Project Three

Studying the role of heme detoxification, iron sequestration and CO emission in bacteriostasis and in urothelial cell viability

PI: Jonathan Barasch, MD, PhD

Similar to the lower tracts of the kidney, the urinary bladder expresses Tfcp2l1 and putative targets that have bacteriostatic functions. In the best known example, the lab identified Lcn2 (NGAL), a beta barrel protein that binds bacterial siderophores and limits bacterial growth (with Dr. R. Strong). Expression of NGAL within hours of urothelial or kidney injury, but not volume depletion (with P. Devarajan), has been recognized worldwide as useful attributes of a biomarker of injury. In a second example, the lab has been evaluating a presumptive pathway of heme transport (slc48a1), heme detoxification (HMOX1), Carbon Monoxide emission, iron storage (Fth/Ftl), and iron export (Slc40a1) in urothelial cells, active in steady state but massively upregulated by urinary infection and by hematuria. Using novel chemical analysis and novel reporter mice and knockouts, Barasch lab is studying the role of heme detoxification, iron sequestration and CO emission in bacteriostasis and in urothelial cell viability. 

The Latest “Hot Stuff”

#1 HMOX is Critical for Urinary Defense

The urothelium expresses all of the components for heme metabolism. To determine if this system is necessary in the urinary defense, we succeeded in deletion of HMOX and found disruption of the urothelium. Subtle changes were noted at baseline, but after introduction of UPEC, the urothelium underwent immediate shedding. The data indicate the interplay between bacteria and epithelial cell, and that heme detoxification is a critical metabolic function of the bladder. 

#2 Snapshots of Nascent RNA Reveal Cell- and Stimulus- Specific Responses

We have adapted a method from Gay, Cleary, Doe permitting the isolation of newly synthesized RNA from any specific cell type at any time of choosing after tissue injury. We express a Cre activated enzyme in a cell type of interest. The enzyme labels only newly synthesized RNA. RNA labeling commences upon inoculation of 4-thio-uracil. Hence, cell specific and time specific, nascent RNA is obtained by extracting the organ of interest and purifying thio-labeled RNA without need for tissue disruption or cell isolation prior to RNA capture. Using this technique, we have identified time dependent changes in gene expression with UTI.