Pigment Epithelium-Derived Factor (PEDF) Inhibits Progression of Liver Metastasis in a Mouse Model of Uveal Melanoma Open Access

Lattier, John Michael (2014)

Permanent URL: https://etd.library.emory.edu/concern/etds/73666457w?locale=en
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Abstract

Uveal melanoma is the most common form of adult eye cancer in the United States, forming in the middle layer of the eye and originating in pigmented melanocytes. While the primary eye tumor is treatable with high success, patients often succumb to liver metastasis within 2 to 5 years. Metastatic progression is highly dependent on angiogenesis, or the formation of new blood vessels, to nourish the growing tumor with oxygen and nutrients. Pigment epithelium-derived factor (PEDF) is one of the most potent inhibitors of angiogenesis, and it is naturally produced by liver hepatocytes. I hypothesized that PEDF inhibits metastatic progression via a mechanism involving angiogenesis. I used PEDF null mice, injected melanoma cells into the posterior compartment of the eye, and measured liver metastasis after 4 weeks. PEDF null mice exhibited a 34.6-fold increase in liver metastasis area and a much greater population of macrometastases (>200 microns in diameter) when compared to controls. Vascular density was also significantly increased in these metastases, and various other characteristics were measured such as stromagenic factors and the effects of fatty liver. I concluded that PEDF is protective against metastatic progression of ocular melanoma in the liver. These findings may prove useful for future therapies.

Table of Contents

Chapter 1--Introduction to Uveal Melanoma and PEDF ......................................... 1

1.1--Purpose and Central Hypothesis...................................................................... 3

1.2--Uveal Melanoma........................................................................................... 5

1.2.1--Biology of the Primary Uveal Melanoma Tumor................................................. 5

1.2.2--Diagnosis, Treatment, and Prognosis............................................................. 8

1.2.3--Epidemiology of Uveal Melanoma................................................................. 10

1.2.4--Etiology and Risk Factors........................................................................... 11

1.3--Metastasis of Uveal Melanoma...................................................................... 14

1.3.1--Epidemiology of UM Metastasis................................................................... 14

1.3.2--Dissemination of Tumor Cells to the Liver...................................................... 15

1.3.3--Progression from Micro to Macrometastases.................................................. 18

1.3.4--Biology of the Metastatic Liver Microniche.................................................... 21

1.3.5--Portal versus Lobular Pattern of Metastatic Growth........................................ 24

1.3.6--Mouse Models of Metastatic Uveal Melanoma................................................ 28

1.4--Properties Regulating Met. Progression: Angiogenesis........................................ 30

1.4.1--Metastatic Tumor Hypoxia......................................................................... 30

1.4.2--Tumor Angiogenesis and the Angiogenic Switch............................................. 31

1.4.3--Factors Regulating Tumor Angiogenesis........................................................ 35

1.4.4--Vasculogenic Mimicry................................................................................ 38

1.5--Properties Regulating Met. Progression: Stromagenesis...................................... 40

1.5.1--Stromagenesis and the Hepatic Stellate Cell................................................. 40

1.5.2--Metastatic Tumor Stromagenesis................................................................ 41

1.5.3--Factors Regulating Tumor Stromagenesis...................................................... 44

1.6--Pigment Epithelium-Derived Factor (PEDF)....................................................... 45

1.6.1--History and Properties of PEDF................................................................... 45

1.6.2--The Role of PEDF in Angiogenesis................................................................ 51

1.6.3--The Role of PEDF in Stromagenesis.............................................................. 52

1.6.4--The Role of PEDF in Lipid Metabolism........................................................... 53

1.6.5--PEDF and Tumor Biology........................................................................... 54

1.6.6--The PEDF Null Mouse................................................................................ 55

1.7--Introduction Summary................................................................................. 56

Chapter 2--Host Pigment Epithelium-Derived Factor (PEDF) Prevents

Progression of Liver Metastasis in a Mouse Model of Uveal Melanoma .................. 57

2.1--Abstract ................................................................................................. 61

2.2--Introduction............................................................................................. 62

2.3--Methods.................................................................................................. 67

2.4--Results ................................................................................................... 72

2.5--Discussion............................................................................................... 87

Chapter 3--Further Investigation of Angiogenesis in our

Mouse Model of Metastatic Uveal Melanoma..................................................... 90

3.1--Further Characterization of Angiogenesis: Rationale......................................... 92

3.2--Further Characterization of Angiogenesis: Methods.......................................... 95

3.3--Further Characterization of Angiogenesis: Results............................................ 97

3.4--Further Characterization of Angiogenesis: Discussion...................................... 105

Chapter 4--Effects of Fatty Liver on Metastasis Progression............................ 108

4.1--Fatty Liver & Metastasis: Rationale............................................................. 110

4.2--Fatty Liver & Metastasis: Methods.............................................................. 111

4.3--Fatty Liver & Metastasis: Results................................................................ 114

4.4--Fatty Liver & Metastasis: Discussion........................................................... 119

Chapter 5--Summary and Conclusions........................................................... 120

5.1--Summary of Results................................................................................. 121

5.2--Discussion of Results................................................................................ 123

5.3--Future Directions..................................................................................... 130

5.4--Conclusion.............................................................................................. 132

References.................................................................................................... 133

End ............................................................................................................. 151

List of Figures

Chapter 1--Introduction to Uveal Melanoma and PEDF

Fig. 1--Uveal Melanoma in the Eye....................................................................... 7

Fig. 2--Metastasis of Melanoma from the Eye to the Liver....................................... 17

Fig. 3--Metastasis Progression in the Liver........................................................... 19

Fig. 4--Portal versus Lobular Pattern of Metastatic Growth...................................... 26

Fig. 5--The Angiogenic Switch........................................................................... 34

Fig. 6--Stromagenesis in the Liver...................................................................... 43

Fig. 7--Pigment Epithelium-Derived Factor (PEDF).................................................. 49

Chapter 2--Host Pigment Epithelium-Derived Factor (PEDF) Prevents

Progression of Liver Metastasis in a Mouse Model of Uveal Melanoma

Fig. 8--PEDF mRNA in PEDF Wt & Null Mouse Liver.................................................. 73

Fig. 9--PEDF Protein in PEDF Wt & Null Mouse Liver................................................ 74

Fig. 10--PEDF Protein in B16-LS9 Cells................................................................. 75

Fig. 11--Eye Tumor Size in PEDF Wt/Het/Null Mouse............................................... 76

Fig. 12--Area of Liver Metastasis in PEDF Wt/Het/Null Mice...................................... 78

Fig. 13--Number of Metastases in PEDF Wt/Het/Null Mice........................................ 79

Fig. 14--Vascular Density in PEDF Wt/Het/Null Mets and Liver.................................. 82

Fig. 15--Hepatic Stellate Cell Density in PEDF Wt/Het/Null Mets................................ 85

Fig. 16--Type III Collagen Density in PEDF Wt/Het/Null Mets.................................... 86

Chapter 3--Further Investigation of Angiogenesis in our

Mouse Model of Uveal Melanoma

Fig. 17--Number of Immature Blood Vessels in Intermediate Mets.............................. 98

Fig. 18--CD31 Immunostain Intensity in PEDF Wt/Het/Null Mets................................ 99

Fig. 19--CD105 Immunostain Intensity in PEDF Wt/Het/Null Mets............................. 101

Fig. 20--Portal Pattern Growth of PEDF Wt/Het/Null Mets....................................... 102

Fig. 21--Vascular Density in PEDF Wt/Het/Null Eyes.............................................. 103

Chapter 4--Effects of Fatty Liver on Metastasis Progression

Fig. 22--Timeline of High Fat or Control Diet in Mouse Model of UM.......................... 112

Fig. 23--Body Weight in High Fat vs. Control Mice................................................ 115

Fig. 24--Hepatocyte Lipid Droplets in High Fat vs. Control Mice............................... 116

Fig. 25--Metastasis Number and Size in High Fat vs. Control Mice............................ 117

Chapter 5--Summary and Conclusions

Fig. 26--Model of the Two Patterns of Metastatic Growth in the Liver...................... 126

Fig. 27--Hypothetical Microniche of Metastatic UM in the Liver................................ 128

End ............................................................................................................. 151

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