Characterization of the Poly-T Variant in the TOMM40 Gene in Diverse Populations

We previously discovered that a polymorphic, deoxythymidine-homopolymer (poly-T, rs10524523) in intron 6 of the TOMM40 gene is associated with age-of-onset of Alzheimer's disease and with cognitive performance in elderly. Three allele groups were defined for rs10524523, hereafter ‘523’, based on the number of ‘T’-residues: ‘Short’ (S, T≤19), ‘Long’ (L, 20≤T≤29) and ‘Very Long’ (VL, T≥30). Homopolymers, particularly long homopolymers like ‘523’, are difficult to genotype because ‘slippage’ occurs during PCR-amplification. We initially genotyped this locus by PCR-amplification followed by Sanger-sequencing. However, we recognized the need to develop a higher-throughput genotyping method that is also accurate and reliable. Here we describe a new ‘523’ genotyping assay that is simple and inexpensive to perform in a standard molecular genetics laboratory. The assay is based on the detection of differences in PCR-fragment length using capillary electrophoresis. We discuss technical problems, solutions, and the steps taken for validation. We employed the novel assay to investigate the ‘523’ allele frequencies in different ethnicities. Whites and Hispanics have similar frequencies of S/L/VL alleles (0.45/0.11/0.44 and 0.43/0.09/0.48, respectively). In African-Americans, the frequency of the L-allele (0.10) is similar to Whites and Hispanics; however, the S-allele is more prevalent (0.65) and the VL-allele is concomitantly less frequent (0.25). The allele frequencies determined using the new methodology are compared to previous reports for Ghanaian, Japanese, Korean and Han Chinese cohorts. Finally, we studied the linkage pattern between TOMM40-‘523’ and APOE alleles. In Whites and Hispanics, consistent with previous reports, the L is primarily linked to ε4, while the majority of the VL and S are linked to ε3. Interestingly, in African-Americans, Ghanaians and Japanese, there is an increased frequency of the ‘523’S-APOEε4 haplotype. These data may be used as references for ‘523’ allele and ‘523’-APOE haplotype frequencies in diverse populations for the design of research studies and clinical trials

Prebriefing for Cultural Humility

•Conducting simulations to address sensitive topics such as bias or racism can be daunting.•Prebriefing for Cultural Humility sets the tone for a safe learning experience.•The mnemonic of Ground Rules, Acknowledge, Safe Psychological Environment, and Define may be followed to guide prebriefing.•Prebriefing for Cultural Humility may assist learners to embrace uncomfortable dialogue.Conducting simulations to address sensitive topics such as bias or racism in healthcare can be daunting or lead to unintended adverse consequences. The content and nature of the prebrief is critical to set the tone for a rich learning experience.This method is based on the Theory of Cultural Humility, aligns with the Healthcare Simulation Standards of Best Practice™ Prebriefing: Preparation and Briefing, and may be used in conjunction with the method of Debriefing for Cultural Humility©.The method of Prebriefing for Cultural Humility© is described. Prebriefing for Cultural Humility entails use of the following:Ground Rules, Acknowledge, Safe Psychological Environment, and Define (GRASPED).Prebriefing for Cultural Humility may assist educators and learners to embrace potentially uncomfortable dialogue to foster learning and appreciation for diversity and cultural humility

Effects of low doses of pioglitazone on resting-state functional connectivity in conscious rat brain.

Pioglitazone (PIO) is a peroxisome proliferator-activated receptor-γ (PPARγ) agonist in clinical use for treatment of type 2 diabetes (T2DM). Accumulating evidence suggests PPARγ agonists may be useful for treating or delaying the onset of Alzheimer's disease (AD), possibly via actions on mitochondria, and that dose strengths lower than those clinically used for T2DM may be efficacious. Our major objective was to determine if low doses of pioglitazone, administered orally, impacted brain activity. We measured blood-oxygenation-level dependent (BOLD) low-frequency fluctuations in conscious rats to map changes in brain resting-state functional connectivity due to daily, oral dosing with low-dose PIO. The connectivity in two neural circuits exhibited significant changes compared with vehicle after two days of treatment with PIO at 0.08 mg/kg/day. After 7 days of treatment with a range of PIO dose-strengths, connections between 17 pairs of brain regions were significantly affected. Functional connectivity with the CA1 region of the hippocampus, a region that is involved in memory and is affected early in the progression of AD, was specifically investigated in a seed-based analysis. This approach revealed that the spatial pattern of CA1 connectivity was consistent among all dose groups at baseline, prior to treatment with PIO, and in the control group imaged on day 7. Compared to baseline and controls, increased connectivity to CA1 was observed regionally in the hypothalamus and ventral thalamus in all PIO-treated groups, but was least pronounced in the group treated with the highest dose of PIO. These data support our hypothesis that PIO modulates neuronal and/or cerebrovascular function at dose strengths significantly lower than those used to treat T2DM and therefore may be a useful therapy for neurodegenerative diseases including AD

The 57 ROIs used in the assessment of whole brain functional connectivity.

<p>The regions were identified according to a fully segmented rat brain atlas [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0117973#pone.0117973.ref039" target="_blank">39</a>].</p

Functional connectivity map from the hippocampus seed for each PIO dose group from the Sub-chronic arm of the study on SD7.

<p>Functional connectivity strength is presented as a heat map of the cross-correlation coefficient. PIO treatment most noticeably influences the functional connections between regions broadly indicated by the white arrows, including the hypothalamus and ventral thalamus, and the hippocampal CA1 region.</p