Fig.3 Drugs with hepatotoxicity modulated by gut microbiota:
acetaminophen (A), tacrine (B), triptolide (C), and geniposide (D). Red
structures signify hepatotoxic drugs or metabolites; green ones signify
gut-dervied metabolites that confers hepatoprotective effects against
relevant drugs. A: PPD is a gut-derived metabolites that exacerbates
APAP-induced hepatotoxicity by competing for GSH and so doesp-cresol which competes for sulfotransferase. DOPA, 4-HPA, and
fructose are prebiotics that ameliorate APAP-induced hepatotoxicity. B:
β-glucuronidase producing bacteria including Bacteroides ,Enterobacteriaceae and Lactobacillus aggravate tacrine
hepatotoxicity. C: Gut-derived propionate ameliorates triptolide-induced
hepatotoxicity. D: Geniposide transforms into hepatotoxic genipin under
the action of gut micorbiota. Abbreviations: 4-HPA,
4-hydroxyphenylacetic acid; APAP, acetaminophe; CYPs, cytochrome 450
proteins; DOPA, 3,4-dihydroxyphenylacetic acid; GSH, glutathione; NAPQI,
N-acetyl-p-benzoquinone imine; SULTs, sulfotransferases; UGTs,
UDP-glucuronosyltransferases.
3.1 Acetaminophen
Acetaminophen
(APAP) is a representative hepatotoxic drug accounting for most cases of
DILI in the United States (Chao et al., 2018). When administered orally,
it mainly undergoes two ways of metabolism: (a) sulfonated or
glucuronidated by phase Ⅱ enzymes and (b) first converted into the
hepatotoxic N-acetyl-p-benzoquinone imine (NAPQI) by CYPs and then
conjugated with glutathione (GSH) (Chang et al., 2020c). Both ways of
metabolism function as detoxification and thus any factor that
influences them could have an impact on APAP-related hepatotoxicity. A
human experiment demonstrated that p-cresol , microbiota-derived
metabolite from tyrosine, competed for sulfation with APAP, which might
induce APAP accumulation and result in hepatotoxicity (Clayton et al.,
2009). This experiment introduced gut microbiota into the understanding
of APAP-related hepatotoxicity for the first time. Indeed, germ-free
mice are less sensitive to APAP-induced hepatotoxicity compared with
conventional mice (Possamai et al., 2015) and antibiotic treatment
decreases urinary sulfonated APAP (Malfatti et al., 2020), suggesting
that intestinal microbiota is implicated in APAP-related hepatotoxicity
and metabolism. It has long been known that APAP-induced liver injury
exhibits diurnal variation, which is more severe in the night than
during the daytime (Kim and Lee, 1998). Despite the hepatic GSH level
and drug metabolic activity mentioned as contributing factors of this
phenomenon (Kim and Lee, 1998), the gut microbiota is thought to play a
pivotal role therein. Evidence suggests that gut microbiota biogeography
and metabolome undergo diurnal fluctuation, which consequently regulates
rhythmic liver transcriptome and detoxification pattern as proved by
APAP-induced circadian hepatotoxicity (Thaiss et al., 2016). The role of
intestinal microbiota in rhythmic hepatotoxicity of APAP is further
confirmed by another study and it also reported that
1-phenyl-1,2-propanedione (PPD), a gut microbial metabolite exacerbated
APAP-induced liver injury by depleting hepatic GSH (Gong S, Lan T, Zeng
L, Luo H, Yang X, Li N, Chen X, Liu Z, Li R, Win S, Liu S, Zhou H,
Schnabl B, Jiang Y, Kaplowitz N, 2018). A pharmacokinetic study
demonstrated that treatment with probiotics, especiallyLactobacillus reuteri K8, diminished absorption of orally
administered APAP (Kim et al., 2018), so it is speculated that
pretreatment of moderate probiotics may ameliorate APAP-induced
hepatotoxicity while maintaining efficacy simultaneously.
Based on the gut microbial mechanism of APAP-induced liver injury, some
microbiota, diet-derived substances, and probiotics have been discovered
to exert hepatoprotective effect against APAP by targeting the gut
microbiota. 3,4-dihydroxyphenylacetic acid (DOPA), a microbial
metabolite of quercetin, protects the liver from damage of APAP by
activating nuclear factor erythroid 2-related factor 2 (Nrf-2) and
enhancing phase II and antioxidant enzymes (Xue et al., 2016), so does
4-hydroxyphenylacetic acid (4-HPA), a major microbial metabolite of
polyphenols (Zhao et al., 2018). As for food-derived hepatoprotective
agents, a study found that supplemental fructose respited APAP-induced
liver injury by altering gut microbial community, especially the genusAnaerostipes , which closely relates to hepatic levels of CYPs and
GSH (Cho et al., 2017). Furthermore, probiotics like bacillus spores
have been proven to protect rat from APAP-induced acute liver injury by
altering proinflammatory cytokines levels (Neag et al., 2020). It seems
gut microbial modulation does work in attenuating APAP-triggered liver
injury, mainly by dietary modulation and probiotics intake, but it has
to be verified in human trails for future application.
3.2 Tacrine
Tacrine was the first approved drug prescribed to treat Alzheimer’s
disease but was withdrawn from the market for its large pharmacokinetic
variation (Jarrott, 2017) and consequent unpredictable hepatotoxicity
(Bethesda (MD): National Institute of Diabetes and Digestive and Kidney
Diseases, 2018). Although it is no longer used in clinical, underlying
mechanisms involved in tacrine-induced liver injury is still being
investigated (Park et al., 2015) and it is utilized to establish liver
injury model in cells and animals (Stachlewitz et al., 1997; Choi et
al., 2015). Previous researches mainly focused on the hepatic metabolism
of tacrine and found that tacrine was extensively metabolized by CYP2A1
and one of the hydroxylated metabolites exerted hepatotoxicity (Madden
et al., 1993). Fluvoxamine is a potent inhibitor of CYP2A1 and is
thought to attenuate tacrine-induced hepatotoxicity by reducing the
formation of toxic metabolites but needs further validation (Rasmussen
et al., 1998). A recent comprehensive study combining pharmacokinetics,
metabolomics, and metagenomics revealed that rats responded differently
to tacrine with strong responders exhibiting higher tacrine exposure,
greater deglucuronidation capabilities, and an abundance of
β-glucuronidase
producing bacteria including Bacteroides ,Enterobacteriaceae and Lactobacillus (Yip et al., 2018).
Further validation indicated that coadministration of oral
β-glucuronidase or antibiotics with
tacrine increased or decreased susceptibility to tacrine-induced
hepatotoxicity, indicating that gut microbiota especially the gut
microbial β-glucuronidase plays a pivotal role in tacrine-related
hepatotoxicity (Yip et al., 2018). It implicates the significance of gut
microbiota in interindividually varied toxicity of drugs. While current
researches attempt to decipher individual medication differences by
genome but with limited achievements, the future focus can be shifted to
gut microbiota.
3.3 Triptolide
Hepatotoxicity is one of the major
factors that hamper the application and progress of herbal medicine
nowadays. Triptolide is the major bioactive component of the herbTripterygium wilfordii Hook F which exerts anti-inflammatory and
anti-autoimmune functions but also confers hepatotoxicity. A study
elucidated that triptolide exerted anti-inflammatory effect against
ulcerative colitis with gut microbiota involved (Wu et al., 2020),
indicating an interactive relationship between gut microbiota and
triptolide. Molecular mechanisms of triptolide-induced hepatotoxicity
have been comprehensively delineated, encompassing membrane damage,
mitochondrial disruption, metabolism dysfunction, endoplasmic reticulum
stress, oxidative stress, apoptosis, and autophagy (Xi et al., 2017),
but the role of gut microbiota plays in it is put forward only recently.
Researchers found that pretreatment of antibiotics exacerbated liver
injury induced by triptolide in mice while supplemental
microbiota-derived propionate proffered liver protection against
triptolide (Huang et al., 2020a). It reveals that gut microbiota
protects the liver from triptolide, contrary to the case in tacrine.
Microbial metabolites of the prototype drugs may be an important factor
herein, which indicates that microbial metabolites of triptolide confer
lower hepatotoxicity while those of tacrine higher. Indeed, gut
microbiota plays a unique role in triptolide metabolism since gut
microbiota and liver microsomes produce different metabolites of
triptolide (Peng et al., 2020). It indicates that gut microbial
metabolism constitutes a significant part in drug metabolism and under
some circumstances it influences and even determines toxic outcomes of
drugs. Besides, triptolide treatment induced bile acids accumulation in
rats and it was associated with suppression of FXR and hepatic Sirtuin
(Sirt1) expression, activation of which ameliorated triptolide-induced
hepatotoxicity (Yang et al., 2017).
3.4 Geniposide
Geniposide is the major bioactive
component of herb Gardeniae Fructus which exhibits a wide range
of pharmacological functions including neuroprotective, hypoglycemic,
hepatoprotective, anti-inflammatory, analgesic, cardio-protective,
antioxidant, immune-regulatory, antithrombotic, and antitumoral effects
(Zhou et al., 2019b). The herb has long been used as choleretic to treat
jaundice in Chinese history and modern research also reveals that
geniposide attenuates APAP-induced liver injury in mice (Yang et al.,
2019). Nonetheless, it has been reported that geniposide elevated serum
alanine aminotransferase (ALT) and aspartate aminotransferase (AST)
levels in rats and the conversion of geniposide to genipin made a big
difference therein (Yamano et al., 1990). Genipin, the aglycon of
geniposide, is produced from geniposide by intestinal bacteria but not
by the liver (Aburada and Kobashi, 1994) and displayed much higher
toxicity than geniposide in HepG2 cells (Kang et al., 2012), indicating
the important role of gut microbiota in geniposide-induced liver injury.
Antibiotic treatment of rats enhanced in vivo exposure of
geniposide by suppressing metabolic activities of gut microbiota (Jin et
al., 2014) and it might reduce hepatotoxicity because of a decreased
level of genipin. A further study demonstrating mechanism involved in
geniposide-induced liver injury attributed it to bile acids synthesis
alteration (Tian et al., 2017), but the role of intestinal microbiota
plays in it remains to be investigated. A recent study also proved the
significance of intestinal microbiota in geniposide-related
hepatotoxicity and further demonstrated that genipin dialdehyde
intermediate, which was capable of conjugating with amino acids
spontaneously, was the origin of geniposide-induced hepatotoxicity (Li
et al., 2019).
3.5 Substances other than drugs
Besides drugs, the gut microbiota is also extensively engaged in other
substances-induced hepatotoxicity which will be discussed here as well.
Some of the researches are profound in mechanistic elucidations and can
provide as references for mechanistic studies of drug-induced
hepatotoxicity.
Aflatoxins B1 (AFB1), a kind of mycotoxin, causes liver fibrosis and
hepatocellular carcinoma both in humans and livestock. AFB1 exposure
caused a wide range of metabolic disturbances and particularly elevated
gut microbial cometabolites including phenylacetylglycine and hippurate
in rats (Zhang et al., 2011). Other metabolomics based studies also
found that AFB1 disrupted several important metabolic pathways of gut
microbiota, including synthesis of SCFAs, pyruvic acid-related pathways,
amino acids, bile acids, and long-chain fatty acids metabolism (Zhou et
al., 2018b, 2019a), suggesting that gut microbial metabolism disruption
contributes to the pathogenesis of AFB1-induced diseases. Probiotics
supplementation including Lactobacillus Species (Liew et al.,
2019; Wacoo et al., 2020), Propionibacterium freudenreichii(El-Nezami et al., 2006), Bacillus subtilis , andCandida utilis (Chang et al.,
2020b) exerted protective effects against AFB1 both in animal
experiments and human trials. These studies illustrate that AFB1
perturbs gut microbiota which can be restored by probiotics and they
also help degrade AFB1 and thus lower toxicity.
Some simple substances form another main category of environmental
toxicants that exert hepatotoxicity. Long-term exposure of low-dose
cadmium causes severe liver damage with gut microbiota implicated. It
was found that cadmium induced gut dysbiosis, mainly decreasingFirmicutes and γ-proteobacteria levels, and increasing serum
lipopolysaccharide (LPS) and consequently induced hepatic inflammation
(Zhang et al., 2015). It also increased intestinal permeability and
hence inflicted accumulation in the liver but could be partially
restored by antibiotic treatment (Liu et al., 2020). Arsenic is another
example here. Although it is structurally simple, it undergoes complex
biotransformation and forms a series of products with different
toxicity. In vitro incubation elucidated the unique metabolic
role of gut microbiota and in vivo study illustrated that gut
microbiota reduced arsenic load by promoting methylation and thus
protected the host from hepatotoxicity of arsenic (Chi et al., 2019a).
Similarly, another study also found that a stable gut microbiome was a
key determinant of survival to arsenic exposure, andFaecalibacterium prausnitzii protected it (Coryell et al., 2018).
Other environmental toxicants that are found to elicit hepatotoxicity
with gut microbiota engaged include polychlorinated biphenyl 126 (Chi et
al., 2019b), 2,3,7,8-tetrachlorodibenzo-p -dioxin (Fader et al.,
2017), titanium dioxide nanoparticles (Chen et al., 2019), etc. These
studies share similarities in researching method: first, 16S rRNA
sequencing analysis is utilized to demonstrate gut dysbiosis-induced by
the toxicants and then omics analysis especially metabolomics is used to
elucidate disturbed metabolites or metabolic pathways by focusing on gut
microbial metabolites and further deduce possible mechanisms involved by
analyzing results of both. It is a current mainstream in gut microbiota
associated mechanistic studies particularly in researches of metabolic
diseases (Zeng et al., 2020).