Faculty Bibliography

Background: Null mice raised by mating -/- with -/- showed sustained hyperglycemia & persistent hyperinsulinemia during glucose tolerance test (GTT) vs wild type (wt) raised by mating wt with wt . Insulin resistance by HOMA was increased 8 fold, but HOMA beta cell function was normal. As null mice ate & weighed more, & as caloric restriction abolished these differences, our data support the role of hyperphagia, possibly due to hypothalamic neuron leptin resistance without TRPC1 channels.
Method(s): We studied adipokines, environmental & genetic factors, including gene dosage, in this diabetic phenotype while minimizing epigenetics & hyperphagia in breeders & dams, using only littermates born to +/-breeders.
Result(s): 9-week-old null born to -/-breeders & nursed by -/-dams had severe random hyperglycemia (171 vs. 98 mg %) vs wt born to +/+ breeders & raised by +/+ dams. In contrast, null mice born to +/-breeders & nursed by +/-dams had minimal hyperglycemia (119 vs 171 mg %), implying anti-diabetic effects by a single maternal wt allele. Conversely, wt born to +/-breeders & nursed by +/-dams were hyperglycemic vs wt born to +/+ breeders & nursed by +/+ dams (118 vs 98 mg %), reflecting pro-diabetic effects of maternal haploid deficiency. These wt were as hyperglycemic as null littermates born to the same +/-breeders & nursed by the same +/-dams (118 vs 119 mg %). These data support the role of nongenetic parental influences. From 5th to 30th week, null but not +/-mice were obese vs wt. At 29 weeks, during GTT, both null & +/-mice were equally diabetic, with glucose (in mg %), respectively of 230 & 234 vs. 183 in wt at 20 min, 219 & 229 vs. 155 in wt at 60 min, & 179 & 188 vs 134 in wt at 90 min. These data support the role of both alleles in glucose homeostasis. In null mice, adiponectin was down (5.7) vs. wt (6.4) & +/-(6.2 mug/ml), but leptin up (2.3 vs. 1.3 in wt & 1.8 ng/ml in +/-).
Conclusion(s): We conclude: 1. Diploid TRPC1 gene deletion produces hyperphagia & obesity. 2. Haploid deficiency suffices to produce diabetes, associated with reduced adiponectin & increased leptin. 3. Non-genetic parental factors, via epigenetics & hyperphagia, markedly alter glucose homeostasis independent of genotypes

BACKGROUND:There is growing evidence that the accumulation of protein- bound uremic retention solutes, such as indoxyl sulfate, p-cresyl sulfate and kynurenic acid, play a role in the accelerated cardiovascular disease seen in patients undergoing chronic hemodialysis. Protein-binding, presumably to albumin, renders these solutes poor-dialyzable. We previously observed that the free fraction of indoxyl sulfate was markedly reduced at the end of hemodialysis. We hypothesized that solute binding might be pH-dependent and attributed the changes in free solute concentration to the higher serum pH observed at the end of standard hemodialysis with dialysis buffer bicarbonate concentration greater than 35 mmol/L. We observed that acidification of uremic plasma to pH 6 in vitro greatly increased the proportion of freeIS. METHODS:We tested our hypothesis by reducing the dialysate bicarbonate buffer concentration to 25 mmol/L for the initial half of the hemodialysis treatment ("isohydric dialysis"). Eight stable hemodialysis patients underwent "isohydric dialysis" for 90 minutes and then were switched to standard buffer (bicarbonate = 37mmol/L). A second dialysis, 2 days later, employed standard buffer throughout. RESULTS:We found a clearcut separation of blood pH and bicarbonate concentrations after 90 minutes of "isohydric dialysis" (pH = 7.37, bicarbonate = 22.4 mmol/L) and standard dialysis (pH = 7.49, bicarbonate = 29.0 mmol/L). Binding affinity varied widely among the 10 uremic retention solutes analyzed. Kynurenic acid (0.05 free), p-cresyl sulfate (0.12 free) and indoxyl sulfate (0.13 free) demonstrated the greatest degree of binding. Three solutes (indoxyl glucuronide, p-cresyl glucuronide, and phenyl glucuronide) were virtually unbound. Analysis of free and bound concentrations of uremic retention solutes confirmed our prediction that binding of solute is affected by pH. However, in a mixed models analysis, we found that the reduction in total uremic solute concentration during dialysis accounted for a greater proportion of the variation in free concentration, presumably an effect of saturation binding to albumin, than did the relatively small change in pH produced by isohydric dialysis. The effect of pH on binding appeared to be restricted to those solutes most highly protein-bound. The solutes most tightly bound exhibited the lowest dialyzer clearances. An increase in dialyzer clearance during isohydric and standard dialyses was statistically significant only for kynurenic acid. CONCLUSION/CONCLUSIONS:These findings provide evidence that the binding of uremic retention solutes is influenced by pH. The effect of reducing buffer bicarbonate concentration ("isohydric dialysis:"), though significant, was small but may be taken to suggest that further modification of dialysis technique that would expose blood to a greater decrease in pH would lead to a greater increase the free fraction of solute and enhance the efficacy of hemodialysis in the removal of highly protein-bound uremic retention solutes.