For decades, researchers working on neurodegenerative disorders (like Parkinson’s and Alzheimer’s) focused on figuring out what happens in the neurons (brain cells). Brain research is replete with molecular and system-based studies, with neurons as the centerpiece. Now, data from next-generation sequencing technology has shed light on a group of cells, often overlooked in research, which may hold the key to fighting neurodegenerative disorders.
Neurons are specialized cells in the nervous system, tasked with transmitting information to neighboring neurons, muscle or gland cells. They consist of a cell body, an axon and dendrites, the ends of which contain synapses (connections to other cells). Neurons are essential for our brain’s functioning: for instance, as you read this article, your neurons transmit electrical impulses, or “fire” in different regions of your brain, allowing you to recognize words and process information. Virtually all our activities, be it walking, talking or thinking, is enabled by neurons.
Due to their importance, the loss of neurons through cell death or damage manifests as problems in mental functioning: a loss of memory, motor skills and/or cognitive skills. These problems are grouped under the term neurodegenerative disorders. The type of disorder depends on the type of neurons affected. For example, in Parkinson’s disease (PD), the SNc (substantia nigra pars compacta) neurons in the basal ganglia region (which controls bodily movement) of the brain are affected first, followed by other neurons in higher cortical regions. This is correlated with symptoms which include unwanted limb movements, and in later stages, memory problems. In Alzheimer’s (AD), the hippocampal neurons in the temporal lobe of the brain are affected first, followed by other regions of the brain. This is seen in the initial stage as problems with short-term memory, followed by a loss of motor control. Doctors note that the first symptoms of PD and AD manifest at approximately 60 – 65 years of age, when around 50 – 60% of the neurons have died. However, the warning signs of sleeping problems, constipation problems, low libido and/or a diminished sense of smell are often seen at 35 to 40 years, when the neurons presumably start to die. Treatment options are limited and focus on relieving the symptoms of the disease. Neurotransmitters (like acetylcholine for AD and dopamine for PD) are found to be at low levels and are therefore supplemented through medication: levodopa or acetylcholine. Patients often overdose on the medication to feel ‘better’ for longer, and do not take it regularly as prescribed by their doctor. In the long term, this causes the aggravation of dyskinesia (uncontrollable limb movements), the very symptom it was intended to prevent. At this point, patients need expensive deep brain stimulation to alleviate their worsening condition.
Neurodegenerative disorders affect approximately 45-50 million people worldwide. PD affects around 1% of the global over-60 population. In India alone, it is estimated that over 7 million patients suffer with PD. The age-specific prevalence of PD is estimated to be 76 to 148 per 100,000 in India. Certain ethnic groups seem to be affected disproportionately, pointing to a strong genetic component in the illness. A study on the Parsi community in Mumbai showed a prevalence of 328 per 100,000. In the case of Alzheimer’s, there are an estimated 4 million patients in India, with the number of cases expected to double by 2030. Neurodegenerative diseases not only affect the patients, but also their families, who face added stress and anxiety due to behavioral changes exhibited by the patients, resulting in emotional and social costs, apart from the financial cost of treatment. On a positive note, the prevalence of neurodegenerative disorders is much lower in India compared to global statistics. However, due to the large, rapidly ageing population, the disease burden in India is high.
NGS Data Analysis
Certain types of neurons are more vulnerable in neurodegenerative diseases. For instance, consider SNc neurons in PD. These neurons are dopaminergic, which means that they are the source of dopamine, an important neurotransmitter associated with movement, memory and reward-based behavior. Although the brain harbors other types of dopaminergic neurons (such as ventral tegmental area neurons), only SNc neurons are affected in the initial stages of Parkinson’s disease. A similar phenomenon is observed in AD as well, with hippocampal neurons affected first.
What makes this neuronal subpopulation vulnerable?
To find a concrete answer to this question, Dr. M. Michael Gromiha and Akila Parvathy Dharshini S, a Ph.D. scholar at the Protein Bioinformatics Lab, decided to work with next-generation sequencing (NGS) transcriptome data, in collaboration with Prof. Y-h. Taguchi of Chuo University, Japan. Akila also had a number of discussions with Prof. Srinivasa Chakravarthy, head of the Computational Neuroscience Lab.
NGS data analysis was chosen for its potential applications in personalized medicine. NGS refers to the highly efficient, revolutionary technique of sequencing the genome (i.e. the entire DNA sequence of any organism). In this technique, the DNA of the organism is split into millions of fragments, which are then multiplied millions of times using a chain reaction. Each fragment is therefore read millions of times, eliminating any reading errors. The individual fragments are then pieced together using advanced software. It is considerably quicker, cheaper, highly reliable and more accurate than other sequencing techniques. NGS data is available in public repositories, including NCBI’s SRA (Sequence Read Archive from the National Center for Biotechnology Information).
What is the transcriptome, and why do we use it? A transcriptome represents the sum of gene transcripts in a given cell/tissue sample. Genes are made up of DNA sequences, a biomolecule made up of 4 nucleotide bases represented by the letters A (adenine), G (guanine), C (cytosine) and T (thymine). So, DNA (and therefore, the gene) consists of a long sequence of A, G, C and T. In all cells, genes have to be transcribed into another biomolecule called RNA before they can be used to make proteins. RNA (ribonucleic acid), like DNA, is a sequence of A (adenine), G (guanine), C (cytosine) and U (uracil instead of thymine, T). So, gene transcripts are essentially the RNA version of the genes. Why are gene transcripts important? In every cell, only certain genes may be transcribed, and some genes may be transcribed more often than others. This is why, a neuron is very different from skin cells, both in terms of appearance and its internal working. While a neuron and a skin cell from an individual will have the same DNA, their transcriptomes will be vastly different. Transcriptomes are more useful when comparing patients with healthy individuals, as they can be used to identify differences which would otherwise be missed if we just compared their DNA.
Akila worked with transcriptome data taken from the temporal lobes and frontal lobes of patients in the advanced stages of Alzheimer’s disease as well as healthy subjects. Three types of analyses were carried out on the data: SNP (single nucleotide polymorphism) analysis, differential gene expression analysis and network analysis. SNP analysis finds single-letter differences between the patients and healthy individuals. In differential gene expression analysis, differences in gene transcription are studied. The differences were then mapped to the products of the genes, i.e. proteins. From these proteins, the affected pathways (which can be understood as a set of proteins performing a related function in the cell) are identified. Thus, an accurate snapshot of the affected cells could be constructed from the variations present in the transcriptome data.
Decades of research have uncovered many possible contributing factors in neurodegenerative disorders, including energy imbalance, calcium load and protein aggregation. These studies focus on the neurons, and this is understandable for two reasons: scientists presumed that the problem was in the neurons, and neurons are easy to study because their electrical activity could be measured easily. However, the transcriptome data analysis suggests that glial cells, not just neurons, may be vulnerable in neurodegenerative disorders.
Glial cells (also referred to as glia or neuroglial cells) constitute a significant percentage (60 – 80%) of the brain and spinal cord. The word “glia” comes from the Greek word for “glue”, from the belief that these cells hold the brain together in some way. They come in four distinct types: astrocytes (star-shaped, hence the name), which maintain the chemical environment of the neurons; oligodendrocytes, which wrap the neurons in a myelin sheath; microglia, which remove cell debris, much like a macrophage engulfing a pathogen; and NG-2 glia, which generate new oligodendrocytes and regulate the synapses between neurons. Additionally, glial cells supply energy to the neurons by acting as a go-between for the neurons and the blood vessels. Neurons “tell” the glial cell how much energy they need, and the glial cells obtain this energy from the bloodstream, convert it into lactate, and transport it to the neurons. In fact, 60-70% of the energy needed by neurons comes from glial cells. The importance of glial cells – the “other brain*” – has only been realized in the past 3-4 years.
Energy imbalance has been cited as a cause for neuron cell damage and/or death in PD. Insufficient energy supply may result in cell damage and death. NGS data analysis shows that glial cells associated with vulnerable neurons seem to lose their structural integrity, and consequently could not keep up with the energy demands of the neurons. It also identified a communication gap between the neurons and the glial cells; the neurons were no longer able to tell the glial cell how much energy was needed.
The Road Ahead
The NGS data analysis hints at the role of glial cells in neurodegenerative disorders and may provide clues to answer questions on the selective vulnerability of the neurons. The research work is due to be published soon.
As with most diseases, emerging evidence suggests that our lifestyle choices (in terms of diet, exercise and mental stimulation) play a prominent role in lowering the risk and delaying the onset and progression of neurodegenerative diseases. Various studies correlate regular physical activity with improved neuron health. Learning multiple languages may also help delay the onset of the disease. For patients and their families, good coping strategies include learning about the disease, physical therapy and involvement with support groups. A healthy lifestyle is key to overcoming neurodegenerative disorders.
Sherlyn Jemimah is a research scholar at the Department of Biotechnology, IIT Madras, and works on the computational analysis of mutant protein-protein interactions. When she is not doing research, she devours popular science articles and enjoys playing music.
Meet the Professor
Dr. M. Michael Gromiha is an Associate Professor at the Department of Biotechnology, IIT Madras and heads the Protein Bioinformatics lab. His research interests include protein stability and folding, protein interactions, protein aggregation, mutation studies, structure-based drug design, deep learning and NGS data analysis. He obtained his Ph.D. in Computational Molecular Biophysics from Bharathidasan University, India in 1989. His post-doctoral experience includes the International Center for Genetic Engineering and Biotechnology (ICGEB), Italy and the Institute of Physical and Chemical Research (RIKEN), Japan. He has also served as a Senior Research Scientist at the National Institute of Advanced Industrial Science and Technology (AIST), Japan. You can learn more at https://www.iitm.ac.in/bioinfo/Gromiha/index.html.
Prof. V. Srinivas Chakravarthy obtained his M.S. and Ph.D. from the University of Texas, heads the Computational Neuroscience lab at the Department of Biotechnology, IIT Madras. His research interests include computational neuroscience, computational cardiology, biomedical engineering, and pattern recognition. You can learn more at https://biotech.iitm.ac.in/Faculty/CNS_LAB/home.html.
*The Other Brain: The Scientific and Medical Breakthroughs That Will Heal Our Brains and Revolutionize Our Health is a highly acclaimed scientific work by Dr. R. Douglas Fields, detailing the role of glial cells in neurodegenerative disorders and everyday neuron activities. It is published by Simon and Schuster.