Discussion
VVC is a mucocutaneous mycosis caused by Candida that has a significant
impact on women’s quality of life and leads to increased healthcare
costs due to high recurrence rates and increased antifungal
resistance[34]. C. albicans is the most common opportunistic
pathogen in VVC and is present in the respiratory, gastrointestinal, and
genitourinary tract of more than 30 percent of healthy individuals
during their lifetime[2]. Usually, yeast-phase C. albicans is
tolerated by the vaginal epithelium. However, in VVC, C. albicans
exhibits an aggressive form of hyphae, producing various extracellular
enzymes, such as secretory aspartate protease (SAP), phospholipase, and
hemolysin, which invade tissues through adhesion, penetration, hyphae
invasion, and endothelial colonization of vaginal epithelial
cells[35].
An increasing number of metabolites have been discovered and defined
with the development of metabolomics research. Metabolomics studies can
investigate a variety of biological processes and phenotypes by means of
metabolites[36], such as amino acid and lipid metabolites[37].
Metabolomics has been extensively studied in inflammation-related
diseases such as hyperuricemia[38], inflammatory bowel
disease[39], rheumatoid arthritis[40], sepsis[41], liver
failure[42], and endometriosis[43]. Research evaluating cervical
and vaginal lavage fluids in patients with inflammatory vaginal disease
has found associations between the composition of vaginal microflora and
metabolite profiles[32]. Furthermore, a lipidomics study of VVC and
cellolytic vaginopathy (CV) have shown significant differences in lipid
composition. Lipids play an essential role in maintaining the
homeostasis of the vaginal microenvironment[44]. However, few
studies have comprehensively studied the metabolomics profile of VVC.
In the present study, untargeted metabolomic analysis was performed to
further explore the potential biological functions of these metabolites
and their role in VVC genesis following C. albicans infection, and to
determine the differential metabolic profiles between the VVC and CTL
groups. The metabolic profiles of patients with VVC were significantly
different from those without. In total, 211 differential metabolites
were identified, including 128 upregulated metabolites and 83
downregulated metabolites in VVC patients compared with healthy
individuals.
Among these differential metabolites, significant increases in sugar
alcohols, amino acids, unsaturated fatty acids, and pyrimidines were
observed in the VVC group. Conversely, higher levels of organic acids
and fatty acyl groups were observed in the CTL group. An analysis of the
diagnostic capability of well-characterized biomarkers (ROC) showed that
L-(-)-arabitol, L-tyrosine, D-(+)-arabitol, and psychosine could
effectively distinguish VVC from healthy people (AUC>0.97).
Additionally, more than ten metabolites (AUC>0.8),
including linoleic acid, arachidonic acid, L-pyrrolysine, and uracil,
were found to have good discrimination capacity, which are potential
biomarkers to distinguish VVC from CTL patients. Tyrosine was thought to
be neurotoxic and capable of causing neurological disorders such as
encephalitis[45]. Arabitol in serum[46] and urine[47] has
been used as a biomarker to rapidly diagnose invasive candidiasis and
can be used to inform antifungal therapy and prognosis.
Amino acid metabolism plays an important role in the activation of
immune cells and the production of antibodies, and excess amino acids
may be detrimental to the immune system[48]. In this study, amino
acid metabolism was one of the major metabolic pathways, including
phenylalanine, tyrosine, and tryptophan biosynthesis, phenylalanine
metabolism, and taurine metabolism. Similarly, one research reported
that the same metabolic pathways were enriched in sepsis[41],
indicating that amino acid metabolism was inextricably linked to the
occurrence of inflammation. Moreover, phenylalanine metabolism promotes
the neutrophil-evasive state[49], and excess L-phenylalanine was
found to affect antibody production by inhibiting protein
synthesis[50]. Similar metabolic changes were observed in our study,
and excess L-phenylalanine may contribute to the development of VVC.
Furthermore, linoleic acid metabolism also played a key role, as it
showed the most obvious and broadest variation in the VVC group.
Linoleic acid and arachidonic acid in the VVC group both increased
significantly. Linoleic acid (LA; ω-6,18:2) and arachidonic acid (AA;
ω-6,20:4) are essential fatty acids for the human body and are two of
the most abundant polyunsaturated fatty acids (PUFAs). AA can be
obtained directly from the diet or can be synthesized from LA. One study
found that a larger amount of AA was synthesized from LA than from
dietary sources[51]. Therefore, the significant elevation of LA
caught our attention. Although LA was associated with a reduced risk of
cardiovascular disease[52], studies have shown that excess LA can
promote the occurrence of inflammatory events[53, 54]. Similarly,
metabolites of LA mediated inflammation through oxidative stress, and
oxidized linoleic acid (LA) metabolites (OXLAMs) induced mitochondrial
dysfunction, apoptosis, and activation of NLRP3 inflammasome by means of
oxidative stress, promoting the development of nonalcoholic
steatohepatitis (NASH)[55]. LA-derived hydroxyoctadecenoic acids
(HODEs) have been found to contribute to the progression of
atherosclerosis as inflammatory regulators[56]. Additionally, the LA
metabolites epoxyoctadecenoic acids (EpOMEs) and dihydroxyoctadecenoic
acids (DiHOMEs) activate NF-κB and AP-1 transcription factors to induce
an inflammatory reaction[57]. Therefore, some lipid metabolites of
VVC may be related to inflammation, which was consistent with previous
findings[44]. LA, found in the VVC group, may act as a
pro-inflammatory factor in the response of the vaginal mucosa to C.
albicans.
IPA network analysis was performed to further explore the potential role
of metabolites in VVC, revealing that the changes in differential
metabolites were mainly related to MAPK signaling, NF-κB signaling,
Dectin-1/Syk signaling, and IL-17 signaling pathways. The study found
that C. albicans significantly activated the epidermal growth factor
receptor (EGFR) in human vaginal epithelial cells, activating the
inflammatory response through several pathways, including
mitogen-activated protein kinase (MAPK) and nuclear factor
kappa-light-chain-enhancer of activated B cells (NF-ĸB)[58, 59].
Among them, Candida mainly initiated the mucosal immunity of the vaginal
epithelium through the MAPK signaling pathway and activated MAPK
phosphatase 1 (MKP1) and c-Fos by activating MAPK-associated proteins
(p38 and ERK1/2)[60, 61]. The activation of MKP1 marked the
transition of Candida from the colonizing yeast phase to the invasive
toxin-producing hyphal phase.
Further molecular docking experiments showed that LA could bind to
ACSL1, suggesting that ACSL1 may be the target protein of LA. As a
member of the family of long-chain acyl-CoA synthetase genes, ACSL1
acted on long-chain fatty acids (FAs), shuttling FAs into mitochondria
in heart and adipose tissue for β-oxidation and lipid synthesis[62,
63]. Moreover, the potential role of ACSL1 in sepsis has been
reported[64]. One study confirmed that inhibition of ACSL1 activity
attenuated phosphorylation of p38 MAPK, ERK1/2, and NF-κB, suggesting
that ACSL1 was an upstream regulator of MAPK and NF-κB signaling
pathways to mediate inflammation[65]. LA may promote the occurrence
of VVC by binding to ACSL1 to regulate the MAPK and NF-κB signaling
pathways. Therefore, LA has the potential to become a new method for the
clinical treatment of VVC, and further research should explore its
function and mechanism. This study had certain limitations. In the
future, we will further validate cell and animal samples and refine in
vivo and in vitro experiments to confirm our findings.