Mitochondrial Function Is Impaired in a Subset of Aged Haematopoietic Stem Cells in Response to Infection

Introduction

The burden of age-related diseases is steadily increasing as our population ages. A better understanding of the physiological changes that occur with age may provide an insight into the processes that promote disease development which in turn may help to identify potential targets for the management of these diseases. Haematopoietic stem cells (HSCs) play a vital role in the bone marrow response to infection and previous work has shown that emergency granulopoiesis is supported by increased mitochondrial function in HSCs (1). HSC numbers have previously been shown to increase with age whilst mitochondrial function is known to decline with age. Here we investigate how mitochondrial health changes in ageing HSCs and how this impacts on HSC function and their ability to respond to stress.

Methods

C57Bl/6 mice were aged to 18-24 months and 8-12-week-old mice were used as controls. Both aged and young mice were sacrificed, and their BM was isolated. Flow cytometry was used to measure numbers of HSCs and mitochondrial content and function using MitoTracker Green and TMRM. Seahorse metabolic flux analysis was used to assess metabolic changes in young and aged HSC populations. Next young and aged C57Bl/6 mice were treated with lipopolycaccharide (LPS) and sacrificed after 16 hours. Their BM was isolated HSC populations were quantified and mitochondrial health was assessed using flow cytometry. Finally, ABT-263 was used to deplete senescent cells in the BM microenvironment. C57Bl/6 mice were treated with 100mg/kg of ABT-263 or vehicle control by oral gavage for 17 days, they were then treated with 0.4mg/kg of LPS and sacrificed after 12 hours. They BM was isolated and analysed by flow cytometry as described above.

Results

In aged mice HSC numbers were increase and this was associated with an increase in mitochondrial content but a decrease in mitochondrial function. Levels of oxidative phosphorylation (OXPHOS) or glycolysis were not significantly changed in aged HSCs in a steady state. In response to infection, however, in aged mice HSC mitochondrial content and function was not shown to increase following LPS treatment when compared to HSCs from young mice. Moreover, LPS significantly increased OXPHOS in young HSCs whilst aged HSCs were shown to favour glycolysis. Finally, aged HSCs were shown to have increased expression of the anti-apoptotic protein BCL-XL but not BCL-2. In vitro analysis of aged HSC showed the BCL-XL inhibitor ABT-263 but not the BCL-2 inhibitor ABT-199 induced HSC apoptosis. Finally, to determine the effect of targeting BCL-XL on HSCs in vivo we treated aged animals with ABT-263 daily for 17 days and then treated these animals with LPS. Results show that treating aged animals with ABT-263 allowed the HSC to use OXPHOS instead of glycolysis in response to stress.

Conclusion

Aged HSCs have altered mitochondrial content and function and this impairs their ability to respond to infection through changes in OXPHOS. Instead aged HSC have increased reliance on glycolysis in response to LPS treatment, which is less efficient, and this may impact on HSC function. Targeting BCL-XL with ABT-263 allows elimination of aged HSCs within the BM, reverses some of the age-related mitochondrial changes observed and improves the HSCP ability to respond to stress.

References

1. Mistry JJ, Marlein CR, Moore JA, Hellmich C, Wojtowicz EE, Smith JGW, et al. ROS-mediated PI3K activation drives mitochondrial transfer from stromal cells to hematopoietic stem cells in response to infection. Proc Natl Acad Sci U S A. 2019;116(49):24610-9.