“Mounting evidence indicates that the effectiveness of therapeutic modalities critically depends on early diagnosis of Alzheimer’s disease, before massive neuron loss.”
Alzheimer’s disease: challenges & concepts
The neurodegenerative disease described by Alois Alzheimer in 1906 is now recognized as a major aging-related dementia worldwide, affecting more than 35 million individuals. With the demographic shift toward aging societies, this number is expected to double every 20 years, to an estimated 135.5 million worldwide afflicted with the disease by 2050 [Citation1]. The global costs related to diminished social functioning in AD patients are similar to the financial burdens of heart disease and cancer, placing Alzheimer’s disease (AD) among major public health concerns [Citation2].
AD develops for tens of years in the preclinical phase before the onset of clinical symptoms, such as cognitive, memory and behavioral impairments associated with the progressive and irreversible loss of brain neurons [Citation3]. Mounting evidence indicates that the effectiveness of therapeutic modalities critically depends on early diagnosis of AD, before massive neuron loss. In 2013, this conclusion was highlighted by the US FDA guidelines for new AD drugs. However, despite intense research worldwide, no diagnostic methods are currently available for preclinical AD, and existing AD treatments are only symptomatic. This reflects the fact that the cause(s) of AD and its molecular pathomechanism are far from being understood. The main concept, known as the amyloid cascade hypothesis, refers to Aβ deposits in the AD brain [Citation4,Citation5]. This hypothesis has been put at center-stage by the identification of rare familial AD cases linked to mutations in one of the three genes encoding proteins involved in the amyloidogenic production of Aβ: APP and presenilins [Citation6]. Thus, it is suggested that Aβ is the primary and main cause of the disease, and Aβ aggregation initiates a cascade of pathological events characteristic for AD, such as hyperphosphorylation of neuronal tau protein, microglial activation and finally synapse and cell loss [Citation4–6]. However, mounting evidence indicates that in sporadic AD Aβ aggregation is preceded by earlier molecular changes, and that only the final step of the sporadic AD and familial AD pathogeneses are common and dominated by the overproduction of Aβ. Multiple recent data suggest that Aβ and tau pathology may appear in neurons in response to various types of stress or brain damage, such as infections, vascular damage or diabetes, and that the pathology involves several mechanisms including Ca2+ dyshomeostasis, oxidative stress, mitochondria impairment, aberrant cell cycle and inflammation [Citation7–11]. Thus, the difficulty in deciphering early AD molecular events lies in AD being an extremely complex, heterogeneous and multifactorial process. Compared with monogenous or infectious diseases caused by a single pathogenic factor, AD sustains a disease model of a much higher complexity and, therefore, requires equally complex research strategies and paradigms. These efforts are necessary because progress in AD diagnostics and therapy depends on elucidating the disease’s molecular mechanism(s), which ultimately will lead to the identification of novel therapeutic targets and biomarkers for prognosis and monitoring of AD progression.
Current Alzheimer’s disease diagnostic criteria & the quest for biomarkers
Conceptualization of the disease’s stages, together with the growing appreciation of biomarkers as key tools for tracking the pathology, led recently to revision of AD diagnostic criteria. The most up-to-date recommendations for AD diagnosis incorporating biomarkers have come from the US National Institute of Ageing and Alzheimer’s Disease Association, National Institute of Neurological and Communicative Disorders and Stroke and Alzheimer’s Disease and Related Disorders Association, which currently represent the gold standard [Citation12–14]. These criteria distinguish the following AD phases: asymptomatic preclinical stage, symptomatic predementia and dementia. As stated earlier, no tools are sensitive enough to diagnose preclinical AD. Diagnostic criteria refer to AD dementia and mild cognitive impairment (MCI) that is attributable to AD, and are based on cognitive testing and cerebrospinal fluid (CSF) biomarkers: total and hyperphosphorylated tau protein, and the Aβ peptides. Genetic tests for mutated APP and presenilins, in addition to brain imaging, support the diagnosis. The major disadvantages of CSF biomarkers are lumbar puncture required for CSF withdrawal and substantial biological variability of CSF biomarkers across various diagnostic centers due to preanalytical, analytical and postanalytical factors. The necessity for extensive standardization of materials and procedures was addressed by several initiatives, including the recent international project ‘BIOMARKAPD’ of the EU Joint Programme–Neurodegenerative Disease Research, the largest global research initiative aimed at tackling the challenge of neurodegenerative diseases. Overall, it becomes clear that progress in AD diagnostics relies to a great extent on the identification of novel biomarkers for preclinical AD, in addition to further development of early AD clinical biomarkers of improved sensitivity and specificity, preferably in more easily available diagnostic materials, such as blood.
Molecular alterations in Alzheimer’s disease blood lymphocytes: potential for biomarkers
Increasing evidence indicates that, in AD, molecular changes occur not only in the CNS, but also in peripheral cells, such as fibroblasts and blood cells, mainly platelets and lymphocytes. Thus, peripheral cells do represent an easily available potential material for diagnosis and drug screening. The ultimate goal is to develop simple, noninvasive and inexpensive diagnostic tests based on blood cells and/or molecules present in blood for the early detection of AD.
While preclinical AD remains undefined, a search for early molecular changes in amnesic patients with MCI is a good alternative, assuming that potential biomarkers will be validated in follow-up studies [Citation5,Citation7]. Consistently, multiple early investigations employed only AD lymphocytes, but in the last few years, molecular changes in MCI lymphocytes have become a matter of interest.
Analyses of oxidative stress and related molecular changes were particularly well-represented in research employing AD lymphocytes. Among molecular changes detected consistently by several groups, both in AD brains as well as AD and MCI blood lymphocytes or mononuclear cells, are those related to impaired Ca2+ homeostasis and endoplasmic reticulum stress [Citation15]. Elevated reactive oxygen species levels, altered levels of antioxidant enzymes, elevated hydroxyl radical-induced DNA oxidation, DNA damage response markers, increased mitochondrial susceptibility and basal apoptosis were found by various groups in T and B AD lymphocytes, compared with healthy cells (recently reviewed in [Citation16]). Thus, peripheral AD lymphocytes reflect the pathological oxidative stress conditions known to characterize the brains of AD patients. These findings were recapitulated in MCI lymphocytes; increased levels of such oxidative stress markers as 8-hydroxy-2′-deoxyguanosine, malondialdehyde and reduced levels of GR activity were observed, in addition to increased levels of antioxidant enzymes, such as SOD [Citation16]. Consistently, substantial oxidative stress and mitochondrial dysfunction were observed in lymphocytes of both AD and MCI patients, without any Aβ or tau pathology. These findings emphasized the systemic nature of oxidative stress in AD and strongly support the hypothesis that oxidative stress is an early factor in AD development.
Another feature of early AD pathogenesis manifested in lymphocytes is cell cycle dysfunction. Re-expression of cell cycle proteins was observed in postmitotic brain neurons of patients with AD and MCI; while cell cycle dysregulation was also observed in lymphocytes (reviewed in [Citation16]). Potential protein markers related to cell cycle changes identified in proliferating AD lymphoblasts were key proteins that regulate G1/S cell cycle progression, such as p53, p21 and cyclinD1. Cell cycle dysregulation seems to be intertwined at the molecular level with oxidative stress [Citation17].
Biochemical alterations detected in lymphocytes seem to affect their physiological immune functions [Citation18,Citation19]. This subject has been the matter of intense studies in recent years, which demonstrated the contribution of peripheral immune responses to the progression of AD neurodegeneration. As the blood–brain barrier is impaired in AD, a growing body of evidence suggests a role for peripheral immune cells in the pathological events that occur in the brain, in addition to the activation of immune resident brain cells [Citation18–20].
Conclusion & future perspective
Molecular alterations in blood lymphocytes at the early stages of AD provide insight into the mechanisms of AD development and potential biochemical biomarkers for early AD detection, in addition to monitoring disease progression and its responses to therapeutics. In light of recent progress in biomarker development for various other pathologies, including cancer, putting special emphasis on biochemical indicators of the disease process has become a universal perspective. This concept may revolutionize future diagnostics, shifting its focus from symptoms to molecular signatures.
In light of the complexity and heterogeneity of AD, the future blood molecular signature of AD will most probably be developed based on an integrated panel of different molecules in blood cells, in addition to blood plasma, including proteins, nucleic acids such as miRNA and/or lipids. Such a combined approach may allow for precise stratification of AD stages and forms, and for individualized therapy. Moreover, the correlation of blood biochemical biomarkers, brain imaging and neuropsychological tests seems necessary for further progress in deciphering the disease’s mechanisms, in addition to diagnostics and therapy.
Acknowledgements
The author thanks J Chlebowska and A Piotrowska for excellent editorial help.
Financial & competing interests disclosure
This work was supported by the Nencki Institute statutory funds, the EU JPND grant 2/BIOMARKAPD/JPND/2012, and the Polish National Science Centre grant 2014/15/D/NZ4/04361. The author has no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript.
No writing assistance was utilized in the production of this manuscript.
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