Tissue-Specific Analysis of UDP-sugar Levels and Ratios in a Rat Model of Classic Galactosemia Öffentlichkeit

Abstract

Classic galactosemia (CG) is a rare, autosomal recessive disorder that results from deficiency of galactose-1-phosphate uridylyltransferase (GALT), the second enzyme in the Leloir pathway. Despite improved survival outcomes in neonates following newborn screening and rapid dietary restriction of galactose, a variety of long-term complications continue to persist in many treated patients, including speech delay, cognitive delay, growth delay, and primary ovarian insufficiency within females. One proposed explanation for the long-term outcomes associated with the disease is the perturbation of UDP-sugar pools hypothesized to underlie alterations of glycosylation. However, conflicting reports of how UDP-sugar levels and ratios are impacted by CG, as well as a lack of studies in primary tissues, have made answering this question difficult. Recently, our lab reported a GALT-null rat model that accurately reflects many important long-term phenotypes associated with CG. In the current study, we quantified levels of UDP-sugars in liver and brain homogenate samples from  ten-day old GALT-null rat pups as well as controls. Our results indicate that UDP-sugar levels and ratios vary across tissue type as well as genotype. Despite the small sample size limiting the statistical power of this study, our results clearly establish the importance of UDP-sugar analysis in understanding CG, as well as highlight the need for further study within this area of research. In the future, we hope to expand this study to include red blood cells (which are clinically significant due to their use in diagnosis within human patients) and an adult rat cohort in order to test for age and diet-dependent differences.

Table of Contents

Table of Contents

Chapter 1: Introduction to Galactose Metabolism and Classic Galactosemia…………………… p. 1

Chapter 2: UDP-sugar levels and ratios are perturbed on a tissue-specific basis in GALT-null

  rats………………………………………………………………………………………………………………….           p. 6

Chapter 3: Discussion, Limitations, and Future Directions……………………………………………….. p. 30

              

Tables

Table 1: Description of rat cohorts by genotype…………………………………………………………….. p. 6

Table 2: Summary statistics and tests for metabolite concentrations (pmol/mg tissue) for          

M3/M3 and WT/WT rat brain homogenate and liver tissue……………………………..........p. 15

Table 3: Summary statistics for UDP-sugar concentrations (pmol/mg tissue) and ratios for         

 M3/M3 and WT/WT rat liver tissue………………………………………………………………….. p. 17

Table 4: Summary statistics for UDP-sugar concentrations (pmol/mg tissue) and ratios for         

 M3/M3 and WT/WT rat brain homogenate tissue……………………………………………......p. 25

Figures

 Figure 1. Galactose metabolism via the Leloir pathway.………………………………………………….. p. 1

Figure 2. Alternative pathways of galactose metabolism…………………………………………………..p. 3

Figure 3. Differences in rat liver glucose-1-phosphate, galactose-1-phosphate, galactitol, and     

  galactose levels, by genotype………………………………………………………………………....p. 12

Figure 4. Differences in rat brain homogenate glucose-1-phosphate, galactose-1-phosphate,

               galactitol, and galactose levels by genotype.……………………………………………………...p. 14

Figure 5. Genotype-specific differences in rat liver UDP-galactose absolute levels…………….......p. 18

Figure 6. Differences in rat liver absolute UDP-glucose concentrations by genotype and             

 litter…………………………………………………………………………………………………………...p. 19

Figure 7. Genotype-specific differences in rat liver UDP-glucose/UDP-galactose relative              

ratios.………………………………………………………………………………………………………….p. 21

Figure 8. Differences in rat liver UDP-N-acetylglucosamine levels, UDP-N-acetylgalactosamine     

levels, and UDP-glcNAc/UDP-galNAc by genotype..……………………………………………...p. 23

Figure 9. Differences in rat brain homogenate UDP-galactose levels, UDP-glucose levels, and       

UDP-glucose/UDP-galactose ratio..……………………………………………………………………p. 27

Figure 10. Differences in rat brain homogenate UDP-N-acetylgalactosamine levels, UDP-N           

-acetylglucosamine levels, and UDP-glcNAc/UDP-galNAc ratio...………………..…….......p. 29

About this Honors Thesis

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School	Emory College
Department	Biology
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Genetics
Stichwort	UDP-sugars Classic Galactosemia
Committee Chair / Thesis Advisor	Fridovich-Keil, Judith, Emory University
Committee Members	Taliaferro-Smith, LaTonia, Emory University Carter, Eloise, Emory University Hickman, Meleah, Emory University

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