RT-qPCR study on post-mortem brain samples from patients with major psychiatric disorders: Reference genes and specimen characteristics

Abasolo, Nerea; Torrell, Helena; Roig, Bárbara; Moyano, Sílvia; Vilella, Elisabet; Martorell, Lourdes

doi:10.1016/j.jpsychires.2011.06.001

Cited by 22 publications

(20 citation statements)

References 31 publications

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“…For this reason, it is important to understand possible sources of variability in the postmortem preservation of RNA. The published data have not been consistent, but overall, there are convincing studies showing that for reliable gene expression data from microarray or qPCR, RNA quality affects the validity of findings (Abasolo et al 2011; Popova et al 2008; Atz et al 2007). Although recent advances in expression profiling using RNA sequencing methods, particular using degraded RNA extracted from formalin fixed-paraffin embedded tissues, have suggested reliable expression profiling results can be obtained, it is noticeable that the manufacturers of RNA sequencing platforms still recommend the use of RNA with high degree of intactness as starting material (Hedegaard et al 2014).…”

Section: Discussionmentioning

confidence: 99%

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Characterization of RNA isolated from eighteen different human tissues: results from a rapid human autopsy program

et al. 2016

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Many factors affect the integrity of messenger RNA from human autopsy tissues including postmortem interval (PMI) between death and tissue preservation and the pre-mortem agonal and disease states. In this communication, we describe RNA isolation and characterization of 389 samples from 18 different tissues from elderly donors who were participants in a rapid whole-body autopsy program located in Sun City, Arizona (www.brainandbodydonationprogram.org). Most tissues were collected within a PMI of 2–6 h (median 3.15 h; N = 455), but for this study, tissue from cases with longer PMIs (1.25–29.25 h) were included. RNA quality was assessed by RNA integrity number (RIN) and total yield (ng RNA/mg tissue). RIN correlated with PMI for heart (r = −0.531, p = 0.009) and liver (r = −558, p = 0.0017), while RNA yield correlated with PMI for colon (r = −485, p = 0.016) and skin (r = −0.460, p = 0.031). RNAs with the lowest integrity were from skin and cervix where 22.7 and 31.4 % of samples respectively failed to produce intact RNA; by contrast all samples from esophagus, lymph node, jejunum, lung, stomach, submandibular gland and kidney produced RNA with measurable RINs. Expression levels in heart RNA of 4 common housekeeping normalization genes showed significant correlations of Ct values with RIN, but only one gene, glyceraldehyde-3 phosphate dehydrogenase, showed a correlation of Ct with PMI. There were no correlations between RIN values obtained for liver, adrenal, cervix, esophagus and lymph node and those obtained from corresponding brain samples. We show that high quality RNA can be produced from most human autopsy tissues, though with significant differences between tissues and donors. The RNA stability and yield did not depend solely on PMI; other undetermined factors are involved, but these do not include the age of the donor.

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Section: Discussionmentioning

confidence: 99%

“…Our preliminary estimation showed no correlation between brain RNA RIN values and those from 6 other tissues. It remains an accepted practice and goal to prepare RNA of high integrity for downstream studies, and having as short a PMI as possible is still advisable for this purpose (Abasolo et al 2011; Popova et al 2008; Atz et al 2007). …”

Section: Discussionmentioning

confidence: 99%

Characterization of RNA isolated from eighteen different human tissues: results from a rapid human autopsy program

et al. 2016

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“…Since it has been reported that GAPDH expression is altered in neurodegenerative diseases [11], we evaluated several endogenous controls by qRT-PCR in the frontal cortex regions and RPLP0 was chosen because this transcript presented overall low C t values and lowest C t variation (unpublished data). In addition, RPLP0 has been used as a stable reference gene in previous studies [1, 16, 31]. …”

Section: Methodsmentioning

confidence: 99%

Aggregation-prone c9FTD/ALS poly(GA) RAN-translated proteins cause neurotoxicity by inducing ER stress

et al. 2014

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The occurrence of repeat-associated non-ATG (RAN) translation, an atypical form of translation of expanded repeats that results in the synthesis of homopolymeric expansion proteins, is becoming more widely appreciated among microsatellite expansion disorders. Such disorders include amyotrophic lateral sclerosis and frontotemporal dementia caused by a hexanucleotide repeat expansion in the C9ORF72 gene (c9FTD/ALS). We and others have recently shown that this bidirectionally transcribed repeat is RAN translated, and the “c9RAN proteins” thusly produced form neuronal inclusions throughout the central nervous system of c9FTD/ALS patients. Nonetheless, the potential contribution of c9RAN proteins to disease pathogenesis remains poorly understood. In the present study, we demonstrate that poly(GA) c9RAN proteins are neurotoxic and may be implicated in the neurodegenerative processes of c9FTD/ALS. Specifically, we show that expression of poly(GA) proteins in cultured cells and primary neurons leads to the formation of soluble and insoluble high molecular weight species, as well as inclusions composed of filaments similar to those observed in c9FTD/ALS brain tissues. The expression of poly(GA) proteins is accompanied by caspase-3 activation, impaired neurite outgrowth, inhibition of proteasome activity, and evidence of endoplasmic reticulum (ER) stress. Of importance, ER stress inhibitors, salubrinal and TUDCA, provide protection against poly(GA)-induced toxicity. Taken together, our data provide compelling evidence towards establishing RAN translation as a pathogenic mechanism of c9FTD/ALS, and suggest that targeting the ER using small molecules may be a promising therapeutic approach for these devastating diseases.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-014-1336-5) contains supplementary material, which is available to authorized users.

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“…This gene was chosen as the reference gene due to general stability and minimal variation across diagnostic groups [30]. We found no significant differences in gene expression of GAPDH in BA9 [F(2, 16) = 0.96, p = 0.40] or in BA24 [F(2, 15) = 3.1, p = 0.08] across diagnostic groups in this cohort of subjects.…”

Section: Methodsmentioning

confidence: 83%

SNAP-25a/b Isoform Levels in Human Brain Dorsolateral Prefrontal Cortex and Anterior Cingulate Cortex

et al. 2015

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SNAP-25 is a neurotransmitter vesicular docking protein which has been associated with brain disorders such as attention deficit hyperactivity disorder, bipolar disorder and schizophrenia. In this project, we were interested if clinical factors are associated with differential SNAP-25 expression. We examined the SNAP-25 isoform mRNA and protein levels in postmortem cortex Brodmann's area 9 (BA9) and BA24 (n = 29). Subjects were divided by psychiatric diagnosis, clinical variables including mood state in the last week of life and lifetime impulsiveness. We found affected subjects with a diagnosis of alcohol use disorder (AUD) had a lower level of SNAP-25b BA24 protein compared to those without AUD. Hispanic subjects had lower levels of SNAP-25a, b and BA9 mRNA than Anglo-American subjects. Subjects who smoked had a total pan (total) SNAP-25 BA9/BA24 ratio. Subjects in the group with a low level of anxious-psychotic symptoms had higher SNAP-25a BA24 mRNA compared to normal controls, and both the high and low symptoms groups had higher pan (total) SNAP-25 BA9/BA24 ratios than normal controls. These data expand our understanding of clinical factors associated with SNAP-25. They suggest that SNAP-25 total and isoform levels may be useful biomarkers beyond limited neurological and psychiatric diagnostic categories.

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RT-qPCR study on post-mortem brain samples from patients with major psychiatric disorders: Reference genes and specimen characteristics

Cited by 22 publications

References 31 publications

Characterization of RNA isolated from eighteen different human tissues: results from a rapid human autopsy program

Characterization of RNA isolated from eighteen different human tissues: results from a rapid human autopsy program

Aggregation-prone c9FTD/ALS poly(GA) RAN-translated proteins cause neurotoxicity by inducing ER stress

SNAP-25a/b Isoform Levels in Human Brain Dorsolateral Prefrontal Cortex and Anterior Cingulate Cortex

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