Cognitive defect, alteration of synaptic function, neuronal death brought by accumulation of Aβ in brain parenchyma, and neurofibrillary tangles (NFT) in neurons are major hallmarks of Alzheimer’s disease (AD) [1]. Current medicine for AD have almost no effect on severe and terminal cases, only can ameliorate early and middle stage. However, indisposition is difficult to discover until the disease shifts to extremely serious stage when substantial neurons are lost and irreversible neuropathological lesions occur, meaning that the brain suffers severe trauma [2]. There are 47 million people living with dementia globally and the number is expected to triple (~ 151 million) by 2050 [3]. Aging societies have made the incidence more inclined to extreme [4].
There are many inducers to cause AD, including Aβ deposition, hyperphosphorylated tau, oxidative stress, metabolic functions, obesity, gastrointestinal microorganism, etc. [5], among which, Aβ deposition, as the dominating therapeutic target, has attracted more attention in recent years [6].
Aβ deposition is owed to unbalance of generation and clearance of Aβ peptide produced from transmembrane protein APP by sequential cleaving reaction of β and γ-secretase [7, 8], which is defined as an amyloidogenic metabolism of APP. In this process, APP is split into a C99 transmembrane protein and a longer N-terminal fragment sAPPβ under catalysis of β-secretase. The intramembrane region of C99 is further incised by γ-secretase to form intracellular fragment AICD and Aβ. Aβ monomer gradually changes to oligomer and eventually forms sedimentary plaque. Nevertheless, non-Amyloidogenic metabolic pathway is the metabolism of APP to produce soluble protein and sAPPα by α-secretase without aggregation or oligomer [9, 10]. Thus, blocking expression of β-secretase would decrease production of Aβ and ultimately reduce Aβ plaque.
β-site APP cleaving enzyme 1 (BACE1), a kind of β-secretase, is required for generation of all monomeric forms of Aβ [11]. Although BACE1 and the homologue BACE2 are expressed in same cell types in brain, BACE2 being much less abundant [12, 13]. Therefore, Aβ production can be modulated by regulating the expression of BACE1.
MicroRNA (miRNA) is a short (19 ~ 25 nt) endogenous noncoding RNA involved in post-transcriptional regulating of target genes, and therefore related to pathogenesis and remedy of disease [14]. Accumulating evidence has revealed that a variety of miRNA(s) are responsible for the occurrence of AD [15], some miRNA(s) participate in Aβ aggregation by regulating the expression of APP and PS1 genes, among which the miRNA-29 family has been elaborated clearly. Clinical trials have found that miRNA-29a and miRNA-29b significantly decline in the parietal cortex of patients compared to the same age group [16]. Zong and his team found that the expression of BACE1 is also regulated by miR-29c but not only by miR-29a and miR-29b [17]. So miRNA-29 family can inhibit the expression of BACE1, thereby decreasing the production of Aβ and finally eliminating aggregated Aβ plaque, in addition, miR-29a-3p is also predicted to be involved in regulating post-transcriptional modification of BACE1 gene, but substantial evidence is lacking, so this paper intends to treat APPswe-SH-SY5Y cells by HuMSCs-derived exosomes loaded with hsa-miR-29a-3p to verify their therapeutic effect on AD.
Recently, stem cells are extensively studied as a potential therapy for neurodegenerative diseases, but their application in AD has been refrained because underlying cognitive benefits have not been illuminated [18], but it is verified that transplanting stem cells can ameliorate the cognitive deficit of AD mouse model [19–21]. Human umbilical cord mesenchymal stem cells (HuMSCs) are considered to be the dominant candidate due to their wide sources, low immunogenicity, and better repair ability. HuMSCs have been reported to reduce Aβ1−42 deposition and improve cognitive disorders [22]. However, clinical application of HuMSCs is certainly limited because of ethics and difficulty to maintain cell viability. Therefore, exosomes derived from HuMSCs have emerged as the first choice in AD treatment. Accumulating evidence has indicated that exosomes play a crucial role in relieving APP/PS1 mouse models [23, 24] and can be a drug carrier to deliver medicine across the blood-brain barrier (BBB) [25].
Exosomes derived from HuMSCs that are cultured in three-dimensional (3D) materials can better improve recovery of APP/PS1 mouse, mainly because of the affluence of microRNAs in exosomes derived from these 3D cultured HuMSCs [24]. If the amount of miRNAs in exosomes could be improved by a different approach, can they achieve the same effect or even better? Our previous work reported a new strategy to efficiently load a small-molecule chemotherapeutic drug into exosomes by utilizing micro/nanofluidic technology [26]. The reported microfluidic system, i.e. exosome nanoengineering platform (ExoNP), consists of an array of nanochannels with height comparable to exosomes dimension. Exosome membranes are permeabilized by mechanical compression and fluid shear when being transported through the nanochannels, allowing the influx of cargo molecules into the exosomes from the surrounding solution while maintaining the exosomes’ integrity. Herein, we firstly transfect the Swedish mutant gene APPswe through lentivirus into SH-SY5Y cells to construct APPswe-SH-SY5Y cells and then quantify the expression of hsa-miR-29a in APPswe-SH-SY5Y. Then, HuMSCs-derived exosomes are treated by ExoNP and loaded with hsa-miR-29a to treat APPswe-SH-SY5Y cells, the expression of Aβ1−42 and BACE1 is studied, aiming to clarify whether hsa-miR-29a-3p regulates Aβ1−42 production by modulating BACE1 expression, also to verify the encapsulation of hsa-miR-29a in exosomes by the microfluidic systems for intracellular delivery.