Age Differences in the Stimulation of Murine Hepatic XBP‐1 Splicing in Response to the Anti‐HIV Drug Efavirenz

Efavirenz is a non‐nucleoside reverse transcriptase inhibitor used in combination with other antiretroviral drugs to treat HIV infection. Efavirenz is one of the most commonly prescribed antiretrovirals and is used in both adult and pediatric populations. Efavirenz treatment has been associated clinically with hepatotoxicity; however, the mechanism underlying this is not fully understood and work in this area has largely been focused on adults. With this in mind, using primary mouse hepatocytes, we probed whether the activation status of transcription factors, such as X‐box binding protein‐1 (XBP‐1), that play a crucial role in regulating cell survival are differentially impacted by efavirenz as a function of age. To do so, the ratio of spliced (activated) to unspliced XBP‐1 mRNA was measured following efavirenz treatment. Primary mouse hepatocytes were isolated from pediatric (3–4 week old) and adult (8–12 week old) male C57Bl/6J mice. XBP‐1 splicing following incubation with efavirenz for 4 hours was measured using reverse transcriptase PCR. In 3–4 week old mice, treatment with 10 μM efavirenz for 4 hours resulted in a 3.4‐fold increase in the ratio of spliced to unspliced XBP‐1 as compared to vehicle controls; however, XBP‐1 splicing was not elevated in response to this same treatment in adult mice. In fact, a 30 μM concentration of efavirenz was required in adult mice to stimulate a comparable (3.9‐fold) increase. Because activation of XBP‐1 splicing can affect cell survival, a cell death assay using ethidium bromide and acridine orange was performed and these experiments were confirmed using a TUNEL assay. In primary hepatocytes isolated from 3–4 week old mice, a 4.8‐fold increase in cell death was observed following treatment with 20 μM efavirenz. Similarly, using hepatocytes from adult mice, cell death was stimulated 4‐fold by treatment with 20 μM efavirenz. In the presence of STF080310, a known inhibitor of XBP‐1 splicing, cell death in response to 20 μM efavirenz was reduced to 2.5‐fold and 1.8‐fold over controls in 3–4 week old and adult mice, respectively. These results were commensurate with data obtained using TUNEL staining. In conclusion, hepatocytes from mice at early stages of development may be more sensitive to activation of XBP‐1 splicing than adult mice. Furthermore, cell death was similar in the 3–4 week old mice and adult mice suggesting that age may not result in differential susceptibility to hepatocyte death in response to efavirenz. Support or Funding Information This research is funded by NIH R01 GM103853. This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .