الفهرس 
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Abstract
Hepatotoxicity Implies Chemical-Driven Liver Damage. Drug-Induced Liver Injury Is A Cause Of Acute And chronic Liver Disease.
Acute Liver Failure (ALF) Is A Condition Resulting from Sudden, Severe Damage To The Liver, Mainly Due To Adverse Reactions To Drugs, Autoimmune Hepatitis, And Viral Infections
Acute Liver Failure Is Loss Of Liver Function That Occurs Rapidly In Days Or Weeks Usually In A Person Who Has No Pre-Existing Liver Disease.
Concomitant Administration Of Lipopolysaccharide And D-Galactosamine Can Lead To An Experimental Model Of Acute Liver Failure, The Main Reason Of The Resulting Dramatic Liver Damage May Be Oxidative Stress And Its Destructive Changes Which Happen After The Administration Of LPS/D-Galn.
One Of The Revolutionary Transcription Factors Is Foxo3a As It Was Found The Great Role It Plays In Oxidative Stress Resistance.
Forkhead Proteins Have A Significant Regulatory Role In Many Cellular Processes Including, Metabolism, Proliferation, Development, Migration, Invasion, Apoptosis And Longevity.
Foxo3a Elicits Its Effect On The Cellular Phenotype By Activating The Transcription Of Their Target Genes, Which Differentially Effect Cell Fate In A Downstream Target-Specific Manner.
FOXO3a Has Also Been Found To Be Key Sensor Of Oxidative Stress Through The Regulation Of Genes Involved In Detoxification Reactions And DNA Repair. ROS Has Been Shown To Be Important In Some Cell Signaling Pathways And Cell Cycle Progression, But Excessive ROS Accumulation Can Be Detrimental To The Cell And Can Lead To The Induction Of Apoptosis And Cell Death. If These Cellular Processes Are Perturbed Disproportionate ROS Accumulation Can Damage Proteins, Lipids And DNA Leading To Cellular Transformation, Tumorigenesis And Cancer. FOXO3a Is Able To Upregulate The Mrna And Protein Expression Of Manganese Superoxide Dismutase (Mnsod) In Response To ROS Accumulation, Promoting Cell Survival. Subsequently, FOXO3a Has Been Found To Be Upregulated In Response To ROS Accumulation And This Coincided With An Increase In The Expression Of Its Downstream Transcriptional Targets.
FOXO3a Is Also Found To Regulate Catalase In Response To Oxidative Stress. Catalase, As The Name Suggests, Catalyzes The Reaction Of Breaking Down Hydrogen Peroxide Into Oxygen And Water. It Was Subsequently Shown That Hydrogen Peroxide Treatment In Mammalian Cells Resulted In The Activation Of FOXO3a And An Increase In Transcriptional Activity. Moreover, FOXO3a Binds To The Catalase Promoter, Activating Its Transcription Overexpressed.
Although Mnsod And Catalase Are Located In Different Cellular Compartments, The Mitochondria And Peroxisomes Respectively, Suggesting That They Have Distinct Roles In Regulating Cellular Stress, They Are The Key Transcriptional Targets Of FOXO3a Proteins And Highlight The Concerted Role Of FOXO3a In Regulating ROS Scavengers, Driving Detoxification Reactions And Promoting Survival
FOXO3a Transcription Factors Have A Diverse Range Of Transcriptional Targets Involved In Different Cellular Processes Such As, Cell Cycle Regulation, Apoptosis (E.G., Bim, Fas-Ligand And TRAIL) And Oxidative Stress (E.G., Mnsod And Catalase). To Achieve Successful Regulation Of This Repertoire Of Target Genes, The Activity Of FOXO3a Transcription Factors Is Regulated By A Range Of Posttranslational Processes Including Phosphorylation, Ubiquitination, Acetylation, Which Allow The Activation Of Specific Subsets Of Genes That Influence Cell Fate.
Phosphorylation Is The Chemical Modification Of Serine, Threonine And Tyrosine Residues By The Addition Of A Phosphate Group. The Posttranslational Status Of Proteins Is A Dynamic Process, The Modification Of Residues Is Regulated By Kinases And Phosphatases Which Phosphorylate And Dephosphorylate Protein Substrates, Respectively. The Phosphorylation Status Of Proteins Is Predominantly Linked Heavily With Their Activity, Phosphorylation Promoting Its Activity Or Reducing It, In A Substrate Specific Manner. FOXO3a Is Known To Be Phosphorylated By The Oncogenic Kinases AKT, IKK And ERK, This Leads To Rapid Export Of FOXO3a from The Nucleus Into The Cytoplasm, Inhibiting FOXO3a Activity.
AMPK Directly Regulates Mammalian FOXO3a, A Member Of The FOXO Family Of Forkhead
Transcription Factors Known To Promote Resistance To Oxidative Stress, Tumor Suppression, And Longevity. Phosphorylation By AMPK Leads To The Activation Of FOXO3a Transcriptional Activity Without Affecting FOXO3a Subcellular Localization. The Regulation Of FOXO3a By AMPK May Play A Crucial Role In Fine Tuning Gene Expression Programs That Control Energy Balance And Stress Resistance In Cells Throughout Life.
Sirtuins Are The Most Recently Discovered Histone Deacetylase ”HDAC” Class And Are Different To Other Families Due To The Fact That They Require NAD+ And Are Resistant To Compounds That Target And Inhibit Class I And II Hdacs. Moreover, Hdacs Are Known Not To Solely Regulate The Acetylation Status Of Histones But Also Regulate The Acetylation Status Of Non-Histone Proteins. Sirtuins Are A Perfect Example Of This, Influencing The Posttranslational Status And Activity Of Key Proteins To Maintain Cell Viability, Such As P53, FOXO Family Members And The DNA-Repair Protein Ku70.
Sirtuins Are Ubiquitously Expressed Although They Occupy Distinct Subcellular Localizations And Consequently Interact With Different Intracellular Targets – Suggesting That Their Main Role Is Mediating The Acetylation Status Of Non-Histone Proteins, Otherwise They Would All Be Located In The Nucleus.
Phosphorylated Foxo3a, Deacetylated Foxo3a, AMPK, SIRT-1, PARP, TLR-4 Together With Liver Function Tests And Antioxidants Were Estimated Under The Influence Of LPS/D-Galn And Drugs (Silibinin And Vitamin E) Treatments In order To Estimate Their Prophylactic And Curative Effectiveness In Modifying The Oxidative Stress Which Has The Principle Role In Acute Liver Failure.
The Present Study Focused On Studying The Changes Observed In Activities And Levels Of These Parameters And How Could Changes In Their Levels Affect The Activation Of Foxo3a Transcription Factor.
Sixty Male Wister Rats Were Used In This Study. The Rats Were Randomly Divided Into Six Groups Of Ten Rats Each And Treated As Follows:
group 1 (Normal Control): Consisted Of Ten Rats Which Were Fed Normal Diet And Received A Single Dose Of Sterile Physiological Saline Through Intraperitoneal Route In A Dose Of 0.5 Ml Per Kg.
group 2 (LPS/D-Galn): Contained 10 Rats Which Were Fed Normal Diet And Were Exposed To Single Intraperitoneal Injection Of D-Galn In A Dose Of 500 Mg/Kg, Followed By A Single Intraperitoneal Injection Of LPS At A Dose Of 50 µg/Kg.
group 3 (Curative Silibinin): Consisted Of 10 Rats That Were Fed Normal Diet And Were Exposed To A Single Intraperitoneal Injection Of Of D-Galn In A Dose Of 500 Mg/Kg And LPS At A Dose Of 50 µg/Kg. Followed By Intraperitoneal Injection Of Silibinin At A Dose Of 100 Mg/Kg For Two Weeks
group 4 (Curative Vitamin E): Consisted Of 10 Rats That Were Fed Normal Diet And Were Exposed To A Single Intraperitoneal Injection Of D-Galn In A Dose Of 500mg/Kg And LPS At A Dose Of 50 µg/Kg, Followed By Oral Administration Of Vitamin E In A Dose Of 400mg/Kg For Two Weeks.
group 5 (Prophylactic Silibinin): Composed Of 10 Rats That Were Fed Normal Diet And Were Administered Silibinin At A Dose Of 100 Mg/Kg For Two Weeks Through The Intraperitoneal Injection, Followed By A Single Intraperitoneal Injection Of D-Galn/LPS With Above Mentioned Doses.
group 6 (Prophylactic Vitamin E): Composed Of 10 Rats That Were Fed Normal Diet And Were Administered Vitamin E With Above Mentioned Dose For Two Weeks Then They Were Exposed To A Single Intraperitoneal Injection Of D-Galn/LPS With Above Mentioned Doses.
After Overnight Fasting, Blood Samples Were Collected from Medial Canthus Blood Capillaries Of The Eye from Each Rat In All Groups Into Sterilized Tube To Be Used For Separation Of Serum For Determination Of ALT And AST.
After That Rats Were Sacrificed, Their Livers Were Eradicated Quickly, Washed With Normal Physiological Saline And Divided Into Four Parts, The First Part Of Them Were Placed In Proteinase Inhibitor And Preserved At -80°C For Western Blot To Determine Pfoxo3a, Deacetylated FOXO3a, AMPK And Mnsod. The Second Part Of Liver Was Placed In Rnase Inhibitor And Preserved At -80°C For RT-PCR To Investigate The Activity Of PARP, TLR-4 And SIRT-1. The Third Part Of The Liver Was Homogenized In 5 Ml Phosphate Buffered Saline By Using Tissue Homogenizer. The Liver Tissue Homogenates Were Centrifuged. The Supernatants Were Collected And Used Directly For Measurements Of MDA, GSH, CAT And SOD Levels. The Last Part Of The Liver Was Preserved In 10% Formalin Solution For Histopathological Examination. Sections from The Liver Were Prepared And Stained With Routine Hematoxylin And Eosin (H&E) Staining.
The Results Revealed That
In LPS/D-Galn Treated group There Were A Significant Decrease In Activity Of Liver Mitochondrial Antioxidant (Mnsod) And Liver Non-Mitochondrial Antioxidants (SOD, CAT And GSH) With Subsequent Increase In Liver MDA Activity Compared To The Normal Control group , In Addition LPS/D-Galn Treated group Showing A Significant Decrease In The Expression Level Of Liver Deacetylated Foxo3a And The Liver Phosphorylated FOXO3a With A Significant Decrease In Their Upstreaming Activatiors Liver SIRT-1 And AMPK Respectively, On The Other Hand Their Administration Significantly Increased The Activity Of TLR-4 And PARP. These Changes Happened After The Administration Of LPS/D-Galn Were Supported By The Histopathological Examination Of Liver Of Rats Intoxicated With Them Which Showed A Slightly Congested Central Vein S And Vacuolated Hepatocytes With Pyknotic Nuclei, Some Of The Hepatocytes Lost Their Architecture With Apparent Cellular Damage And Cellular Death As Well As Most Of The Hepatic Sinusoids Appeared Degenerated.
On The Other Hand, Administration Of Either Silibinin Or Vitamin E To The LPS/D-Galn Treated Rats Showed A Significant Restoration Of Liver Mitochondrial (Mnsod) And Liver Non- Mitochondrial (CAT, SOD And GSH) Antioxidant Levels With Subsequent Decrease In Liver MDA Content Compared To LPS/D-Galn Treated Group. ALT And AST Activities Showed A Significant Reduction In Groups Treated With Silibinin Or Vitamin E In A Prophylactic Or A Curative Manner Compared To That Treated With LPS/D-Galn Group. The Expression Levels Of Liver Deacetylated FOXO3a And Liver Phosphorylated FOXO3a Showed A Significant Increase And This May Be Due To Increase In The Expression Levels Of Their Up-Streaming Stimulators Liver SIRT-1 And AMPK Respectively. And The Activity Of PARP And TLR-4 Showed A Significant Reduction.
These Results Were Supported With The Histopathological Examination Which Showed That The Liver Of The Treated Groups Resembling Normal Ones With An Improvement In The Liver Architecture Compared To LPS/Dgaln Treated Group.