Oxidative stress goes hand in hand with inflammation and their underlying mechanisms are inextricably interconnected [
1]. Growing evidence suggest that oxidative stress and neuroinflammation underpin a diverse range of CNS diseases, including stroke, traumatic brain injury, multiple sclerosis, Alzheimer’s, Parkinson’s, and other neurodegenerative diseases [
2‐
5]. The brain is particularly vulnerable to oxidative stress due to an abundance of iron and polyunsaturated fatty acids which are susceptible to lipid peroxidation [
6,
7], thus underscoring the importance of maintaining redox balance for normal brain functioning. One mechanism by which the brain defends itself against oxidative insults is via the upregulation of antioxidant molecules. Interestingly, oxidative stress strongly induces expression of heme oxygenase-1 (HO-1) in the CNS, a system not actively involved in red blood cell metabolism [
8]. Indeed, while HO-1’s primary function is the catalysis of heme, leading to the eventual formation of antioxidant bilirubin, carbon monoxide, and ferrous iron (Fe
2+), and in the process, prevents heme-mediated free radical production [
9], HO-1 is also found to have anti-neuroinflammatory and neuroprotective properties in the CNS [
8,
10]. Furthermore, HO-1 upregulation has been reported to be a mechanism by which certain isoflavone metabolites protect astrocytes from hydrogen peroxide-induced reactive oxygen species [
11]. Together with their well-recognized roles in facilitating neuronal trophic support, biochemical homeostasis, blood brain barrier integrity, response to neuroinflammatory signals and scar formation, astrocytes play a critical role in redox homeostasis and are the major source of antioxidant molecules and enzymes which protect them and the neurons they support from oxidative stresses [
12,
13]. The critical involvement of astrocytes in neuroinflammation and oxidative stress protection as well as the pathogenicity of astrocyte dysregulation in various CNS diseases [
14‐
16] gave research impetus to discover and characterize novel anti-neuroinflammatory/antioxidant therapeutics with efficacy on astrocytes. Besides the aforementioned work on isoflavone metabolites [
11], a wide range of other bioactive molecules have been studied. Of these, andrographolide is a labdane diterpenoid derived from the herbaceous
Andrographis paniculata, which has been traditionally used in Asia to treat a variety of ailments, including fever, cough, tuberculosis, snake bites, respiratory tract, and urinary tract infections [
17,
18]. Andrographolide has been reported to exhibit anti-inflammatory and antioxidant activities in peripheral tissues [
19]. Furthermore, we have previously shown that the brain-penetrant andrographolide attenuates IL-1β or lipopolysaccharide-stimulated upregulation of the C–C and C–X–C chemokines in the brain cortex as well as in cultured astrocytes [
20,
21]. However, while the efficacy of andrographolide in reducing oxidative stress in the CNS has been demonstrated in several studies [
22‐
24], the underlying molecular mechanisms have not been thoroughly ascertained. Furthermore, andrographolide seems to have pleiotropic effects on signaling pathways involved in inflammatory and oxidative stress responses, but the mechanisms underlying these effects appear different in various cell types [
20,
21,
25‐
27]. In this study, our focus is to investigate the effects of andrographolide on HO-1 expression in astrocytes. Because HO-1 is a known gene target of transcription factor NF-E2-related factor 2 (Nrf2), which is critically involved in cellular defense against oxidative stress [
11,
28,
29], we also studied andrographolide effects on astroglial Nrf2 regulation.