Taurine-Conjugated Bile Acids Protect Expanding Hematopoietic Stem/Progenitor Cells from Unfolded Protein Stress As Natural Chaperones

Hematopoietic stem cells (HSCs) give rise to all lineages of hematopoietic cells in the body for entire life span and are thus protected from risk factors by multiple defense systems. We have recently discovered that HSCs are highly susceptible to stress caused by accumulation of mis-/un-folded proteins, so called endoplasmic reticulum (ER) stress upon enhanced growth conditions, and addition of a specific type of bile acid (BA), Tauroursodeoxycholic acid (TUDCA), known as a chemical chaperone can maintain functional murine HSCs for 2 weeks in vitro, by reducing ER stress (Miharada et al., Cell Rep. 2014). This work depicts the importance of proper protein quality control in HSC maintenance, particularly during the expansion.

HSCs are kept in dormant state in the adult body, but actively expanding in the fetal liver. BAs are synthesized from cholesterol in the liver. Interestingly, bile acid synthesis is highly up-regulated in the fetal liver during embryogenesis and the composition of fetal BAs gradually reduces after birth. In addition, composition of bile acids in the fetus is different from adult liver, with the vast majority of fetal BAs are of Taurine-conjugated form that is more stable and non-toxic. Of note, hematopoietic cells and hepatocytes producing BAs are in close contact in the fetal liver and HSCs are therefore exposed to BAs, whereas the adult liver has anatomically isolated bile duct structures that separate blood flow and bile flow. However the role for these fetal BAs has been unknown.

Here we report that bile acids support expansion of hematopoietic stem and progenitor cells (HSPCs) in the fetal liver and ex vivo. Since TUDCA is a rare component in human and mouse BAs, even in the fetal liver, we sought analogue(s) that similarly function as ER stress inhibitors. We identified that Taurocholic acid (TCA), one of the main components of fetal BA, and Tauro-alpha-muricholic acid (TαMCA) that is a rodent specific BA have a potential to reduce ER stress, similar to TUDCA. Mouse HSCs cultured with TCA or TαMCA in vitro for 2 weeks showed a robust increase in the reconstitution level compared to non-treated cells (14-fold, n=14, p<0.001 and 13-fold, n=9, p<0.05, respectively), which has comparable or even better potential to support HSC function than TUDCA.

To study physiological roles of BA in ER stress reduction and HSC expansion in the fetal liver, an inhibitor of BA synthesis, GW4064 (an agonist of a nuclear receptor FxR that negatively regulates key enzymes in the BA biosynthesis, CYP7A1 and CYP8B1), was intraperitoneally injected into pregnant mice. E16.5 fetuses derived from GW4064-injected pregnant mice showed severe decrease in the number of HSPCs (0.40-fold, n=28-33, p<0.001) in the fetal liver, due to increased apoptosis triggered by elevated ER stress levels. Importantly, co-injection of TCA or Salubrinal (inhibitor of the ER stress-induced apoptosis signal) rescued the effects of GW4064 on cellularity of the fetal liver and levels of ER stress, confirming that the phenotype seen here is due to increased ER stress resulting from lowered levels of BA.

Analyses of CYP27A1 knockout (KO) mice that have reduced BA synthesis observed decreased HSC number and increased ER stress in the fetal liver, whereas CYP8B1 KO mice that have increased Tα/βMCA synthesis instead of lack of TCA didn’t show any difference.

These findings strongly suggest that fetal BA, particularly TCA and TαMCA, supports the expansion of HSPCs in the fetal liver and the ex vivo culture as chemical chaperones by lowering ER stress levels. Our findings propose a new role of bile acids in hematopoiesis as natural chaperones and provide a novel connection between hematopoiesis and fetal liver.