During the summer of 1998, between graduating and starting my PhD, I spent two months at SmithKline Beecham (now GlaxoSmithKline) Pharmaceuticals in Harlow, Essex, as a research assistant in the Department of Molecular Neurobiology.
The project I was working on involved the analysis of transcripts of genes associated with Alzheimer’s Disease, and I spent those months isolating nucleic acids from chunks of brains of people who had died from the condition.
I googled myself recently, and discovered, to my surprise, that I am the co-author of 3 papers in Molecular Brain Research. The papers were published more than 5 years after I’d done the work. Here are the abstracts:
Insoluble β-amyloid deposits in Alzheimer’s disease (AD) brain are proteolytically derived from the membrane bound amyloid precursor protein (APP). The APP gene is differentially spliced to produce isoforms that can be classified into those containing a Kunitz-type serine protease inhibitor domain (K+, APP751, APP770, APRP365 and APRP563), and those without (K−, APP695 and APP714,). Given the hypothesis that Aβ is a result of aberrant catabolism of APP, differential expression of mRNA isoforms containing protease inhibitors might play an active role in the pathology of AD. We took 513 cerebral cortex samples from 90 AD and 81 control brains and quantified the mRNA isoforms of APP with TaqMan™ real-time RT-PCR. After adjustment for age at death, brain pH and gender we found a change in the ratio of KPI+ to KPI− mRNA isoforms of APP. Three separate probes, designed to recognise only KPI+ mRNA species, gave increases of between 28% and 50% in AD brains relative to controls (p=0.002). There was no change in the mRNA levels of KPI-(APP 695) (p=0.898). Therefore, whilst KPI-mRNA levels remained stable the KPI+species increased specifically in the AD brains.
Quantitative human mRNA data are derived from post-mortem or biopsied tissue. RNA degradation, poor replication, a large mRNA variance and confounding factors such as brain pH and age of death are often cited, however, as objections to the data’s reliability. A central question is whether post-mortem human mRNA can be treated as a statistically ordered system. TaqMan™ real-time RT-PCR was used to measure seven mRNAs in 513 cortical samples taken from 90 Alzheimer’s disease and 81 control brains. Despite a high mRNA variance strong correlations were found between the mRNA transcripts in a single brain. Where a brain has a high/low level of one mRNA, the same brain invariably has a high/low level of other mRNAs; correlated order is present and allows removal of that source of variation common to all genes. Although levels of mRNA are highly variable between subjects (>1000-fold), quantitative order is present in post-mortem human mRNA, allowing effects due to pathology or gender to be isolated and tested for significance.
β-Secretase (BACE) and glycogen synthase kinase (GSK 3) are two enzymes thought to play a role in Alzheimer’s disease. We extracted mRNA from 90 Alzheimer and 81 control brains. Levels of mRNA were quantified for BACE and GSK 3 with TaqMan™ real-time RT-PCR. We found no change in the Alzheimer’s disease brains relative to controls for either the BACE or the GSK 3α mRNA levels.
- An apple a day keeps amnesia at bay
- Broken down protein degradation could lead to neurodegeneration
- Vaccine reverses cognitive impairment associated with Alzheimer’s
- New PET method may enable early diagnosis of Alzhemier’s
- Disentangling Alzheimer’s
- Alzheimer’s Disease: early signs & a possible therapy