The Heritability of Kidney Function Using an Older Australian Twin Population

Introduction: Twin studies are unique population models which estimate observed rather than inferred genetic components of complex traits. Nonmonogenic chronic kidney disease (CKD) is a complex disease process with strong genetic and environmental influences, amenable to twin studies. We aimed to assess the heritability of CKD using twin analysis and modeling within Older Australian Twin Study (OATS) data. Methods: OATS had 109 dizygotic (DZ) and 126 monozygotic (MZ) twin pairs with paired serum creatinine levels. Heritability of kidney function as estimated glomerular filtration rate (eGFR CKD Epidemiology Collaboration [CKD-EPI]) was modeled using the ACE model to estimate additive heritability (A), common (C), and unique (E) environmental factors. Intratwin pair analysis using mixed effects logistic regression allowed analysis of variation in eGFR from established CKD risk factors. Results: The median age was 69.71 (interquartile range 78.4–83.0) years, with 65% female, and a mean CKD-EPI of 82.8 ml/min (SD 6.7). The unadjusted ACE model determined kidney function to be 33% genetically determined (A), 18% shared genetic-environmental (C), and 49% because of unique environment (E). This remained unchanged when adjusted for age, hypertension, and sex. Hypertension was associated with eGFR; however, intertwin variance in hypertension did not explain variance in eGFR. Two or more hypertension medications were associated with decreased eGFR (P = 0.009). Conclusion: This study estimates observed heritability at 33%, notably higher than inferred heritability in genome-wide association study (GWAS) (7.1%–18%). Epigenetics and other genomic phenomena may explain this heritability gap. Difference in antihypertension medications explains part of unique environmental exposures, though discordance in hypertension and diabetes does not

Apoptosis and schizophrenia: a pilot study based on dermal fibroblast cell lines

Introduction: The aim of this study was to investigate whether there is an increased susceptibility to apoptosis in cultured fibroblasts from patients with schizophrenia

Gene Expression Analysis Implicates a Death Receptor Pathway in Schizophrenia Pathology

An increase in apoptotic events may underlie neuropathology in schizophrenia. By data-mining approaches, we identified significant expression changes in death receptor signaling pathways in the dorsolateral prefrontal cortex (DLPFC) of patients with schizophrenia, particularly implicating the Tumor Necrosis Factor Superfamily member 6 (FAS) receptor and the Tumor Necrosis Factor [ligand] Superfamily member 13 (TNFSF13) in schizophrenia. We sought to confirm and replicate in an independent tissue collection the noted mRNA changes with quantitative real-time RT-PCR. To test for regional and diagnostic specificity, tissue from orbital frontal cortex (OFC) was examined and a bipolar disorder group included. In schizophrenia, we confirmed and replicated significantly increased expression of TNFSF13 mRNA in the DLPFC. Also, a significantly larger proportion of subjects in the schizophrenia group had elevated FAS receptor expression in the DLPFC relative to unaffected controls. These changes were not observed in the bipolar disorder group. In the OFC, there were no significant differences in TNFSF13 or FAS receptor mRNA expression. Decreases in BH3 interacting domain death agonist (BID) mRNA transcript levels were found in the schizophrenia and bipolar disorder groups affecting both the DLPFC and the OFC. We tested if TNFSF13 mRNA expression correlated with neuronal mRNAs in the DLPFC, and found significant negative correlations with interneuron markers, parvalbumin and somatostatin, and a positive correlation with PPP1R9B (spinophilin), but not DLG4 (PSD-95). The expression of TNFSF13 mRNA in DLPFC correlated negatively with tissue pH, but decreasing pH in cultured cells did not cause increased TNFSF13 mRNA nor did exogenous TNFSF13 decrease pH. We concluded that increased TNFSF13 expression may be one of several cell-death cytokine abnormalities that contribute to the observed brain pathology in schizophrenia, and while increased TNFSF13 may be associated with lower brain pH, the change is not necessarily causally related to brain pH

CELL DEATH AND PROLIFERATION IN MENTAL DISORDER

The aetiology of two groups of psychiatric disorders, schizophrenia and the major depressive disorders, are poorly understood. Unlike neurodegenerative conditions, these two disorders have no obvious neuropathology and clues to their aetiology must be gleaned from other fields of research. The dominant models to date have been neurochemical in nature based on known actions of drugs that improve or mimic these conditions. However, the resulting monoaminergic hypotheses have been found to be over-simplistic and unable to comprehensively account for these complex psychiatric disorders. Based on epidemiological findings concerning perinatal factors and the typical adolescent age-of-onset, as well as the results of neuroimaging and neuropathological studies, research interest is now shifting to the processes involved in neurodevelopment, brain maturation and homeostasis. This thesis explores two such processes apoptosis in relation to schizophrenia, and neurogenesis in relation to depressive disorders. The negative associations between cancer, rheumatoid arthritis, and schizophrenia have led to the hypothesis that patients with schizophrenia have an increased susceptibility to programmed cell death (apoptosis). A study was carried out assessing several apoptotic markers in dermal fibroblast cell lines from groups of patients with schizophrenia, patients with non-schizophrenic psychotic disorder, and healthy comparison subjects. Apoptotic nuclei, caspase-3 activity, and protein levels of Bcl-2, Bax and P53P392Ser were quantified. Apoptosis was studied under basal cell culture conditions and after induction using the protein synthesis inhibitor, cycloheximide. The results suggested significant abnormalities in the regulation of apoptosis in schizophrenia that do not occur in non-schizophrenic psychotic disorder. It has been proposed that the therapeutic action of antidepressants is related to their effect on hippocampal neurogenesis mediated by brain-derived neurotrophic factor (BDNF). This hypothesis arose from the finding that blocking the action of BDNF in rodents leads to decreased neurogenesis and increased depressive behaviour. It has also been demonstrated that the neurotrophin receptor, p75NTR, is required for BDNF-induced differentiation of neural precursor cells in vitro, although no in vivo data have been published confirming this. A study was conducted to investigate the effect of running and treatment with the selective serotonin reuptake inhibitor fluoxetine (both positive modulators of neurogenesis) on the pool of hippocampal progenitor cells from which new neurons are derived. The pool of hippocampal progenitor cells was found to be a dynamic population, which varies in proliferative potential with the duration of antidepressant treatment. This variation may be attributable to changes in the number of progenitor cells occurring with prolonged induction of neurogenesis. Whether p75NTR plays a role in the regulation of neurogenesis was then determined. Neurogenesis was found to be reduced in p75NTR knock-out mice, providing us with a mouse model for the study of the role of neurogenesis in depression. Future work will depend on the identification of an appropriate behavioural model of depression that responds to chronic, and not acute, administration of antidepressant medication. A third stream of work involved application of high-throughput technology to investigate gene expression in psychiatric disorders. Clues to the aetiology of psychiatric disorders have been sought from microarray analysis of post-mortem brain tissue. In preparation for a microarray study of post-mortem tissue from a large sample of patients with schizophrenia and depressive disorder, post-mortem mRNA degradation was investigated in mouse brain tissue. A subgroup of mammalian mRNA transcripts was found to be particularly susceptible to post-mortem-related degradation, and to be more likely to carry the AUUUA motif in the 3 untranslated region. As transcription factors are more likely to carry this motif, this finding brings into question the suitability of post-mortem tissue for the study of apoptosis and cell proliferation. In conclusion, there is epidemiological, neuroimaging and neuropathological evidence that neurodevelopmental and brain maturational processes are involved in schizophrenia and depressive disorders. Supporting this view, the literature review identified a substantial number of laboratory studies implicating apoptosis in schizophrenia, and neurogenesis in depressive disorder. The series of studies conducted for this thesis added to this body of evidence. However, specific pathway abnormalities are yet to be determined. The complex multi-factorial nature of psychiatric disorder suggests that multiple molecular pathways will be implicated, and that future research needs to be conducted on much larger samples using multi-level assays (genotype - gene expression - protein levels - cellular function). Neurobiological studies of schizophrenia and major depression using large sample sizes, convergent experimental approaches, and employing high-throughput technologies will be required to achieve this aim

FAS receptor pathway in peripheral cells [<b>76</b>].

<p>The membrane FAS-R (blue) forms a trimer upon ligand binding (TNFSF13, red). The adaptor molecule FADD (green) then binds to the death domain of FAS-R through its own death domain. The amino terminus of FADD facilitates binding to caspase-8 (maroon oval). Caspase-8 self-activates through proteolytic cleavage into active caspase subunits, which can cleave other effector caspases (3, 6, 9), eventually leading to DNA degradation, membrane blebbing, and other hallmarks of apoptosis. In most cell types (Type II), caspase-8 catalyzes the cleavage of the pro-apoptotic BH3-only protein BID (orange) into its truncated form, tBID. tBID engages anti-apoptotic members of the family (e.g. BCL-2), allowing pro-apoptotic members (e.g. BAX) to translocate to the outer mitochondrial membrane, thus permeabilizing the membrane and facilitating release of pro-apoptotic proteins such as cytochrome c (tan). Cytochrome c release reinforces the cellular apoptotic drive through the intrinsic apoptotic pathway. Death receptors 4/5 (TNFSF10) appear to have identical intracellular pathway signaling to FAS receptor <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035511#pone.0035511-Lavrik1" target="_blank">[76]</a>.</p

Gene expression levels of TNFSF13-FAS receptor pathway genes of interest relative to three housekeeping genes in dorsolateral prefrontal cortex from patients with schizophrenia.

<p>cDNA samples obtained from the combined SMRI and NSW TRC collections from individuals with schizophrenia and unaffected controls were subjected to qRT-PCR. Expressions of TNFSF13 (A), FAS receptor (C), CFLAR_pan (E), CFLAR_long (F) and BID (G) relative to three housekeeping genes (β-actin, TATA box binding protein and ubiquitin C) were calculated using the deltadelta Ct method. Horizontal lines indicate the population median, except for panel (G) where they indicate the mean as those data were normally distributed. Frequency distributions for TNFSF13, FAS receptor and BID are displayed in panels (B), (D), and (H). ** p<0.01.</p

Gene expression levels of TNFSF13, FAS receptor and BID relative to three housekeeping genes in OFC from patients with schizophrenia, patients with bipolar disorder and unaffected controls.

<p>cDNA samples from individuals with schizophrenia, bipolar disorder and unaffected controls were subjected to qRT-PCR. Expressions of TNFSF13 (A), FAS receptor (B) and BID (C) relative to three housekeeping genes (β-actin, TATA box binding protein and ubiquitin C) were calculated using the deltadelta Ct method. Horizontal lines indicate the population median, except for panel (C) where they indicate the mean as those data were normally distributed.</p

Demographic and clinical variables of the combined SMRI and NSW TRC collection.

<p>The combined SMRI and NSW TRC collections were utilized for analysis of gene expression in the DLPFC only. Data are provided as means ± the standard deviation or number of individuals in each category.</p>a<p>A/B/C/D: European/Asian/Hispanic.</p

The effect of TNFSF13 exposure on intracellular pH, and of decreased intracellular pH on TNFSF13 and FAS receptor gene expression.

<p>Intracellular pH of U-87 MG cells was artificially decreased using nigericin and phosphate buffer to pH 6.4, 6.9 and 7.3 for 0.5, 3, 12 or 24 hours and transcript levels of the TNFSF13 (A) or FAS receptor (B) genes measured by qRT-PCR. Expression levels were calculated relative to three housekeeping genes (β-actin, TATA box binding protein and ubiquitin C) using the deltadelta Ct method. ^aTranscript levels at the 0.5 hours time point were significantly greater than at all other time points, ^bDecreasing pH significantly suppressed transcript levels of TNFSF13, ^cThere was no significant effect of pH on FAS receptor transcripts. U-87 MG glioblastoma cell cultures were treated with TNFSF13 (1 pg or 100 pg TNFSF13/microliter of culture media) for up to 48 hours and the intracellular pH determined (C). *p<0.05, ***p<0.001.</p