Effect of CNTF Derived Peptide, P021 on Cognition and Pathology in 3xTG-AD Mouse Model of Alzheimer\u27s Disease

Studies described in this thesis deal with the preventive effects of a neurogenic/neurotropic peptidergic compound, P021, on neurogenesis and synaptic deficits, neurodegeneration, cognitive impairment, and Ab and tau pathologies in a 3xTg-AD mouse model of Alzheimer’s disease (AD). Background: AD is a chronic progressive neurodegenerative disease. Its multifactorial nature and the heterogeneity make its treatment especially challenging. Although it is a major burden in society, at present there is no drug that can stop or slow down the progression of the disease. Currently, the only available treatments are symptomatic and for mild to severe stages. The development of a drug that can prevent AD at its early stages would be of major importance. The use of neurotrophic factors mimetic for the treatment of AD is an exciting therapeutic strategy. It focuses mainly on boosting synaptic plasticity and neurogenesis which lead to shifting the balance from neurodegeneration to regeneration of the brain as well as preventing Ab and tau pathologies. Herein, I present the preventive effects of P021 treatment in 3xTg-AD mice that is initiated very early in the disease during the period of synaptic compensation and is continued for the lifespan of the animal. Aims: The specific aims were: 1) to study the presence of the synaptic compensation phenomenon in the brain as a self-repairing mechanism in 3xTg-AD mice; 2) to study the preventive effects of P021 on cognitive deterioration when administered during the compensation period; 3) to study the preventive effects of P021 on amyloid beta (Ab) and tau pathologies; and 4) to study the effect of P021 on synaptic deficit, neuronal degeneration and neurogenesis. Methods: A battery of behavioral tests was conducted to assess the cognitive performance in 3xTg-AD mice with and without treatment with P021 at different disease stages. Immunohistochemical and biochemical analyses were performed to determine the levels of synaptic protein expression as well as Ab and tau pathologies at different time points that correspond to different stages of disease progression. Neurodegeneration was studied immunohistochemically with Fluorojade C staining. Neurogenesis was studied immunohistochemically with DCX (double cortin) and Ki-67 staining. Results: The 3xTg-AD mice at the age of 12 weeks were found to be cognitively impaired and showed a decrease in multiple synaptic and neuronal markers. This decrease was compensated by the brain until ~16 weeks of age. Beyond 16 weeks the brain was found to fail to compensate for the synaptic deficit. P021 intervention, started at 14 weeks of age, prevented cognitive impairment 9 months post-treatment, as tested by the Morris Water maze task. At 15-17 months post-treatment P021 was able to rescue short-term spatial reference memory as well as episodic memory, as determined by the novel object location and the novel object recognition tasks. The treatment with P021 also prevented Ab and tau pathologies during 9-18 months post-treatment. P021 was able to rescue synaptic deficit and neurodegeneration 9-18 months post-treatment and boost neurogenesis at 9 months post-treatment. P021 treatment increased survival from 41% in 3xTg-AD-vh to 87% in 3xTg-AD-P021 mice. In the entire study I did not find any severe side effects of P021 including loss of appetite or body weight. Conclusions: Early intervention with P021 during the period of synaptic compensation of the brain was successful in preventing cognitive impairment in 3xTg-AD mice. The P021 treatment prevented synaptic and neurogenesis deficits, neurodegeneration, and Ab and tau pathologies. These findings provide a proof of principle of the potential therapeutic effect of P021 on several major features of AD

Prevention of dendritic and synaptic deficits and cognitive impairment with a neurotrophic compound

Background: The use of neurotrophic factors to treat Alzheimer’s disease (AD) is hindered by their blood–brain barrier impermeability, short half-life, and severe side effects. Peptide 021 (P021) is a neurotrophic/neurogenic tetrapeptide that was derived from the most active region of the ciliary neurotrophic factor (CNTF) by epitope mapping. Admantylated glycine was added to its C-terminal to increase its blood–brain barrier permeability and decrease its degradation by exopeptidases to make it druggable. Here, we report on the preventive effect of P021 in 3 × Tg-AD, a transgenic mouse model of AD. Methods: P021 was administered in the diet at 3 months, i.e., 6–9 months before any overt amyloid beta (Aβ) or tau pathology, and during the period of synaptic compensation, and was continued until 21 months in 3 × Tg-AD mice. The 3 × Tg-AD mice and wild-type (WT) mice were treated identically but with a vehicle-only diet serving as controls. The effects of P021 on neurogenesis, dendritic and synaptic markers, and cognitive performance were investigated. Results: We found that P021 treatment was able to rescue dendritic and synaptic deficits, boost neurogenesis, and reverse cognitive impairment in 3 × Tg-AD mice. Conclusions: Availability of appropriate neurotrophic support during the period of synaptic compensation can prevent synaptic deficit and cognitive impairment, and P021 is a promising neurotrophic compound for this purpose

Dissecting the Seed Maturation and Germination Processes in the Non-Orthodox Quercus ilex Species Based on Protein Signatures as Revealed by 2-DE Coupled to MALDI-TOF/TOF Proteomics Strategy

Unlike orthodox species, seed recalcitrance is poorly understood, especially at the molecular level. In this regard, seed maturation and germination were studied in the non-orthodox Quercus ilex by using a proteomics strategy based on two-dimensional gel electrophoresis coupled to matrix-assisted laser desorption ionization/time of flight (2-DE-MALDI-TOF).Cotyledons and embryo/radicle were sampled at different developmental stages, including early (M1–M3), middle (M4–M7), and late (M8–M9) seed maturation, and early (G1–G3) and late (G4–G5) germination. Samples corresponding to non-germinating, inviable, seeds were also included. Protein extracts were subjected to 2-dimensional gel electrophoresis (2-DE) and changes in the protein profiles were analyzed. Identified variable proteins were grouped according to their function, being the energy, carbohydrate, lipid, and amino acid metabolisms, together with protein fate, redox homeostasis, and response to stress are the most represented groups. Beyond the visual aspect, morphometry, weight, and water content, each stage had a specific protein signature. Clear tendencies for the different protein groups throughout the maturation and germination stages were observed for, respectively, cotyledon and the embryo axis. Proteins related to metabolism, translation, legumins, proteases, proteasome, and those stress related were less abundant in non-germinating seeds, it related to the loss of viability. Cotyledons were enriched with reserve proteins and protein-degrading enzymes, while the embryo axis was enriched with proteins of cell defense and rescue, including heat-shock proteins (HSPs) and antioxidants. The peaks of enzyme proteins occurred at the middle stages (M6–M7) in cotyledons and at late ones (M8–M9) in the embryo axis. Unlike orthodox seeds, proteins associated with glycolysis, tricarboxylic acid cycle, carbohydrate, amino acid and lipid metabolism are present at high levels in the mature seed and were maintained throughout the germination stages. The lack of desiccation tolerance in Q. ilex seeds may be associated with the repression of some genes, late embryogenesis abundant proteins being one of the candidates

The Regulation of Ion Homeostasis, Growth, and Biomass Allocation in Date Palm Ex Vitro Plants Depends on the Level of Water Salinity

The date palm, a central plant in the fragile oasis ecosystem, is considered one of the fruit species most tolerant to salt stress. However, the tolerance mechanisms involved are yet to be addressed and their evaluation until now was mainly based on heterogenous plant material such as seedlings or limited to in vitro experiment conditions. For these reasons, we propose to deepen our knowledge of the morphological and physiological responses to salt stress using acclimated ex vitro plants resulting from the propagation of a single genotype. The plants were irrigated with 0, 150, 300, or 450 mM NaCl solutions for four months. Our results showed that the influence of water salinity on growth and ion-homeostasis regulation was very dependent on stress levels. The 150 mM NaCl concentration was found to improve dry biomass by about 35%, but at higher salt concentrations (300 and 450 mM) it decreased by 40–65%. The shoot:root dry mass ratio decreased significantly at the 150 mM NaCl water concentration and then increased with increasing water salt concentration. The leaf:root ratio for Na+ and Cl− decreased significantly with increasing water salinity up to a concentration of 300 mM NaCl, and then stabilized with similar values for 300 mM and 450 mM NaCl. In contrast to Na+ and Cl−, leaf K+ content was significantly higher in the leaf than in the root for all salt treatments. Unlike Na+ and K+, Cl− was expelled to the surface of leaves in response to increased water salinity. Overall, date palm plants appear to be more capable of excluding Cl− than Na+ and of changing biomass allocation according to salt-stress level, and their leaves and roots both appear to play an important role in this tolerance strategy.All authors are funded through the Small Research group project from the Deanship of Scientific Research at King Khalid University under research grant number (R.G.P.1/295/43).Peer reviewe

Differential effect of water salinity levels on gas exchange, chlorophyll fluorescence and antioxidant compounds in ex vitro date palm plants

In this study, the response to salt stress was evaluated in ex vitro acclimated date palm plants, regenerated from in vitro culture multiplication. The plants, eighteen-month-old, were irrigated with 0 (control), 150, 300 or 450 mM NaCl solutions (high to very high-water salinity). Photosynthesis parameters and antioxidant compounds were determined at the end of the experiment in leaves. At 150 mM NaCl, net CO2 assimilation rate and internal CO2 concentration were not impaired; while transpiration and stomatal conductance decreased by 60 and 70%, respectively. By increasing salt concentrations, all gas exchanges parameters were decreased. Measurement of chlorophyll fluorescence and P700 redox state showed that PSII and PSI machineries were significantly enhanced under 150 mM NaCl, conditions. With the 300 mM NaCl, the PSI parameters remained unchanged compared to control, while some of the PSII parameters, such as NPQ and Y (NPQ), were increased. At 450 mM NaCl, photosystems functionality was light intensity (PAR) dependent. Only at low PAR, a significant increase of some PSI and PSII parameters was observed. At the contrary, with high PAR, most of the energy conversion functions were significantly reduced, especially those related to PSI, indicating that PSI was more susceptible for damage by salinity than PSII. To overcome high salinity stress, ex vitro date palm plants mobilized a cascade of physio-biochemical pathways including the antioxidant activity and proline biosynthesis. Overall, the salinity of irrigation water, and up to 150 mM, improves the physiological performance of ex vitro date palm plants, which manage to tolerate very high levels of this stress.The authors extend their appreciation to the Deanship of Scientific Research at King Khalid University for funding this work through the large research Groups projects (Project under grant number (RGP. 2/73/44)).Peer reviewe

Additional file 1: of Prevention of dendritic and synaptic deficits and cognitive impairment with a neurotrophic compound

Table S1. Number of animals used. Table S2. Antibodies used. Figure S1. Rescue of synaptic deficit with P021 in 3 × Tg-AD mice at 9 months posttreatment. Western blots: a) cortex; b) hippocampus; and c) relative quantification of the blots. All blots were normalized against the loading control GAPDH. One-way ANOVA post-hoc test. *p < 0.05. WT, n = 5; Tg-AD-vh, n = 5; Tg-AD-P021, n = 5. Quantification data are shown as mean ± SD. Figure S2. Preventive effect of P021 on levels of dendritic and synaptic markers in 3 × Tg-AD mice at 15 months posttreatment. Western blots: a) cortex; b) hippocampus; and c) relative quantification of the blots. The 3 × Tg-AD-vh show a decrease in the level of synaptic markers and P021 was able to rescue this deficit. All blots were normalized against the loading control GAPDH. One-way ANOVA post-hoc test. **p < 0.01. WT, n = 5; Tg-AD-vh, n = 3; Tg-AD-P021, n = 5. Quantification data are shown as mean ± SD. (DOCX 5530 kb

Vegetative Growth Dynamic and Its Impact on the Flowering Intensity of the Following Season Depend on Water Availability and Bearing Status of the Olive Tree

A sufficient and mature vegetative growth is an essential condition for production in the following season and consequently affects the alternate bearing (AB) behavior. However, little is known about its interaction with the crop load and water supply. Herein, we studied the effect of different water regimes and bearing status on the vegetative intensity and flush and its consequence on the flowering parameters of the following season. Rainfed (RF) and fully irrigated (FI) treatments were applied for bearing (ON) and non-bearing (OFF) trees of the &lsquo;Zalmati&rsquo; olive orchard in south Tunisia during 2018. The water deficit condition (RF) and the high crop load (ON) have caused a similar decrease in the total node number and shoots length by about 65%. Furthermore, the flowering parameters in 2019, especially the percentage of floral buds (i.e., floral induction intensity), were differentially affected by water supply according to the bearing status of the previous season. FI reduced the percentage of floral buds in 2019 by about 37% for OFF trees, while it increased it by more than four times for the ON trees. Concerning the growth flush, the second (i.e., autumnal) flush seems to respond better to water supply than the first (i.e., spring) one, for both vegetative and flowering parameters. The second flush of growth provided 28% of the total vegetative growth but contributed about 35% to the total number of floral buds for RF OFF trees. Besides, FI makes the first and the second flushes contribute equally to total vegetative growth and to the flowering parameters of the following season. On the other hand, the bearing status of the trees does not affect the contribution of each flush to the total vegetative growth

Vegetative Growth Dynamic and Its Impact on the Flowering Intensity of the Following Season Depend on Water Availability and Bearing Status of the Olive Tree

A sufficient and mature vegetative growth is an essential condition for production in the following season and consequently affects the alternate bearing (AB) behavior. However, little is known about its interaction with the crop load and water supply. Herein, we studied the effect of different water regimes and bearing status on the vegetative intensity and flush and its consequence on the flowering parameters of the following season. Rainfed (RF) and fully irrigated (FI) treatments were applied for bearing (ON) and non-bearing (OFF) trees of the ‘Zalmati’ olive orchard in south Tunisia during 2018. The water deficit condition (RF) and the high crop load (ON) have caused a similar decrease in the total node number and shoots length by about 65%. Furthermore, the flowering parameters in 2019, especially the percentage of floral buds (i.e., floral induction intensity), were differentially affected by water supply according to the bearing status of the previous season. FI reduced the percentage of floral buds in 2019 by about 37% for OFF trees, while it increased it by more than four times for the ON trees. Concerning the growth flush, the second (i.e., autumnal) flush seems to respond better to water supply than the first (i.e., spring) one, for both vegetative and flowering parameters. The second flush of growth provided 28% of the total vegetative growth but contributed about 35% to the total number of floral buds for RF OFF trees. Besides, FI makes the first and the second flushes contribute equally to total vegetative growth and to the flowering parameters of the following season. On the other hand, the bearing status of the trees does not affect the contribution of each flush to the total vegetative growth

Effect of the exposure period to different water salt levels on the morphological behavior of olive plants

International audienceThe increasing salinity of water and soil is one of the environmental factors that most threatens the sustainability of olive cultivation systems in the Mediterranean basin. The identification of plant material with high tolerance to this stress would be one of the ways to solve this challenge, but it is generally a slow and expensive process. The selection of the most reliably parameters involved in the response of the plant to salinity and that are easy to evaluate, could help to speed up the identification of the most tolerant genotypes. The objective of this study is to determine the most interesting morphological characters which could be used in future as early criteria in the selection process of olive tolerant genotypes. For that, young plants, three-month-old, were exposed to salinity (0, 4 and 8 g/L NaCl) during different periods of time (30, 50 and 70 days), and several morphological parameters were assessed. The obtained results showed that most of the parameters were affected by the concentration 8 g/L of NaCl after a treatment period of 50 days. The most affected parameters by this level of salinity were the plant height, the leaf number and the number of lateral shoots. The PCA analysis showed that the number of lateral shoots was poorly correlated with the other parameters, but the height and the leaf number were highly correlated with each other

Assessment of the Cadmium and Copper Phytoremediation Potential of the Lobularia maritima&nbsp;Thioredoxin 2 Gene Using Genetically Engineered Tobacco

Rapid global modernization, urbanization, and industrialization have accelerated the release of heavy metals, causing soil pollution. These highly noxious environmental pollutants induce oxidative stress in plants via stimulation of the production of reactive oxygen species (ROS). Thioredoxin (Trxs) is a highly conserved disulfide reductase that plays a crucial role in intracellular redox homeostasis in both eukaryotes and prokaryotes. Herein, the presence of heavy metals highly upregulated LmTrxh2 transcription in Lobularia maritima seedlings and its overexpression-conferred tolerance to Cd, Cu, Mn, and Zn in Saccharomyces cerevisiae. In addition, LmTrxh2-overexpressing tobacco plants had higher seedling survival rates than non-transgenic plants (NT), with enhanced root length and biomass production and reduced ROS accumulation, following Cd and Cu stress. These plants also accumulated more Cd, Cu, and Mn than the NT plants. Moreover, LmTrxh2 overexpression stimulated the transcription of genes encoding metallothioneins (Met-1, Met-2, Met-3, and Met-4), a copper transport protein, a Snakin/GASA protein (Snakin-2), and ROS-scavenging enzymes (SOD, APX1, and CAT), which might contribute to heavy metal tolerance in tobacco plants. These results suggest that LmTrxh2 overexpression helps to improve heavy metal tolerance by stimulating antioxidant capacities and the expression of several stress-responsive genes in plants