Impact of dietary changes on retinal neuronal plasticity in rodent models of physical and psychological trauma.

Introduction: Blast injury has been implicated as the major cause of traumatic brain injury (TBI) and ocular system injury, in military operations in Iraq and Afghanistan. Soldiers exposed to traumatic stress also have undiagnosed, chronic vision problems. Here we hypothesize that excessive intake of ω-6 fatty acid linoleic acid (LA) and insufficiency of dietary long chain ω-3 polyunsaturated fatty acids (PUFAs, e.g., docosahexaenoic acid; DHA) would dysregulate endocannabinoid-mediated neuronal plasticity and immune response. The study objective was to determine the effect of blast-TBI and traumatic stress on retinal gene expression and assess the role of dietary deficiency of long chain ω-3 PUFAs on the vulnerability to these injury models. Methods: Linoleic acid was used as an independent variable to reflect the dietary increase in LA from 1 percent of energy (en%) to 8 en% present in the current western diets, and these custom LA diets were also devoid of long chain ω-3 PUFAs. Animals were exposed to a simulated blast overpressure wave followed by a weight drop head-concussion to induce TBI. A Separate group of rats were subjected to traumatic stress by a forced immersion underwater. Results: Our findings showed that blast-TBI exposure, post 14 days, produced significant neuropathological changes such as axonal degeneration in the brain optic tracts from all the three diet groups, especially in rats fed the DHA-deprived 1 en% LA diet. Transcriptomic analysis showed that presence of DHA in the house chow diet prevented blast-induced disruption of neuronal plasticity by activating molecular networks like SNARE signaling, endocannabinoid pathway, and synaptic long-term depression when compared to DHA-deprived 8 en% LA diet group. Under traumatic stress, retinal synaptic function, neurovascular coupling, and opioid signaling mechanisms were dysregulated in rodents fed DHA-deficient diets (i.e., 8 en% LA and 1 en% LA), where reducing the levels of ω-6 linoleic acid from 8 en% to 1 en% was associated with increased neuronal plasticity and suppressed immune signaling. Conclusion: The findings of our study suggest that deprivation of long chain ω-3 PUFAs in the diet affects endocannabinoid-mediated neuronal plasticity, vascular function and inflammatory response that could influence the resistance of veterans to TBI and psychological trauma.

Upregulation of multiple toll-like receptors in ferret brain after blast exposure: Potential targets for treatment.

Although blast-induced traumatic brain injury (bTBI) has been designated as the signature injury of recent combat operations, its precise pathological mechanism(s) has not been identified thus far. Prior preclinical studies on bTBI demonstrated acute neuroinflammatory cascades which are known to be contributing to neurodegeneration. Danger-associated chemical patterns are released from the injured cells, which activate non-specific pattern recognition receptors, such as toll-like receptors (TLRs) leading to increased expression of inflammatory genes and release of cytokines. Upregulation of specific TLRs in the brain has been described as a mechanism of injury in diverse brain injury models unrelated to blast exposure. However, the expression profile of various TLRs in bTBI has not been investigated thus far. Hence, we have evaluated the expression of transcripts for TLR1-TLR10 in the brain of a gyrencephalic animal model of bTBI. We exposed ferrets to tightly coupled repeated blasts and determined the differential expression of TLRs (TLR1-10) by quantitative RT-PCR in multiple brain regions at 4 hr, 24 hr, 7 days and 28 days post-blast injury. The results obtained indicate that multiple TLRs are upregulated in the brain at 4 hr, 24 hr, 7 days and 28 days post-blast. Specifically, upregulation of TLR2, TLR4 and TLR9 was noted in different brain regions, suggesting that multiple TLRs might play a role in the pathophysiology of bTBI and that drugs that can inhibit multiple TLRs might have enhanced efficacy to attenuate brain damage and thereby improve bTBI outcome. Taken together, these results suggest that several TLRs are upregulated in the brain after bTBI and participate in the inflammatory response and thereby provide new insights into the disease pathogenesis. Therefore, inhibition of multiple TLRs, including TLR2, 4 and 9, simultaneously might be a potential therapeutic strategy for the treatment of bTBI.

Blast Exposure Dysregulates Nighttime Melatonin Synthesis and Signaling in the Pineal Gland: A Potential Mechanism of Blast-Induced Sleep Disruptions.

Blast-induced traumatic brain injury (bTBI) frequently results in sleep-wake disturbances. However, limited studies have investigated the molecular signaling mechanisms underlying these sleep disturbances, and potentially efficacious therapies are lacking. We investigated the levels of melatonin and genes involved in melatonin synthesis pathway in the pineal glands of Sprague Dawley rats exposed to single and tightly coupled repeated blasts during the night and daytime. Rats were exposed to single and tightly coupled repeated blasts using an advanced blast simulator. The plasma, cerebrospinal fluid (CSF), and pineal gland were collected at 6 h, 24 h, or 1 month postblast at two different time points: one during the day (1000 h) and one at night (2200 h). Differential expressions of genes involved in pineal melatonin synthesis were quantified using quantitative real-time polymerase chain reaction (qRT-PCR). Plasma and CSF melatonin levels were assessed using a commercial melatonin ELISA kit. The plasma and CSF melatonin levels showed statistically significant decreases at 6 h and 24 h in the blast-exposed rats euthanized in the night (in dim light), with no significant alterations noted in rats euthanized in the morning (daylight) at all three-time points. Blast-exposed rats showed statistically significant decreases in Tph1, Aanat, Asmt, and Mtnr1b mRNA levels, along with increased Tph2 mRNA, in the pineal gland samples collected at night at 6 h and 24 h. No significant changes in the mRNA levels of these genes were noted at 1 month. These findings imply that the melatonin circadian rhythm is disrupted following blast exposure, which may be a factor in the sleep disturbances that blast victims frequently experience.

The intrinsic apoptotic pathway lies upstream of oxidative stress in multiple organs.

Oxidative stress increases with age in multiple organ systems and is implicated in the development of age-related pathologies in them. Studies from our laboratory show that the intrinsic pathway proapoptotic proteins BAX and caspase-3 (CASP3) lie upstream of mitochondrial production of oxidative stress-inducing reactive species (RS) such as reactive oxygen and reactive nitrogen species (ROS and RNS) in apoptotic and nonapoptotic neurons in cell culture. Our objective in this study was to determine if these findings could be generalized to the development of oxidative stress in nonneuronal tissues in vivo. We first investigated the effect of genetic deletion of Bax on DNA damage in the liver, heart and kidneys of female mice of increasing ages (5, 14, 22 months). The organs of the aged mice showed increased oxidative DNA strand breaks compared to young animals (5 month). Ablation of Bax greatly reduced this damage. We next assessed lipid peroxidation, DNA oxidation, and protein tyrosine nitration to determine whether Casp3 deletion reduces oxidative stress in the hearts, livers, and kidneys of 12-month-old female mice. Lipid peroxides and 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels were much lower in organs from mice with depleted Casp3 than in those of wild type animals. Nitration of protein tyrosine residues, caused by RNS, was also significantly suppressed in the tissues of Casp3 null mice compared to those in wild type mice. Our findings indicate that BAX and CASP3 have a vital role in the generation of oxidative stress in organs of aged mice.

The neuroprotective effect of fisetin in the MPTP model of Parkinson's disease.

BACKGROUND: Parkinson's disease (PD) is characterized by excessive deposition of neuritic plaques known as Lewy bodies of which α-synuclein is the major contributor to neuronal death. Both oxidative stress and cytokines signaling have been proposed to play an important role in α-synuclein-induced neuronal death in MPTP and PD-related neuronal cell death. Fisetin, a natural polyphenol, possesses antioxidant, anti-inflammatory, and anti-apoptotic properties. However, the molecular neuroprotective mechanisms of fisetin against MPTP-induced cytotoxicity are still unknown. OBJECTIVE: The present study investigated the inhibitory effect of fisetin on MPTP/MPP+-induced neurotoxicity in PC12 cells. METHODS: Cells were pretreated with varying concentrations of fisetin prior exposure to MPTP/MPP+. Cell viability and apoptosis were investigated using MTT assay and DNA fragmentation. The expression and release of transcription factor, pro-inflammatory cytokines, and apoptotic mediators were assessed using western blot analysis and ELISA. RESULTS: Results showed that a pre-treatment with fisetin before exposure to MPTP/MPP+ significantly decreased MPTP/MPP+-induced cytotoxicity and cell death probably by decreasing α-synuclein expression. Mechanisms study showed that fisetin has the potential to inhibit several apoptotic and inflammatory pathways, which play important roles in the initiation and progression of PD. CONCLUSIONS: Altogether, these observations indicate that fisetin is capable of attenuating α-synuclein levels and promoting neuroprotective effects, meanwhile also present some insights into the potential signaling pathways that are involved. Thus, these findings support the role of natural polyphenols in preventive and/or complementary therapies for neurodegenerative diseases.