Alzheimers Disease Drug Discovery Review

Brain Research Reviews 50 (2005) 361 – 376 www.elsevier.com/locate/brainresrev

Review

Can herbs provide a new generation of drugs for treating Alzheimer’s disease? Thimmappa S. Anekonda, P. Hemachandra Reddy* Neurogenetics Laboratory, Neurological Sciences Institute, Oregon Health and Science University, 505 NW 185th Avenue, Beaverton, OR 97006, USA Accepted 16 September 2005 Available online 2 November 2005

Abstract The overall aim of this review is to discuss cellular mechanisms at work in the progression of AD and current therapeutic strategies for treating AD, with a focus on the potential efficacy of herbal treatments. Recent advances in molecular, cellular, and animal model studies have revealed that formation of the 4-kDa amyloid beta peptide is a key factor in the development and progression of AD. Several cellular changes have been identified that are related to amyloid beta plaques and neurofibrillary tangles found in the autopsied brains of AD patients and in AD animal models. Several therapeutic strategies have been developed to treat AD, including anti-inflammatory, anti-oxidant, and anti-amyloid approaches. Recently, herbal treatments have been tested in animal and cellular models of AD and in clinical trials with AD subjects. In AD animal models and cell models, herbal extracts appear to have fewer adverse effects than beneficial effects on Ah and cognitive functions. These extracts have multi-functional properties (pro-cholinergic, anti-oxidant, anti-amyloid, and anti-inflammatory), and their use in the treatment of AD patients looks promising. The chemical compositions of herbs and their potential for alleviating or reducing symptoms of AD or for affecting the disease mechanism need to be further studied. D 2005 Elsevier B.V. All rights reserved. Theme: Disorders of the nervous system Topic: Degenerative disease: Alzheimer’s—miscellaneous Keywords: Alzheimer’s disease; Animal model; Bioavailability; Herbal drug; In vitro model; Mitochondria

Contents 1. 2. 3. 4.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Cellular changes in AD progression . . . . . . . . . . . . . . . . Therapeutic strategies . . . . . . . . . . . . . . . . . . . . . . . . Herbal drugs . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4.1. Herbs tested for anti-Ah and related effects in AD models . 4.1.1. Animal models . . . . . . . . . . . . . . . . . . . 4.1.2. In vitro models . . . . . . . . . . . . . . . . . . . 4.2. Herbs tested for anti-oxidant or anti-apoptotic effects in AD 4.2.1. Animal models . . . . . . . . . . . . . . . . . . . 4.2.2. In vitro models . . . . . . . . . . . . . . . . . . .

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Abbreviations: Ah, amyloid beta; AChE, acetylcholinesterase; AD, Alzheimer’s disease; APP, amyloid precursor protein; ATP, adenosine triphosphate; BBB, blood – brain barrier; FAD, familial Alzheimer’s disease; FDA, Federal Drug Administration; NFTs, neurofibrillary tangles; PC12 cells, pheochromocytoma cells; ROS, reactive oxygen species; SAD, sporadic Alzheimer’s disease; NO, nitric oxide; TrkA, tyrosine kinase receptor A * Corresponding author. Fax: +1 503 418 2501. E-mail address: [email protected] (P.H. Reddy). 0165-0173/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.brainresrev.2005.09.001

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4.3.

Herbs tested for inhibiting AChE or NMDA receptors and enhancing 4.3.1. Animal models . . . . . . . . . . . . . . . . . . . . . . . . 4.3.2. In vitro models. . . . . . . . . . . . . . . . . . . . . . . . 4.4. Herbs tested for anti-inflammatory effects in AD models . . . . . . . 4.5. Mixtures of herbs for treating AD. . . . . . . . . . . . . . . . . . . 5. Clinical trials on herbal drugs, using AD patients . . . . . . . . . . . . . . 6. What makes herbs particularly suitable for treating AD?. . . . . . . . . . . 6.1. Bioavailability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6.2. The blood – brain barrier. . . . . . . . . . . . . . . . . . . . . . . . 6.3. Toxic and adverse drug effects and drug-drug interaction. . . . . . . 6.4. Synergistic interactions . . . . . . . . . . . . . . . . . . . . . . . . 7. Concluding remarks . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Acknowledgments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Introduction Alzheimer’s disease (AD) is a complex, multifactorial, heterogeneous mental illness, which is characterized by an age-dependent loss of memory and an impairment of multiple cognitive functions [82,112,113,126,139]. AD is associated with the presence of intracellular neurofibrillary tangles (NFTs) and extracellular amyloid beta (Ah) plaques, loss of neuronal subpopulations, synaptophysin immunoreactivity of presynaptic terminals, loss of cholinergic fibers, proliferation of reactive astrocytes and microglia, and mitochondrial dysfunction [43,56,79,112, 114,115,126,127,138,142]. With human life span increasing and with decreasing cognitive functions in elderly individuals with AD-related dementia, AD has become a major health problem in society. Early detection, prevention, and therapeutic interventions are urgently needed to minimize the ill effects of this devastating disease [113]. Based on a survey of PubMed literature on herbal medicines used in cellular studies of AD, studies of animal models of AD, and clinical trials using AD patients, we investigated herbal medicines as an intervention for treating AD patients. This review begins with a discussion of cellular mechanisms that are involved in AD development and progression and then reviews current therapeutic strategies for AD that involve herbal medicines.

2. Cellular changes in AD progression AD occurs in both familial and sporadic forms. In familial AD (FAD), mutations in the amyloid precursor protein (APP), presenilin 1, and presenilin 2 genes are the currently known causal factors. These genetic mutations inherit in an autosomal dominant fashion. FAD constitutes only 2– 3% of the total number of AD patients [112], and it has an early age of onset (younger than 65 years of age). Sporadic AD (SAD) constitutes the vast majority of AD cases, and it has a late age of onset (65 years of age and older). The causes of SAD are still unknown [126].

synaptic functions in AD models . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

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366 366 367 369 369 370 370 370 370 371 371 371 371 371

Histological, pathological, molecular, cellular, and gene expression studies of AD have revealed that multiple cellular pathways are involved in AD progression [113]. Pathologically, there are no differences between FAD and SAD [126]. In patients with SAD, pathological changes including Abeta production and deposits, NFTs, synaptic damage, and neuronal loss occur latter than in patients with FAD [43,56,79,112,114,115,126,127,138,142]. In FAD, genetic mutations accelerate the disease process [126], whereas in SAD, in the absence of genetic mutation, cellular changes that control AD progression take more time to develop [113]. It is possible that several factors are involved in causing SAD, the major one of which is aging [126]. Other factors that have been implicated are the apolipoprotein genotype (ApoE4) [108,109], mitochondrial defects [112], insulin-dependent diabetes [26,107], environmental conditions [56], and diet [56]. In FAD, recent molecular, cellular, and animal model studies have provided evidence that a 4-kDa peptide, a cleavage product of APP due to h and g secretases, is a key factor in AD development and progression [113,126]. The formation of the 4-kDa Ah peptide in the brains of AD patients is a progressive and sequential process. Initially, soluble monomeric and oligomeric forms of 40 – 42-amino acid residues (Ah1 – 40, shorter form; and Ah1 –42, longer form) accumulate and later become insoluble fibrils and Ah deposits. In recent studies of triple transgenic mice that express 3 transgenes related to AD (AD-PS1, AD-APP, and FTD-tau), Ah plaques were found in mice at 5 months of age, and NFTs were found at 12 months, suggesting that Ah production is critical and may facilitate tau pathology [94]. Further, synaptic changes that occur in the triple transgenic mouse line have been directly associated with Ah production [94,95]. In addition, Ah immunotherapy studies of triple transgenic mice showed a reduction in not only extracellular Ah plaques but also in intracellular Ah accumulation, which led to the clearance of early tau pathology, suggesting that early Ah production is critical for subsequent cellular changes seen in these mice, including the synaptic damage, hyperphosphorylation of tau and NFTs [94,95].

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The Ah plaques in the AD transgenic mice were also found to be associated with activated microglia and astrocytes and to trigger inflammatory responses [56]. However, astrocytes and microglia were found to proliferate in the vicinity of Ah and to clear Ah deposits [56]. Yet in other studies, interactions among Ah, glia, and astrocytes were found to cause inflammation in the AD brain, which can lead to altered neuronal homeostasis and oxidative injury. Based on this last set of evidence, Ah oligomers have been hypothesized to cause oxidative injury, which can lead to altered kinases and phosphatases [56]. In addition to findings of inflammatory changes in AD, recent molecular, cellular, and animal model studies have revealed that mutant APP and/or Ah enters mitochondria and interacts with the Ah-induced alcohol dehydrogenase protein, disrupts electron transport, generates reactive oxygen species (ROS, a term used to describe free radicals derived from molecular oxygen in the mitochondria), and inhibits cellular ATP [112]. These results suggest that mutant APP and Ah interactions with mitochondrial proteins cause mitochondrial dysfunction in AD [7,74,112]. In SAD, aging plays a significant role in AD progression [126]. In addition, mitochondrial defects [24,79,112] and ApoE4 allele are major initiating factors [108,109] of AD progression. ApoE4 and ROS (generated by mitochondrial defects) activate h and g secretases of APP and generate Ah peptides [61,108]. It has been proposed that chronic ROS exposure can result in oxidative damage to mitochondrial and cellular proteins, lipids, and nucleic acids, resulting in a shut-down of mitochondrial energy production [112]. Defective mitochondria in AD neurons may not move effectively and may not supply necessary cellular ATP at nerve terminals (such as dendritic spines and synapses) for normal neural communication. The low levels of cellular ATP at nerve terminals may lead to the loss of synapses and synaptic function and may ultimately cause cognitive decline in AD patients [112].

3. Therapeutic strategies There is a large body of evidence suggesting that the accumulation of Ah is a major causative factor in AD pathogenesis. As a result, therapeutic strategies aiming to decrease mutant APP and/or Ah levels are currently a major focus in AD research. Approaches for decreasing Ah levels include inhibiting the generation of Ah [57], reducing soluble Ah levels [97,149], and enhancing Ah clearance from the brain [27,28,34,88,95,124]. Molecular, cellular, and animal model studies revealed that AD progression involves such cellular changes as inflammatory responses, mitochondrial dysfunction, oxidative damage, synaptic failure, and hyperphosphorylation of tau, all of which are directly related to Ah production and aging [56,66,94,95,97,112,113,149]. Both passive and active immunization of Ah in AD transgenic mouse models have promised that Ah levels can

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be reduced in the brains of AD mice [27,28,34,97,124]. With encouraging results from in vivo studies that have aimed to abolish Ah in cellular and animal models of AD, immunotherapy research has moved quickly to human clinical trials by Elan Pharmaceuticals [123]. Unfortunately, their phase II clinical trials with AD patients as subjects were stopped because a small percentage developed symptoms of aseptic meningoencephalitis [123]. Before resuming immunotherapy in clinical trials, several issues need to be addressed: (1) the long-term consequences of Ah immunization for the AD brain (2) while clearing Ah deposits in the AD brain, the consequences of glial and Ah interactions, and the downstream effects of AD progression, and (3) the relationship between the clearing of Ah and the improvement of cognitive functions in AD patients. Anti-inflammatory therapy has been used to treat AD patients. Inflammation is an important component in the pathogenesis of AD, consisting of the activation of both microglia and astrocytes. Recent histological studies revealed the presence of activated microglia and reactive astrocytes in and around extraneuronal Ah plaques, which are thought to facilitate the clearing of Ah deposits from the brain parenchyma [56]. In Ah-induced inflammation in AD, microglia can activate and differentiate into phagocytic CD11b+ cells that in turn secrete IL-1h, TNF-a, nitric oxide (NO), free radicals, and chemokines, and that activate complement via an innate pathway [86]. Thus generated, NO can cause T cell apoptosis. Microglia can also differentiate into CD11c antigens presenting both Th1 and Th2 cells via an adaptive pathway, which in AD can suppress the innate pathway by secreting anti-inflammatory cytokines (IL-4, IL-10, TGF-h) [86]. However, there is increasing evidence to suggest that the chronic activation of microglia, presumably via the secretion of cytokines and reactive molecules [1,140], may exacerbate plaque pathology as well as enhance the hyperphosphorylation of tau and the formation of NFTs [94,95]. Thus, the suppression of microglial activity in the AD brain has been considered a possible therapeutic strategy to treat AD patients [56]. Along these lines, anti-inflammatory drugs, particularly non-steroidal anti-inflammatory drugs, have shown to lessen the effects of AD pathology [32,33,62]. Oxidative stress is a major factor involved in the development and progression of AD and other forms of dementia. A large body of data suggests that free radical oxidative damage—particularly of neuronal lipids [72,80, 81], proteins [17,75], and nucleic acids [75]—is extensive in the brains of AD patients. Increased oxidative stress is thought to result in the generation of free radicals and ROS, which is reported to be released by microglia activated by Ah [84,106]. Using a Tg2576 mouse model of AD and treating the Tg2576 mice with a vitamin E-supplemented diet, in vivo studies reported decreased levels of Ah1 – 40 and Ah1– 42, suggesting that vitamin E may have a direct effect on AD pathology. Several recent anti-oxidant studies using AD patients revealed beneficial effects of diets

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supplemented with vitamin E [89,90,161]. The pathological effects of oxidative stress are yet to be assessed in patients treated with anti-oxidants. Other studies have shown beneficial effects of several other anti-oxidants, such as melatonin, Gingko, and alpha lipoic acid, supplemented in the water or in the diet of transgenic mouse models of AD [31,147]. These studies found that the anti-oxidant therapies are safe and produce no adverse effects. Using in vitro cell culture and transgenic mouse models of AD, several laboratories around the world are currently involved in developing anti-oxidant therapies. There are only four drugs that the Federal Drug Administration (FDA) has approved and that are currently available for treating AD patients in the United States. Three of the drugs—Tacrine (CognexR), Donepezil (AriceptR), and Rivastigmine (ReminylR)—inhibit acetylcholinesterase (AChEI) either selectively or non-selectively, but they have resulted in various adverse drug effects [6]. In two recent studies, AD patients treated with Donepezil showed rescued APP metabolism [40,168] or a slow-down in the progression of hippocampal atrophy, a surrogate of disease progression. Thus, Donepezil was shown to provide neuroprotective effects [40]. Memantine (NamendaR), the fourth and most recently approved drug, non-competitively inhibits NMDA receptors, prevents glutamate excitotoxicity, and shows minimal adverse drug effects in AD patients [6]. All four of these drugs improved the cognitive functions of AD patients symptomatically and have thus improved the quality of life for these patients; however, these drugs do not modify the disease mechanism in the long run. Thus, when patients no longer take the drugs, their symptoms of AD return. The paucity of drugs currently available for treating AD and their limited targets in AD pathology call for the development of a new generation of drugs that not only affect cholinergic functions associated with AD but also target other cellular pathways in AD pathogenesis.

batches of the drugs since their bioactivity varies considerably due to differences in plant-growth environments [83]. This limitation has prompted pharmaceutical companies to use single molecules of synthetic compounds in drug therapies. Although herbal drugs promise significant health benefits, they have been found to be either ineffective or effective but showing excessive adverse drug effects, especially when administered to treat complex diseases, such as cancer, osteoporosis, and AD. A large segment of the public finds solace in herbs, in part believing that herbs are natural and hence safer than synthetic drugs, and that a complex mixture of herbs can effectively treat complex diseases. These beliefs may account for the sudden increase in herbal use in the last decade [111]. The United States market for just herbal supplements now exceeds $7 billion per year [36]. In 2002, the projected worldwide sales of plant-derived pharmaceuticals and their precursors exceeded $30 billion [111]. Today, one in three Americans uses herbal supplements, with consumption much greater among women [87,141], patients undergoing surgery [9], and the elderly population. The complex pathology of AD and heterogeneous pharmacological effects of herbal extracts pose difficult challenges in the development of herbal drugs for AD treatment [45]. However, the number and quality of recent studies suggest that herbal drugs and AD pathology are at a new crossroad. Here, we identify herbal extracts that have been found to affect AD pathomechanisms, highlighting interactions of Ah, mitochondrial anti-oxidant mechanisms, inflammatory pathways, and cholinergic and glutamergic functions in presynaptic and postsynaptic neurons. We draw on recent reviews of herbal extracts affecting the central nervous system and age-related dysfunctions [2,3,8,18, 39,46 – 48,58,59,77,101,102,119,121,131,134,135,137, 144,159]. 4.1. Herbs tested for anti-Ab and related effects in AD models

4. Herbal drugs Over 35,000 plant species currently used for medicinal purposes around the world possess more than 4000 flavonoid (polyphenolic) structures, terpenes, and phytochemicals, such as alkaloids [76 – 78,154]. These plant drugs provide numerous health benefits, including antipsychotic, anti-fatigue, anti-depressant, anxiolytic, hypnotic, anti-inflammatory, anti-oxidant, anti-neoplastic, anti-arthritic, anti-diabetic, and anti-lipogenic effects [29,92,152]. The drugs showing anti-depression, antiinflammatory, anti-oxidant, and anti-psychotic benefits may be particularly beneficial to AD patients because in the late stages of disease progression, AD patients exhibit psychotic changes in addition to cellular changes relating to inflammation, oxidation, and infection. However, there are problems surrounding the preparation of herbal drugs, including significant variations across

4.1.1. Animal models Table 1A summarizes studies of herbal extract/chemical treatments that have anti-amyloid effects, including anti-h and anti-g secretases and pro a-secretase, and that used mouse and rat models of AD. Curcumin, a bioactive compound present in the Indian spice turmeric (Curcuma longa), reduced accumulations of Ah plaques in the brains of aged Tg2576 mice [157]. In the same mouse model, a Ginkgo biloba extract prevented an age-dependent decline in spatial cognition and enhanced the metabolic rate of the Tg2576 brain via increased levels of protein carbonyls, although the extract did not change Ah plaque burdens or protein oxidation levels [136]. In a gene expression study with adult C57BL6 mice, EGb 761, a common extract of G. biloba, increased the mRNA expression of transthyretin (a Ah sequester) in the hippocampus and increased tyrosine/threonine phosphatase

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Table 1 Herbal extracts/chemicals tested for their anti-amyloid effects in animal and in vitro models of Alzheimer’s disease Plant extracts (A) Animal models Curcumina Ginkgo biloba extract EGb 761b

Models and oxidants

Effects of plant extracts

References

Tg2576 mice; Ah1 – 40, Ah1 – 42 Tg2576 mice and wild-type mice

Blocked the aggregation, oligomer, and fibril formation in vivo and in vitro Prevented age-dependent decline in spatial cognition Increased the mRNA expression of transthyretin, tyrosine/threonine phosphatase 1 and microtubule-associated tau Increased the release of a-APPs Reversed the Ah-induced down-regulation of APP secretion and protein kinase C Ameliorated the performance impairment in a passive avoidance task and suppressed the over-expression of hippocampal Ah Increased the expression of transthyretin in the brainstem and hippocampus Reduced insoluble amyloid Ah1 – 40 and Ah1 – 42 peptides in the brain cortex

Yang et al. [157]

C57BL6 mice

EGb761 Huperzine Ac

Sprague – Dawley rats Sprague – Dawley rats; Ah1 – 40

Dipsacus asper extract

Sprague – Dawley rats; aluminum chloride-induced Ah

Nicotined

Holtzman rats

Nicotined

APPsw mice

(B) In vitro models Curucuma longa compounds Curcumin Eugenol and h-asaronee

Rat pheochromocytoma cells; Ah25 – 35, Ah1 – 42 Biochemical assay; Ah1 – 40, Ah1 – 42 Rat PC12 cells; Ah1 – 40

Tenuigeninf

Neuroblastoma cells

Indirubinsg

Insect Sf9 cells and tau phosphorylation in vitro; slices from adult mouse brain striatum

a b c d e f g

Stackman et al. [136] Rimbach et al. [118]; Watanabe et al. [151] Colciaghi et al. [23] Zhang et al. [166] Zhang et al. [165]

Li et al. [65] Hellstrom-Lindahl et al. [41]; Nordberg et al. [93]

Protected against Ah insult

Park and Kim [98]

Inhibited Ah fibril formation Attenuated cell death by blocking Ah-induced Ca2+ intake Suppressed the secretion of Ah by inhibiting BACE1 or h-secretase Inhibited glycogen synthase kinase-3h and cyclin-dependent kinase-5

Ono et al. [96] Irie and Keung [49] Jia et al. [51] Leclerc et al. [64]

Curcumin, a bioactive compound from the rhizome of Indian spice, turmeric (Curcuma longa). EGb 761, a standard total extract from the leaves of Ginkgo biloba. Huperzine A, an alkaloid derived from a Chinese herb, club moss (Huperzia serrata). Nicotine, a bioactive compound from the leaves of tobacco (Nicotiana tabaccum). Eugenol and h-asarone, essential oil from Rhizoma acori graminei. Tenuigenin, a bioactive compound from Polygala tenuifolia. Indirubins, extracted from Qing Dai (Indigo naturalis), and isatan plants or molluscs.

1 and microtubule-associated tau in the cortex [151]. Transthyretin is known to participate in the transport of thyroxin and in retina-binding proteins, to function as the carrier of Ah in cerebrospinal fluid, and to prevent Ah aggregation and fibril formation [85]. Tyrosine/threonine phosphatase 1 and tau are involved in the formation and disintegration of NFTs in the AD brain. An increase in tyrosine/threonine phosphatase 1 may play a role in dephosphorylating the hyperphosphorylated microtubule that is associated with tau [118,151]. In a Sprague – Dawley rat model of AD, EGb 761 increased the release of a-secretase of APPs in a PKCindependent manner by affecting the cleavage of APP asecretase [23]. In this same rat model, huperzine A (a potent cholinesterase inhibitor), an extract of club moss (Huperzia serrata), reversed the Ah-induced down-regulation of APP secretion and the protein kinase C [166]. The root extract of Dispacus asper, another Chinese herb, ameliorated the impairment of cognitive dysfunction in a passive avoidance task and suppressed the over-expression of hippocampal Ah that had been induced by aluminum chloride [165].

Treatments with nicotine, a bioactive compound found in tobacco, not only increased the expression of transthyretin in the brainstems and hippocampi of Holtzman rats [65] but also attenuated insoluble amyloid Ah1 –40 and Ah1 – 42 peptides in the brain cortices of APPsw mice [41,93]. 4.1.2. In vitro models Herbal treatments in vitro have also conferred protection against Ah-induced toxicity in various cell culture systems (Table 1B). Curucuma longa extracts prevented Ah fibril formation [96] and protected pheochromocytoma cells (PC12 cells) [98] from the insults caused by Ah oligomers and fibrils. In brain sections from AD patients and Tg2576 mice [157], curcumin effectively blocked Ah1– 40 aggregation and Ah1– 42 fibril and oligomer formation. Eugenol and hasarone, derived from Rhizoma acori graminei, rescued PC12 cells from death by blocking Ah-induced Ca2+ intake [49]. EGb 761-treated hippocampal slices from rat brains showed an increased release of soluble APPs (sAPPs) [23], and mutant embryonic kidney cells from humans that were treated with huperzine A reversed the Ah-induced down-

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regulation of APP secretion and PKC activity [166]. In addition, tenuigenin (Polygala tenuifolia) suppressed the secretion of Ah in neuroblastoma cells by inhibiting BACE1 or h-secretase [51]. In insect Sf9 cells and in the brain striatum of adult mice, indirubins, derived from Qing Dai (Indigo naturalis) and from isatan plants or molluscs, inhibited glycogen synthase kinase-3h and cyclin-dependent kinase-5, which are responsible for the abnormal hyperphosphorylation of tau [64]. Thus, the findings from cell culture systems generally support those of animal model studies in terms of the influence of herbal drugs on preventing Ah formation and aggregation. 4.2. Herbs tested for anti-oxidant or anti-apoptotic effects in AD models 4.2.1. Animal models Oxidative events in mitochondria are known to generate accumulations of ROS in several age-related diseases including AD. Recent studies strongly suggest that such events are the primary factors that initiate SAD [105]. As shown in Table 2A, only a limited number of studies have used animal models for testing anti-oxidant effects of herbs compared to animal models used for testing other effects of herbs. In a morphometric study using Wistar rats, EGb 761treated, vitamin E-deficient rats showed increased populations and increased densities of synaptic mitochondria, a disproportionate number of small-sized synapses and improved physiological adaptive capacity [15]. An antioxidant treatment combining G. biloba, vitamin E, pycnogenol, and ascorbyl palmitate reduced periodic acid Schiffpositive inclusion bodies and reduced apoptotic cells in the hippocampus of ApoE-deficient mice on a C57B1/6J hybrid background. This anti-oxidant treatment also increased the life span of the mice [148]. EGb 761 increased the resistance of both wild-type and aging mutant Caenorhabditis elegans (worms) to acute oxidative and thermal stress and increased their lifespan [156]. Egb761 also attenuated an age-related accumulation of H2O2-related ROS [131]. In male Wistar rats, resveratrol, a bioactive compound in red wine, and Centella asiatica, an Ayurvedic Indian medicinal herb, both prevented intracerebroventrical, straptozotocin-induced cognitive impairment and oxidative stress [128,147]. This rat model for memory impairment is well known for SAD, as it directly alters glucose levels in the brain and energy metabolism in the mitochondria. 4.2.2. In vitro models Table 2B summarizes herbs tested for their anti-oxidant and anti-apoptotic effects in cell culture systems. EGb 761 and its bioactive compounds appear to be tested the most frequently in several systems, such as PC12 cells [30,132, 158,167], hippocampal cells from Sprague – Dawley rats [11,12,13], human neuroblastoma cells [14,73], and postmortem AD brain slices [110]. In these systems, anti-oxidants

against several oxidants—including Ah peptide(s) (25 – 35, 1 – 40, 1 –42), H2O2, antimycin, xanthine, serum deprivation, staurosporine, sodium nitroprusside, or prion protein, and EGb 761—showed classic anti-oxidant effects. The antioxidants prevented the toxic effects of Ah fibrils; decreased ROS-induced c-Myc, p53, Bax, and caspase-3 activity, which led to reduced apoptosis, prevented a reduction in cytochrome c levels, attenuated DNA fragmentation, restored mitochondrial function, reduced the formation of toxic cyclooxygenases, and protected cells against lipid oxidation. Similar anti-oxidant effects were also conferred by C. longa [60] and aged garlic extracts [99] on PC12 cells subjected to oxidant assaults. In other studies, ginsenoside Rg1 (a ginseng extract) [20], red-wine crude extracts [120], resveretrol [50], Bacopa monniera [16], and epigallocatechum gallate (a green tea extract) [21], showed strong anti-oxidant effects and protected cell cultures from cytotoxic oxidants. 4.3. Herbs tested for inhibiting AChE or NMDA receptors and enhancing synaptic functions in AD models 4.3.1. Animal models Inhibition of AChE and NMDA receptors is one of the main therapeutic strategies for treating AD patients. Indeed, three of the four FDA-approved AD drugs were designed, based on their AChE inhibitory effects, and the fourth FDAapproved AD drug, mematine, was developed primarily to attenuate the expression of NMDA receptors [100]. Interestingly, few herbs seem to inhibit the expression both AChE and NMDA receptors [37,155,163]. Using the Sprague – Dawley rat cortex, Liang and Tang [67] compared the in vivo effects of huperzine A with Donepezil and Rivastigmine in terms of their effects on acetylcholine and the activity of acetylcholinesterase (Table 3A). They found that huperzine A increased the concentration of acetylcholine and inhibited acetylcholinesterase more efficiently than did injections of Donepezil and Rivastigmine. They also found that huperzine A not only penetrated the blood –brain barrier (BBB) more efficiently but also showed long-lasting inhibitory effects on AChE. Wang et al. [150] reported that anisodamine, extracted from the Chinese herb Anisodus tanguticus, produced cholinomimetic effects in mice when combined with the peripheral muscarinic blocker pilocarpine. When pilocarpine was administered alone, anisodamine effectively blocked cholineacetylesterase activity, but it initiated typical cholinergic side effects, such as diarrhea, hypersalivation, and bradycardia [150]. These adverse effects were nearly eliminated with the administration of anisodamine in combination with pilocarpine. In another recent in vivo study with an ICR mice model of AD, scopolamine-induced memory deficits caused by acetylcholinesterase activity were reversed with green tea extract [55]. Nearly 50% of the cortical tyrosine kinase receptor A (TrkA) is lost in the early stages of AD progression [25]. Nicotine treatment of Wistar rats increased the expression of

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Table 2 Herbs tested for antioxidant- or antiapoptosis-related effects in AD models Plant extracts

Models and oxidants

Effects of plant extracts

References

(A) Animal models EGb 761a

Wistar rats deficient in vitamin E

Increased the proportion of small-sized synapses and mitochondrial density Increased the life span and reduced periodic acid Schiffpositive inclusion bodies and apoptotic cells Attenuated age-related accumulation of ROS Increased resistance to acute oxidative and thermal stress, and increased life span Prevented ICV STZ-induced cognitive impairment and oxidative stress Increased cognitive behavior and prevented oxidative stress

Bertoni-Freddari et al. [15]

Ginkgo biloba, Vitamin E, Pycnogenol, Ascorbyl palmitate

ApoE-deficient mice

EGb 761 Kaempferol Quercetin EGb 761

Mutant C. elegans worm

Resveratrolb

Male Wistar rats; Intracerebroventrical straptozotocin model of SAD

Centella asiatica extract

Male Wistar rats; Intracerebroventrical straptozotocin model of SAD

(B) In vitro models EGb 761 and its active compounds

C. elegans worm

Rat PC12 cells; hippocampal cells from Sprague – Dawley rats; neuroblastoma cells from human AD brains

Curcuma longa extract

Rat PC12 cells; Pyrogallol, H2O2

Aged garlic extract and S-allylcysteinec

Rat PC12 cells; Ah25 – 35

Ginseoside Rg1d Red wine crude extract and resveratrol Bacopa monniera extract

Cortical cells from Sprague – Dawley rats human umbilical vein endothelial cells and PC12 cells; Ah25 – 35, Ah1 – 42 Astrocytes from Wistar albino rat brains; S-nitroso-N-penicillamine Hippocampal neurons from Sprague – Dawley rats; Ah25 – 35

Epigallocatechin gallatee a b c d e

Prevented the formation of Ah-derived diffusible ligands or toxic fibrils; decreased ROS-induced c-Myc, p53, Bax, and caspase-3 activity leading to reduced apoptosis; prevented a reduction in cytochrome c levels; attenuated DNA fragmentation; restored mitochondrial function; reduced formation of toxic cyclo-oxigenases; protected cells against lipid oxidation Rescued cells from cell death and increased anti-oxidant enzyme activity Suppressed ROS, caspase-3, and DNA fragmentation; protected cells from apoptosis Reduced apoptosis Protected cells from ROS; prevented DNA fragmentation Inhibited DNA fragmentation and ROS formation Protected cells against apoptosis

Veurink et al. [148]

Smith and Luo [131] Wu et al. [156]

Sharma and Gupta [128]

Veerendra Kumar and Gupta [147]

Eckert et al. [30]; Smith et al. [132]; Yao et al. [158]; Zhou and Zhu [167]; Bastianetto and Quirion [11]; Bastianetto et al. [12,13]; Bate et al. [14]; Luo et al. [73]; Ramassamy et al. [110]

Koo et al. [60]

Peng et al. [99]

Chen et al. [20] Russo et al. [120]; Jang and Surh [50] Bhattacharya et al. [16] Choi et al. [21]

EGb 761, a standard total extract from the leaves of Ginkgo biloba. Aged garlic extract and S-allylcysteine derived from the bulbs of garlic (Allium sativum). Ginseoside Rg1, a bioactive compound from the roots of ginseng (Panax ginseng). Resveretrol, a bioactive compound from the seeds of red grapes (Vitis vinifera). Epigallocatechin gallate, a bioactive compound from the leaves of green tea (Camellia sinensis).

TrkA in the hippocampus [52]. In the cholinergic neurons of the basal forebrain from AD patients, TrkA was found to serve as a receptor for the nerve growth factor, a critical trophic factor for the survival of neurons. Based on the effects of nicotine on acetylcholine-receptor antagonists, Jonnala et al. [52] suggest that the neuroprotective action of nicotine may be mediated via a central a7 acetylcholine receptor. In a recent study of the cerebral cortex and hippocampus of wild-type mice, Ah25– 35 treatment caused impaired learning memory and a reduction in the expression of

phosphorylated neurofilament H, an axonal marker, and synaptophysin, a synaptic marker [145]. These mice recovered these functions when treated with Rb1 (a protopanaxadiol-type saponin) and MI (a derivative of Rb1), both of which are extracted from the Vietnamese ginseng (Panax vietnamensis). 4.3.2. In vitro models Table 3B summarizes in vitro effects of herbs on the inhibition of AChE and NMDA receptors, which are

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Table 3 Herbs tested in animal models for inhibitory effects on cholinesterase and N-methyl-d-aspartate receptors Plant extracts

Models and oxidants

Effects

References

(A) Animal models Huperzine Aa

Male Sprague – Dawley rats

Liang and Tang [67]

Anisodamineb

Kunming mice

Green tea extract

ICR mice; Scopolamine

Nicotinec Ginsenoside Rb1 and MId

Wistar rats Mouse model of AD

Increased the concentration of acetylcholine; inhibited acetylcholine esterase Provided cholinomimetic effects when anisodamine was combined with peripheral muscarinic blockers Reversed memory deficits by inhibiting AChE activity Increased the expression of TrkA receptors Recovered impaired learning and memory; increased axonal density and synaptophysin expression in cerebral cortex and hippocampus

(B) In vitro models Huperzine A

In vitro cholinesterase inhibition assay

Huperzine A dimers inhibited AChE more potently than they inhibited the monomers Inhibited NMDA-induced toxicity Inhibited an NMDA receptor-induced current

Wong et al. [155]

Huperzine A Huperzine A EGb 761e Ginkgolides A, B, C, and J, and bilobalide Ginkgo biloba Ptychopetalum olacoides extract Alkaloids and plant extracts from narcissus Zeatinf Salvia lavandulaefolia and other Salvia species

Cortex or synaptic plasma membranes Hippocampal neurons from Sprague – Dawley rats Hippocampal and cerebellar neurons from Wistar rats Rat PC12 cells; Ah1 – 42 Frontal cortex, hippocampus, and striatal neurons of male Wistar rats (in vitro) and of male Swiss albino mice (ex vivo) Microplate assay Rat PC12 cells In vitro studies

Wang et al. [150]

Kim et al. [55] Jonnala et al. [52] Tohda et al. [145]

Gordon et al. [37] Zhang and Hu [163]

Blocked glycine-activated chloride channels; weakly inhibited an NMDA receptor-activated current Inhibited Ah-derived diffusible ligands and the formation of oligomers Inhibited AChE in vitro and ex vivo

Chatterjee et al. [19]

Seven alkaloids showed AChE inhibitory activity

Lopez et al. [71]

Inhibited AChE activity Inhibited AChE activity

Heo et al. [42] Perry et al. [103,104], Ren et al. [116], Savelev et al. [122]

Chromy et al. [22] Siqueira et al. [129]

a

Huperzine A, an alkaloid derived from a Chinese herb, club moss (Huperzia serrata). Anisodamine, a bioactive compound from anisodamine (Anisodus tanguticus). c Nicotine, a bioactive compound from the leaves of tobacco (Nicotiana tabaccum). d Ginseoside Rb1, a bioactive compound from the roots of Vietnamese ginseng (Panax vietnamensis); MI = 20-O – d-glucopyranosyl-20(S)-protopanaxadiol, a metabolite of Rb1. e EGb 761, a standard total extract from the leaves of Ginkgo biloba. f Zeatin, a bioactive compound derived from the dried plants of Fiatoua villosa. b

associated with AD progression. Huperzine A dimers rather than monomers more potently inhibited AChE [155]. Huperzine A inhibited NMDA-induced toxicity in the cortex, synaptic plasma membranes [37], and hippocampal neurons [163]. The in vivo effects of the ginsenosidederivative MI were repeated in cultured rat cortical neurons, where MI treatment exerted axonal extension of the neurons in an in vitro cell culture system [145]. Similar to the in vivo effect of huperzine A, nicotine treatment of PC12 neuronal cells showed an increased expression of TrkA receptors [52]. Chatterjee et al. [19] studied G. biloba constituents for their impact on ion channels. They showed that bilobalide weakly inhibited NMDA receptor-activated currents and the ginkgolides A, B, C, and J, and blocked glycine-activated chloride channels in the pyramidal hippocampal neurons of the rat. At a low concentration (1 Ag/ml), G. biloba was also

found to protect PC12 cells from spontaneously formed Ahderived diffusible ligands [22]. These ligands attenuate oxidative metabolism and vesicle trafficking, alter NGFdependent ERK stimulation, and activate Rac 1 stimulation. All these events play a critical role in hippocampal longterm potentiation. Ptychopetalum olacoides, a traditional Amazonian herb, inhibited AChE activity in the frontal cortex, hippocampi, and striatal neurons of 3-month-old male Wistar rats and of 12-month-old male Swiss albino mice [129]. In a microplate assay that measured AChE activity, 23 pure alkaloids and plant extracts from 26 species of the genus Narcissus from Amaryllidaceae were tested [71]. In this study, seven alkaloids belonging to galantamine and lycorine skeletontype, as well as three Narcissus species (N. confusus, N. perez-chiscanoi, and N. Assoanus), showed AChE inhibitory activity. Zeatin, derived from Fiatoua villosa, also inhibited

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AChE activity in PC12 cells of the rat [42]. In addition, several species of Salvia (S. lavendulefolia, S. officinalis, and S. multiorrhiza) showed both AChE inhibitory and antioxidant activities [103,116,122], thus suggesting they may be useful for dementia therapy [102,105]. 4.4. Herbs tested for anti-inflammatory effects in AD models Non-steroidal anti-inflammatory drugs are known to slow down cognitive impairment in patients with mild and moderate AD. Many herbs are known for their NSAID activity. Curcumin treatment of Tg2576 mice suppressed the activity of pro-inflammatory cytokine IL-1h and the astrocyte inflammatory marker GFAP and reduced oxidative damage and plaque burdens [68]. In THP-1 and peripheral blood monocytes, Giri et al. [35] showed that curcumin treatment inhibited Ah1 – 40, and Ah1 – 42-induced the activation of EGR-1, Erk1/2, Elk-1, and the expression of cytokines (TNF-a and IL-1h), chemokines (MIP-1h, MCP1 and IL-8), chemokine receptor-5, and MAP kinase. In human peripheral blood mononuclear cells, Nelumbo nucifera, a Chinese herb, suppressed phytohemagglutinininduced activated PBMC proliferation by arresting the transition from G1 to the S phase of the cell cycle, reduced the expression of cyclin-dependent kinase-4 following PHA treatment, and suppressed the expression of IL-2, IL-4, IL10, and IFN-g [69]. Overall, these studies provide evidence of the positive effects of herbal extracts on inflammation in AD models.

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4.5. Mixtures of herbs for treating AD Table 4 lists mixtures of herbs and their uses. Besides complex extracts and single-herb pure bioactive compounds, mixtures of several herbs have been traditionally used for treating dementia. More recently, these mixtures have shown potential for AD treatment. In a recent study, Rho et al. [117] treated male Sprague – Dawley rats with yukmijihwang-tang, a traditional Chinese medicine containing six different herbs. Yukmijihwang-tang increased the expression of transthyretin and PEP-19, a neuron-specific protein that inhibits apoptosis in the hippocampus. Another traditional Chinese medicine, naoweikang, a combination of G. biloba and Panax ginseng, increased the level of AChE in the brains of Sprague – Dawley rats following an Ah1– 40 insult [70]. The Korean herbal medicine ESP-2, which contains a combination of extracts from three herbs, effectively inhibited AChE activity, alleviated scopalamine-induced memory impairment in ICR mice, and protected rat neurons from Ah or glutamate-induced neurotoxicity [53]. Two traditional Chinese and Japanese herbs (called ‘‘Kampo’’) have been studied for their effects in AD mouse models. In the ddY mouse model of AD, Kami-untan-to, a mixture of 13 herbs used in Chinese Japanese herbal medicine, inhibited thiamine-deficient, feeding-induced learning and memory impairment, increased choline acetyl transferase activity, and increased the survival rate of the mice [91]. In the same mouse model, Zhokumei-to, a

Table 4 Herbal mixtures studied as potential treatments for AD Plant extracts

Models and oxidants

Drug effects

References

Yukmijihwang-tanga (6 herbs). Chinese traditional medicine

Male Sprague – Dawley rats

Rho et al. [117]

ESP-102b, a combined extract (3 herbs); Korean herbal medicine

Male ICR mice; Mixed cortical cells from Sprague – Dawley rats; Scopalamine, Ah25 – 35, Glutamate

Naoweikangc (2 herbs). Chinese traditional medicine ‘‘Kami-untan-to’’d, Kampo medicine (13 herbs). Traditional Chinese and Japanese medicine Zhokumei-toe, a Kampo formula (9 herbs). Traditional Chinese and Japanese medicine

Male Sprague – Dawley rats; Ah1 – 40

Increased the expression of transthyretin and PEP-19, a neuron-specific protein that inhibits apoptosis Alleviated scopolamine-induced memory impairment; inhibited AChE activity in mice; protected neurons from Ah or glutamate-induced neurotoxicity Increased the level of AChE in the whole brain Increased the survival rate of mice and inhibited TD-induced learning and memory impairment and ChAT activity Repaired Ah-induced memory impairment; increased the expression of synaptophysin levels in the cortex and hippocampus

Male ddY mice; Thiamine-deficient (TD) feeding Male ddY mice; Ah25 – 35

Kang et al. [53]

Liu et al. [70] Nakagawasai et al. [91]

Tohda et al. [145]

a Yukmijihwang-tang, a mixture of 6 herbs: Rehmannia radix (19.83%), Discoreae radix (20.05%), Corni fructus (41.64%), Hoelen (1.11%), Mountain cortex radicis (21.45%), and Alismatis radix (20.92%). b ESP-102, a standardized combined extract of Angelica gigas, Saururus chinensis, and Schizandra chinensis in a 8:1:1 ratio. c Naoweikang, a mixture of 3herbs: ginseng (Ginsenosides Rg1 and Re, 35%), Ginkgo biloba (Ginkgolides, 20%), and Ginkgoflavones (16%). d Kami-untan-to, a combination of 13 types of dried medicinal herbs; daily dosage: Pinellia ternate Breit (3.0 g of tuber), Poria cocos (3.0 g of fungus), Citrus unshiu (3.0 g of peel), Phyllostachys nigra (3.0 g of stalk), Zizyphus jujuba (2.0 g of seed), Scrophularia ningpoensis (2.0 g of root), Polygala tenuifolia (2.0 g of root), Panax ginseng (2.0 g of root), Rehmanii glutinosa (2.0 g of root), Zizyphus jujuba (2.0 g of fruit), Citrus aurantium (2.0 g of immature fruit), Glycyrrhiza glabra (2.0 g of root), and Zingiber officinable (0.5 g of rhizome). e Zhokumei-to, a combination of crude drugs from 9 herbs: Prunus armeniaca (4 g), Ephedra sinica (3 g), Cinnamomum cassica (3 g), Panax ginseng (3 g), Angelica autiloba (3 g), Cnidium officinale (2 g), Zingiber officinale (2 g), Glycyrrhiza uralensis (2 g), and Gypsum fibrosum (6 g).

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mixture of nine herbs, repaired Ah-induced memory impairment and increased the expression of synaptophysin in the cortex and hippocampus [145].

5. Clinical trials on herbal drugs, using AD patients Of the 40 or so clinical trials conducted for treating cerebral insufficiency with G. biloba, only eight were judged adequate in terms of appropriateness of experimental design [3]. Among these eight, seven studies showed positive effects of EGb 761. Even in widely investigated EGb 761, only a few clinical trials specifically focused on AD patients [46,125]. We discuss some of the recent studies designed specifically to determine the effects of herbal drugs on AD patients. In a randomized, double-blind, placebo-controlled study, Le Bars [63] reported that AD patients who were administered EGb 761 (240 mg/day) for 52 weeks showed improvements in visual constructional impairment, a lesser degree of worsening in verbal deficits, and minimal improvement in both visual and verbal deficits. Similarly, AD patients with presenile and senile primary degenerative dementia, and multi-infarct dementia of mild to moderate severity showed cognitive improvements when treated with EGb 761 [54]. In contrast, AD patients (66 – 76 years of age) who were treated with EGb 761 (240 or 160 mg/day) for 24 weeks showed no improvement in vascular dementia or in age-associated memory impairment compared to AD patients treated with placebos [146]. Thus, there is some disagreement about the therapeutic effects of EGb 761 on AD patients. In another clinical study, huperzine A was administered to AD patients in 300 Ag/day doses for the first 2 – 3 weeks and then 400 Ag/day for the next 4 –12 weeks. These patients significantly improved in their cognitive, non-cognitive, and ADL functions [164]. In placebocontrolled, double-blind, randomized clinical trials, Melissa officinalis and Salvia officinalis administered to patients with mild and moderate AD significantly improved their cognitive functions [4,5]. In addition, Melissa oil (M. officinalis) and lavender oil (Lavendula officinalis), forms of aromatherapies, also improve behavioral and psychological symptoms in severe cases of dementia [10,38,44, 130,133,143]. Most of the huperzine A clinical trials have been conducted in China thus far. However, recently in the United States, to determine the effectiveness of huperzine A on AD patients, the National Institute on Aging and Alzheimer’s Disease Cooperative Study have collaboratively initiated a phase II clinical trial (http://www.ClinicalTrials.gov). More recently, the John Douglas French Foundation Institute for the Study of Aging has initiated a phase II clinical trial to determine the effects of curcumin on AD patients. Slowly but steadily, herbal drugs are entering AD clinical trials in the United States.

6. What makes herbs particularly suitable for treating AD? The three most important criteria in selecting drugs for treating AD also apply to herbal drugs: the bioavailability of herbals, the ability of herbs to cross the BBB, and the lack of adverse effects associated with the herbal treatments. In addition, herbal drugs appear to meet a fourth criterion: they result in a synergistic effect. 6.1. Bioavailability Bioavailability has been broadly defined as ‘‘absorption and utilization of a nutrient’’ [61]. Herbal extracts, once consumed, must penetrate the intestinal barrier and enter the systemic circulation system. There is growing evidence of the bioavailability and bioefficacy of plant flavonoids (flava-based herbs), but that the bioavailability of herbs varies considerably across different types of flavonoids and that the most abundantly consumed polyphenol is not necessarily the most readily bioavailable [78,154]. According to these studies, isoflavones (e.g., soybeans, grape seeds, and red clover) and gallic acid (walnuts) are the most readily bioavailable, followed by catechins (green and black tea), flavones (cocoa, chocolate, red wine), and quercetin glucosides (onion, apple, tea, broccoli, red wine, and ginkgo). The least absorbed polyphenols are proanthocyanidins (e.g., pine bark, grape seeds, cranberries), galloylated tea catechins, and anthocyanins (black currant, elderberries, red grapes, strawberries, blueberries). The extent to which the human colon can absorb plant drugs depends on the metabolic activity of microflora in the intestine and hepatic activity. There is considerable personto-person variation in these processes [154]. 6.2. The blood – brain barrier Herbal extracts, once administered, must pass through the BBB to be effective in the central nervous system. The BBB is made of a dense layer of endothelial cells that create a barrier between the blood and brain parenchyma, which primarily consists of astrocytes and microglia. In the BBB, a layer of endothelial cells is different from a layer of endothelial cells in other tissues. The layer of endothelial cells in the BBB has a low density of pinocytotic vessels and contains brain microvessels and specific efflux transporters that selectively control the flow of molecules from cerebrovascular circulation into the brain [160]. In addition, the BBB expresses numerous types of efflux transporters, such as P-glycoprotein, multi-drug resistance associated protein, and monocarboxylic acid transporters. To gain entry into different parts of the brain, flavonoids exhibit either stimulatory or inhibitory interactions with one or many of these transporters directly or indirectly [160]. For example, quercetin and kaempferol, both bioactive compounds found in ginkgo, stimulate P-glycoprotein transporters, while resveratrol, found in grape seeds, inhibits

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them. The extent to which an herbal drug can readily penetrate the BBB determines its bioavailability. The herbal drug must interact with specific brain cells or must be able to flow through intercellular space in order to manifest its desired effects. 6.3. Toxic and adverse drug effects and drug-drug interaction Few clinical and toxicological studies have been conducted to determine the adverse effects of herbal treatments even in the most widely used herbal treatments. Assessing the adverse effects of herbal treatments is affected by the conditions in which an herb is administered. For example, certain herbal drugs taken before surgery may adversely affect perioperative patient care [9]. Eight commonly used herbs in the United States have been identified as having adverse perioperative affects: echinacea, ephedra, garlic, ginkgo, ginseng, kava, St. John’s Wort, and valerian. Of these, as discussed, garlic, ginkgo, and ginseng may have a role in treating AD. The main concern with garlic, ginkgo, and ginseng is that they can inhibit platelet formation, activate other platelet inhibitors, and prevent blood clotting in humans. Thus, 2 –7 days prior to surgery, patients are counseled not to consume these herbs [9]. In clinical trials investigating the effects of ginkgo treatment on AD, ginkgo was found not to have any serious adverse effects [77], but there were non-serious side effects, including mild skin allergies, gastro-intestinal upset, and headaches. In a recent clinical study of 50 AD patients, ginkgo treatments were found generally safe. Mortality rates of AD patients treated with ginkgo were no different from mortality rates of patients treated with a placebo [146]. However, the AD patients treated with ginkgo reported marginal adverse effects, such as dizziness, nervousness, and headaches. In a clinical trial to determine the effects of M. officinalis on AD patients, AD patients treated with this herb exhibited mild but relatively stronger adverse effects in terms of vomiting, dizziness, wheezing, abdominal pain, and nausea, than did AD patients treated with a placebo [5]. Studies have also found relatively fewer adverse side effects with huperzine A (an acetylcholinesterase inhibitor) compared to commercial cholinesterase inhibitors [162]. Although these studies suggest that herbal treatments may result in only mild discomfiture, the potential toxicological adverse effects of the herb need to be further assessed. 6.4. Synergistic interactions A synergy is the interaction of two or more agents or forces, the combined effect of which is greater than the sum of their individual effects. Synergistic interactions can occur in a single herb due to the presence of dozens of bioactive compounds. For example, G. biloba possesses several ginkgolides and bilobalides. Eastern herbal medicines, including traditional Chinese and Indian Ayurveda medical

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approaches, are based on the synergistic interactions among constituent bioactive compounds. Table 4 lists synergistic effects flowing from different combinations of herbs. Recently, in a study of AD patients treated with phytomedicines, Williamson [153] provided many examples of synergistic interactions that result in both positive and adverse effects. It is difficult to assess synergy in an herbal treatment because of the large number of constituent herbs or active compounds in a single herb. Perhaps one of the greatest challenges in determining the efficacy of herbal treatments is to prove the existence of synergy. Increasing numbers of studies are employing more advanced tools and techniques to unravel the secrets of cellular pathways in disease progression and pathology of AD patients.

7. Concluding remarks Tremendous progress has been made in developing strategies to treat AD. Some of these strategies include anti-inflammatory, anti-amyloid, anti-oxidant, and pro-cholinergic medicines. A successful application of a therapeutic strategy in clinical trials requires a clearer understanding of both the adverse and beneficial effects of the drugs. Currently available FDA-approved drugs treat AD symptomatically and provide temporary relief from dementia. However, these drugs are frequently associated with adverse drug effects and do not cure the disease by modifying its pathology. There remains an urgent need for developing alternative approaches to AD therapeutics. Recently, herbal drugs have been systematically tested in animal and cell models of AD and, to lesser extent, in clinical trials. Herbal drugs are relatively less toxic, can readily cross the BBB, and are bioavailable to exert multiple synergistic effects, including improved cognitive and cholinergic functions. Thus, herbal drugs appear to be a promising alternative medicine in treating AD patients. However, to determine their adverse effects in AD patients, we need further research on each herb in terms of pathology and phenotypic behavior in well-designed clinical trails.

Acknowledgments The authors thank Sandra Oster, Neurological Sciences Institute, Oregon Health and Science University, for editing the manuscript. This research was supported, in part, by the American Federation for Aging Research and NIH #AG22643.

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