent, with therapeutic potential in fibrotic lung disease.

Transforming growth factor β suppresses glutamate–cysteine ligase gene expression and induces oxidative stress in a lung fibrosis model

The concentration of glutathione (GSH), the most abundant intracellular free thiol and an important antioxidant, is decreased in the lung in both fibrotic diseases and experimental fibrosis models. The underlying mechanisms and biological significance of GSH depletion, however, remain unclear. Transforming growth factor β (TGF-β) is the most potent and ubiquitous profibrogenic cytokine and its expression is increased in almost all fibrotic diseases. In this study, we show that increasing TGF-β1 expression in mouse lung to a level comparable to those found in lung fibrotic diseases by intranasal instillation of AdTGF-β1(223/225), an adenovirus expressing constitutively active TGF-β1, suppressed the expression of both catalytic and modifier subunits of glutamate-cysteine ligase (GCL), the rate-limiting enzyme in de novo GSH synthesis, decreased GSH concentration, and increased protein and lipid peroxidation in mouse lung. Furthermore, we show that increasing TGF-β1 expression activated JNK and induced activating transcription factor 3, a transcriptional repressor involved in the regulation of the catalytic subunit of GCL, in mouse lung. Control virus (AdDL70-3) had no significant effect on any of these parameters, compared to saline-treated control. Concurrent with GSH depletion, TGF-β1 induced lung epithelial apoptosis and robust pulmonary fibrosis. Importantly, lung GSH levels returned to normal, whereas fibrosis persisted at least 21 days after TGF-β1 instillation. Together, the data suggest that increased TGF-β1 expression may contribute to the GSH depletion observed in pulmonary fibrosis diseases and that GSH depletion may be an early event in, rather than a consequence of, fibrosis development.

Rasayanas: Evidence for the Concept of Prevention of Diseases

Rasayanas are non-toxic Ayurvedic complex herbal preparations or individual herbs used to rejuvenate or attain the complete potential of an individual in order to prevent diseases and degenerative changes that leads to disease. The present paper reviews various activities of rasayanas to support the above concept, its role as a prophylactic medication and significance in the prevention of diseases in both healthy as well as diseased individuals. The emerging data suggest that the possible mechanisms may be by immunostimulation, quenching free radicals, enhancing cellular detoxification mechanisms, repair damaged non-proliferating cells, inducing cell proliferation and self-renewal of damaged proliferating tissues, and replenishing them by eliminating damaged or mutated cells with fresh cells.

Protective Effects of Rasayanas on Cyclophosphamide- and Radiation-Induced Damage

Rasayanas are a group of herbal drug preparations widely used in Ayurveda to improve the general health of the body. In mice, Rasayanas are potent myeloprotective agents against chemotherapeutic agents and radiation. Rasayanas are also effective myeloprotectors in patients with cancer undergoing chemotherapy and/or radiotherapy. In this study, we provide further evidence to support the chemoprotective and radioprotective efficacy of four Rasayanas in mice. Rasayanas were found to reduce the loss of body weight and organ weight induced by cyclophosphamide and radiation significantly. Rasayanas were also found to protect tissue from cytotoxic injury associated with reduced serum and liver lipid peroxides, alkaline phosphatase, and glutamate pyruvate transaminase in cyclophosphamide- and radiation-treated animals. These results suggest the potential chemoprotective and radioprotective effects of Rasayanas, which require further study to explore their complete usefulness in cancer therapy.

Detection of electrophile-sensitive proteins

BACKGROUND Redox signaling is an important emerging mechanism of cellular function. Dysfunctional redox signaling is increasingly implicated in numerous pathologies, including atherosclerosis, diabetes, and cancer. The molecular messengers in this type of signaling are reactive species which can mediate the post-translational modification of specific groups of proteins, thereby effecting functional changes in the modified proteins. Electrophilic compounds comprise one class of reactive species which can participate in redox signaling. Electrophiles modulate cell function via formation of covalent adducts with proteins, particularly cysteine residues. SCOPE OF REVIEW This review will discuss the commonly used methods of detection for electrophile-sensitive proteins, and will highlight the importance of identifying these proteins for studying redox signaling and developing novel therapeutics. MAJOR CONCLUSIONS There are several methods which can be used to detect electrophile-sensitive proteins. These include the use of tagged model electrophiles, as well as derivatization of endogenous electrophile-protein adducts. GENERAL SIGNIFICANCE In order to understand the mechanisms by which electrophiles mediate redox signaling, it is necessary to identify electrophile-sensitive proteins and quantitatively assess adduct formation. Strengths and limitations of these methods will be discussed. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn.

Mitochondrial Bioenergetics of Metastatic Breast Cancer Cells in Response to Dynamic Changes in Oxygen Tension: Effects of HIF-1α

Solid tumors are characterized by regions of low oxygen tension (OT), which play a central role in tumor progression and resistance to therapy. Low OT affects mitochondrial function and for the cells to survive, mitochondria must functionally adapt to low OT to maintain the cellular bioenergetics. In this study, a novel experimental approach was developed to examine the real-time bioenergetic changes in breast cancer cells (BCCs) during adaptation to OT (from 20% to <1% oxygen) using sensitive extracellular flux technology. Oxygen was gradually removed from the medium, and the bioenergetics of metastatic BCCs (MDA-MB-231 and MCF10CA clones) was compared with non-tumorigenic (MCF10A) cells. BCCs, but not MCF10A, rapidly responded to low OT by stabilizing HIF-1α and increasing HIF-1α responsive gene expression and glucose uptake. BCCs also increased extracellular acidification rate (ECAR), which was markedly lower in MCF10A. Interestingly, BCCs exhibited a biphasic response in basal respiration as the OT was reduced from 20% to <1%. The initial stimulation of oxygen consumption is found to be due to increased mitochondrial respiration. This effect was HIF-1α-dependent, as silencing HIF-1α abolished the biphasic response. During hypoxia and reoxygenation, BCCs also maintained oxygen consumption rates at specific OT; however, HIF-1α silenced BCC were less responsive to changes in OT. Our results suggest that HIF-1α provides a high degree of bioenergetic flexibility under different OT which may confer an adaptive advantage for BCC survival in the tumor microenvironment and during invasion and metastasis. This study thus provides direct evidence for the cross-talk between HIF-1α and mitochondria during adaptation to low OT by BCCs and may be useful in identifying novel therapeutic agents that target the bioenergetics of BCCs in response to low OT.

Mitochondrial thiol modification by a targeted electrophile inhibits metabolism in breast adenocarcinoma cells by inhibiting enzyme activity and protein levels.

Many cancer cells follow an aberrant metabolic program to maintain energy for rapid cell proliferation. Metabolic reprogramming often involves the upregulation of glutaminolysis to generate reducing equivalents for the electron transport chain and amino acids for protein synthesis. Critical enzymes involved in metabolism possess a reactive thiolate group, which can be modified by certain oxidants. In the current study, we show that modification of mitochondrial protein thiols by a model compound, iodobutyl triphenylphosphonium (IBTP), decreased mitochondrial metabolism and ATP in MDA-MB 231 (MB231) breast adenocarcinoma cells up to 6 days after an initial 24 h treatment. Mitochondrial thiol modification also depressed oxygen consumption rates (OCR) in a dose-dependent manner to a greater extent than a non-thiol modifying analog, suggesting that thiol reactivity is an important factor in the inhibition of cancer cell metabolism. In non-tumorigenic MCF-10A cells, IBTP also decreased OCR; however the extracellular acidification rate was significantly increased at all but the highest concentration (10 µM) of IBTP indicating that thiol modification can have significantly different effects on bioenergetics in tumorigenic versus non-tumorigenic cells. ATP and other adenonucleotide levels were also decreased by thiol modification up to 6 days post-treatment, indicating a decreased overall energetic state in MB231 cells. Cellular proliferation of MB231 cells was also inhibited up to 6 days post-treatment with little change to cell viability. Targeted metabolomic analyses revealed that thiol modification caused depletion of both Krebs cycle and glutaminolysis intermediates. Further experiments revealed that the activity of the Krebs cycle enzyme, aconitase, was attenuated in response to thiol modification. Additionally, the inhibition of glutaminolysis corresponded to decreased glutaminase C (GAC) protein levels, although other protein levels were unaffected. This study demonstrates for the first time that mitochondrial thiol modification inhibits metabolism via inhibition of both aconitase and GAC in a breast cancer cell model.

A novel class of mitochondria-targeted soft electrophiles modifies mitochondrial proteins and inhibits mitochondrial metabolism in breast cancer cells through redox mechanisms.

Despite advances in screening and treatment over the past several years, breast cancer remains a leading cause of cancer-related death among women in the United States. A major goal in breast cancer treatment is to develop safe and clinically useful therapeutic agents that will prevent the recurrence of breast cancers after front-line therapeutics have failed. Ideally, these agents would have relatively low toxicity against normal cells, and will specifically inhibit the growth and proliferation of cancer cells. Our group and others have previously demonstrated that breast cancer cells exhibit increased mitochondrial oxygen consumption compared with non-tumorigenic breast epithelial cells. This suggests that it may be possible to deliver redox active compounds to the mitochondria to selectively inhibit cancer cell metabolism. To demonstrate proof-of-principle, a series of mitochondria-targeted soft electrophiles (MTSEs) has been designed which selectively accumulate within the mitochondria of highly energetic breast cancer cells and modify mitochondrial proteins. A prototype MTSE, IBTP, significantly inhibits mitochondrial oxidative phosphorylation, resulting in decreased breast cancer cell proliferation, cell attachment, and migration in vitro. These results suggest MTSEs may represent a novel class of anti-cancer agents that prevent cancer cell growth by modification of specific mitochondrial proteins.

Mitochondrial oncobioenergetic index: A potential biomarker to predict progression from indolent to aggressive prostate cancer

Mitochondrial function is influenced by alterations in oncogenes and tumor suppressor genes and changes in the microenvironment occurring during tumorigenesis. Therefore, we hypothesized that mitochondrial function will be stably and dynamically altered at each stage of the prostate tumor development. We tested this hypothesis in RWPE-1 cells and its tumorigenic clones with progressive malignant characteristics (RWPE-1 < WPE-NA22 < WPE-NB14 < WPE-NB11 < WPE-NB26) using high-throughput respirometry. Our studies demonstrate that mitochondrial content do not change with increasing malignancy. In premalignant cells (WPE-NA22 and WPE-NB14), OXPHOS is elevated in presence of glucose or glutamine alone or in combination compared to RWPE-1 cells and decreases with increasing malignancy. Glutamine maintained higher OXPHOS than glucose and suggests that it may be an important substrate for the growth and proliferation of prostate epithelial cells. Glycolysis significantly increases with malignancy and follow a classical Warburg phenomenon. Fatty acid oxidation (FAO) is significantly lower in tumorigenic clones and invasive WPE-NB26 does not utilize FAO at all. In this paper, we introduce for the first time the mitochondrial oncobioenergetic index (MOBI), a mathematical representation of oncobioenergetic profile of a cancer cell, which increases significantly upon transformation into localized premalignant form and rapidly falls below the normal as they become aggressive in prostate tumorigenesis. We have validated this in five prostate cancer cell lines and MOBI appears to be not related to androgen dependence or mitochondrial content, but rather dependent on the stage of the cancer. Altogether, we propose that MOBI could be a potential biomarker to distinguish aggressive cancer from that of indolent disease.