Multiple beta cell-independent mechanisms drive hypoglycemia in Timothy syndrome.

The canonical G406R mutation that increases Ca2+ influx through the CACNA1C-encoded CaV1.2 Ca2+ channel underlies the multisystem disorder Timothy syndrome (TS), characterized by life-threatening arrhythmias. Severe episodic hypoglycemia is among the poorly characterized non-cardiac TS pathologies. While hypothesized from increased Ca2+ influx in pancreatic beta cells and consequent hyperinsulinism, this hypoglycemia mechanism is undemonstrated because of limited clinical data and lack of animal models. We generated a CaV1.2 G406R knockin mouse model that recapitulates key TS features, including hypoglycemia. Unexpectedly, these mice do not show hyperactive beta cells or hyperinsulinism in the setting of normal intrinsic beta cell function, suggesting dysregulated glucose homeostasis. Patient data confirm the absence of hyperinsulinism. We discover multiple alternative contributors, including perturbed counterregulatory hormone responses with defects in glucagon secretion and abnormal hypothalamic control of glucose homeostasis. These data provide new insights into contributions of CaV1.2 channels and reveal integrated consequences of the mutant channels driving life-threatening events in TS.

Cholestasis impairs gut microbiota development and bile salt hydrolase activity in preterm neonates.

Cholestasis refers to impaired bile flow from the liver to the intestine. In neonates, cholestasis causes poor growth and may progress to liver failure and death. Normal bile flow requires an intact liver-gut-microbiome axis, whereby liver-derived primary bile acids are transformed into secondary bile acids. Microbial bile salt hydrolase (BSH) enzymes are responsible for the first step, deconjugating glycine- and taurine-conjugated primary bile acids. Cholestatic neonates often are treated with the potent choleretic bile acid ursodeoxycholic acid (UDCA), although interactions between UDCA, gut microbes, and other bile acids are poorly understood. To gain insight into how the liver-gut-microbiome axis develops in extreme prematurity and how cholestasis alters this maturation, we conducted a nested case-control study collecting 124 stool samples longitudinally from 24 preterm infants born at mean 27.2 ± 1.8 weeks gestation and 946 ± 249.6 g, half of whom developed physiologic cholestasis. Samples were analyzed by whole metagenomic sequencing, in vitro BSH enzyme activity assays optimized for low biomass fecal samples, and quantitative mass spectrometry to measure the bile acid metabolome. In extremely preterm neonates, acquisition of the secondary bile acid biosynthesis pathway and BSH genes carried by Clostridium perfringens are the most prominent features of early microbiome development. Cholestasis interrupts this developmental pattern. BSH gene abundance and enzyme activity are profoundly reduced in cholestatic neonates, resulting in decreased quantities of unconjugated bile acids. UDCA restores total fecal bile acid levels in cholestatic neonates, but this is due to a 522-fold increase in fecal UDCA. A majority of bile acids in early development are atypical positional and stereo-isomers of bile acids. We report novel associations linking isomeric bile acids and BSH activity to neonatal growth trajectories. These data highlight deconjugation of bile acids as a key microbial function that is acquired in early neonatal development and impaired by cholestasis.