Abstract
Prefrontal cortical regions play a key role in behavioural regulation, which is profoundly disturbed in suicide. The study was carried out on frozen cortical samples from the anterior cingulate cortex (dorsal and ventral parts, ACd and ACv), the orbitofrontal cortex (OFC), and the dorsolateral cortex (DLC) obtained from 20 suicide completers (predominantly violent) with unknown psychiatric diagnosis and 21 non-suicidal controls. The relative level of ribosomal RNA (rRNA) as a marker of the transcriptional activity of ribosomal DNA (rDNA) was evaluated bilaterally in prefrontal regions mentioned above (i.e. in eight regions of interest, ROIs) by reverse transcription and quantitative polymerase chain reaction (RT-qPCR). The overall statistical analysis revealed a decrease in rDNA activity in suicide victims versus controls, particularly in male subjects. Further ROI-specific post hoc analyses revealed a significant decrease in this activity in suicides compared to non-suicides in five ROIs. This effect was accentuated in the ACv, where it was observed bilaterally. Our findings suggest that decreased rDNA transcription in the prefrontal cortex plays an important role in suicide pathogenesis and corresponds with our previous morphometric analyses of AgNOR-stained neurons.
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Introduction
Prefrontal cortex (PFC) regions play a key role in behavioural regulation, which is profoundly disturbed in suicide victims. Impaired executive functions with poor impulse control, problem-solving and decision-making are the outstanding manifestations of PFC dysfunction preceding suicidal behaviour (for reviews see: [1, 2]). Numerous neurobiological data are suggestive of the distinctness of suicide in mental disorders (for reviews see: [1, 2]), which has been considered in the fifth edition of the Diagnostic and Statistical Manual of Mental Disorders—DSM V [3]. Neuropathological studies of our workgroup have also suggested this distinctness [4,5,6,7] (for a review of our previous studies see: [8]).
The transcription of ribosomal genes is crucial for key cellular functions in the brain, inter alia, the outgrowth of neurites [9,10,11] and myelination process [12], both deteriorated in the PFC of suicide victims ([13] and [14], respectively). Disturbed ribosomal function in the PFC in suicide has been highlighted in recent exploratory microarray analysis of genetic variants in suicide completers [15]. As revealed by molecular research, rDNA transcription is attenuated in the hippocampus in cases with a history of child abuse and this effect is related to epigenetic modulations [16], which are also observed in the PFC [17, 18]. Similar results were obtained in the animal model of early-life stress [19], which is one of well-established and strongest distal factors in stress-diathesis model of suicidal behaviour [1, 2].
As rDNA transcription is affected by multiple factors [20], there are numerous molecular signatures involved in the dysregulation of this key cellular process in suicide, which have been revealed recently. Besides epigenetic phenomena, current molecular suicide research points to disturbances of stress axis components [21, 22] (for a review see: [23]), malfunction of trophic factors [21] (for a review see: [24]), glutamatergic system diathesis [25,26,27], abnormal glial–neuronal interaction [14, 28,29,30,31] and cross-talk between immune system and brain [32, 33].
Our previous morphological studies by the AgNOR staining method have suggested the attenuated rDNA transcription in prefrontal pyramidal neurons as a phenomenon specific for suicidal patients with established diagnoses of unipolar or bipolar depression [34, 35]. Our recent AgNOR study has suggested the same effect in forensic cohort of suicide victims with unknown psychiatric diagnosis [4]. Silver-stained nucleolar organising regions (AgNORs) clustered together in the nucleolus in the AgNOR area, which is observed by light microscopy, represent the site of rDNA transcription in human interphase cells [4, 34, 35]. The transcriptional activity of rDNA can be assessed by measuring AgNOR parameters, among them the AgNOR area, which is decreased in prefrontal pyramidal neurons of suicide completers [4].
The distinctness of suicide-specific abnormalities in the PFC has been supported by a study on microglial reaction from a workgroup in Magdeburg, which was found to be increased in suicidal patients from different groups of mental disorders [28]. This effect has been verified by studies of other workgroups [29, 30]. The oxidative stress as a deleterious consequence of microglia hyperactivity in turn may lead to the decreased rDNA transcription in affected cellular elements of the PFC [12, 36, 37].
Therefore, in the present study, we hypothesized a decreased rDNA transcriptional activity in prefrontal regions of suicide completers regardless of their underlying psychiatric diagnosis (i.e. independent of psychiatric comorbidity) and tested this hypothesis by the application of reverse transcription and quantitative polymerase chain reaction (RT-qPCR) in forensic postmortem material. We aimed at both basic research on the neurobiology of suicide and the informative comparison between molecular and morphological evaluation of rDNA transcription in the same pool of brain samples [4], which has not been reported previously.
Materials and methods
Human brain tissue
Prefrontal parts of both hemispheres of 20 suicide victims (14 males/6 females) with unknown data both on psychiatric comorbidity and on possible psychotropic medication preceding death (typical for most of suicide cases autopsied in our Department of Forensic Medicine) and 21 (17 males/4 females) controls were obtained during routine forensic autopsies in accordance with existing EU law regulations. The study has been approved by the local ethics committee of the Medical University of Gdańsk as performed in accordance with the ethical standards laid down in the Declaration of Helsinki of 1989.
The detailed diagnostic and demographic data of investigated cases are present in the Supplementary Table. Violent suicide methods prevailed in the suicide cohort (13 out of 20), which is representative for our autopsy material. Control cases of unnatural manner of death were more numerous than those of natural manner (13 and 8, respectively). Only sudden death cases were investigated in suicide and control cohorts. All brains were free of gross neuropathology suggestive of vascular, traumatic, inflammatory, neoplastic and neurodegenerative processes. Macroscopic evaluation of brains was confirmed by histopathological investigation in cases, where the cause of death was unclear at autopsy and the routine histopathological evaluation of internal organs was necessary for the forensic diagnosis (i.e. in seven non-violent suicide cases of self-poisoning by medication overdose and eight control cases of natural manner of death). Neither chronic nor acute pathological processes were observed microscopically in these cases in neocortical areas and other brain regions in hematoxylin–eosin-stained sections. Among others, neuronal necrosis as a consequence of protracted antemortem hypoxia was excluded by histopathological investigation. Neurodegenerative changes, such as amyloid plaques, perivascular amyloid deposits and neurofibrillary tangles, were not observed microscopically in the AgNOR staining in prefrontal regions in the same cohorts investigated previously [4]. Blood and urine were tested for the presence of ethanol at each autopsy. The majority of investigated cases (17 suicide victims and 15 controls) revealed the blood alcohol concentration (BAC) below the limit of quantification (LOQ), i.e. < 0.2 g/l according to internationally accepted analytical guidelines. The remaining three suicide victims and 6 controls revealed BAC in the range of 0.59–3.4 g/l (the highest value in a case of self-poisoning by quetiapine) and 0.74–3.15 g/l (the highest value in one of stabbed cases), respectively. Other substances of abuse, antidepressant and antipsychotic drugs, as well as their metabolites were investigated when intoxication was suggested by the scene inspection and/or other available information sources prior to the autopsy, i.e. in seven cases. These cases constituted the non-violent suicide subgroup.
Prefrontal parts of the brains were separated at forensic autopsies from both hemispheres by coronal sections at the level of temporal poles. Immediately after the separation, cortical samples for the molecular rRNA assays were isolated closely to the section plane from the following prefrontal regions: dorsolateral prefrontal cortex (DLC), anterior cingulate cortex (dorsal (ACd) and ventral (ACv) parts) and orbitofrontal cortex (OFC) (see Fig. 1). After isolation of the samples, the remaining prefrontal parts were fixed for the morphological investigations of the same cortical regions by the AgNOR staining method, which were presented previously [4].
rRNA isolation and reverse transcription
Immediately after the separation from the brain, cortical samples were vortexed, double-centrifuged (60 s, 5 000 RCF, room temperature) and incubated overnight at 4–8 °C in RNA-stabilising RNAlater Solution (Thermo Fisher Scientific, USA). Subsequently, they were stored in this solution at – 54 °C until RNA extraction. All samples were blinded by pseudonymization through assigning consecutive case numbers regardless of the forensic diagnosis of suicide vs. non-suicide.
Total RNA present in the frozen samples was extracted with a GeneMATRIX Universal DNA/RNA/Protein Purification Kit (EURx, Poland) with an additional step of treatment of RNA isolate with desoxyribonuclease I (DNAse I). RNA concentration was measured spectrophotometrically in a NanoDrop ND-1000 UV–Vis spectrophotometer (NanoDrop Technologies, USA). RNA (100 ng) was reversely transcribed with a TranScriba Kit (A&A Biotechnology, Poland) with an RNase inhibitor and random hexamer oligonucleotides as primers according the producer’s instructions. As opposed to oligo(dT) primers commonly used in routine reverse transcription, random hexanucleotides enable cDNA synthesis for rRNA, which is not stabilised in the cell via polyadenylation. Final cDNA concentration of 5 ng/µl after the reverse transcription step was sufficient for quantitative PCR.
Real-time quantitative polymerase chain reaction
Fragments of RNA18S5 (RNA, 18S ribosomal 5) as a target gene and GAPDH (glyceraldehyde 3-phosphate dehydrogenase) as a reference gene were amplified simultaneously in one 5 µL reaction by duplex qPCR in a 7900HT Fast Real-Time PCR System instrument (Applied Biosystems, USA) with the use of TaqMan Gene Expression Master Mix (Thermo Fisher Scientific) and predesigned TaqMan Gene Expression Assays: Hs03928990 with a FAM-labelled TaqMan probe specific for RNA18S5, and Hs03929097 with a VIC-labelled TaqMan probe specific for GAPDH (Thermo Fisher Scientific). The housekeeping gene GAPDH was proven to be one of the most stable reference genes [38] used for normalisation of gene expression data also in current molecular suicide research on prefrontal samples [39, 40]. All prefrontal samples and non-template contamination controls were analysed in four replicates.
Amplification was performed in standard mode thermal cycling conditions with initial 2-min uracil-DNA glycosylase (UDG) incubation (50 °C) for prevention of carry-over PCR contamination and with 10-min AmpliTaq Gold DNA polymerase activation (95 °C). Due to the late signal of amplification of the reference GAPDH gene (resulting from the high cellular expression level of rDNA), real-time amplification data were collected for as many as 80 qPCR cycles. PCR parameters were set according to the manufacturer’s instructions: 15 s of denaturation at 95 °C followed by 1 min of annealing and elongation at 60 °C.
Data analysis
The relative expression level of rDNA was estimated by comparison of the expression level of RNA18S5 to the expression level of GAPDH from values of a threshold cycle (CT) in which fluorescence of fluorophore released during qPCR exceeded the background level. The analysis was performed with the use of SDS v2.2 software (Applied Biosystems) with automatic setting of the baseline and threshold values for the determination of CT from amplification curves.
Statistical analyses were performed with the data analysis software system STATISTICA version 10 (StatSoft®, Inc. 2011, www.statsoft.com). As normal distribution was not given for analysed data (i.e. significant values of Kolmogorov–Smirnov tests were obtained), non-parametric statistical procedures were used in hierarchic mode.
First, STATISTICA generalized linear/nonlinear models (GLZ) module containing general custom designs (GCD) procedure was applied as an omnibus method to analyse associations between dependent variable, i.e. the relative rRNA level and independent categorical variables (i.e. suicidal/control group, PFC region, cortical layer, and sex as the categorical confounding variable). The results of the GCD analyses were reported automatically including the Wald statistic value, degrees of freedom, and the respective P value.
Age, postmortem interval, brain weight and BAC (values below LOQ were accounted null values in statistical analysis) were considered as numerical confounding variables. Therefore, the subsequent GCD procedure was applied in each of ROIs to analyse associations between the relative rRNA level and these variables. Supplementary to GCD analyses in ROIs, Spearman’s correlation coefficients were calculated to determine the impact of numerical variables which might confound the dependent variable. These coefficients were also calculated to determine correlations between rRNA levels and AgNOR area values in prefrontal pyramidal neurons found previously [4].
Following the GCD analyses, unadjusted two-way post hoc comparisons with Mann–Whitney U test and the χ2 test were used to detect the possible differences between the studied groups with respect to the variables mentioned above (i.e. rRNA level and confounders). All statistical tests were two-tailed. In general, P values of < 0.05 were accepted as statistically significant.
Kruskal–Wallis analysis of the variance of ranks (H test) with subsequent U tests was performed for the evaluation of differences in the rRNA level related to sex between suicides and controls, which was suggested by the GCD analysis; in this procedure, U test P values were adjusted to multiple comparisons according to the Bonferroni correction. The same procedure was applied for the evaluation of differences in dependent variable between subgroups of suicide victims (violent vs. non-violent) and controls (unnatural vs. natural manner of death).
Results
The analysis of rRNA relative levels
Cumulative analysis of results from all 8 investigated ROIs (i.e. from all cortical samples obtained from 4 evaluated PFC regions bilaterally, 159 suicidal and 167 control values) by GCD procedure revealed the association of forensic diagnosis (i.e. suicide vs. non-suicide) with the rRNA relative level (Wald statistic = 14.951, df = 1, P = 0.000110, median values 6.309 and 9.153 for suicides and non-suicidal controls, respectively, see Table 1). The effect of diagnosis was not associated with the brain hemisphere or investigated PFC region. Instead, it was associated with sex, i.e. mainly driven by male subjects (see Tables 1 and 2, and the next paragraph).
Further ROI-specific analyses by U tests revealed rRNA decrease in suicides compared to non-suicides in all eight ROIs. This effect was significant in five of them and was accentuated in the ACv, where it was observed bilaterally (see Table 2).
No significant differences existed in the rRNA level between violent (n = 13) and non-violent (n = 7) suicide victims as well as between control cases of unnatural (n = 13) and natural manner of death (n = 8) in any of ROIs (non-significant U test P values corrected for multiple comparisons).
Confounders
Suicidal and control groups were matched by gender (non-significant χ2 test P value, Table 1 and Supplementary Table) and no significant intra-group differences between sexes were found in the rRNA relative level in any of analysed ROIs (non-significant U test P values corrected for multiple comparisons). However, according to the associated effect of diagnosis and sex suggested by the initial GCD procedure, only males revealed an association between forensic diagnosis and rRNA level in the cumulative analysis of all investigated ROIs (see Table 1). Following this initial evaluation, only male suicide victims revealed significant decreases in the rRNA level compared to male controls in ROI-specific analyses (significant U test P values corrected for multiple comparisons in the right ACv and OFC, see Table 2). Therefore, the observed phenomenon was specific for males.
Age and PMI revealed no significant differences between suicides and controls (non-significant U test P values, Table 1 and Supplementary Table), similar to brain weight and BAC. ROI-specific analyses by GCD procedure revealed no associated impact of any of those numerical confounders and forensic diagnosis on rRNA levels (non-significant Wald statistic P values) in any of investigated ROIs. Correspondingly, no relevant correlations were found between numerical confounders and rRNA level in any ROI in both groups (non-significant Spearman’s correlation P values and/or irrelevant r values). Therefore, the observed differences in the rRNA level between suicides and controls both in the cumulative and ROI-specific analyses were not confounded by these variables.
Correlation with previous AgNOR findings
An additional correlation analysis was performed between rRNA levels and mean values of AgNOR area calculated from the values of this parameter in prefrontal pyramidal neurons in layers III and IV, which were obtained previously [4]. This analysis revealed significant negative correlations in control group, whereas no correlations between current molecular and previous morphological results were found in any ROI in suicide victims (non-significant Spearman’s correlation P values and irrelevant r values, see Table 3).
Discussion
We have found the decreased rDNA transcription in PFC regions in suicide victims, which was specific for male suicides, similarly to our previous study of prefrontal pyramidal neurons by the AgNOR staining method [4]. There are different possible interpretations of this finding. Experimental studies in animal models of chronic stress, which is an established proximal factor in suicide [1, 2], revealed in prefrontal pyramidal neurons sex-specific differences in the formation of dendrites (for a review see: [41]), i.e. in the process closely related to rDNA activity [9,10,11]. Correspondingly, different expression of genes related to the formation of neuronal processes has been observed in the PFC of male compared to female suicide completers [42, 43]. However, the small sample size of female subjects may lead to an underpowered analysis and there is a chance of false negative results. Therefore, the results should be replicated in larger cohorts with more numerous female samples. Other variables which may confound present results, among them postmortem interval, did not influence our current results. The observed accentuated intra-group differences in rDNA expression profiles in bulk cortical tissue may be a consequence of differences in cellular composition related to the variation in gray/white matter ratios in the extracted tissue samples and/or inter-subject variability [44]. Our method does not allow to explain these differences.
The rDNA transcription was globally decreased in suicide in analysed prefrontal regions, which was accentuated in the cumulative evaluation of all ROIs simultaneously (see Table 1) and this effect was associated with neither the hemisphere nor the prefrontal region. This finding could be explained by close reciprocal connectivity between PFC areas which constitute a functional syncytium [45]. However, further region-specific post hoc analyses revealed significant bilateral decrease in ribosomal gene expression only in the ACv (see Table 1), i.e. in prefrontal region crucial for behavioural regulation [46]. Therefore, both our current molecular findings and previous morphological reports [4, 34, 35, 47] are in line with the aberrant regulatory function of the ventromedial PFC accentuated in suicidal behaviour [1, 2].
Our previous AgNOR studies of cohorts with both established diagnoses of affective disorders [34, 35] and unknown psychiatric comorbidity [4] suggested a decreased rDNA transcription in prefrontal pyramidal neurons as a phenomenon specific for suicide completers and our current results correspond with them. However, we have not found positive correlations between AgNOR area in these neurons and rRNA levels in samples of bulk cortical tissue in both controls and suicide victims. There are some possible explanations of this discrepancy. Pyramidal neurons constitute roughly 30% of prefrontal cells, and layer III and V neurons investigated in our study [4] are only part of them, whereas other cellular populations in the PFC include inhibitory interneurons (10%), oligodendrocytes (45%), astrocytes (12%) and microglia (3%) [48, 49]. Thus, abnormalities in other cell types besides pyramidal neurons, predominantly in oligodendrocytes, may also contribute to the decrease of rRNA synthesis in bulk PFC tissue observed in suicide victims. This hypothesis corresponds with previous morphological reports on cohorts including suicides with established diagnoses of affective disorders or schizophrenia, where decreased numbers of oligodendrocytes were found in the PFC [50, 51]. Interestingly, negative correlations between AgNOR area and rRNA level were found in controls, whereas they were lacking in suicide victims (see Table 3). The observed effect could suggest a reciprocal regulation of rRNA synthesis in prefrontal pyramidal neurons and oligodendrocytes in normal human brain. According to recent research, a molecular cross-talk exists between these cell types (for a review see: [52]). However, further molecular analysis of distinct cellular populations in the PFC is needed to explain the relation between rDNA transcriptional activity in pyramidal neurons and oligodendrocytes.
The silver staining in AgNOR areas in interphase cells is related predominantly to the multifunctional protein nucleolin (important for the function of RNA-polymerase I) but not directly to the rRNA amount [53, 54]. This could be a further possible explanation of the discrepancy between our current molecular and previous morphological study. As numerous factors are involved in the regulation of rRNA synthesis crucial for cellular functions [20], the amount of nucleolin may not correlate directly with the level of rDNA transcription. On the one hand, molecular suicide research suggests that rDNA transcription in the PFC may be downregulated more directly by the hypermethylation of rDNA promoter region, i.e. by the epigenetic phenomenon [17, 18]. On the other hand, however, the key node in the intracellular regulation of rDNA transcription is the mammalian target to rapamycin (mTOR) [55], which is disturbed in the PFC in suicide [56]. Moreover, suicide research revealed abnormalities of different molecular factors related to the function of mTOR.
Among the most important are molecular components of stress axis, which is profoundly disturbed in suicidal behaviour [1, 2]. Both corticotropin-releasing hormone receptor 1 [22] and glucocorticoid receptor (for a review see: [23]) are decreased in the PFC in suicide, which may impact rDNA transcription [57,58,59]. The abnormalities in molecules regulating glucocorticoid receptor function, which have been found in suicide [21, 60, 61], may also contribute to the attenuated rRNA synthesis in prefrontal pyramidal cells and oligodendrocytes [21].
The stress axis function is closely related to glutamatergic neurotransmission [57], which is disturbed in suicidal behaviour in the PFC [25,26,27]. Impaired function of brain-derived neurotrophic factor (BDNF) is an important molecular effect of this disturbed neurotransmission [62]. The decrease of BDNF has been found in prefrontal regions of suicide victims (for a review see: [24]), which may inhibit rDNA transcription in neurons [62] and oligodendrocytes [12].
Besides glucocorticoids, glutamatergic neurotransmission is profoundly influenced by activated microglia producing pro-inflammatory cytokines [63]. Both microglial reaction [28,29,30] and these cytokine levels [32, 33] are increased in the PFC of suicide victims. Subsequent oxidative stress may attenuate rRNA synthesis in pyramidal neurons [64, 65] and oligodendrocytes [12, 66] leading to disturbed synaptic formation [65] and deteriorated myelin production [14], respectively.
Limitations
The present study has certain limitations that have to be considered: (1) A relatively small number of predominantly male cases was analyzed. Therefore, results have to be confirmed in a larger sample with more numerous female subjects. (2) The psychiatric diagnoses (also including substance use disorders) and the data on possible psychotropic medication preceding death were not available. The levels of psychotropic drugs were established only in seven suicide victims where medication overdose constituted a cause of death. However, our current study did not aim at analysis of relation between suicide and other mental disorders and our previous studies did not suggest that the decreased rDNA activity in prefrontal pyramidal neurons may be related to the medication used in the last three months of life [13, 15]. (3) As we used bulk tissue homogenates, our method does not allow to differentiate between cell types, in which the observed phenomenon of decreased rDNA transcription occurs. (4) Our relative rDNA expression estimates are based on assumption of stable housekeeping GAPDH gene expression across subjects and samples.
Conclusion
In summary, our results suggest decreased rDNA transcription in the PFC in male suicide victims as a presumable consequence of multiple molecular events. The molecular results correspond with our previous morphological research on PFC pyramidal neurons in suicide based on the AgNOR staining method.
Data availability
On behalf of all authors, the corresponding author states that the data being reported are accurate and are coming from the official source.
References
van Heeringen K, Mann JJ (2014) The neurobiology of suicide. Lancet Psychiatry 1:63–72
Turecki G, Brent DA (2016) Suicide and suicidal behaviour. Lancet 387:1227–1239
American Psychiatric Association (2013) Diagnostic and statistical manual of mental disorders, 5th edn. American Psychiatric Publishing, Arlington
Krzyżanowska M, Steiner J, Pieśniak D, Karnecki K, Kaliszan M, Wiergowski M, Rębała K, Brisch R, Braun K, Jankowski Z, Kosmowska M, Chociej J, Gos T (2020) Ribosomal DNA transcription in prefrontal pyramidal neurons is decreased in suicide. Eur Arch Psychiatry Clin Neurosci 270:859–867
Brisch R, Steiner J, Mawrin C, Krzyżanowska M, Jankowski Z, Gos T (2017) Microglia in the dorsal raphe nucleus plays a potential role in both suicide facilitation and prevention in affective disorders. Eur Arch Psychiatry Clin Neurosci 267:403–415
Krzyżanowska M, Steiner J, Brisch R, Mawrin C, Busse S, Karnecki K, Jankowski Z, Gos T (2016) Decreased ribosomal DNA transcription in dorsal raphe nucleus neurons is specific for suicide regardless of psychiatric diagnosis. Psychiatry Res 241:43–46
Krzyżanowska M, Steiner J, Karnecki K, Kaliszan M, Brisch R, Wiergowski M, Braun K, Jankowski Z, Gos T (2016) Decreased ribosomal DNA transcription in dorsal raphe nucleus neurons differentiates between suicidal and non-suicidal death. Eur Arch Psychiatry Clin Neurosci 266:217–224
Gos T, Steiner J, Jankowski Z, Bogerts B (2013) Postmortem-assessed impairment of neuronal activity in depression: the dominant impact of suicide. CNS Neurol Disord Drug Targets 12:930–935
Gomes C, Smith SC, Youssef MN, Zheng JJ, Hagg T, Hetman M (2011) RNA polymerase 1-driven transcription as a mediator of BDNF-induced neurite outgrowth. J Biol Chem 286:4357–4363
Slomnicki LP, Pietrzak M, Vashishta A, Jones J, Lynch N, Elliot S, Poulos E, Malicote D, Morris BE, Hallgren J, Hetman M (2016) Requirement of neuronal ribosome synthesis for growth and maintenance of the dendritic tree. J Biol Chem 291:5721–5739
Vashishta A, Slomnicki LP, Pietrzak M, Smith SC, Kolikonda M, Naik SP, Parlato R, Hetman M (2018) RNA polymerase 1 is transiently regulated by seizures and plays a role in a pharmacological kindling model of epilepsy. Mol Neurobiol 55:8374–8387
Figlia G, Gerber D, Suter U (2018) Myelination and mTOR. Glia 66:693–707
Hercher C, Canetti L, Turecki G, Mechawar N (2010) Anterior cingulate pyramidal neurons display altered dendritic branching in depressed suicides. J Psychiatr Res 44:286–293
Lutz PE, Tanti A, Gasecka A, Barnett-Burns S, Kim JJ, Zhou Y, Chen GG, Wakid M, Shaw M, Almeida D, Chay MA, Yang J, Larivière V, M’Boutchou MN, van Kempen LC, Yerko V, Prud’homme J, Davoli MA, Vaillancourt K, Théroux JF, Bramoullé A, Zhang TY, Meaney MJ, Ernst C, Côté D, Mechawar N, Turecki G (2017) Association of a history of child abuse with impaired myelination in the anterior cingulate cortex: convergent epigenetic, transcriptional, and morphological evidence. Am J Psychiatry 174:1185–1194
Rodríguez-López ML, Martínez-Magaña JJ, Cabrera-Mendoza B, Genis-Mendoza AD, García-Dolores F, López-Armenta M, Flores G, Vázquez-Roque RA, Nicolini H (2019) Exploratory analysis of genetic variants influencing molecular traits in cerebral cortex of suicide completers. Am J Med Genet B Neuropsychiatr Genet 183:26–37
McGowan PO, Szyf M (2010) The epigenetics of social adversity in early life: Implications for mental health outcomes. Neurobiol Dis 39:66–72
Haghighi F, Xin Y, Chanrion B, O’Donnell AH, Ge Y, Dwork AJ, Arango V, Mann JJ (2014) Increased DNA methylation in the suicide brain. Dialogues Clin Neurosci 16:430–438
Schneider E, El Hajj N, Müller F, Navarro B, Haaf T (2015) Epigenetic dysregulation in the prefrontal cortex of suicide completers. Cytogenet Genome Res 146:19–27
Wei L, Hao J, Kaffman A (2014) Early life stress inhibits expression of ribosomal RNA in the developing hippocampus. PLoS ONE 9:e115283
Neumüller RA, Gross T, Samsonova AA, Vinayagam A, Buckner M, Founk K, Hu Y, Sharifpoor S, Rosebrock AP, Andrews B, Winston F, Perrimon N (2013) Conserved regulators of nucleolar size revealed by global phenotypic analyses. Sci Signal 6:70
Nagy C, Maitra M, Tanti A, Suderman M, Théroux JF, Davoli MA, Perlman K, Yerko V, Wang YC, Tripathy SJ, Pavlidis P, Mechawar N, Ragoussis J, Turecki G (2020) Single-nucleus transcriptomics of the prefrontal cortex in major depressive disorder implicates oligodendrocyte precursor cells and excitatory neurons. Nat Neurosci 23:771–781
Pandey GN, Rizavi HS, Bhaumik R, Ren X (2019) Increased protein and mRNA expression of corticotropin-releasing factor (CRF), decreased CRF receptors and CRF binding protein in specific postmortem brain areas of teenage suicide subjects. Psychoneuroendocrinology 106:233–243
Pandey GN (2013) Biological basis of suicide and suicidal behavior. Bipolar Disord 15:524–541
Dwivedi Y (2012) Brain-derived neurotrophic factor in suicide pathophysiology. In: Dwivedi Y (ed) The neurobiological basis of suicide. CRC Press, New York, pp 139–158
Sequeira A, Mamdani F, Ernst C, Vawter MP, Bunney WE, Lebel V, Rehal S, Klempan T, Gratton A, Benkelfat C, Rouleau GA, Mechawar N, Turecki G (2009) Global brain gene expression analysis links glutamatergic and GABAergic alterations to suicide and major depression. PLoS ONE 4:e6585
Dean B, Gibbons AS, Boer S, Uezato A, Meador-Woodruff J, Scarr E, McCullumsmith RE (2016) Changes in cortical N-methyl-D-aspartate receptors and post-synaptic density protein 95 in schizophrenia, mood disorders and suicide. Aust N Z J Psychiatry 50:275–283
Zhao J, Verwer RWH, Gao SF, Qi XR, Lucassen PJ, Kessels HW, Swaab DF (2018) Prefrontal alterations in GABAergic and glutamatergic gene expression in relation to depression and suicide. J Psychiatr Res 102:261–274
Steiner J, Bielau H, Brisch R, Danos P, Ullrich O, Mawrin C, Bernstein HG, Bogerts B (2008) Immunological aspects in the neurobiology of suicide: elevated microglial density in schizophrenia and depression is associated with suicide. J Psychiatr Res 42:151–157
Schnieder TP, Trencevska I, Rosoklija G, Stankov A, Mann JJ, Smiley J, Dwork AJ (2014) Microglia of prefrontal white matter in suicide. J Neuropathol Exp Neurol 73:880–890
Torres-Platas SG, Cruceanu C, Chen GG, Turecki G, Mechawar N (2014) Evidence for increased microglial priming and macrophage recruitment in the dorsal anterior cingulate white matter of depressed suicides. Brain Behav Immun 42:50–59
Tanti A, Lutz PE, Kim J, O’Leary L, Théroux JF, Turecki G, Mechawar N (2019) Evidence of decreased gap junction coupling between astrocytes and oligodendrocytes in the anterior cingulate cortex of depressed suicides. Neuropsychopharmacology 44:2099–2111
Brundin L, Erhardt S, Bryleva EY, Achtyes ED, Postolache TT (2015) The role of inflammation in suicidal behaviour. Acta Psychiatr Scand 132:192–203
Pandey GN, Rizavi HS, Zhang H, Bhaumik R, Ren X (2018) Abnormal protein and mRNA expression of inflammatory cytokines in the prefrontal cortex of depressed individuals who died by suicide. J Psychiatry Neurosci 43:376–385
Gos T, Krell D, Brisch R, Bielau H, Trübner K, Bernstein HG, Bogerts B (2007) The changes of AgNOR parameters of anterior cingulate pyramidal neurons are region-specific in suicidal and non-suicidal depressive patients. World J Biol Psychiatry 8:245–255
Gos T, Krell D, Bielau H, Steiner J, Mawrin C, Trübner K, Brisch R, Bernstein HG, Jankowski Z, Bogerts B (2009) Demonstration of disturbed activity of orbitofrontal pyramidal neurons in depressed patients by the AgNOR staining method. J Affect Disord 118:131–138
Chen SH, Oyarzabal EA, Hong JS (2016) Critical role of the Mac1/NOX2 pathway in mediating reactive microgliosis-generated chronic neuroinflammation and progressive neurodegeneration. Curr Opin Pharmacol 26:54–60
Mayer C, Bierhoff H, Grummt I (2005) The nucleolus as a stress sensor: JNK2 inactivates the transcription factor TIF-IA and down-regulates rRNA synthesis. Genes Dev 19:933–941
Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3:research0034
Belzeaux R, Gorgievski V, Fiori LM, Lopez JP, Grenier J, Lin R, Nagy C, Ibrahim EC, Gascon E, Courtet P, Richard-Devantoy S, Berlim M, Chachamovich E, Théroux JF, Dumas S, Giros B, Rotzinger S, Soares CN, Foster JA, Mechawar N, Tall GG, Tzavara ET, Kennedy SH, Turecki G (2020) GPR56/ADGRG1 is associated with response to antidepressant treatment. Nat Commun 11:1635
Yoshino Y, Dwivedi Y (2020) Elevated expression of unfolded protein response genes in the prefrontal cortex of depressed subjects: effect of suicide. J Affect Disord 262:229–236
Hyer MM, Phillips LL, Neigh GN (2018) Sex differences in synaptic plasticity: hormones and beyond. Front Mol Neurosci 11:266
Labonté B, Engmann O, Purushothaman I, Menard C, Wang J, Tan C, Scarpa JR, Moy G, Loh YE, Cahill M, Lorsch ZS, Hamilton PJ, Calipari ES, Hodes GE, Issler O, Kronman H, Pfau M, Obradovic ALJ, Dong Y, Neve RL, Russo S, Kasarskis A, Tamminga C, Mechawar N, Turecki G, Zhang B, Shen L, Nestler EJ (2017) Sex-specific transcriptional signatures in human depression. Nat Med 23:1102–1111
Cabrera B, Monroy-Jaramillo N, Fries GR, Mendoza-Morales RC, García-Dolores F, Mendoza-Larios A, Diaz-Otañez C, Walss-Bass C, Glahn DC, Ostrosky-Wegman P, Fresno C, Nicolini H (2019) Brain gene expression pattern of subjects with completed suicide and comorbid substance use disorder. Mol Neuropsychiatry 5:60–73
Nido GS, Dick F, Toker L, Petersen K, Alves G, Tysnes OB, Jonassen I, Haugarvoll K, Tzoulis C (2020) Common gene expression signatures in Parkinson’s disease are driven by changes in cell composition. Acta Neuropathol Commun 8:55
Price JL, Drevets WC (2010) Neurocircuitry of mood disorders. Neuropsychopharmacology 35:192–216
Bush G, Luu P, Posner MI (2000) Cognitive and emotional influences in anterior cingulate cortex. Trends Cogn Sci 4:215–222
Steiner J, Walter M, Gos T, Guillemin GJ, Bernstein HG, Sarnyai Z, Mawrin C, Brisch R, Bielau H, Meyer zu Schwabedissen L, Bogerts B, Myint AM (2011) Severe depression is associated with increased microglial quinolinic acid in subregions of the anterior cingulate gyrus: evidence for an immune-modulated glutamatergic neurotransmission? J Neuroinflammation 8:94
Pelvig DP, Pakkenberg H, Stark AK, Pakkenberg B (2008) Neocortical glial cell numbers in human brains. Neurobiol Aging 29:1754–1762
Zikopoulos B, Barbas H (2013) Altered neural connectivity in excitatory and inhibitory cortical circuits in autism. Front Hum Neurosci 7:609
Uranova NA, Vostrikov VM, Orlovskaya DD, Rachmanova VI (2004) Oligodendroglial density in the prefrontal cortex in schizophrenia and mood disorders: a study from the Stanley Neuropathology Consortium. Schizophr Res 67:269–275
Vostrikov VM, Uranova NA, Orlovskaya DD (2007) Deficit of perineuronal oligodendrocytes in the prefrontal cortex in schizophrenia and mood disorders. Schizophr Res 94:273–280
Raabe FJ, Slapakova L, Rossner MJ, Cantuti-Castelvetri L, Simons M, Falkai PG, Schmitt A (2019) Oligodendrocytes as a new therapeutic target in schizophrenia: from histopathological findings to neuron-oligodendrocyte interaction. Cells 8:1496
Roussel P, Hernandez-Verdun D (1994) Identification of Ag-NOR proteins, markers of proliferation related to ribosomal gene activity. Exp Cell Res 214:465–472
Jia W, Yao Z, Zhao J, Guan Q, Gao L (2017) New perspectives of physiological and pathological functions of nucleolin (NCL). Life Sci 186:1–10
Mayer C, Grummt I (2006) Ribosome biogenesis and cell growth: mTOR coordinates transcription by all three classes of nuclear RNA polymerases. Oncogene 25:6384–6391
Jernigan CS, Goswami DB, Austin MC, Iyo AH, Chandran A, Stockmeier CA, Karolewicz B (2011) The mTOR signaling pathway in the prefrontal cortex is compromised in major depressive disorder. Prog Neuropsychopharmacol Biol Psychiatry 35:1774–1779
Polman JA, Hunter RG, Speksnijder N, van den Oever JM, Korobko OB, McEwen BS, de Kloet ER, Datson NA (2012) Glucocorticoids modulate the mTOR pathway in the hippocampus: differential effects depending on stress history. Endocrinology 153:4317–4327
Refojo D, Echenique C, Müller MB, Reul JM, Deussing JM, Wurst W, Sillaber I, Paez-Pereda M, Holsboer F, Arzt E (2005) Corticotropin-releasing hormone activates ERK1/2 MAPK in specific brain areas. Proc Natl Acad Sci U S A 102:6183–6188
Laplante M, Sabatini DM (2012) mTOR signaling in growth control and disease. Cell 149:274–293
Pandey GN, Rizavi HS, Zhang H, Bhaumik R, Ren X (2016) The expression of the suicide-associated gene SKA2 is decreased in the prefrontal cortex of suicide victims but not of nonsuicidal patients. Int J Neuropsychopharmacol 19:pyw015
Chen GG, Fiori LM, Moquin L, Gratton A, Mamer O, Mechawar N, Turecki G (2010) Evidence of altered polyamine concentrations in cerebral cortex of suicide completers. Neuropsychopharmacology 35:1477–1484
Ignácio ZM, Réus GZ, Arent CO, Abelaira HM, Pitcher MR, Quevedo J (2016) New perspectives on the involvement of mTOR in depression as well as in the action of antidepressant drugs. Br J Clin Pharmacol 82:1280–1290
Haroon E, Miller AH, Sanacora G (2017) Inflammation, glutamate, and glia: a trio of trouble in mood disorders. Neuropsychopharmacology 42:193–215
Choi HS, Ahn JH, Park JH, Won MH, Lee CH (2016) Age-dependent changes in the protein expression levels of Redd1 and mTOR in the gerbil hippocampus during normal aging. Mol Med Rep 13:2409–2414
Ota KT, Liu RJ, Voleti B, Maldonado-Aviles JG, Duric V, Iwata M, Dutheil S, Duman C, Boikess S, Lewis DA, Stockmeier CA, DiLeone RJ, Rex C, Aghajanian GK, Duman RS (2014) REDD1 is essential for stress-induced synaptic loss and depressive behavior. Nat Med 20:531–535
Maas DA, Vallès A, Martens GJM (2017) Oxidative stress, prefrontal cortex hypomyelination and cognitive symptoms in schizophrenia. Transl Psychiatry 7:e1171
Steiner J, Mawrin C, Ziegeler A, Bielau H, Ullrich O, Bernstein HG, Bogerts B (2006) Distribution of HLA-DR-positive microglia in schizophrenia reflects impaired cerebral lateralization. Acta Neuropathol 112:305–316
Acknowledgements
The research was supported by the Polish Ministry of Science and Higher Education. The authors would like to thank L. Cybulska and E. Kapińska for their excellent technical assistance.
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The study has been approved by the local ethics committee of the Medical University of Gdańsk as performed in accordance with the ethical standards laid down in the Declaration of Helsinki of 1989 (Approval No. NKBBN/195-319/2015).
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The authors dedicate this work to the memory of Professor Roman Hauser.
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Krzyżanowska, M., Rębała, K., Steiner, J. et al. Reduced ribosomal DNA transcription in the prefrontal cortex of suicide victims: consistence of new molecular RT-qPCR findings with previous morphometric data from AgNOR-stained pyramidal neurons. Eur Arch Psychiatry Clin Neurosci 271, 567–576 (2021). https://doi.org/10.1007/s00406-021-01232-4
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DOI: https://doi.org/10.1007/s00406-021-01232-4