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TITLE
DECLARATION
SUPERVISORS NOTE
ACKNOWLEDGEMENT
CONTENTS
ABBREVIATIONS
1. INTRODUCTION
2. REVIEW OF LITERATURE
2.1 REVIEW OF REACTIVE OXYGEN SPECIES
2.2 ANTIOXIDANT DEFENCE SYSTEM
2.2.1 Superoxide dismutase (SOD)
2.2.2 Catalase (CAT)
2.2.3 Glutathione peroxidase (GPx)
2.2.4 Reduced glutathione
2.3 WHY IS BRAIN PRONE TO FREE RADICAL DAMAGE?
2.4 WHAT IS HYPOXIA AND WHY IS HYPOXIA OF THE BRAIN IMPORTANT?
2.5 HYPOXIA AND FREE RADICAL GENERATION
2.6 RESPONSE OF ANTIOXIDANT DEFENCE SYSTEM TO HYPOXIC CONDITIONS IN TISSUES
2.7 PATHOPHYSIOLOGICAL ALTERATIONS IN BRAIN IN RESPONSE TO HYPOXIA
2. 7.1 Cellular acidosis
2.7.2 Energy depletion
2.7.3 Intracellular calcium overload
2.7.4 Accumulation of free fatty acids
2.7.5 Lipid peroxidation
2.7.6 Changes in lysosomal enzyme levels
2.7.7 Impairment of protein synthesis
2.7.8 Changes in blood brain barrier permeability
2.8 ULTRASTRUCTURAL CHANGES OBSERVED IN BRAIN UNDER HYPOXIC CONDITIONS
2.9 SELECTIVE VULNERABILITY OF HIPPOCAMPAL NEURONS TO HYPOXIA
2.10 ANTIOXIDANT THERAPY
2.10.1 Liposomes
Table 1 Methods of liposome preparation
2.10.2 Liposomes as a multifunctional carrier system
3. MATERIAL AND METHODS
3.1 MATERIALS
3.1. I Experimental design
Table: 2 Experimental Design Animal pool
3.2.1.1 Estimation of protein
Fig: 2 Standard curve- Protein
3.2.1.2 Determination of superoxide dismutase activity
3.2.1.3 Biochemical assay for catalase
3.2.1.4 Biochemical assay for glutathione peroxidase
3.2.1.5 Estimation of reduced glutathione
3.2.1.6 Biochemical assay for acid phosphatase
3.2.1.7 Estimation of malondialdehyde
3.2.2 Liposome preparation and entrapment of SOD
3.2.3 Confirmation of liposome formation by electron microscopy
3.2.4 Estimation of protein entrapment in liposomes
3.2.5 SOD entrapped liposome administration
Fig: 5 Superoxide dismutase administration through jugular vein
3.2.6 Electron microscopic studies
3.2.7 Statistical analysis
4. OBSERVATIONS
4.1 PROTEIN CONTENT IN THE CONTROL RAT BRAIN
Table: 3 Protein content in the control rat brain regions
Table: 4 Result of one-way ANOVA used to compare the regional variation in the protein content in the young rat brain
Table: 5 Result of one-way ANOVA used to compare the regional variation in the protein content in the adult rat brain
Fig: 6 Protein content in the young and adult rat brain regions
4.2 SUPEROXIDE DISMUTASE ACTIVITY IN THE CONTROL RAT BRAIN
Fig.SOD activity in the young and the adult rat brain regions
Table 6 Superoxide dismutase activity in the control rat brain regions
Table 7 Result of one-way ANOVA used to compare thr regional variation in the SOD activity in the young rat brain
Table 8 Result of one-way ANOVA used to compare regional variation in the SOD activity in the adult rat brain
4.3 CATALASE ACTIVITY IN THE CONTROL RAT BRAIN
Table: 9 Catalase activity in the control rat brain regions
Table: 10 Result of one-way ANOVA used to compare the regional variation in the CAT activity in the young rat brain
Table: 11 Result of one-way ANOVA used to compare the regional variation in the CAT activity in the adult rat brain
4.4 GLUTATHIONE PEROXIDASE ACTIVITY IN THE CONTROL RAT BRAIN
Table: 12 Glutathione peroxidase activity in the control rat brain regions
Table: 13 Result of one-way ANOVA used to compare the regional variation in the GPx activity in the young rat brain
Table: 14 Result of one-way ANOVA used to compare the regional variation in the GPx activity in the adult rat brain
4.5 REDUCED GLUTEIHIONE CONTENT IN THE CONTROL RAT BRAIN
Table: 15 Reduced glutathione content in the control rat brain regions
Table: 16 Result of one-way ANOVA used to compare the regional variation in the GSH content in the young rat brain
Table: 17 Result of one way ANOVA used to compare the regional variation in the GSH content i the adult rat brain
4.6 MALONDIALDEHYDE CONTENT IN THE CONTROL RAT BRAIN
Table: 18 Malondialdehyde content in the control rat brain regions
Table: 19 Result of one-way ANOVA used to compare the regional variation in the MDA content in the young rat brain
Table: 20 Result of one-way ANOVA used to compare the regional variation in the MDA content in the adult rat brain
4.7 ACID PHOSPHATASE ACTIVITY IN THE CONTROL RAT BRAIN
Table: 21 Acid phosphatase activity in the control rat brain regions
Table: 22 Result of one-way ANOVA used to compare the regional variation in the ACP activity in the young rat brain
Table: 23 Result of one-way ANOVA used to compare the regional variation in the ACP activity in the adult rat brain
4.8 EFFECT OF ISOBARIC HYPOXIA ON THE RAT BRAIN
4.8.1. Effect of isobaric hypoxia on protein content in the rat brain
Table: 24 Changes in the protein content in the rat brain after hypoxia for 1 hour
Table: 25 Changes the protein content in the rat brain after hypoxia for 3 hours
4.8..2 Effect of isobaric hypoxia on superoxide dismutase activity in the rat brain
Table: 26 Changes in the SOD activity in the rat brain after hypoxia for 1 hour
Table: 27 Changes in the SOD activity in the rat brain after hypoxia for 3 hours
4.8.3 Effect of isobaric hypoxia on catalase activity in the rat brain
Table: 28 Changes in the CAT activity in the rat brain after hypoxia for 1 hour
Table: 29 Changes in the CAT activity in the rat brain after hypoxia for 3 hours
4.8.4 Effect of isobaric hypoxia on glutathione peroxidase activity in the rat brain
Fig: 21 (GPx activity in the rat brain after hypoxia for 3 hours
Table: 30 Changes in the GPx activity in the rat brain after hypoxia of 1 hour
Table: 31Changes in the GPx activity in the rat brain after hypoxia for 3 hours
4.8.5 Effect of isobaric hypoxia on reduced glutathione content in the rat brain
4.8.6 Effect of isobaric hypoxia on malondialdehyde content in the rat brain
Table: 32 Changes in the GSH content in the rat brain after hypoxia for 1 hour
Table: 33 Changes in the GSH content in the rat brain after hypoxia for 3 hours
Fig.27 Percentage increase in MDA content in young and adult rat brain after hypoxia for 3 hours
Table: 34 Changes in the MDA content in the rat brain after hypoxia for 1 hour
Table: 35 Changes in the MDA content in the rat brain after hypoxia for 3 hours
4.8.7 Effect of isobaric hypoxia on acid phosphatase activity in the rat brain
Table: 36 Changes in the ACP activity in the rat brain after hypoxia for 1 hour
Table: 37 Changes in the ACP activity in the rat brain after hypoxia for 3 hours
4.9 ULTRASTRUCTLRAL CHANGES IN RAT BRAIN AFTER SUBJECTING TO HYPOXIA
4.9.1 Changes in the ultrastructure of mitochondria
4.9.2 Vacuolization
4.9.3 Accumulation of lipid granules
4.9.4 Damage to the ultrastructure of protein synthesizing apparatus
4.9.5 Changes in brain cell processes
4.9.6 Changes in neuroglial cells
4.9.7 Changes in capillary morphology
4.10 SUPEROXIDE DISMUTASE ADMINISTRATION
4.10.1 Detection of liposome in rat brain by electron microscopy
4. 10.2 Biochemical changes in the young rat brain after the administration of SOD entrapped in liposomes under hypoxic conditions
Table: 38 Changes in the protein content in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions 3 hours)
Table: 39 Changes in the SOD activity in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions (3 hours)
Table: 40 Changes in the CAT activity in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions (3 hours)
Table: 41Changes in the Gpx activity in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions (3 hours)
Table: 42 Changes in the GSH content in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions (3 hours)
Table: 43 Changes in the MDA content in the young rat brain regions after the administration of liposome entrapped SOD under hypoxic conditions (3 hours)
4 10.3 Structural modifications in the young rat brain after administration of SOD entrapped in liposomes under hypoxic conditions
Fig.57 ACP activity in the rat brain after SOD administration
4.10.3.1 Control
4.10.3.2 Experimental
5. DISCUSSION
5.1 ANTIOXIDANT DEFENCE ENZYMES IN RAT BRAIN -THE REGIONAL VARIATION
5.2 INFLUENCE OF AGE ON ANTIOXIDANT ENZYME ACTIVITIES IN THE RAT BRAIN
5.3 EFFECT OF HYPOXIA ON ANTIOXIDANT DEFENCE SYSTEM IN THE RAT BRAIN
5.4 DOES A DECREASE IN THE PROTEIN CONTENT IN THE BRAIN FOLLOWING HYPOXIA EXPLAIN THE DECREASE IN ANTIOXIDANT ENZYME ACTIVITIES?
5.5 CASH DEPLETION IN THE RAT BRAIN AFTER HYPOXIA
5.6 LIPID PEROXIDATION - AN INDICATOR OF FREE RADICAL DAMAGE IN THE RAT BRAIN
5.7 ACID PHOSPHATASE - AN INDICATOR OF PATHOLOGICAL CONDITION IN TISSUES
5.8 HIPPOCAMPUS AS THE REGION MOST VULNERABLE TO HYPOXIA
5.9 IS ADULT RAT BRAIN LESS SUSCEPTABLE TO HYPOXIA THAN THE YOUNG?
5.10 VACUOLIZATION AND DAMAGED MITOCHONDRIA - A COMMON FEATURE IN ULTRATHIN SECTIONS OF THE RAT BRAIN ALTER HYPOXIA
5.11 CHANGES IN THE CAPILLARY ULTRASTRUCTURE [N THE HIPPOCAMPAL SLICES OF THE YOUNG RAT BRAIN AFTER HYPOXIA
5.12 SOD THERAPY
5.13 SOD ADMINISTRATION PREVENTED/ REDUCED OXIDATIVE INJURY IN RAT BRAIN FOLLOWING HYPOX IA-REOXYGENATION
6. SUMMARY AND CONCLUSION
6.1 COMPONENTS OF ANTIOXIDANT DEFENCE SYSTEM SHOWED REGIONAL VARIATIONS IN RAT BRAIN
6.2 ADULT RAT BRAIN SHOWED A DECREASE IN THE DEFENCE ENZYME ACTIVITIES IN COMPARISON WITH THE YOUNG
6.3 HYPOXlA [MPARTED BIOCHEMICAL AND ULTRASTRUCTURAL ALTERATIONS IN RAT BRAIN LEADING TO OXIDATIVE INJURY
6.4 SOD ADMINISTRATION LEAD TO A DECREASE IN OXIDATIVE INJURY IMPARTED BY HYPOXIA
7. REFERENCES
8. LIST OF PUBLICATIONS