Publications

BACKGROUND: Low frequency coding variants in TREM2 are associated with Alzheimer disease (AD) risk and cerebrospinal fluid (CSF) TREM2 protein levels are different between AD cases and controls. Similarly, TREM2 risk variant carriers also exhibit differential CSF TREM2 levels. TREM2 has three different alternative transcripts, but most of the functional studies only model the longest transcript. No studies have analyzed TREM2 expression levels or alternative splicing in brains from AD and cognitively normal individuals. We wanted to determine whether there was differential expression of TREM2 in sporadic-AD cases versus AD-TREM2 carriers vs sex- and aged-matched normal controls; and if this differential expression was due to a particular TREM2 transcript.

METHODS: We analyzed RNA-Seq data from parietal lobe brain tissue from AD cases with TREM2 variants (n = 33), AD cases (n = 195) and healthy controls (n = 118), from three independent datasets using Kallisto and the R package tximport to determine the read count for each transcript and quantified transcript abundance as transcripts per million.

RESULTS: The three TREM2 transcripts were expressed in brain cortex in the three datasets. We demonstrate for the first time that the transcript that lacks the transmembrane domain and encodes a soluble form of TREM2 (sTREM2) has an expression level around 60% of the canonical transcript, suggesting that around 25% of the sTREM2 protein levels could be explained by this transcript. We did not observe a difference in the overall TREM2 expression level between cases and controls. However, the isoform which lacks the 5' exon, but includes the transmembrane domain, was significantly lower in TREM2- p.R62H carriers than in AD cases (p = 0.007).

CONCLUSION: Using bulk RNA-Seq data from three different cohorts, we were able to quantify the expression level of the three TREM2 transcripts, demonstrating: (1) all three transcripts of them are highly expressed in the human cortex, (2) that up to 25% of the sTREM2 may be due to the expression of a specific isoform and not TREM2 cleavage; and (3) that TREM2 risk variants do not affect expression levels, suggesting that the effect of the TREM2 variants on CSF levels occurs at post-transcriptional level.

BACKGROUND: Alzheimer's disease (AD) is the most common form of dementia. This neurodegenerative disorder is associated with neuronal death and gliosis heavily impacting the cerebral cortex. AD has a substantial but heterogeneous genetic component, presenting both Mendelian and complex genetic architectures. Using bulk RNA-seq from the parietal lobes and deconvolution methods, we previously reported that brains exhibiting different AD genetic architecture exhibit different cellular proportions. Here, we sought to directly investigate AD brain changes in cell proportion and gene expression using single-cell resolution.

METHODS: We generated unsorted single-nuclei RNA sequencing data from brain tissue. We leveraged the tissue donated from a carrier of a Mendelian genetic mutation, PSEN1 p.A79V, and two family members who suffer from sporadic AD, but do not carry any autosomal mutations. We evaluated alternative alignment approaches to maximize the titer of reads, genes, and cells with high quality. In addition, we employed distinct clustering strategies to determine the best approach to identify cell clusters that reveal neuronal and glial cell types and avoid artifacts such as sample and batch effects. We propose an approach to cluster cells that reduces biases and enable further analyses.

RESULTS: We identified distinct types of neurons, both excitatory and inhibitory, and glial cells, including astrocytes, oligodendrocytes, and microglia, among others. In particular, we identified a reduced proportion of excitatory neurons in the Mendelian mutation carrier, but a similar distribution of inhibitory neurons. Furthermore, we investigated whether single-nuclei RNA-seq from the human brains recapitulate the expression profile of disease-associated microglia (DAM) discovered in mouse models. We also determined that when analyzing human single-nuclei data, it is critical to control for biases introduced by donor-specific expression profiles.

CONCLUSION: We propose a collection of best practices to generate a highly detailed molecular cell atlas of highly informative frozen tissue stored in brain banks. Importantly, we have developed a new web application to make this unique single-nuclei molecular atlas publicly available.

Infantile globoid cell leukodystrophy (GLD, Krabbe disease) is a fatal demyelinating disorder caused by a deficiency in the lysosomal enzyme galactosylceramidase (GALC). GALC deficiency leads to the accumulation of the cytotoxic glycolipid, galactosylsphingosine (psychosine). Complementary evidence suggested that psychosine is synthesized via an anabolic pathway. Here, we show instead that psychosine is generated catabolically through the deacylation of galactosylceramide by acid ceramidase (ACDase). This reaction uncouples GALC deficiency from psychosine accumulation, allowing us to test the long-standing "psychosine hypothesis." We demonstrate that genetic loss of ACDase activity (Farber disease) in the GALC-deficient mouse model of human GLD (twitcher) eliminates psychosine accumulation and cures GLD. These data suggest that ACDase could be a target for substrate reduction therapy (SRT) in Krabbe patients. We show that pharmacological inhibition of ACDase activity with carmofur significantly decreases psychosine accumulation in cells from a Krabbe patient and prolongs the life span of the twitcher (Twi) mouse. Previous SRT experiments in the Twi mouse utilized l-cycloserine, which inhibits an enzyme several steps upstream of psychosine synthesis, thus altering the balance of other important lipids. Drugs that directly inhibit ACDase may have a more acceptable safety profile due to their mechanistic proximity to psychosine biogenesis. In total, these data clarify our understanding of psychosine synthesis, confirm the long-held psychosine hypothesis, and provide the impetus to discover safe and effective inhibitors of ACDase to treat Krabbe disease.

There are currently no effective therapeutics for Alzheimer disease (AD). Clinical trials targeting amyloid beta thus far have shown very little benefit and only in the earliest stages of disease. These limitations have driven research to identify alternative therapeutic targets, one of the most promising is the triggering receptor expressed on myeloid cells 2 (TREM2). Areas covered: Here, we review the literature to-date and discuss the potentials and pitfalls for targeting TREM2 as a potential therapeutic for AD. We focus on research in animal and cell models for AD and central nervous system injury models which may help in understanding the role of TREM2 in disease. Expert opinion: Studies suggest TREM2 plays a key role in AD pathology; however, results have been conflicting about whether TREM2 is beneficial or harmful. More research is necessary before designing TREM2-targeting therapies. Successful therapeutics will most likely be administered early in disease.

Eighteen loci and several susceptibility genes have been related to Parkinson's disease (PD). However, most studies focus on single genes in small PD series. Our aim was to establish the genetic background of a large Spanish PD sample. Pooled-DNA target sequencing of 7 major PD genes (SNCA, PARK2, PINK1, DJ-1, LRRK2, GBA, and MAPT) was performed in 562 PD cases. Forty-four variants were found among 114 individuals (20.28%, p<0.05). Among these variants, 30 were found in Mendelian genes (68.18%) and 14 in PD susceptibility genes (31.82%). Seven novel variants were identified. Interestingly, most variants were found in PARK2 and PINK1 genes, whereas SNCA and DJ-1 variants were rare. Validated variants were also genotyped in Spanish healthy controls (n = 597). Carriers of heterozygous PARK2 variants presented earlier disease onset and showed dementia more frequently. PD subjects carrying 2 variants at different genes (1.42%) had an earlier age of onset and a predominantly akinetic-rigid PD phenotype (55.6%, p < 0.05), suggesting that the accumulation of genetic risk variants could modify PD phenotype.

A study on familial Parkinson disease (PD) described 4 variants in the gene TMEM230 (Chr. 20p13) as the cause of PD. The aim of this study was to test if variants in the TMEM230 gene are associated with PD in 2 independent American European data sets. No variants in the TMEM230 region were found associated with PD, age at onset, or cerebrospinal fluid α-synuclein levels.

BACKGROUND: Attention deficit and hyperactive disorder (ADHD) is highly prevalent among children in Bogota City. Both genetic and environmental factors play a very important role in the etiology of ADHD. However, to date few studies have addressed the association of genetic variants and ADHD in the Colombian population.

OBJECTIVES: To test the genetic association between polymorphisms in the DAT1, HTTLPR, COMT and BDNF genes and ADHD in a sample from Bogota City.

METHODS: We genotyped the most common polymorphisms in DAT1, SERT, COMT and BDNF genes associated with ADHD using conventional PCR followed by restriction fragment length polymorphism (RFLP) in 97 trios recruited in a medical center in Bogota. The transmission disequilibrium test (TDT) was used to determine the association between such genetic variants and ADHD.

RESULTS: The TDT analysis showed that no individual allele of any variant studied has a preferential transmission.

CONCLUSIONS: Our results suggest that the etiology of the ADHD may be complex and involves several genetic factors. Further studies in other candidate polymorphisms in a larger sample size will improve our knowledge of the ADHD in Colombian population.