CHUNG, SOONKYU, Ph. D. Mechanisms by Which Conjugated ...

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Figure 4.3. LPS stimulates inflammatory cytokine gene expression predominantly in the stromal vascular fraction (SVF) obtained from primary cultures of newly differentiated human adipocytes. Differentiated cultures of human adipocytes were fractionated using 6% Iodixanol (1.03g/ml). Lipid-laden adipocytes fraction (ADF �) were floated, leaving the stromal vascular fraction (SVF �) as pellets. A: Fractionations were verified <strong>by</strong> measuring the gene expression of aP2 and adipocyte enhancer-binding protein1 (AEBP-1) using real time qPCR and Pref-1 using RT-PCR, B: Cultures of differentiated human adipocytes (day 14) were incubated in the presence or absence of LPS (10 ng/ml) for 3 h before fractionation. Relative mRNA expression of interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-α (TNF-α), interleukin-1β (IL- 1β), cyclooxygenase 2 (COX2), adiponectin (APM-1), and PPARγ were investigated using qPCR. Means (+ SEM, n=4) not sharing a common superscript differ significantly (p
Figure 4.4 LPS induction of cytokine gene expression decreases as the degree of adipocyte differentiation increases. Three human (pre)adipocyte cell models containing ~0 (AD0), ~50% (AD50) or ~90% (AD 90) adipocytes were established <strong>by</strong> modulating exposure to differentiation medium (DM1; see Methods for details). A: On day 14, each culture was stained with Oil-red-O to show neutral lipid accumulation, B: On day 14, cultures of human (pre)adipocyts were incubated in the absence (�) or presence(�) with LPS for 3 h and total RNA were harvested for the analyses of TNF-α, IL-6, IL-8 mRNA expression using qPCR in each cell model. Means (+ SEM, n=4) not sharing a common superscript differ significantly (p
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CHUNG, SOONKYU, Ph. D. Mechanisms b
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MECHANISMS BY WHICH CONJUGATED LINO
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APPROVAL PAGE This Dissertation has
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I also would like to thank The Univ
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IV. PREADIPOCYTES MEDIATE LPS-INDUC
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LIST OF FIGURES ix Page Figure 1.1.
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Figure 4.7. LPS-induced NFκB activ
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in animals and some humans (reviewe
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CLA is potentially effective in: 1)
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Anti-adipogenic Mechanisms of CLA T
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2005). Chapter II of this dissertat
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Figure 1. 2. Multiple mechanisms by
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In contrast, studies conducted in a
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activation. Similarly, Chen et al.
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Central Hypothesis and Specific Obj
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CHAPTER II TRANS-10, CIS-12 CLA INC
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of MEK/ERK signaling by trans-10, c
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Materials and Methods Materials All
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the cultures for additional 12 h. A
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precipitated with ethanol, dried, a
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or for 30 min with 10 µM isoproter
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either 30 µM cis-9, trans-11 CLA o
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dependent effects of CLA on the pho
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Rapamycin Blocks CLA’s Increase i
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Figure 2. 2. Trans-10, cis-12 CLA a
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Figure 2. 4. Trans-10, cis-12 CLA a
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Figure 2. 6. Trans-10, cis-12 CLA i
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Figure 2. 8. Trans-10, cis-12 CLA-i
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Discussion Feeding mixed CLA isomer
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in lipolysis may occur via an ERK-d
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espectively, by the MEK inhibitor U
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Furthermore, the CLA-mediated incre
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proteins. Inhibition of NFκB activ
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Sopasakis et al. 2004) and insulin
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antibodies for anti-glyceraldehydre
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pM of human insulin in the presence
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Page 75 and 76: Transfection efficiency was examine
Page 77 and 78: treatment (Brown et al. 2004), we d
Page 79 and 80: myotubes (Sinha et al. 2004; Weiger
Page 81 and 82: controls was found in cytosol (Fig
Page 83 and 84: Depletion of NFκB p65 by RNAi Atte
Page 85 and 86: eports of cytokine-mediated insulin
Page 87 and 88: Figure 3. 2. Trans-10, cis-12 CLA i
Page 89 and 90: Figure 3. 4. Trans-10, cis-12 CLA a
Page 91 and 92: Figure 3. 6. Trans-10, cis-12 CLA-i
Page 93 and 94: Figure 3. 8. Specific depletion of
Page 95 and 96: Figure 3. 10. Working model: Trans-
Page 97 and 98: Based on our working model (Fig 3.1
Page 99 and 100: mRNA levels of TNF-α (Fig. 3). How
Page 101 and 102: humans (Riserus et al. 2004a), and
Page 103 and 104: most likely not impairing the diffe
Page 105 and 106: 1) decreased adipogenic gene expres
Page 107 and 108: adipocytes from WAT in inflammation
Page 109 and 110: differentiation, cultures of SV cel
Page 111 and 112: Immunoblotting and 4MUrea-SDS-PAGE
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Page 117 and 118: cultures, insulin’s stimulation o
Page 119 and 120: AD90 cultures treated with LPS. Con
Page 121 and 122: Table 4.1. List of human-gene speci
Page 123: Figure 4.2. Primary cultures of new
Page 127 and 128: Figure 4. 6. LPS suppresses PPARγ
Page 129 and 130: Figure 4. 8. Inhibitors of NFκB at
Page 131 and 132: Discussion Adipose tissue is a sour
Page 133 and 134: To determine if non-adipocytes othe
Page 135 and 136: attenuation of insulin sensitivity
Page 137 and 138: epression of NFκB target gene expr
Page 139 and 140: EPILOGUE The “Obesity epidemic’
Page 141 and 142: Apart from our initial goal for thi
Page 143 and 144: Concerning membrane specific recept
Page 145 and 146: esearch topic I would like to exami
Page 147 and 148: Berg, A.H., Lin, Y., Lisanti, M.P.,
Page 149 and 150: Chen, C., Koh, A.J., Datta, N.S., Z
Page 151 and 152: Diradourian, C., Girard, J., and Pe
Page 153 and 154: Granlund, L., Juvet, L.K., Pedersen
Page 155 and 156: Imamura, M., Inoguchi, T., Ikuyama,
Page 157 and 158: Loscher, C.E., Draper, E., Leavy, O
Page 159 and 160: Piper, R.C., Hess, L.J., and James,
Page 161 and 162: Sinha, S., Perdomo, G., Brown, N.F.
Page 163 and 164: Vanden Berghe, W., Vermeulen, L., D
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