A Glutamate Scavenging Protocol Combined with Deanna Protocol in SOD1-G93A Mouse Model of ALS.

Amyotrophic lateral sclerosis (ALS) is a progressive disease of neuronal degeneration in the motor cortex, brainstem, and spinal cord, resulting in impaired motor function and premature demise as a result of insufficient respiratory drive. ALS is associated with dysfunctions in neurons, neuroglia, muscle cells, energy metabolism, and glutamate balance. Currently, there is not a widely accepted, effective treatment for this condition. Prior work from our lab has demonstrated the efficacy of supplemental nutrition with the Deanna Protocol (DP). In the present study, we tested the effects of three different treatments in a mouse model of ALS. These treatments were the DP alone, a glutamate scavenging protocol (GSP) alone, and a combination of the two treatments. Outcome measures included body weight, food intake, behavioral assessments, neurological score, and lifespan. Compared to the control group, DP had a significantly slower decline in neurological score, strength, endurance, and coordination, with a trend toward increased lifespan despite a greater loss of weight. GSP had a significantly slower decline in neurological score, strength, endurance, and coordination, with a trend toward increased lifespan. DP+GSP had a significantly slower decline in neurological score with a trend toward increased lifespan, despite a greater loss of weight. While each of the treatment groups fared better than the control group, the combination of the DP+GSP was not better than either of the individual treatments. We conclude that the beneficial effects of the DP and the GSP in this ALS mouse model are distinct, and appear to offer no additional benefit when combined.

Cell-Wall-Degrading Enzymes-Related Genes Originating from Rhizoctonia solani Increase Sugar Beet Root Damage in the Presence of Leuconostoc mesenteroides.

Sugar beet crown and root rot caused by Rhizoctonia solani is a major yield constraint. Root rot is highly increased when R. solani and Leuconostoc mesenteroides co-infect roots. We hypothesized that the absence of plant cell-wall-degrading enzymes in L. mesenteroides and their supply by R. solani during close contact, causes increased damage. In planta root inoculation with or without cell-wall-degrading enzymes showed greater rot when L. mesenteroides was combined with cellulase (22 mm rot), polygalacturonase (47 mm), and pectin lyase (57 mm) versus these enzymes (0-26 mm), R. solani (20 mm), and L. mesenteroides (13 mm) individually. Carbohydrate analysis revealed increased simpler carbohydrates (namely glucose + galactose, and fructose) in the infected roots versus mock control, possibly due to the degradation of complex cell wall carbohydrates. Expression of R. solani cellulase, polygalacturonase, and pectin lyase genes during root infection corroborated well with the enzyme data. Global mRNAseq analysis identified candidate genes and highly co-expressed gene modules in all three organisms that might be critical in host plant defense and pathogenesis. Targeting R. solani cell-wall-degrading enzymes in the future could be an effective strategy to mitigate root damage during its interaction with L. mesenteroides.

Exogenous ketone ester delays CNS oxygen toxicity without impairing cognitive and motor performance in male Sprague-Dawley rats.

Hyperbaric oxygen (HBO2) is breathing >1 atmosphere absolute (ATA; 101.3 kPa) O2 and is used in HBO2 therapy and undersea medicine. What limits the use of HBO2 is the risk of developing central nervous system (CNS) oxygen toxicity (CNS-OT). A promising therapy for delaying CNS-OT is ketone metabolic therapy either through diet or exogenous ketone ester (KE) supplement. Previous studies indicate that KE induces ketosis and delays the onset of CNS-OT; however, the effects of exogeneous KE on cognition and performance are understudied. Accordingly, we tested the hypothesis that oral gavage with 7.5 g/kg induces ketosis and increases the latency time to seizure (LSz) without impairing cognition and performance. A single oral dose of 7.5 g/kg KE increases systemic ß-hydroxybutyrate (BHB) levels within 0.5 h and remains elevated for 4 h. Male rats were separated into three groups: control (no gavage), water-gavage, or KE-gavage, and were subjected to behavioral testing while breathing 1 ATA (101.3 kPa) of air. Testing included the following: DigiGait (DG), light/dark (LD), open field (OF), and novel object recognition (NOR). There were no adverse effects of KE on gait or motor performance (DG), cognition (NOR), and anxiety (LD, OF). In fact, KE had an anxiolytic effect (OF, LD). The LSz during exposure to 5 ATA (506.6 kPa) O2 (≤90 min) increased 307% in KE-treated rats compared with control rats. In addition, KE prevented seizures in some animals. We conclude that 7.5 g/kg is an optimal dose of KE in the male Sprague-Dawley rat model of CNS-OT.

Dose response of a novel exogenous ketone supplement on physiological, perceptual and performance parameters.

BACKGROUND: Interest into the health, disease, and performance impact of exogenous ketone bodies has rapidly expanded due to their multifaceted physiological and signaling properties but limiting our understanding is the isolated analyses of individual types and dose/dosing protocols. METHODS: Thirteen recreational male distance runners (24.8 ± 9.6 years, 72.5 ± 8.3 kg, VO2max 60.1 ± 5.4 ml/kg/min) participated in this randomized, double-blind, crossover design study. The first two sessions consisted of a 5-km running time trial familiarization and a VO2max test. During subsequent trials, subjects were randomly assigned to one (KS1: 22.1 g) or two (KS2: 44.2 g) doses of beta-hydroxybutyrate (ßHB) and medium chain triglycerides (MCTs) or flavor matched placebo (PLA). Blood R-ßHB, glucose, and lactate concentrations were measured at baseline (0-min), post-supplement (30 and 60 min), post-exercise (+ 0 min, + 15 min). Time, heart rate (HR), rating of perceived exertion (RPE), affect, respiratory exchange ratio, oxygen consumption (VO2), carbon dioxide production, and ventilation were measured during exercise. Cognitive performance was evaluated prior to and post-exercise. RESULTS: KS significantly increased R-ßHB, with more potent and prolonged elevations in KS2, illustrating an administrative and dosing effect. R-ßHB was significantly decreased in KS1 compared to KS2 illustrating a dosing and exercise interaction effect. Blood glucose elevated post-exercise but was unchanged across groups. Blood lactate significantly increased post-exercise but was augmented by KS administration. Gaseous exchange, respiration, HR, affect, RPE, and exercise performance was unaltered with KS administration. However, clear responders and none-responders were indicated. KS2 significantly augmented cognitive function in pre-exercise conditions, while exercise increased cognitive performance for KS1 and PLA to pre-exercise KS2 levels. CONCLUSION: Novel ßHB + MCT formulation had a dosing effect on R-ßHB and cognitive performance, an administrative response on blood lactate, while not influencing gaseous exchange, respiration, HR, affect, RPE, and exercise performance.

Exogenous Ketones Lower Blood Glucose Level in Rested and Exercised Rodent Models.

Diseases involving inflammation and oxidative stress can be exacerbated by high blood glucose levels. Due to tight metabolic regulation, safely reducing blood glucose can prove difficult. The ketogenic diet (KD) reduces absolute glucose and insulin, while increasing fatty acid oxidation, ketogenesis, and circulating levels of ß-hydroxybutyrate (ßHB), acetoacetate (AcAc), and acetone. Compliance to KD can be difficult, so alternative therapies that help reduce glucose levels are needed. Exogenous ketones provide an alternative method to elevate blood ketone levels without strict dietary requirements. In this study, we tested the changes in blood glucose and ketone (ßHB) levels in response to acute, sub-chronic, and chronic administration of various ketogenic compounds in either a post-exercise or rested state. WAG/Rij (WR) rats, a rodent model of human absence epilepsy, GLUT1 deficiency syndrome mice (GLUT1D), and wild type Sprague Dawley rats (SPD) were assessed. Non-pathological animals were also assessed across different age ranges. Experimental groups included KD, standard diet (SD) supplemented with water (Control, C) or with exogenous ketones: 1, 3-butanediol (BD), ßHB mineral salt (KS), KS with medium chain triglyceride/MCT (KSMCT), BD acetoacetate diester (KE), KE with MCT (KEMCT), and KE with KS (KEKS). In rested WR rats, the KE, KS, KSMCT groups had lower blood glucose level after 1 h of treatment, and in KE and KSMCT groups after 24 h. After exercise, the KE, KSMCT, KEKS, and KEMCT groups had lowered glucose levels after 1 h, and in the KEKS and KEMCT groups after 7 days, compared to control. In GLUT1D mice without exercise, only KE resulted in significantly lower glucose levels at week 2 and week 6 during a 10 weeks long chronic feeding study. In 4-month and 1-year-old SPD rats in the post-exercise trials, blood glucose was significantly lower in KD and KE, and in KEMCT groups, respectively. After seven days, the KSMCT group had the most significantly reduced blood glucose levels, compared to control. These results indicate that exogenous ketones were efficacious in reducing blood glucose levels within and outside the context of exercise in various rodent models of different ages, with and without pathology.

Delaying latency to hyperbaric oxygen-induced CNS oxygen toxicity seizures by combinations of exogenous ketone supplements.

Central nervous system oxygen toxicity (CNS-OT) manifests as tonic-clonic seizures and is a limitation of hyperbaric oxygen therapy (HBOT), as well as of recreational and technical diving associated with elevated partial pressure of oxygen. A previous study showed that ketone ester (1,3-butanediol acetoacetate diester, KE) administration delayed latency to seizures (LS) in 3-month-old Sprague-Dawley (SD) rats. This study explores the effect of exogenous ketone supplements in additional dosages and formulations on CNS-OT seizures in 18 months old SD rats, an age group correlating to human middle age. Ketogenic agents were given orally 60 min prior to exposure to hyperbaric oxygen and included control (water), KE (10 g/kg), KE/2 (KE 5 g/kg + water 5 g/kg), KE + medium-chain triglycerides (KE 5 g/kg + MCT 5 g/kg), and ketone salt (Na+ /K+ ßHB, KS) + MCT (KS 5 g/kg + MCT 5 g/kg). Rats were exposed to 100% oxygen at 5 atmospheres absolute (ATA). Upon seizure presentation (tonic-clonic movements) experiments were immediately terminated and blood was tested for glucose and D-beta-hydroxybutyrate (D-ßHB) levels. While blood D-ßHB levels were significantly elevated post-dive in all treatment groups, LS was significantly delayed only in KE (P = 0.0003), KE/2 (P = 0.023), and KE + MCT (P = 0.028) groups. In these groups, the severity of seizures appeared to be reduced, although these changes were significant only in KE-treated animals (P = 0.015). Acetoacetate (AcAc) levels were also significantly elevated in KE-treated animals. The LS in 18-month-old rats was delayed by 179% in KE, 219% in KE + MCT, and 55% in KE/2 groups, while only by 29% in KS + MCT. In conclusion, KE supplementation given alone and in combination with MCT elevated both ßHB and AcAc, and delayed CNS-OT seizures.

Identification of crystals forming on porcine articular cartilage: a new method for the estimation of the postmortem interval.

Articular cartilage was examined to determine its decomposition sequence and its potential for assessing the postmortem interval. Scanning electron microscopy of articular cartilage from buried porcine trotters showed the presence of microcrystals on the synovial surface. These orthorhombic pyramidal or "coffin"-shaped crystals, appeared at 3 weeks (22 days) after interment and disappeared after 6 weeks. The disappearance of these crystals was linked to decompositional changes to the integrity of the synovial joint. The formation and disappearance of these crystals was associated with a pH change at the cartilage surface. Scanning electron microscopy-energy-dispersive X-ray (SEM-EDX) analysis showed that the five main elements contained within these crystals were carbon, nitrogen, oxygen, magnesium, and phosphorous. Such elemental analysis suggested the crystals may be struvite (MgNH4 PO4 6(H2 O)). Bacteria cultured from the cartilage synovial surface produced struvite crystals when grown in suitable media and were identified by DNA analysis to be Comamonas sp.

Physicochemical effects on uncontaminated kaolinite due to electrokinetic treatment using inert electrodes.

To determine the consequences of applying electrokinetics to clay soils, in terms of mechanisms acting and resulting effects on the clay, tests were conducted in which an electrical gradient was applied across controlled specimens of English China Clay (ECC) using 'inert' electrodes and a 'Reverse Osmosis' water feed to the electrodes (i.e., to mimic electrokinetic stabilisation without the stabiliser added or electrokinetic remediation without the contaminant being present). The specimens in which electromigration was induced over time periods of 3, 7, 14 and 28 days were subsequently tested for Atterberg Limits, undrained shear strength using a hand shear vane, water content, pH, conductivity and zeta potential. Water flowed through the system from anode to cathode and directly affected the undrained shear strength of the clay. Acid and alkali fronts were created around the anode and cathode, respectively, causing changes in the pH, conductivity and zeta potential of the soil. Variations in zeta potential were linked to flocculation and dispersion of the soil particles, thus raising or depressing the Liquid Limit and Plastic Limit, and influencing the undrained shear strength. Initial weakening around the anode and cathode was replaced by a regain of strength at the anode once acidic conditions had been created, while highly alkaline conditions at the cathode induced a marked improvement in strength. A novel means of indicating strength improvement by chemical means, i.e., free from water content effects, is presented to assist in interpretation of the results.