Team:Jiangsu High School/Description
Description
Know about alzheimer's disease
Alzheimer's Disease (AD) is the sixth leading cause of death in the United States and is the third biggest cause of death for seniors (just behind heart disease and cancer) (National Institute on Aging). According to the Alzheimer's Association, though the risk of death by heart disease has been decreased by 9% between the years 2000 to 2017, the risk of one dying of Alzheimer's increased by 145%. Scientists also predict that, by 2030, there would be 75 million AD patients worldwide (Alzheimer's Disease International).
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What is Alzheimer's Disease?
Inspiration And Description
1 What it does
Alzheimer's Disease (AD) is the most common cause of dementia—a loss of cognitive functions—among the senior population (National Institute on Aging). AD's most common symptoms are memory problems, lack of spatial awareness, and impaired reasoning (National Institute on Aging). However, as the disease worsens within the patient, it damages the areas of the brain that control language, conscious thought, and processing (National Institute on Aging). In severe cases of AD, brain tissues will begin to shrink; moreover, the accumulation of amyloid plaques in the brain would increase neurotoxicity. AD patients would not be able to communicate and would be completely dependent on others' care (National Institute on Aging).
2 Causes
As mentioned above, the accumulation of amyloid plaques in the brain appears to be the main cause of AD. Since ß-amyloid peptides are the main constituent of the plaques, figuring out how to reducing the peptides was thought to be the first step of discovering a treatment for AD (Gouras et al. 2015). The plaques constituted of accumulated Aß and neurofibrillary tangles (composed of protein tau); because the latter appears in other neurodegenerative diseases, the former was thought to be directly correlated with and more specific to AD (Gouras et al. 2015). Furthermore, familial AD lead to mutations in enzymes such as presenilin 1 and 2 (both of which responsible for Aß autophagy) whereas familial mutations in tau are more closely related to other forms of dementia rather than AD (Gouras et al. 2015). Tau protein promotes tubulin assembly into microtubules and help stabilize the neuronal structure (Iqbal et al. 2010). Tau, a phosphoprotein, are found on a gene in chromosome 17; if hyperphosphorylated, the protein would accumulate into neurofibrillary tangles (Iqbal et al. 2010). Other than these two theories, scientists also have hypotheses about the root cause of AD. Because the relationship between the hyperphosphorylation of tau protein and accumulation of Aß is not clear, determining an effective treatment of AD is difficult.
3 What is the standard treatment nowadays?
Drug development for dementia had become more and more urgent now that 22% of the world's population is predicted to be over 60 years old by 2050 (WHO). According to Scientific American, of the 244 compounds tested in clinical trials for AD in between 2002 and 2012, only one was approved. This places AD's failure rate above almost all diseases in any area with a 99.6%, even ranking above cancer with a rate of 81% (Burke 2014). Cholinesterase inhibitors are one type of prescription drugs for AD; the drug is designed to prevent acetylcholine breakdown which is a chemical believed to play a role in memory recall (National Institute on Aging). However, as AD progresses to affect the brain, the latter would produce less acetylcholine; thus, cholinesterase inhibitors would then lose their effect.
Inspiration
Our Design Inspiration
From the help of our professors and teachers, our experimental topic was set to be Identify A Potential Compound Which Cures Alzheimer's Disease Through The Autophagy Path. Although there are a lot of Alzheimer's sufferer around the world and in China, there isn't a affective medicine to help them , so our team decided we are going to help. After setting the topic, we are facing other problems. From interviewing and talking to our professors, we found out that we need more support from the society.
Design
Inspiration And Description
Alzheimer's disease is becoming an increasingly severe problem around the world. According to a statistic, globally there will be one new case every 3 seconds. 50 million people living worldwide are already living with Alzheimer's disease, and this number will triple to 152 million by 2030. Yet the current situation of medication development is not promising as well. The failure-to approval ratio for new Alzheimer's medicines is only 30 to 1. From 1998 to 2014, only 3 drugs received approval for treatment of symptoms. With this in mind, we aim to seek a potential compound which can cure this disease through an approach fundamentally different from other medicines': it incorporates autophagy, a biological process inherent in each cell, to let the neurons themselves clear toxins that could induce the development of Alzheimer's disease. Therefore, this approach will less likely bring side effects and since can be approved more easily by drug administration, giving more opportunity for patients to reduce their struggle with the disease. Moreover, since autophagic process exists in virtually every cell of the human body, it can also be applied in other kinds of new medications to remove disease-inducing-toxins. Thus, autophagy is a versatile and powerful solution for new medicine developmen
1 sentence saying what we are accomplishing with our experiments
Our goal is to find a compound that may induce autophagy in the brain to prevent protein accumulation and thus AD.
2 Autophagy treatment; how are we going to prevent it
Since familial AD causes mutations in proteases which would lead to the creation of amyloid plaques, autophagy is essential in the cleavage of Aß and neurofibrillary tangles to prevent neurodegeneration (Rahman et al. 2017). It is very likely that by finding a compound that is effective in neurofibrillary tangle cleavage, we could subdue the effects of AD.
3 What is plasmid DNA?
In molecular biology, a vector is an artificial medium that carries foreign DNA material from one cell to another in order for it to be expressed or replicated in the latter. There are four major types of vectors: plasmids, viral vectors, cosmids, and artificial chromosomes; of which plasmids are the most common.
A couple of characteristics define a vector, such as having an origin of replication (where the vector could replicate itself autonomously inside its host), possessing a compatible site for insertion of genetic material, having the ability to clone in bulk, and retaining genetic markers that would predict if it is inside of its host (and use this to screen for non-recombinant cells). �
Plasmids, the vector we used in this experiment, are generally circular, dual-stranded DNA molecules and are independent from the chromosomal DNA of any cell (because it possesses a piece of genetic material also known as the origin of replication). Plasmids could be in the cytoplasm of the cell but could not survive outside of its host. Plasmids can be transferred from one cell to another by bacterial conjugation. Not only could plasmids replicate itself independently, it could also integrate itself into its host's genomic information and replicate its material through the cell (known as transformation, where a cell adopts new genetic material from a vector).
4 mTagRFP-mWasabi-LC3
mWasabi-LC3 was used because it was more acid sensitive than EGFP-LC3 in lysosomes and brighter when monitoring autolysosomes in autophagy flux. Also, the exposure time needed by mWasabi-LC3 is shorter than the one of EGFP-LC3, which yielded photos of higher quality. Because LC3 could also be present in non-autophagy cells, mWasabi-LC3∆G is more effective at discriminating them during autophagy flux (Zhou 2012).
5 LC3
The protein we tested for to prove the effectiveness of autophagy is LC3-II, a common protein involved in the process. The ratio between LC3-I and LC3-II (a conjugation achieved from LC3-I) would reflect on the amount of autophagosomes present, which might be a good indication of whether the autophagy is happening or not (Kuma et al. 2017). In the fluorescence test, autophagosomes would have their red and green light merged together (a yellow light under the fluorescent microscope) and autolysosomes would be seen as red light. We would test the effectiveness of cell autophagy by incorporating plasmid DNA (TagRPF-mWasabi-LC3) into normal E. coli to turn them into competent cells for eventual autophagy flux testing.
6 H
The participation and help Anti-AD has received from the doctors played a crucial role to the success of our human practices and experiments. At the start, our survey was not in the final yet, but after interviewing Dr.Xia, our team was able to finalize our survey. Dr.Xia provided us with vital data to further improve our survey into something that is not only benefiting our research project, but also something that contains basic information that could help the public better understand Alzheimer. Throughout our journey, not only Dr.Xia lended us a hand, Dr.Zhu also made a big contribution to our work. During the early phases of our experiment, the bacteria we have cultured was too dense and hard to pick out one to continue to our next planned step. As we became stuck on this problem, Dr.Zhu came to the rescue. Dr.Zhu, a seasoned Alzheimer researcher suggested us to lower the concentration when first culturing the bacterias. Though we were lost at first, the glimmer of hope that Dr.Zhu showed us allowed us to successfully continue with our experiment. This magnificent jouney would not be whole without our two beloved doctors.
7 D
Our final version questionnaire include three topics that we are curious about.
1) Have most people met people that have Alzheimer's? Do thier relatives or friends have Alzheimer's?
2) Do most people know about the symptoms of Alzheimer's?
3) Do the public know any way of preventing or curing Alzheimer's
And then we divided those three topics into eleven questions. From handing our the survey, we hope to know whether the public know enough information of Alzheimer's Disease, and from there, we are going to spread the basic knowlege of AD and also about our project.
8 Future Plans
Of course, this would not be the end of AD research we could accomplish; rather, this would be the beginning. Future research might entail experimenters testing the effectiveness of the compound C4-Paroxetine, C13-Amitriptyline with animal models. Testing might include investigating potential side effects of the compound such as level of toxicity within the animal model. Another future possibility may be utilizing proteins that indicate autophagy flux to prove that the compound really incites autophagy. For example, p62 proteins are degraded into autolysosomes (thus, the protein could be seen as a marker for degeneration in autophagy); in other words, decreased levels of p62 is equivalent to autophagy activation (Klionsky et al. 2012). This knowledge obtained would aid further explorations regarding the compound's action mechanism against AD and the plausibility of it becoming a full-fledged medicine to repress the disease's symptoms.
9 Reference
• Burke, Maria. (2014). Why Alzheimer's Drugs Keep Failing. Scientific American. Retrieved from https://www.scientificamerican.com/article/why-alzheimer-s-drugs-keep-failing/.
• Cuihong Zhou, Wu Zhong, Jun Zhou, Fugeng Sheng, Ziyuan Fang, Yue Wei, Yingyu Chen, Xiaoyan Deng, Bin Xia & Jian Lin (2012) Monitoring autophagic flux by an improved tandem fluorescent-tagged LC3 (mTagRFP-mWasabi-LC3) reveals that high-dose rapamycin impairs autophagic flux in cancer cells, Autophagy, 8:8, 1215-1226, DOI: 10.4161/auto.20284
• Dementia Statistics. (n.d.). Retrieved July 21, 2019, from Alzheimer's Disease International: https://www.alz.co.uk/research/statistics.
• Facts and Figures. (2019). Retrieved July 21, 2019, from Alzheimer's Association: https://alz.org/alzheimers-dementia/facts-figures.
• Gouras, G. K., Olsson, T. T., & Hansson, O. (2015). β-Amyloid peptides and amyloid plaques in Alzheimer's disease. Neurotherapeutics, 12(1), 3-11.
• Iqbal, K., Liu, F., Gong, C. X., & Grundke-Iqbal, I. (2010). Tau in Alzheimer disease and related tauopathies. Current Alzheimer Research, 7(8), 656-664.
• Klionsky, Daniel, Abdalla, Fabio, Abeliovich, Hagai et al. (2012). Guidelines for the use and interpretation of assays for monitoring autophagy. Autophagy. 8. 445-544. 10.4161/auto.19496.
• Kuma, Akiko, Komatsu, Masaaki, and Mizushima, Noboru. (2017). Autophagy-monitoring and Autophagy-deficient Mice. Autophagy. 13. 10.1080/15548627.2017.1343770.
• Lee, J. H., Yu, W. H., Kumar, A., et al. (2010). Lysosomal proteolysis and autophagy require presenilin 1 and are disrupted by Alzheimer-related PS1 mutations. Cell, 141(7), 1146-1158.
• National Institute on Aging. (n.d.). Alzheimer's Disease Fact Sheet. National Institutes of Health. Retrieved from https://www.nia.nih.gov/health/alzheimers-disease-fact-sheet.
• National Institute on Aging. (n.d.). How is Alzheimer's Disease Treated. National Institutes of Health. Retrieved from https://www.nia.nih.gov/health/how-alzheimers-disease-treated.
• Rahman, Md. Ataur & Rhim, Hyewhon. (2017). Therapeutic implication of autophagy in neurodegenerative diseases. BMB reports. 50. 10.5483/BMBRep.2017.50.7.069.
• World Health Organization. (2018). Ageing and Health. Retrieved from https://www.who.int/news-room/fact-sheets/detail/ageing-and-health.
• Zhang, X. J., Chen, S., Huang, K. X., and Le, W. D. (2013). Why should autophagic flux be assessed?. Acta pharmacologica Sinica, 34(5), 595–599. doi:10.1038/aps.2012.