Assessment of DNA damage and repair efficiency in drug naïve schizophrenia using comet assay

Highlights

• Drug naïve patients with schizophrenia exhibited greater degree of baseline DNA damage.

• The DNA repair capacity was preserved in the majority of cases.

• Age, gender and duration of untreated illness did not influence genomic damage or repair efficiency.

Abstract

The etiology of schizophrenia continues to be confounding and elusive. Some knowledge gaps exist in the neurodegenerative theory of schizophrenia. Oxidative DNA damage and repair deficits are relevant to the mechanisms of neurodegeneration but have not been studied in drug naïve schizophrenia. The present study used the comet assay technique to study the extent of DNA damage in circulating peripheral lymphocytes of patients with drug naïve schizophrenia (n = 40) along with an age and gender matched control group (n = 40). We also assessed the DNA repair efficiency in cases following incubation in a nutrient medium. All the assayed comet parameters demonstrated significantly greater baseline DNA damage in cases in comparison to the controls except for head diameter (p < 0.001 for all significant results, p = 0.32 for head diameter). Gender, age and duration of illness (p = 0.21, 0.69 and 0.12 respectively for tail length) did not influence any of the parameters significantly. Significant decrease was noted in the comet tail length and percentage of DNA in comet tail (p < 0.001 for both) in cases following incubation suggesting that the DNA repair machinery was preserved. No difference in DNA repair efficiency was noted between the genders (p = 0.23 for tail length). Our findings confirm the presence of significant baseline DNA damage in schizophrenia even prior to the initiation of anti-psychotic treatment. Additionally, intact genomic repair efficiency was noted in this group as a whole. These results provide some evidence for oxidative DNA damage as molecular link underpinning neurodegeneration in drug naïve schizophrenia.

Introduction

Schizophrenia is a chronic, debilitating psychiatric disorder whose etiology has been challenging to decipher. There are, broadly speaking, three hypothesis put forth to explain the etiopathogenesis of this condition. They are the neurodevelopmental (Fatemi and Folsom, 2009), neurodegenerative (Christopoulos et al., 2006) and the progressive neurodevelopmental model (Woods, 1998). The term neurodegeneration refers to chronic, progressive disorders of the nervous system characterized by continuing neuronal loss. The prototype examples of such disorders are Alzheimer's disease and Huntington's disease (Coppedè et al., 2006). Numerous lines of evidence support the neurodegenerative theory of schizophrenia. Longitudinal neuroimaging studies have been able to demonstrate progressive changes in the size and function of frontal and temporal lobes following illness onset lending credence to this theory (Shenton et al., 2001, Kasai et al., 2003, Buckley, 2005). Oxidative stress is a hallmark of any neurodegenerative process and is also believed to be involved in the pathophysiology of schizophrenia. Evidence has accumulated regarding the decreased levels of endogenous anti-oxidants such as superoxide dismutase, glutathione and catalase in people with schizophrenia (Fendri et al., 2006, Dadheech et al., 2008, Tsai et al., 2013). On a conflicting note, gliosis which is another feature of neurodegeneration has been notably absent in histopathological models of schizophrenia indicative of knowledge gaps about this hypothesis (Harrison 1999; Schnieder and Dwork, 2011).

Deoxyribonucleic Acid (DNA) damage can be defined as any alteration in DNA that induces a change in its function or modifies its coding properties (Lindahl, 1993, Rao, 1993). Compelling evidence, from animal and invitro models, suggest that DNA strand breaks are early antecedents of both apoptoptic and necrotic forms of neuronal cell death (Liu and Martin, 2001, Martin et al., 2006). Studies using cultured embryonic mouse cortical neurons have demonstrated the rapid accumulation of increasing DNA-single strand breaks (DNA-SSBs) with increasing levels of oxidative stress prior to eventual cell death. Furthermore, the timing of these genomic strand breaks in neurodegenerative disease models suggests that they are forerunners of a p53-driven apoptotic cascade in the cerebral cortex (Martin et al., 2009). Interestingly, authors who investigated mechanisms of motor neuron (MN) degeneration using transgenic mice expressing human mutant superoxide dismutase-1 (mSOD1) identified novel patterns of MN degeneration that differed from apoptosis structurally and biochemically. They concluded that there is a role for oxidative stress in the process of motor neuron degeneration induced by mSOD1 (Martin et al., 2007). Based on many of these findings, researchers have posited that oxidative stress induced genomic damage is relevant to the mechanisms resulting in neurodegeneration (Martin, 2008). Oxidative stress is thought to activate DNA damage pathways and play a critical role in activating telomere erosion, a key molecular correlate of the aging process (Sahin and DePinho, 2010). Some authors have suggested that this chronic state of oxidative stress on nucleic acids may underpin some of the clinical indicators of early aging in schizophrenia such as mildly accelerated cognitive decline (Jorgensen et al., 2013). DNA repair mechanisms have also been noted to be defective in many neurodegenerative conditions (Kidson and Chen, 1986). In a recent review that specifically focused on consequences of defective DNA repair mechanisms in non-proliferating cells such as neurons, the authors concluded that inefficient DNA repair has a robust link with aging and manifestations of classic neurodegenerative conditions such as Alzheimer's disease (Madabhushi et al., 2014). The evidence for neurodegeneration in schizophrenia remains sketchy. Previous investigators who studied urinary markers of oxidative DNA damage using ultra performance liquid chromatography with tandem mass spectroscopy concluded that there was a statistically significant increase in the excretion of these markers in schizophrenia patients when compared to matched controls (Jorgensen et al., 2013). Evidence for increased levels of oxidative DNA damage markers in schizophrenia have also come from studies on post mortem hippocampi in elderly patients with poor clinical outcomes (Nishioka and Arnold, 2004). By contrast, DNA repair response in schizophrenia has received comparatively less research attention. In a study using immortalized lymphoblasts from schizophrenia patients and healthy controls, the authors investigated the issue of DNA repair by studying temporal change in markers of DNA-DSBs following irradiation and concluded that there was no difference in rates of DNA repair between the two groups (Catts et al., 2012). Single cell gel electrophoresis or “comet” assay is regarded as a sensitive method for estimating the major types of DNA damage lesions in eukaryotic cells (Tice et al., 2000). Few studies have used this technique to measure levels of DNA damage and repair in schizophrenia. Further, most of the available data are from chronic schizophrenia patients who carry a lot of potential confounders that may contaminate these observations, particularly, antipsychotic treatment which has been shown to negatively influence genotoxic parameters in pre-clinical and clinical research (Polischouk et al., 2007, Picada et al., 2011). Hence, we undertook the present study with the twin objectives of assessing DNA damage and repair efficiency in a selected sample of drug naïve schizophrenia and comparing them with a healthy control group. We also aimed at assessing the influence of age, gender, family history of schizophrenia and duration of untreated illness on these parameters.