Mitochondrial oncobioenergetics of prostate tumorigenesis.

Mitochondria are emerging as key players in the tumorigenic process of cells by maintaining the biosynthetic and energetic capabilities of cancer cells. It is now evident that mitochondria are involved in the malignant transformation, cell proliferation, aggression and metastatic behavior of prostate cancer (PC). Recent comprehensive analysis of the mitochondrial oncobioenergetic (MOB) profile of PC cells using microplate-based high resolution respirometry has clearly demonstrated that characteristic MOB alterations occur at different stages of PC development. Additionally, studies have reported that modification of the MOB profile significantly inhibits the growth of malignant cells. This observation suggests that dynamic alterations in the MOB function are a critical component in the development of malignant disease of the prostate. Therefore, quantification of MOB function may be a good marker for the prediction of tumor stage. Future studies may develop novel approaches to measure oncobioenergetics of tumors with minimal invasive procedures effectively in men to determine the general prostate health and tumor staging, and to predict whether a tumor will proceed to malignancy in patients. Additionally, since PC is a slow growing tumor, modulating the MOB profile at specific stages of tumor development may be a novel approach to treat or prevent PC.

Deficiency of tumor suppressor Merlin facilitates metabolic adaptation by co-operative engagement of SMAD-Hippo signaling in breast cancer.

The NF2 gene encodes the tumor and metastasis suppressor protein Merlin. Merlin exerts its tumor suppressive role by inhibiting proliferation and inducing contact-growth inhibition and apoptosis. In the current investigation, we determined that loss of Merlin in breast cancer tissues is concordant with the loss of the inhibitory SMAD, SMAD7, of the TGF-ß pathway. This was reflected as dysregulated activation of TGF-ß signaling that co-operatively engaged with effectors of the Hippo pathway (YAP/TAZ/TEAD). As a consequence, the loss of Merlin in breast cancer resulted in a significant metabolic and bioenergetic adaptation of cells characterized by increased aerobic glycolysis and decreased oxygen consumption. Mechanistically, we determined that the co-operative activity of the Hippo and TGF-ß transcription effectors caused upregulation of the long non-coding RNA Urothelial Cancer-Associated 1 (UCA1) that disengaged Merlin's check on STAT3 activity. The consequent upregulation of Hexokinase 2 (HK2) enabled a metabolic shift towards aerobic glycolysis. In fact, Merlin deficiency engendered cellular dependence on this metabolic adaptation, endorsing a critical role for Merlin in regulating cellular metabolism. This is the first report of Merlin functioning as a molecular restraint on cellular metabolism. Thus, breast cancer patients whose tumors demonstrate concordant loss of Merlin and SMAD7 may benefit from an approach of incorporating STAT3 inhibitors.

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 24h 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 (10µM) 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.

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.

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.

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.

Transforming growth factor ß 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 ß (TGF-ß) 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-ß1 expression in mouse lung to a level comparable to those found in lung fibrotic diseases by intranasal instillation of AdTGF-ß1(223/225), an adenovirus expressing constitutively active TGF-ß1, 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-ß1 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-ß1 induced lung epithelial apoptosis and robust pulmonary fibrosis. Importantly, lung GSH levels returned to normal, whereas fibrosis persisted at least 21 days after TGF-ß1 instillation. Together, the data suggest that increased TGF-ß1 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.

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-ß1 (TGF-ß1), the most potent and ubiquitous profibrogenic cytokine, and in human lung fibroblasts (CCL-210 cells). The results show that an intranasal instillation of AdTGF-ß1(223/225), an adenovirus expressing constitutively active TGF-ß1, 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-ß1(223/225), restored the activities of uPA and tPA and almost completely blocked TGF-ß1-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-ß1-treated) and naive (TGF-ß1-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-ß1 for 6 hours. Together, the results suggest that TM5275 is a relatively safe and potent antifibrotic agent, with therapeutic potential in fibrotic lung disease.

Glutathione suppresses TGF-beta-induced PAI-1 expression by inhibiting p38 and JNK MAPK and the binding of AP-1, SP-1, and Smad to the PAI-1 promoter.

Transforming growth factor (TGF)-beta upregulates plasminogen activator inhibitor type 1 (PAI-1) in a variety of cell types, and PAI-1 is considered to be an essential factor for the development of fibrosis. Our previous studies demonstrated that TGF-beta decreased intracellular glutathione (GSH) content in murine embryonic fibroblasts (NIH/3T3 cells), whereas treatment of the cells with GSH, which restored intracellular GSH concentration, inhibited TGF-beta-induced collagen accumulation by blocking PAI-1 expression and enhancing collagen degradation. In the present study, we demonstrate that GSH blocks TGF-beta-induced PAI-1 promoter activity in NIH/3T3 cells, which is associated with an inhibition of TGF-beta-induced JNK and p38 phosphorylation. Interestingly, although exogenous GSH does not affect phosphorylation and/or nuclear translocation of Smad2/3 and Smad4, it completely eliminates TGF-beta-induced binding of transcription factors to not only AP-1 and SP-1 but also Smad cis elements in the PAI-1 promoter. Decoy oligonucleotides (ODN) studies further demonstrate that AP-1, SP-1, and Smad ODNs abrogate the inhibitory effect of GSH on TGF-beta-induced PAI-1 promoter activity and inhibit TGF-beta-induced expression of endogenous PAI-1. Furthermore, we show that GSH reduces TGF-beta-stimulated reactive oxygen species (ROS) signal. Blocking ROS production with diphenyleneiodonium or scavenging ROS with a superoxide dismutase and catalase mimetic MnTBaP dramatically reduces TGF-beta-induced p38 and JNK phosphorylation as well as PAI-1 gene expression. In composite, these findings suggest that GSH inhibits TGF-beta-stimulated PAI-1 expression in fibroblasts by blocking the JNK/p38 pathway, probably by reducing ROS, which leads to an inhibition of the binding of transcription factors to the AP-1, SP-1, and Smad cis elements in the PAI-1 promoter.

Glutathione restores collagen degradation in TGF-beta-treated fibroblasts by blocking plasminogen activator inhibitor-1 expression and activating plasminogen.

Transforming growth factor (TGF)-beta plays an important role in tissue fibrogenesis. We previously demonstrated that reduced glutathione (GSH) supplementation blocked collagen accumulation induced by TGF-beta in NIH-3T3 cells. In the present study, we show that supplementation of GSH restores the collagen degradation rate in TGF-beta-treated NIH-3T3 cells. Restoration of collagen degradation by GSH is associated with a reduction of type I plasminogen activator inhibitor (PAI)-1 expression/activity as well as recovery of the activities of cell/extracellular matrix-associated tissue-type plasminogen activator and plasmin. Furthermore, we find that NIH-3T3 cells constitutively express plasminogen mRNA and possess plasmin activity. Blockade of cell surface binding of plasminogen/plasminogen activation with tranexamic acid (TXA) or inhibition of plasmin activity with aprotinin significantly reduces the basal level of collagen degradation both in the presence or absence of exogenous plasminogen. Most importantly, addition of TXA or active PAI-1 almost completely eliminates the restorative effects of GSH on collagen degradation in TGF-beta treated cells. Together, our results suggest that the major mechanism by which GSH restores collagen degradation in TGF-beta-treated cells is through blocking PAI-1 expression, leading to increased PA/plasmin activity and consequent proteolytic degradation of collagens. This study provides mechanistic evidence for GSH's putative therapeutic effect in the treatment of fibrotic disorders.

Green tea polyphenols prevent ultraviolet light-induced oxidative damage and matrix metalloproteinases expression in mouse skin.

Chronic exposure of solar ultraviolet (UV) light to human skin results in photoaging. UV-induced oxidative damage and induction of matrix metalloproteinases (MMP) have been implicated in this process. Because polyphenols from green tea (GTP) prevent other cutaneous adverse effects of UV radiation we hypothesized that UV irradiation-induced oxidative damage and induction of MMP might be prevented in vivo in mouse skin by oral administration of GTP. GTP was administered in drinking water (0.2%, wt/vol) to SKH-1 hairless mice, which were then exposed to multiple doses of UVB (90 mJ per cm2, for 2 mo on alternate days) following in vivo photoaging animal protocol. Treatment of GTP resulted in inhibition of UVB-induced protein oxidation in vivo in mouse skin, a hallmark of photoaging, when analyzed biochemically, by immunoblotting, and immunohistochemistry. GTP treatment also inhibited UVB-induced protein oxidation in vitro in human skin fibroblast HS68 cells, which supports in vivo observations. Moreover, oral administration of GTP also resulted in inhibition of UVB-induced expression of matrix degrading MMP, such as MMP-2 (67%), MMP-3 (63%), MMP-7 (62%), and MMP-9 (60%) in hairless mouse skin. These data suggest that GTP as a dietary supplement could be useful to attenuate solar UVB light-induced premature skin aging.

Treatment of epigallocatechin-3-gallate inhibits matrix metalloproteinases-2 and -9 via inhibition of activation of mitogen-activated protein kinases, c-jun and NF-kappaB in human prostate carcinoma DU-145 cells.

BACKGROUND: Matrix metalloproteinases (MMPs) are involved in tumor progression including the carcinoma of the prostate (CaP). Therefore, the effect of (-)-epigallocatechin-3-gallate (EGCG) was determined on the synthesis and activation of tumor invasion-specific MMP-2 and MMP-9 in human prostate carcinoma DU-145 cells. METHODS: MMP-2 and MMP-9 were determined by zymography and Western blot analysis. Since fibroblast conditioned medium (FCM) partially mimics in vivo tumor-host microenvironment, DU145 cells were co-cultured in FCM. RESULTS: Treatment of EGCG to DU-145 cells resulted in dose-dependent inhibition of FCM-induced pro and active both forms of MMP-2 and MMP-9 concomitant with marked inhibition of phosphorylation of ERK1/2 and p38. In identical conditions, treatment of EGCG or inhibitors of MEK or p38 to DU-145 cells inhibited FCM-induced phosphorylation of ERK1/2 and/or p38 concomitant reduction in MMP-2 and -9. EGCG also inhibited androgen-induced pro-MMP-2 expression in LNCaP cells. Further, treatment of EGCG also resulted in inhibition of activation of c-jun and NF-kappaB in in vitro DU-145 cells. CONCLUSIONS: The inhibition of MMP-2 and MMP-9 in DU145 cells by EGCG is mediated via inhibition of phosphorylation of ERK1/2 and p38 pathways, and inhibition of activation of transcription factors c-jun and NF-kappaB. EGCG may play a role in prevention of invasive metastatic processes of both androgen-dependent and -independent prostate carcinoma.

Proanthocyanidins from grape seeds inhibit expression of matrix metalloproteinases in human prostate carcinoma cells, which is associated with the inhibition of activation of MAPK and NF kappa B.

Prostate cancer (PCA) is the second most frequently diagnosed and leading cause of cancer-related deaths in men in the USA. The recognition that matrix metalloproteinases (MMPs) facilitate tumor cell invasion and metastasis of PCA has led to the development of MMP inhibitors as cancer therapeutic agents. As part of our efforts to develop newer and effective chemopreventive agents for PCA, we evaluated the effect of proanthocyanidins from grape seeds (GSP) on metastasis-specific MMP-2 and -9 in human prostate carcinoma DU145 cells by employing western blot and gelatinolytic zymography. Treatment of GSP dose-dependently inhibited cell proliferation (15-100% by 5-80 microg/ml of GSP), viability (30-80% by 20-80 microg/ml of GSP) and fibroblast conditioned medium (FCM)-induced expression of MMP-2 and -9 in DU145 cells. Since the signaling cascade of mitogen-activated protein kinases (MAPK) have been shown to regulate the expression of MMPs in tumor cells, we found that the treatment of DU145 cells with GSP (20-80 microg/ml) resulted in marked inhibition of FCM-induced phosphorylation of extracellular signal regulated kinase (ERK)1/2 and p38 but had little effect on c-Jun N-terminal kinase under similar experimental conditions. GSP treatment (20-80 microg/ml) to DU145 cells also dose-dependently inhibited FCM-induced activation of NF kappa B concomitantly with inhibition of MMP-2 and -9 expression in the same system. Additionally, the treatment of inhibitors of MEK (PD98059) and p38 (SB203580) to DU145 cells resulted in the reduction of FCM-induced phosphorylation of ERK1/2 and p38 concomitantly marked reduction in MMP-2 and -9 expressions. In further studies, treatment of androgen-sensitive LNCaP cells with a synthetic androgen R1881, resulted in an increase of MMP-2 and -9, which were completely abrogated in the presence of GSP (20-60 microg/ml). These data suggest that inhibition of metastasis-specific MMPs in tumor cells by GSP is associated with the inhibition of activation of MAPK and NF kappa B pathways, and thus provides the molecular basis for the development of GSP as a novel chemopreventive agent for both androgen-sensitive and -insensitive prostate cancer therapies.

Treatment of green tea polyphenols in hydrophilic cream prevents UVB-induced oxidation of lipids and proteins, depletion of antioxidant enzymes and phosphorylation of MAPK proteins in SKH-1 hairless mouse skin.

The use of botanical supplements has received immense interest in recent years to protect human skin from adverse biological effects of solar ultraviolet (UV) radiation. The polyphenols from green tea are one of them and have been shown to prevent photocarcinogenesis in animal models but their mechanism of photoprotection is not well understood. To determine the mechanism of photoprotection in in vivo mouse model, topical treatment of polyphenols from green tea (GTP) or its most chemopreventive constituent (-)-epigallocatechin-3-gallate (EGCG) (1 mg/cm(2) skin area) in hydrophilic ointment USP before single (180 mJ/cm(2)) or multiple UVB exposures (180 mJ/cm(2), daily for 10 days) resulted in significant prevention of UVB-induced depletion of antioxidant enzymes such as glutathione peroxidase (78-100%, P < 0.005-0.001), catalase (51-92%, P < 0.001) and glutathione level (87-100%, P < 0.005). Treatment of EGCG or GTP also inhibited UVB-induced oxidative stress when measured in terms of lipid peroxidation (76-95%, P < 0.001), and protein oxidation (67-75%, P > 0.001). Further, to delineate the inhibition of UVB-induced oxidative stress with cell signaling pathways, treatment of EGCG to mouse skin resulted in marked inhibition of a single UVB irradiation-induced phosphorylation of ERK1/2 (16-95%), JNK (46-100%) and p38 (100%) proteins of MAPK family in a time-dependent manner. Identical photoprotective effects of EGCG or GTP were also observed against multiple UVB irradiation-induced phosphorylation of the proteins of MAPK family in vivo mouse skin. Photoprotective efficacy of GTP given in drinking water (d.w.) (0.2%, w/v) was also determined and compared with that of topical treatment of EGCG and GTP. Treatment of GTP in d.w. also significantly prevented single or multiple UVB irradiation-induced depletion of antioxidant enzymes (44-61%, P < 0.01-0.001), oxidative stress (33-71%, P < 0.01) and phosphorylation of ERK1/2, JNK and p38 proteins of MAPK family but the photoprotective efficacy was comparatively less than that of topical treatments of EGCG and GTP. Lesser photoprotective efficacy of GTP in d.w. in comparison with topical application may be due to its less bioavailability in skin target cells. Together, for the first time a cream based formulation of green tea polyphenols was tested in this study to explore the possibility of its use for the humans, and the data obtained from this in vivo study further suggest that GTP could be useful in attenuation of solar UVB light-induced oxidative stress-mediated and MAPK-caused skin disorders in humans.

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.

Antioxidant and antimutagenic properties of aqueous extract of date fruit (Phoenix dactylifera L. Arecaceae).

Fruits of the date palm (Phoenix dactylifera L. Arecaceae) are very commonly consumed in many parts of the world and are a vital component of the diet in most of the Arabian countries. This preliminary study documents for the first time its antioxidant and antimutagenic properties in vitro. There was a dose-dependent inhibition of superoxide and hydroxyl radicals by an aqueous extract of date fruit. The amount of fresh extract required to scavenge 50% of superoxide radicals was equivalent to 0.8 mg/mL of date fruit in the riboflavin photoreduction method. An extract of 2.2 mg/mL of date fruit was needed for 50% hydroxyl-radical-scavenging activity in the deoxyribose degradation method. Concentrations of 1.5 and 4.0 mg/mL completely inhibited superoxide and hydroxyl radicals, respectively. Aqueous date extract was also found to inhibit significantly the lipid peroxidation and protein oxidation in a dose-dependent manner. In an Fe(2+)/ascorbate system, an extract of 1.9 mg/mL of date fruit was needed for 50% inhibition of lipid peroxides. In a time course inhibition study of lipid peroxide, at a 2.0 mg/mL concentration of date extract, there was a complete inhibition of TBARS formation in the early stages of the incubation period that increased during later stages of the incubation. Similarly, in the high Fe(2+)/ascorbate induction system a concentration of 2.3 mg/mL inhibited carbonyl formation measured by DNPH reaction by 50%. Moreover, a concentration of 4.0 mg/mL completely inhibited lipid peroxide and protein carbonyl formation. Date fruit extract also produced a dose-dependent inhibition of benzo(a)pyrene-induced mutagenecity on Salmonella tester strains TA-98 and TA-100 with metabolic activation. Extract from 3.6 mg/plate and 4.3 mg/plate was found required for 50% inhibition of His+ revertant formation in TA-98 and TA-100, respectively. These results indicate that antioxidant and antimutagenic activity in date fruit is quite potent and implicates the presence of compounds with potent free-radical-scavenging activity.

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.