The flower pot method of REM sleep deprivation causes apoptotic cell death in the hepatocytes of rat

Abstract

Short running title: REM sleep deprivation causes liver cell death. Highlights of the study• We observed significant apoptosis in the hepatocytes of REMSD group of rats.• Our expression analysis confirmed altered expression for genes p53, Bcl2, Bax, and Caspase-3 after REMSD.• Protein level analysis supported our gene expression results for p53, Bcl2, Bax, Caspase 3 and Caspase 9 after REMSD.• Sleep recovery improved the respective genes and protein expression levels towards normalcy, signifying the functional role of REM sleep. Abstract Introduction:The rapid eye movement sleep deprivation (REMSD) of rats relates with increased inflammations, acute phase response, oxidative damage, neuronal cell loss, and neurodegenerative diseases. Whereas, its role outside brain are not well studied. This study tried to explore the causal effect of REM sleep loss on hepatocytes. Methods: We deprived the rats of REM sleep using standard flower pot method. We focused on liver to see the REMSD affects which controls most of the metabolic processes of the body. Results: We report here that flower pot induced REMSD causes apoptotic cell death of hepatocytes (~10% by Annexin Assay & ~20% by TUNEL assay). This were further got alleviated up to extent after sleep recovery of 5 days (recovered approximately 8.0% by Annexin Assay & 14% by TUNEL assay). The gene expression and protein level profiling revealed the up-regulation of p53, Bax, Cytochrome c, Caspase 3, and Caspase 9. While, Bcl2 which is an anti-apoptotic protein were down-regulated in response to REMSD. Relentless recovery of 5 days affected the expression pattern of these genes/proteins. Conclusions: Our study offer great pathological and physiological significance for sleep loss, by inferring the apoptotic celldeath in the hepatocytes of rat. This further signifies the functional and preventive role of REM sleep which is unique to mammals and avians with certain exceptions, as its loss can affect the natural well-being and survival of the individuals. IntroductionSleep is an important evolutionary physiological and behavioral process required for the survival and well-being of animals studied, although no predominant hypothesis has emerged to explain its functions [1,2]. In mammals, it is categorize as two types, non-rapid eye movement (NREM) sleep and rapid eye movement (REM) sleep. Evolutionary REM sleep is present in mammals and birds with essential functions related to physiological and ecological success. REM sleep, also called as paradoxical sleep in general associates with memory consolidation, brain maturation, spatial memory acquisition, muscle regeneration and maintenance of body physiology [3][4][5][6][7][8][9]. Functional aspect of REM sleep can inferred by its effect on amygdala activity in response to previous emotional experiences, reorganizing the hippocampal excitability, pruning and maintenance of new synapses in development and learning [10][11][12]. Its prolonged loss can alter blood brain barrier functions and can be fatal [13][14][15]. Some recent stu...

Rapid Eye Movement sleep deprivation of rat generates ROS in the hepatocytes and makes them more susceptible to oxidative stress

“…Previous studies had shown that NO exerts a protective effect by blocking TNF-α induced apoptosis and hepatotoxicity both in vivo and in vitro , by inhibiting the caspase-3-like protease activity 43 . Our unpublished results also show that REMSD can cause apoptosis in hepatocytes 36 . It suggests that might be excess of NO is being produced to counteract the oxidative stress generated due to sleep loss and it protects the hepatocytes from apoptotic death.…”

“…We used LPC group of rats as a reference group for statistical comparisons as CC and LPC groups had no significant differences in observed parameters. Our findings suggest that REM sleep deprivation of rats can increase ROS production in their hepatocytes which ultimately can induce apoptotic cell death 36 . Treatment of hepatocytes with H 2 O 2 or Xanthine/Xanthine oxidase can selectively induce production of ROS inside the hepatocytes and it can be quenched by Catalase or Oxypurinol treatment.…”

“…Our results demonstrate that the REM sleep is very important for the well-being of animals and its loss can't only induce toxicity in the hepatocytes due to the production of ROS but also make them more susceptible to further oxidative stress and death due to apoptosis 36 . Apart from this, REMSD also can induce iNOS circulation in the system which would increase NO production, which may further affect the well-being of the hepatocytes.…”

Differential Gene Expression in Brain and Liver Tissue of Wistar Rats after Rapid Eye Movement Sleep Deprivation

“…In the brain, REM sleep is involved in the reorganization of hippocampal excitability, pruning and maintenance of new synapses during development, and learning and memory consolidation [ 40 , 41 , 42 ]. Some recent studies also suggest that lack of REM sleep may cause cell death of somatic cells and neurons [ 43 , 44 , 45 ]. Outside of the brain, deprivation of the REM sleep was found to be associated with acute phase response in the liver, increased synthesis of pro-inflammatory cytokines such as IL1β, IL-6, and IL-12, and an increase in liver enzymes, alanine transaminase and aspartic transaminase [ 46 ].…”

“…In the present study, we compared the effect of prolonged REM sleep loss in the brain and liver in order to compare and contrast the effects that occur simultaneously on these vital organs, as previous studies have indicated that REM sleep may have drastic effects on the liver [ 44 , 46 , 48 ]. To address this, we used a microarray technique to compare gene expression and identify the processes affected in the brain and liver of a given subject after REM sleep deprivation for nine days.…”