Biochemical and Genetical Evaluation of Pomegranate Impact on Diabetes Mellitus Induced by Alloxan in Female Rats

Abstract: Various food industries explored the possibility of developing a nutritional supplement rich in natural antioxidants from pomegranates. This study has focused on the ability of pomegranate peel and juice to study the antioxidant status. Thirty two rats were allocated in 4 groups as follows: GroupI; control group without any treatment; GroupII: diabetic animals injected with alloxan; Group III: diabetic peel group animals injected with alloxan and then feed on peel pomegranate; GroupIV: diabetic juice group animals injected with alloxan and then gavage with pomegranate juice. After 4 weeks of treatment biochemical analysis were measured such as glucose, insulin, alpha-amylase, lipid profile (cholesterol, triglyceride HDL, LDL and total lipids), total protein, homocysteine, total antioxidant capacity and liver enzymes (AST&ALT). In addition, pancreas and liver tissues were separated for genetic analysis in which pancreatic tissues were used for RAPD-PCR analysis and liver tissues for DNA fragmentation assay. Results showed significant increase in glucose and alpha amylase levels in diabetic group, while insulin decreased. Peel and juice of pomegranate ameliorates this effect and decreased glucose, alpha amylase while insulin level increased. Cholesterol, triglycerides, LDL and total lipids increased while HDL decreased in diabetic group. Peel and juice of pomegranate prevented these changes. The more pronounced effect appeared in group III treated with peel pomegranate. Total protein was not affected by alloxan or pomegranate. Homocysteine was significantly increased while total antioxidant capacity decreased in diabetic group. After treatment by pomegranate peel and juice, these parameters become near to the control values. AST and ALT were significantly increased in diabetic group. But after treatment with peel and juice, AST and ALT levels decreased and become near to the control level especially ALT value. Furthermore, rate of DNA fragmentation and DNA band polymorphism increased significantly in diabetic group. While after treatment by peel and juice rate of DNA band polymorphism and DNA fragmentation were decreased significantly. Pomegranate peel and juice showed significant reduction in LDL oxidative susceptibility and an increase in total antioxidant status. Pomegranate is able to reduce the progression in atherosclerosis. The antioxidant content in foods decreased the oxidative stress related diseases

The Persistence-Inducing Toxin HokB Forms Dynamic Pores That Cause ATP Leakage

Bacterial populations harbor a small fraction of cells that display transient multidrug tolerance. These so-called persister cells are extremely difficult to eradicate and contribute to the recalcitrance of chronic infections. Several signaling pathways leading to persistence have been identified. However, it is poorly understood how the effectors of these pathways function at the molecular level. In a previous study, we reported that the conserved GTPase Obg induces persistence in Escherichia coil via transcriptional upregulation of the toxin HokB. In the present study, we demonstrate that HokB inserts in the cytoplasmic membrane where it forms pores. The pore-forming capacity of the HokB peptide is demonstrated by in vitro conductance measurements on synthetic and natural lipid bilayers, revealing an asymmetrical conductance profile. Pore formation is directly linked to persistence and results in leakage of intracellular ATP. HokB-induced persistence is strongly impeded in the presence of a channel blocker, thereby providing a direct link between pore functioning and persistence. Furthermore, the activity of HokB pores is sensitive to the membrane potential. This sensitivity presumably results from the formation of either intermediate or mature pore types depending on the membrane potential. Taken together, these results provide a detailed view on the mechanistic basis of persister formation through the effector HokB. IMPORTANCE There is increasing awareness of the clinical importance of persistence. Indeed, persistence is linked to the recalcitrance of chronic infections, and evidence is accumulating that persister cells constitute a pool of viable cells from which resistant mutants can emerge. Unfortunately, persistence is a poorly understood process at the mechanistic level. In this study, we unraveled the pore-forming activity of HokB in E. coil and discovered that these pores lead to leakage of intracellular ATP, which is correlated with the induction of persistence. Moreover, we established a link between persistence and pore activity, as the number of HokBinduced persister cells was strongly reduced using a channel blocker. The latter opens opportunities to reduce the number of persister cells in a clinical setting

Development of artificial systems to mimic cellular structures and functions

Membrane models proved its uniqueness as a tool for biophysics studies at single molecule level. Despite impressive progress in the field of biomimetic membrane models, there are serious challenges limiting their wider use in biological application, for example their stability and ability for communication with different environment. This thesis described the development and uses of artificial membrane for biophysical applications. The initial focus was on the investigation and design of a new membrane model platform that mimics biological cell and tissues. In the first study we built a model membrane platform by encapsulating one of membrane model called Droplet Interface Bilayer in hydrogel and we test the function ability of this system by incorporating pore forming proteins. In the second study we characterize amyloid peptides oligomers behavior using planar lipid technique. In the final study of this thesis we designed and construct membrane nanomachine from DNA which it produce nanomechanical motion and transport across the membranestatus: publishe

Multi-compartment encapsulation of communicating droplets and droplet networks in hydrogel as a model for artificial cells

Constructing a cell mimic is a major challenge posed by synthetic biologists. Efforts to this end have been primarily focused on lipid- and polymer-encapsulated containers, liposomes and polymersomes, respectively. Here, we introduce a multi-compartment, nested system comprising aqueous droplets stabilized in an oil/lipid mixture, all encapsulated in hydrogel. Functional capabilities (electrical and chemical communication) were imparted by protein nanopores spanning the lipid bilayer formed at the interface of the encapsulated aqueous droplets and the encasing hydrogel. Crucially, the compartmentalization enabled the formation of two adjoining lipid bilayers in a controlled manner, a requirement for the realization of a functional protocell or prototissue

Autonomous and Active Transport Operated by an Entropic DNA Piston

We present a synthetic nanoscale piston that uses chemical energy to perform molecular transport against an applied bias. Such a device comprises a 13 by 5 nm protein cylinder, embedded in a biological membrane enclosing a single-stranded DNA (ssDNA) rod. Hybridization with DNA cargo rigidifies the rod, allowing for transport of a selected DNA molecule across the nanopore. A strand displacement reaction from ssDNA fuel on the other side of the membrane then liberates the DNA cargo back into solution and regenerates the initial configuration. The entropic penalty of ssDNA confinement inside the nanopore drives DNA transport regardless of the applied bias. Multiple automated and reciprocating cycles are observed, in which the DNA piston moves through the 10 nm length of the nanopore. In every cycle, a single DNA molecule is transported across the nanopore against an external bias force, which is the hallmark of biological transporters

Aβ42 assembles into specific β-barrel pore-forming oligomers in membrane-mimicking environments

The formation of amyloid-β peptide (Aβ) oligomers at the cellular membrane is considered to be a crucial process underlying neurotoxicity in Alzheimer's disease (AD). Therefore, it is critical to characterize the oligomers that form within a membrane environment. To contribute to this characterization, we have applied strategies widely used to examine the structure of membrane proteins to study the two major Aβ variants, Aβ40 and Aβ42. Accordingly, various types of detergent micelles were extensively screened to identify one that preserved the properties of Aβ in lipid environments-namely the formation of oligomers that function as pores. Remarkably, under the optimized detergent micelle conditions, Aβ40 and Aβ42 showed different behavior. Aβ40 aggregated into amyloid fibrils, whereas Aβ42 assembled into oligomers that inserted into lipid bilayers as well-defined pores and adopted a specific structure with characteristics of a β-barrel arrangement that we named β-barrel pore-forming Aβ42 oligomers (βPFOsAβ42). Because Aβ42, relative to Aβ40, has a more prominent role in AD, the higher propensity of Aβ42 to form βPFOs constitutes an indication of their relevance in AD. Moreover, because βPFOsAβ42 adopt a specific structure, this property offers an unprecedented opportunity for testing a hypothesis regarding the involvement of βPFOs and, more generally, membrane-associated Aβ oligomers in AD

The Persistence-Inducing Toxin HokB Forms Dynamic Pores That Cause ATP Leakage

Bacterial populations harbor a small fraction of cells that display transient multidrug tolerance. These so-called persister cells are extremely difficult to eradicate and contribute to the recalcitrance of chronic infections. Several signaling pathways leading to persistence have been identified. However, it is poorly understood how the effectors of these pathways function at the molecular level. In a previous study, we reported that the conserved GTPase Obg induces persistence in Escherichia coli via transcriptional upregulation of the toxin HokB. In the present study, we demonstrate that HokB inserts in the cytoplasmic membrane where it forms pores. The pore-forming capacity of the HokB peptide is demonstrated by in vitro conductance measurements on synthetic and natural lipid bilayers, revealing an asymmetrical conductance profile. Pore formation is directly linked to persistence and results in leakage of intracellular ATP. HokB-induced persistence is strongly impeded in the presence of a channel blocker, thereby providing a direct link between pore functioning and persistence. Furthermore, the activity of HokB pores is sensitive to the membrane potential. This sensitivity presumably results from the formation of either intermediate or mature pore types depending on the membrane potential. Taken together, these results provide a detailed view on the mechanistic basis of persister formation through the effector HokB.status: Published onlin

Aβ(1-42) tetramer and octamer structures reveal edge conductivity pores as a mechanism for membrane damage

International audienceFormation of amyloid-beta (Aβ) oligomer pores in the membrane of neurons has been proposed to explain neurotoxicity in Alzheimer's disease (AD). Here, we present the three-dimensional structure of an Aβ oligomer formed in a membrane mimicking environment, namely an Aβ(1-42) tetramer, which comprises a six stranded β-sheet core. The two faces of the β-sheet core are hydrophobic and surrounded by the membrane-mimicking environment while the edges are hydrophilic and solvent-exposed. By increasing the concentration of Aβ(1-42) in the sample, Aβ(1-42) octamers are also formed, made by two Aβ(1-42) tetramers facing each other forming a β-sandwich structure. Notably, Aβ(1-42) tetramers and octamers inserted into lipid bilayers as well-defined pores. To establish oligomer structure-membrane activity relationships, molecular dynamics simulations were carried out. These studies revealed a mechanism of membrane disruption in which water permeation occurred through lipid-stabilized pores mediated by the hydrophilic residues located on the core β-sheets edges of the oligomers