312 CNS & Neurological Disorders - Drug Targets, 2014, 13, 312-321
A Molecular Bridge: Connecting Type 2 Diabetes and Alzheimer’s Disease
Firoz Ahmed
1,§, Juned Asghar Ansari
2, Zahid Eqbal Ansari
3, Qamre Alam
4, Siew Hua Gan
5, Mohammad A. Kamal
4and Ejaj Ahmad
*,11Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, Uttar Pradesh, India
2Department of Radiology, Brij Lal Hospital & Research Centre, Haldwani, Uttarakhand, India
3Department of Community Medicine, Jawaharlal Nehru Medical College, Aligarh, Uttar Pradesh, India
4King Fahd Medical Research Center, King Abdulaziz University, Jeddah, Saudi Arabia
5Human Genome Centre, School of Medical Sciences, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan, Malaysia
Abstract: Type 2 diabetes (T2D) and Alzheimer's disease (AD) are complex diseases commonly associated with aging.
Accumulating evidence indicates a connection between these two diseases at the molecular level. Much of what we currently know about T2D and AD is derived from in vivo and in vitro studies. However, further research and characterization of molecules is necessary to establish a strong connection between T2D and AD. In silico studies play a major role in finding non-evident patterns of gene expression and gene network connectivity. In this review, we give a brief introduction to T2D and AD and then describe the risk factors and molecules that are commonly associated with these diseases. Finally, we discuss the future directions and applications of bioinformatics that can provide greater insight into the relationship between these two diseases. Analysis and integration of high-throughput data on genomics, transcriptomics, proteomics and metabolomics from normal and disease tissues would be very useful to improve our understanding of the mechanism behind disease initiation and the connection between these two diseases. We encourage researchers to use bioinformatics approaches to identify genes and their regulatory pathways that are commonly affected in T2D and AD, as these genes and pathways could be potential biomarkers and targets for disease treatment.
Keywords: Type 2 diabetes, Alzheimer's disease, bioinformatics, high-throughput sequencing data.
INTRODUCTION
Type 2 Diabetes (T2D) and Alzheimer's disease (AD) are the most common diseases associated with aging. Several studies have indicated a connection between T2D and AD [1-3]. In this review, we focus on molecules that are commonly affected in T2D and AD. We also discuss recent trends in these areas, and the application of bioinformatics approaches that can help to better understand disease initiation and progression and the connection between these two diseases at the molecular level. While doing so, it is necessary to give a brief introduction to T2D and AD.
TYPE 2 DIABETES
Prevalence of disease: Diabetes is a condition of impaired glucose metabolism in which the body’s cells are unable to take up glucose properly, which leads to elevated levels of fasting glucose and postprandial serum glucose [1].
Insulin secreted by the β-cells of the pancreas stimulates
*Address correspondence to this author at the Interdisciplinary Biotechnology Unit, Aligarh Muslim University, Aligarh, Uttar Pradesh, India;
Tel: +91-571-2720388; Fax: +91-571-2721776;
E-mail: [email protected]
§Current address: Bioinformatics Laboratory, Plant Biology Division, The Samuel Roberts Noble Foundation, Ardmore, Oklahoma, United States of America.
glucose uptake in cells. Diabetes has been classified into two major classes based upon the availability of insulin. (1) Type 1 Diabetes (T1D), in which insulin is not secreted by the pancreas due to the destruction of the insulin-producing β- cells, and which is thus termed insulin-dependent diabetes [4], and (2) T2D, in which insulin is secreted by the β-cells but fails to stimulate the cellular uptake of glucose, and which is thus termed insulin-independent diabetes [4].
According to the World Health Organization (WHO;
http://www.who.int/), 346 million people worldwide had diabetes in 2012, which corresponds to ~5% of the total population [5]. This number is increasing at an alarming rate, particularly among adults. According to the International Diabetes Federation (IDF; http://www.idf.org/diabetesatlas/) this number will rise to 552 million by 2030. As per the findings of the WHO, diabetes is among the top ten leading causes of death in the world, creating a heavy health and economic burden on society.
Glucose: Glucose is the primary source of energy for cells and its proper supply is necessary for their survival and functions [6]. A critically low level of glucose (hypoglycemia) causes cell death while higher levels (hyperglycemia) for a prolonged period can damage small blood vessels, leading to several complications, including organ failure [7, 8]. Therefore, to regulate proper levels of glucose in the body, two important hormones work with great coordination: (1) insulin, which is secreted by the β- cells of the pancreas, and (2) glucagon, which is an insulin
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antagonist that is secreted by the pancreatic α-cells. In addition, these hormones work in coordination with the pancreas, liver, skeletal and cardiac muscle and adipose tissues and with other hormones to carefully regulate glucose homeostasis [9]. Glucose is transported from plasma to the cell via glucose transporter proteins (GLUTs), which are present on cell membranes [10, 11]. There are different classes of GLUTs and each has specific physiological functions, tissue distribution and affinity for glucose [12].
Release of insulin: After intake of a meal, the glucose level in the plasma becomes higher than basal levels, which leads to transport of glucose to the pancreatic β-cell through GLUT2 [13]. After phosphorylation of the glucose, it is oxidized through the tricarboxylic acid cycle and the electron transport system to eventually produce ATP, which leads to an increased cellular ATP/ADP ratio [13]. Once the ATP/ADP ratio becomes high, potassium channels (K+ATP- channels) are closed, and the cell becomes depolarized, which leads to opening of voltage-dependent Ca2+-channels (VDCC) [14]. Increased calcium concentration inside the cell stimulates the fusion of insulin-containing vesicles with the plasma membrane, thus releasing insulin outside the cell.
Insulin and glucose metabolism: GLUT4 is an insulin- sensitive glucose transporter expressed in the liver, skeletal and cardiac muscle and adipose tissue, and it is sequestered in the internal cellular membrane [15]. When insulin binds to its receptor on the surface of the cell, it triggers downstream signal transduction pathways that stimulate the exocytosis of the GLUT4 transporter on the cell surface and facilitates glucose uptake, reducing plasma glucose. Endocytosis of GLUT4 reduces the concentration of the transporter on the plasma membrane in the case of low levels of insulin. After uptake into the cell, glucose undergoes phosphorylation. In the case of liver and muscle cells, it is either oxidized to generate ATP, or is stored as glycogen, the secondary long- term energy store, while glucose is converted to lipids for energy storage in adipose tissue. In the liver, insulin inhibits glucose formation by inhibiting gluconeogenesis (the formation of glucose from a non-carbohydrate source), and glycogenolysis (hydrolysis of glycogen to glucose). It inhibits lipolysis and fatty acid release in adipose cells and promotes the formation of triglycerides and accumulation of fat. Insulin also influences the expression of several genes.
When glucose levels in plasma become lower than basal levels, e.g., between meals or during exercise, this stimulates the release of glucagon by the α-cells of the pancreas [16].
Glucagon induces liver cells to convert stored glycogen into glucose and release it into the plasma.
ALZHEIMER'S DISEASE
The pathogenesis of AD is associated with the formation of extracellular amyloid beta (Aβ) plaques and intracellular neurofibrillary tangles (NFTs) in the brain that lead to cognitive decline. AD is the most common type of dementia, in which neurons die or become non-functional resulting in decreased cognitive thinking, impaired memory, altered behavior, and a decrease in the ability to perform everyday activities, eventually leading to death. AD International (http://www.alz.co.uk/) estimates that there were 35.6 million people living with dementia in 2010, and estimates that there will be 65.7 million people with dementia by 2030
worldwide. According to the Alzheimer's Association (http://www.alz.org/downloads/facts_figures_2013.pdf), in the USA alone, AD affected more than 5.2 million people in 2013, most of whom were aged 65 or older, making AD the sixth leading cause of death. Although the causes of AD are not fully understood, there are several hypotheses regarding the etiology of AD: (1) the cholinergic hypothesis, which states that AD is caused by abnormalities in the cholinergic system, which leads to reduced synthesis of the neurotransmitter acetylcholine [17, 18]. (2) The amyloid hypothesis, which proposes that the main cause of AD is the processing of amyloid precursor protein (APP) to form Aβ that aggregates and forms plaques in brain tissue [19].
However, recent clinical-trials of drugs targeting Aβ in Alzheimer’s patients reduced Aβ production but failed to arrest disease progression, creating a new controversy regarding the amyloid hypothesis [20-23]. (3) The tau hypothesis, which postulates that hyper-phosphorylation of tau protein initiates self-assembly resulting in the formation of NFTs inside nerve cell bodies, which ultimately leads to the disintegration of microtubules and impairs the neuron's transport system [24, 25].
Aβ: Aβ is the pathological hallmark of AD and the major component of senile plaques. It is a 39-42 amino acid peptide fragment generated from the amyloid precursor protein (APP), which spans the outer membrane of nerve cells [26, 27]. APP is a synaptic receptor and its full-length protein is necessary for proper migration and positioning of neuronal precursors during brain development in mammals [28]. APP also promotes synapse formation and neuronal connections [29, 30]. In addition, APP can undergo proteolysis through either one of two pathways: (1) non- amyloidogenic and (2) amyloidogenic [31]. Non- amyloidogenic cleavage of APP by α-secretase produces soluble APPα (sAPPα) and C83; subsequent cleavage of C83 by γ -secretase produces p3 and the APP intracellular domain (AICD) [31]. The amyloidogenic cleavage of APP by β- secretase and γ-secretase generates sAPPβ, Aβ and ACID [31]. Aβ accumulates in AD to form insoluble fibrils of packed β-pleated sheets [26]. This fibril forms senile plaques on the cell surface and causes impaired synaptic function of cells and reduces cell survival. The proteolytic cleavage of APP is the normal physiological process and is dominated by the non-amyloidogenic pathway, which produces sAPPα (regulator of cell proliferation, survival, and migration) and inhibits Aβ production [32]. Initially Aβ was considered a toxic product; however, later studies showed that it is a regulator of the expression of ion channels and controls the excitability and survival of neuronal cells [33, 34]. The physiological levels of Aβ are very crucial and are controlled by its production, degradation and clearance, and only excessive amounts are considered part of AD pathology [31].
In addition, AICD moves into the nucleus and regulates gene expression and calcium signaling. The role of numerous other peptides generated during the proteolytic process is not clearly understood [31, 32]. Studies have shown that mutations in genes encoding APP, presenilin 1 or presenilin 2 are associated with abnormal Aβ accumulation and familial AD. However, the occurrence of familial AD is rare; over 90% of known AD is sporadic [27]. Impaired insulin signaling in T2D also affects the expression of several genes and metabolic processes that enhance the production of Aβ
and cause the progression of AD. Although Aβ plaques are
considered as one of the hallmarks of AD pathogenesis, normal people without dementia have also been reported to exhibit elevated Aβ levels [35].
NFTs: Tau is a soluble protein expressed in neurons and is associated with neuronal microtubules. It interacts with tubulin to stabilize the axonal microtubules. Tau protein activity is regulated by kinase-mediated phosphorylation, which leads to disruption of microtubule organization.
Abnormally high phosphorylation of the tau protein leads to a decrease in the extent of the microtubule network and self- assembly of tau, resulting in neurofibrillary tangles, which aggregate in the cell bodies and proximal dendrites, resulting in pathogenesis of AD [36]. Hyper-phosphorylation of tau also occurs due to a decrease in protein phosphatase-2A (PP- 2A) activity, which regulates the normal phosphorylation/
dephosphorylation ratio [37].
CONNECTION BETWEEN T2D AND AD
High blood sugar in T2D affects every tissue and organ of the body, causing other complications, such as cardio- vascular disease, nephropathy, retinopathy, and neuropathy;
however, unlike other complications, the relationship between T2D and AD is not yet fully understood [1]. Studies have indicated that people with T2D have a greater risk of developing AD [38, 39]. The higher risk may be because the high blood sugar levels in T2D damage blood vessels in the brain, which when combined with impaired glucose metabolism leads to mild cognitive impairment (MCI) and cognitive decline (CD), which are hallmarks of AD. In addition, insulin also plays an important role in the biosynthesis of neurotransmitters, which are essential, both for communication between neurons and for memory.
However, whether AD can also influence the progression of insulin resistance or T2D is not known. To understand the connection between T2D and AD, it is imperative to find the
biochemical factors that are involved in the progression of both T2D and AD. Greater insight into how T2D and AD are connected may eventually lead to new strategies to treat both of these diseases. Important molecular events connecting T2D and AD are listed in Table 1, and their implications for disease pathogenesis are discussed.
SHARED RISK FACTORS
Aging: Age progression leads to enhanced oxidative and endoplasmic reticulum (ER) stresses in the cell and the chances of T2D and AD initiation become higher [54-56].
Mitochondria are the powerhouses of the cell; through oxidative phosphorylation they produce ATP and reactive oxygen species (ROS) [57]. The abnormally high levels of ROS due to over-activity of pancreatic and neuronal cells leads to oxidative damage of cellular organelles, enzymes and DNA, and also affects a wide range of metabolic functions which may lead to T2D and AD [1, 55, 57, 58].
The ER plays important roles in protein folding, cholesterol biosynthesis and Ca2+ balance. Cellular stress creates obstacles to protein folding and subsequently causes an accumulation of misfolded or unfolded proteins in the ER lumen, leading to ER stress. ER stress triggers several pathways to manage the stress, and dysfunction of these cellular defense mechanisms is reported to be involved in the pathogenesis of T2D and AD [59, 60].
Obesity: Health problems associated with obesity are numerous; the most devastating may be T2D and AD [43, 61]. Excess caloric intake combined with a sedentary lifestyle results in the storage of energy in the adipose tissue, which makes people obese [62]. Adipocytes secrete different proteins, cytokines (leptin, adiponectin, omentin, visfatin, resistin TNF-α, IL-6 and RBP4) and hormones to regulate glucose homeostasis [62]. The adipose tissue of obese people releases increased amounts of fatty-acids, glycerol,
Table 1. List of Common Molecular Events Reported in T2D and AD
Molecular Events Description References
Aging Cells pass through oxidative and ER stress conditions which disrupt the normal metabolic processes resulting in
T2D and AD pathogenesis. [40, 41]
Obesity
Obesity changes the normal lipid profile and induces ER stress in the liver cells that results in a reduction of protein synthesis and activates the expression of genes responsible for gluconeogenesis through CRTC2/CREB. Obesity
increases insulin resistance and the level of Aβ in the brain. [42-44]
Insulin resistance Insulin resistance results in uncontrolled blood glucose that leads to T2D. T2D damages blood vessels in the brain
and affects the ability of brain cells to use glucose, thus increasing the risk of developing AD. [45]
Amyloid formation Amyloid fibrils in the brain are a hallmark of AD and are also reported in the islets of the pancreas in T2D.
Misfolded Aβ and IAPP proteins are deposited, which leads to the formation of amyloid plaques in AD and T2D, respectively.
[46, 47]
GSK3
GSK3 is a kinase that plays an important role in the regulation of glucose metabolism. It also regulates Aβ production and tau phosphorylation in the brain. Abnormal regulation of GSK3 changes cell metabolism and
contributes to T2D and AD pathogenesis. [48]
CDK5
CDK5 is a multi-faceted kinase that works with its regulatory protein p35 to form the CDK5/p35 complex and promote pancreatic β-cell survival via Fak-Akt signaling pathways. CDK5 also modulates the processing of APP and Aβ generation. Oxidative stress leads to cleavage of p35 to produce p25, which causes constitutive activation of CDK5, which in turn hyper-phosphorylates the tau protein and leads to NFT formation in the brain.
[49, 50]
BchE BchE hydrolyzes acetylcholine, an anti-inflammatory molecule, which results in acetylcholine depletion, which in
turn may trigger inflammation and promote the pathogenesis of T2D and AD. [51-53]
BchE: butyrylcholinesterase; CDK: cyclin-dependent protein kinase 5; ER: endoplasmic reticulum; GSK3: glycogen synthase kinase 3.
hormones, cytokines and inflammatory molecules, which induce insulin resistance and impair β -cell function, ultimately leading to T2D [61]. The inflammatory molecules also induce inflammation and cause neuron damage, contributing to the progression of AD [48]. Moreover, research shows a strong correlation between body mass index and high levels of beta-amyloid in the blood [44, 63].
Although the precise molecular mechanisms of how obesity leads to AD remain to be elucidated [64], studies indicate a role for dysregulated lipid metabolism.
Insulin resistance: Insulin resistance is a condition in which cells or tissues fail to respond to the physiological level of insulin. To maintain glucose homeostasis, β-cells keep producing more insulin, resulting in β-cell deterioration and ultimately leading to T2D. Insulin resistance may arise due to abnormality in insulin signal transduction, glucose metabolism, inflammation or lipid metabolism. High levels of insulin reduce blood sugar levels and stop the burning of fats for fuel, triggering a switch from the use of stored fat to the use of muscle protein as an energy source. This leads to an increase in stored fat and loss of lean muscle. High insulin levels can also cause inflammation of blood vessels and tissue-damage. Insulin resistance affects lipid metabolism and activates multiple pro-ceramide genes, causing toxicity and endoplasmic reticulum stress in the brain and an increase in AD severity [65].
SHARED AMYLOID FORMATION
Amyloid deposition within the islets of Langerhans is also considered a common pathological characteristic of T2D [66], similar to the Aβ deposited in the brains of AD patients. An aggregate of islet amyloid polypeptide (IAPP), or amylin, a 37-residue peptide hormone expressed in β- cells, has been identified as a component of pancreatic amyloid deposits [46]. After processing of the 89-residue proIAPP, IAPP forms and is co-secreted with insulin in a specific ratio under normal physiological conditions. It binds with a receptor present on postrema neurons and controls glucose homeostasis by reducing food intake, postprandial glucagon secretion, and slowing gastric emptying [67, 68].
Recent findings showed that at low concentrations, monomeric human IAPP (hIAPP) binds to the lipid bilayer interface in a α-helical form and causes membrane expansion in β-cells [69]. However, at high concentrations, hIAPP forms β-aggregates, which exit the bilayer and form a large fibril, which interacts electrostatically and hydrophobically with the lipid bilayer and makes the bilayer leaky [69]. The process of amyloid formation by Aβ and IAPP proteins occurs through a nucleation-dependent mechanism which takes place in three phases [46, 47]: (a) the nucleation phase, a rate limiting step where monomeric peptides form unusual associations and form an oligomeric nucleus; (b) the growth phase, in which the oligomeric nucleus rapidly grows in size to form a fibril; and (c) the steady state phase, in which the fibril mass reaches steady-state, and an equilibrium is established between the fibril and the monomer peptides.
The nucleation phase is largely dependent upon protein concentration, temperature and pH of the cell environment, and time taken for its completion may take anywhere from a several days to years [46, 47, 70]. The nucleation phase is the initial and slowest step in amyloid formation, thus
perturbation of this is an important strategy to prevent amyloid formation and disease onset [47]. It is also important to know the critical concentration of Aβ and IAPP below which no nucleus and fibril formation occurs [47].
Aβ and hIAPP have very similar biophysical and biochemical properties [46]. Recent evidence showed that Aβ can directly activate the hIAPP receptor (AMY3) and trigger several signal transduction pathways through Protein kinase A (PKA), Mitogen-activated protein kinase (MAPK), Akt and cFos [2]. This raises the cytosolic cAMP and Ca2+
concentrations which lead to cytotoxicity, including neurotoxicity [2]. The study indicates that AMY3 could serve as a useful therapeutic target to treat Aβ toxicity and AD.
SHARED MOLECULAR FACTORS
Glycogen synthase kinase 3 (GSK3): GSK3 is a serine/threonine kinase involved in the regulation of glucose metabolism through phosphorylation of glycogen synthase (GS). Activated GS converts excess glucose molecules to glycogen polymers (glycogen synthesis) in skeletal muscle.
GS is activated through dephosphorylation due to inactive GSK3. Accumulating evidence points to GSK3 as a key kinase that interacts with different proteins involved in the etiology and progression of T2D and AD pathogenesis [71].
Mammalian GSK3 has two isoforms, GSK3α and GSK3β, and its activity is regulated through different pathways including insulin signaling [72]. Insulin binds to its receptor and activates the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway which phosphorylates GSK3 on serine residues (at position 21 in GSK3α or at position 9 in GSK3β), leading to inhibition of GSK3 activity [73] (Fig.
1). However, phosphorylation of tyrosine 216 in GSK3β or tyrosine 279 in GSK3α through different mechanisms enhances GSK3 activity. The study [73] showed that a decrease in the insulin-induced phosphorylation of GSK3 is found in the skeletal muscle of diabetic humans and the cardiac muscle of diabetic rats and that this leads to enhanced activity of GSK3 [74, 75]. Therefore, insulin stimulates glycogen synthesis in skeletal muscle by inactivating GSK3, leading to a decrease in glucose to an optimum level.
GSK3 can phosphorylate over 50 known substrates and regulate several critical metabolic processes, including gene expression, cell cycle, cell migration, apoptosis and inflammation [48]. GSK3α and GSK3β are tau protein kinases and hyper-phosphorylation at many sites including Thr-231 of tau leads to aggregation and formation of neurofibrillary tangles resulting in progression of AD [76- 79] (Fig. 1). Activated GSK3 also promotes an increase in the splicing of exon 10 in tau mRNA, producing aberrant tau protein [80]. Studies show that GSK3α regulates the processing of APP, can increase Aβ production and contributes to AD pathogenesis [79, 81, 82]. Inflammation is associated with both AD and T2D pathogenesis and is also promoted by GSK3 [48]. The level of different pro- inflammatory cytokines, such as IL-6 and TNF-α, is enhanced through activated GSK3 in AD and T2D [83-86].
Elevated levels of TNF-α impair insulin signaling in cells, and its reduction in obese rodents leads to a significant
increase in insulin-induced glucose uptake [83, 84]. Various studies have suggested that an inhibitor of GSK3 would provide an important therapeutic strategy to treat both T2D and AD by inducing glycogen synthesis and reducing Aβ production, tau phosphorylation and inflammation in both pancreatic and brain cells [71, 87]. Several findings showed that inhibiting the activity of GSK3 using lithium resulted in a reduction of tau phosphorylation and Aβ production, which thereby reduced the formation of both amyloid plaques and neurofibrillary tangles and protected cells from neurotoxicity, indicating a novel point of intervention for AD [81, 88, 89]. Several compelling lines of evidence indicate that long-term use of non-steroidal anti- inflammatory drugs (NSAIDs) also minimizes the risk for developing AD [86].
Cyclin-dependent Protein Kinase 5 (CDK5): There are some specialized genes commonly expressed in both endocrine cells of pancreatic islets and cells of neuronal lineages. The over-activation of CDK5 in the nervous system is also associated with initiation and progression in the
pathogenesis of AD [90]. Protein p35 is the activator of CDK5, however, in the neurons of AD patients, a truncated form, p25, was reported to accumulate, leading to a constitutive increase in CDK5 kinase activity [90]. A change in the cellular localization and substrate specificity was also noticed for the p25/CDK5 pathway, leading to tau hyper- phosphorylation and collapse of the cytoskeleton [90].
The study showed that both p35 and CDK5 genes are expressed in the β-cells of the pancreas and form an active p35/CDK5 complex [91, 92]. An increase in the concentration of extracellular glucose leads to increases in the mRNA and protein levels of p35, resulting in elevated p35/CDK5 kinase activity. Signaling through p35/CDK5 activity affects the promoter region of the insulin gene and increases its transcription [92]. The study [92] indicates that tight regulation is required of the p35/CDK5 complex in the β-cells under extracellular concentration of glucose. Elevated glucose levels in T2D patients leads to over-stimulation of the p35/CDK5 pathway, resulting in pathological abnormalities of IAPP deposition similar to Aβ in AD [92].
Fig. (1). The insulin signal transduction pathway and GSK3 mediate metabolic effects. Binding of insulin to its receptor initiates a cascade of kinase activity through PDK1 and Akt. GSK3 becomes inactive after phosphorylation, resulting in GS activation and synthesis of glycogen in the liver cells. Insulin resistance causes inhibition of downstream signal transduction and GSK3 remains in its active form, which hyper-phosphorylates several substrates, including tau protein, in nerve cells. Hyper-phosphorylation of tau destabilizes the microtubules and induces neurofibrillary tangle formation that leads to AD. IR (insulin receptor), IRS (insulin receptor substrate), PI3-kinase (phosphoinositol phosphate 3-kinase), PDK1 (3-phosphoinositide-dependent protein kinase-1), GS (glycogen synthase).
Cell membrane
P P P P
P
P IRS PI3K
PDK1
Akt
GSK3 GSK3
P
inactive active
GS P
inactive
GS
active
Glucose-6-P Glycogen
tau stabilizes microtubules
tau hyper- phosphorylation
P
P P
P
microtubules depolymerize
P P P P P
P P P P P
aggregates
P P
filaments
tangles insulin
IR
P
P
P
Butyrylcholinesterase (BchE): BchE is the cholinesterase enzyme that is synthesized in the liver and hydrolyzes different choline esters in the plasma. It shows high sequence and structural similarity with acetylcholinesterase (AchE), a neuronal cholinesterase [93].
AchE is mainly involved in the rapid hydrolysis of the neurotransmitter acetylcholine at cholinergic synapses. Loss of AchE function causes muscle paralysis, seizure and death by asphyxiation. BchE is the non-specific cholinesterase that preferentially hydrolyzes butyrylcholine, but it can also act on acetylcholine and other ester-containing drugs and on neuromuscular blocking agents used in anesthesia [94].
Therefore, BchE detoxifies anticholinesterase compounds before they reach AchE [93]. Studies indicate that the variant form of BchE acts synergistically with apolipoprotein E- epsilon 4 in the pathogenesis of AD [95]. A role for BchE is also indicated in the pathogenesis of T2D and metabolic syndrome [96-99].
COMMON MEDICINE FOR TREATMENTS
Thiazolidinediones (TZDs): TZDs are used in the treatment of T2D because they reduce insulin resistance and free fatty acids and thus lower the blood glucose levels.
TZDs activate the nuclear hormone receptor PPARγ (peroxisome proliferator-activated receptors gamma) to form a dimer with another nuclear receptor, RXR. The PPARγ- RXR complex binds to the hormone response elements upstream of the target gene and modulates the expression of various genes involved in glucose and lipid metabolism.
Recent evidence shows that TZDs increase the expression of apolipoprotein E (apoE) in the brain, resulting in enhanced clearance of the soluble forms of Aβ and reduced incidence of AD [100-102].
CONCLUSION, CURRENT CHALLENGES, AND PERSPECTIVES
T2D and AD are very complex diseases that occur due to abnormalities in metabolic pathways. Accumulating evidence points to a link between T2D and AD at the molecular level. Several specialized genes are expressed in both β-cells and neurons, with examples being the temporary expression of insulin and the transcription factor IDX-1/IPF1 in the developing brain [103, 104] and the permanent expression of ATP-sensitive potassium channels [105, 106]
and Isl-1 [107]. Despite several discoveries of genes associated with T2D and AD, we are still far from having a complete understanding of these genes and their regulatory networks. We are still trying to understand the molecular mechanism by which T2D influences the risk of developing AD; whether it is the effects of abnormal glucose and lipid metabolism that cause AD, or that T2D associated disease genes are directly involved in AD development. It is also not clear whether AD can also influence the risk for T2D pathogenesis.
Bioinformatics and systems biology-based approaches are very promising tools to address these challenging questions, and ultimately help to decipher the link between T2D and AD [108, 109]. Biological systems are very complex and the fate of each cell is precisely regulated through a complex molecular network in response to
environmental conditions. To understand the behavior and functions of cells, biological data and computational tools are required. The advent of high-throughput sequencing techniques has led to the generation of huge amounts of data on genes (genomics), transcripts (transcriptomics), proteins (proteomics) and metabolites (metabolomics) at an extraordinary scale and speed from normal and disease- affected cells/tissues. These molecular and clinical data contain important information that needs to be analyzed using bioinformatics approaches to understand how genes and molecules interact to regulate various biological processes and disease traits.
RNA-seq data provide transcriptome information with deep coverage and higher base level resolution [110, 111].
By comparing the transcriptome of normal and diseased cells/tissues, differentially expressed genes associated with a disease can be identified. The expression data can also be useful for the discovery of new genes and pathways through the reconstruction of gene regulatory networks [112, 113]. In addition, it would also be useful to find common gene expression patterns in T2D and AD using clustering methods. This will help to find not only disease-associated genes and molecules but also help to discover intricate pathways responsible for the etiology and connectivity of T2D and AD [114].
Bioinformatics is an emerging field but maintaining and analyzing vast multi-dimensional data for meaningful purposes poses a challenge. Relevant data are available on several public databases, such as the Pancreatic Expression Database (PED; http://www.pancreasexpression.org/), which is a repository for pancreatic-derived omics data [115];
High-throughput data from microarray analyses and next- generation sequencing are available through Gene Expression Omnibus (http://www.ncbi.nlm.nih.gov/geo/) and ArrayExpress (http://www.ebi.ac.uk/arrayexpress/).
Moreover, various software applications are publicly available for data analysis. Galaxy (http://galaxyproject.
org/), DAVID (http://david.niaid.nih.gov), R & Biocon- ductor (www.r-project.org; www.bioconductor.org) are helpful for genome-wide identification of protein binding sites and histone modification, for discovery of different- ially expressed genes in a disease condition, and gene annotation for knowledge discovery [116]. Cytoscape (http://www.cyto scape.org) is useful for visualizing molecular interaction networks and biological pathways and for integrating these networks with annotations and gene expression profiles [117, 118]. Several hundred candidate genes with disease associ-ation can be further prioritized by tools such as ToppGene (http://toppgene.
cchmc.org/) [119] and ENDEAVOUR: (http://www.esat.
kuleuven.be/endeavou rweb [108, 120, 121]. Networks constructed using metabolomics and proteomics data have discovered new biomarkers for T2D, and similar approaches can also be used to discover more biomarkers for AD [122]. Genetic as well as environmental factors also influence disease outcome. Genetic variation in the form of single-nucleotide polymorphisms (SNPs) associated with T2D or AD can be identified using genome-wide association study (GWAS) approaches [123, 124]. Thousands of SNPs associated with T2D and AD, in addition to those associated with various other human
traits and diseases, are in GWAS Central (http://www.gwa
scentral.org/) and GWASdb (http://jjwanglab.org/gwasdb) [125].
Discovery of RNA interference (RNAi) mechanisms in eukaryotes has completely changed our way of thinking about gene regulation, and has also revolutionized technical advancements for the study of functional genomics [126]. RNAi triggers the formation of small RNA molecules, microRNAs (miRNAs) or small interfering RNAs (siRNAs), which bind to their target mRNAs through base-pairing interaction and lead to gene silencing [127, 128]. miRNA is an important regulator of gene expression and also considered as a promising biomarker to detect several pathological conditions [127, 129-131]. A study reported that over-expression of miRNA-146a in human brain cells with AD leads to a decrease in the expression of several inflammatory genes, including complement factor-H (CFH), the interleukin-1 receptor associated kinase-1 (IRAK-1), and tetraspanin-12 (TSPAN12) [132]. Several miRNAs involved in the pathogenesis of T2D and AD have been discovered and compiled into a database http://cmbi.bjmu.edu.
cn/hmdd [133, 134]. Moreover, eukaryotic mRNA contains several functional elements that influence gene expression [129]. The disease might also result from mutations in the coding regions or functional elements of mRNA, which may lead to expression of abnormal protein and/or an uncontrolled rate of expression. Mutations in the functional elements, such as miRNA binding sites or polyA signals cause aberrant gene structure and expression [135, 136]. Dicer cleavage sites in pre- miRNA can be predicted using PHDcleav [137]. This web- based tool is available at http://www.imtech.res.in/ raghava/
phdcleav/ and would be very useful in genome-wide investigat- ions of miRNA-related polymorphisms (SNPs) and their consequences at Dicer processing sites [137]. Furthermore, siRNA has been emerging as a potential therapeutic intervention to treat infectious and metabolic diseases by modulating the expression of disease-causing genes [138-140]. Gene mutations causing disease can also be selectively silenced by siRNAs
designed using the desiRm tool
(http://www.imtech.res.in/raghava/desirm/) [141].
Moreover, advancements in molecular imaging techniques such as positron emission tomography (PET) have made it possible to detect Aβ deposition in the living human brain [35, 142]. Amyloid PET imaging not only improves our understanding of amyloid formation as a biomarker but also facilitates potential drug development. However, the application of amyloid PET imaging and its associated safety issues have not been fully evaluated for the diagnosis of AD patients, though a guideline was issued by the Amyloid Imaging Taskforce (AIT) [35]. High-throughput techniques, molecular imaging and bioinformatics resources would be excellent sources from which to gain a deep understanding of the link between T2D and AD, thereby helping to provide improved diagnosis, prognosis and treatment of these complex diseases.
LIST OF ABBREVIATIONS Aβ = Amyloid beta AD = Alzheimer's disease NFTs = Neurofibrillary tangles T2D = Type 2 Diabetes
CONFLICT OF INTEREST
The authors confirm that this article content has no conflict of interest.
ACKNOWLEDGEMENTS
The authors would like to thank Saleha Malik and Someswar Sagurthi for critical reading of the manuscript and providing valuable comments. No financial support was taken for this work.
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