PCOS is a heterogeneous disorder characterized by hyperandrogenemia, ovulatory dysfunction, and it is a common reproductive and endocrine disorder that mainly occurs in puberty and childbearing age, affecting 6%–20% of women (Rotterdam, 2004; Azziz et al., 2009; Rocha et al., 2019). Furthermore, PCOS is associated with multiple metabolic disorders, including insulin resistance (IR), obesity, dyslipidemia, type 2 diabetes (T2DM), and cardiovascular disease (Goyal and Dawood, 2017). PCOS is the most common cause of anovulatory infertility, and its high incidence has brought huge health and economic costs to the family and society (Homburg and obstetrics, 2004).

NAFLD as one of the most prevalent chronic liver diseases globally is a type of metabolic-related liver disease closely related to IR and genetic susceptibility that includes non-alcoholic simple hepatic steatosis (NAFL), non-alcoholic steatosis Hepatitis (NASH), cirrhosis and hepatocellular carcinoma (HCC) (Diehl and Day, 2017). NAFLD is commonly recognized as a hepatic manifestation of the metabolic syndrome (MS), and it is frequently accompanied by T2DM, obesity, and dyslipidemia (Wattacheril, 2020). According to statistics, the global prevalence of NAFLD was estimated to be 25% (Younossi et al., 2016). Therefore, NAFLD has become a health problem and an economic burden in the world.

In 2005, Brown et al. first found a potential relationship between PCOS and NAFLD (Brown et al., 2005). To date, there are more and more that studies have substantiated a significant link between NAFLD and PCOS (Kelley et al., 2014). Some reports have suggested that 68 the prevalence of NAFLD in women with PCOS is significantly higher than that in 69 non-PCOS women (Falzarano et al., 2022). PCOS patients can promote the occurrence and development of NAFLD through various factors such as hyperandrogenism, IR, obesity, dyslipidemia, chronic low-grade inflammation, and intestinal flora imbalance (Vassilatou, 2014; Boursier et al., 2016; Kim et al., 2017; Yurtda ş and Akdevelioğlu, 2020). Although the relationship between NAFLD and PCOS has attracted substantial interest recently, leading to a vast amount of research related to this topic, the related genetics research is still limited and needs further exploration. There is a need for further studies to investigate NAFLD in the context of PCOS, as well as potential therapeutic options.

In recent years, microarray technology and bioinformatic analysis have been widely used to screen genetic alterations at the genome level. In the present study, we for the first time analyzed two original microarray datasets that were downloaded from Gene Expression Omnibus (GEO, http://www.ncbi.nlm.nih.gov/geo) to obtain DEGs between NAFLD and PCOS. Subsequently, the identified DEGs were analyzed by Gene Ontology (GO) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis and protein-protein interaction (PPI) analysis. Then, we predicted the potential target miRNAs of DEGs, and constructed a visualized miRNA-gene interaction network through Cytoscape software (http://www.cytoscape.org/). The present study aimed to identify hub genes and hot research topic.

Nowadays, more and more studies have confirmed the correlation between PCOS and NAFLD. A previous study showed that the prevalence of NAFLD was significantly higher in women with PCOS than in those without, with a prevalence of 51.56 and 29.64%, respectively (Falzarano et al., 2022). Meanwhile, a study revealed that PCOS was also more prevalent in female NAFLD patients with a prevalence of 50% up to a high of 70%, which is significantly higher than in those without NAFLD (Georgescu, 2022). Furthermore, studies have shown that both PCOS patients and NAFLD patients can increase the risk of cardiovascular disease (Dokras, 2019), and metabolic abnormalities were more pronounced in the PCOS patients with concurring NAFLD (Sarkar et al., 2020). In addition, cardiovascular disease is the most common cause of death in NAFLD patients (Söderberg et al., 2010). So far, the mechanism of the link between NAFLD and PCOS is not fully understood. Therefore, exploring the molecular mechanisms between PCOS and NAFLD, early identification and intervention, undoubtedly have important clinical significance.

In the present study, a series of bioinformatics analyses were performed on two independent gene chip databases of PCOS and NAFLD, and 52 common DEGs between PCOS and NAFLD were obtained based on the GEO database. The result of GO enrichment analysis indicated that the DEGs were mainly enriched in immune response, neutrophil activation, neutrophil mediated immunity, and leukocyte migration. This result suggests that the DEGs between PCOS and PCOS are related to inflammation and immune response, which is consistent with previous studies (Moulana, 2019; Oates et al., 2019). In addition, KEGG analysis showed that the DEGs were mainly enriched in Staphylococcus aureus infection, TNF signaling pathway, Tuberculosis, Osteoclast differentiation, and Fluid shear stress and atherosclerosis. We further classified these pathways according to the KEGG pathway Database. We found that they are mainly related to immunity and inflammation. Many studies have shown that immune inflammatory processes and atherosclerosis play a critical role in cardiovascular diseases (Zhang et al., 2020). Moreover, PCOS and NAFLD are strongly associated with cardiovascular diseases (Scicchitano et al., 2012; Niederseer et al., 2021). Our findings provide new ideas that patients with PCOS and NAFLD are more prone to cardiovascular disease.

Subsequently, according to the MCODE plug-in and cytoHubba plug-in of Cytoscape, we screened nine overlapping DEGs as hub genes in the PPI network, including TREM1, S100A9, FPR1, NCF2, FCER1G, CCR1, S100A12, MMP9, and IL1RN. These nine genes were all upregulated in both PCOS patients and NAFLD patients, suggesting these genes may play important role in the mechanism of PCOS and NAFLD.

S100A9 and S100A12 belong to a family of 25 homologous low‐molecular‐weight intracellular calcium‐binding proteins produced by cells of myeloid origin (Donato and biology, 2001). S100A9 is mainly released by activated granulocytes and associated with various types of inflammation-related pathways (Wang et al., 2018). A previous study showed that S100A9 could increase the production of inflammatory cytokines and disturb steroidogenesis via activating nuclear factor kappa B (NF‐κB) signaling pathway in PCOS (Li et al., 2020). Furthermore, S100A9 promotes inflammation and lipolysis in the liver during NAFLD progression (Wu et al., 2020). Another study demonstrated that S100A9 can differentiate hepatic and metabolic progression in NAFLD as a biomarker in NAFLD progression (Liu et al., 2015). Additionally, studies have shown that S100A12 interacts with S100A9 (Hatakeyama et al., 2004) and also has proinflammatory activity (Miranda et al., 2001). There are no reports of S100A12 in PCOS and NAFLD. We speculate that S100A9 and S100A12 synergistically promote inflammation in PCOS and NAFLD. Interestingly, S100A12 has also been shown to block the activity of S100A9 (Katte and Yu, 2018).

NCF2 is a rate-limiting cofactor of NADPH oxidase that is necessary for reactive oxygen species (ROS) production in phagocytes, which plays a critical role in innate immunity and phagocytic microbicidal activity (Muise et al., 2012). NCF2 is highly expressed in PCOS and NAFLD and contributes to oxidative stress in PCOS and NAFLD (Kaur et al., 2012; Garcia-Jaramillo et al., 2019).

IL1RN is originally discovered as a natural antagonist of IL-1 (Arend, 1991), which encodes the IL1 antagonist protein (IL1RA) (Gabay et al., 2010). And IL1RA has been reported as a critical mediator of inflammatory processes and plays a vital role in the pathogenesis of PCOS (Xia et al., 2013). IL1RN also plays an important role in the development of NAFLD (Wolfs et al., 2015).

MMP9 is one of the zinc-ion-dependent metalloproteinase family, playing a pathogenic role in chronic inflammatory autoimmune diseases (Akter et al., 2015). MMP9 was shown to be overexpressed in PCOS and involved in the pathogenesis of PCOS (Ranjbaran et al., 2016). MMP9 could increase fibrosis severity in NAFLD and be involved in the development and progression of human HCC metastasis (Shi et al., 2010; Goyale et al., 2021). TREM1 is expressed on myeloid lineage cells and played an important role in maintaining homeostasis and the normal immune responses. Dou et al. demonstrated that TREM1 in the liver was involved in the pathological process and therapeutic outcome of NASH (Dou et al., 2016). In addition, TREM1 promotes atherosclerosis by inducing monocytosis and foam cell formation (Zysset et al., 2016). FPR1 is a member of 7-transmembrane G protein-coupled receptors (GPCRs), which is mostly expressed by myeloid cells and is a key regulator of the inflammatory environment (Dahlgren et al., 2016; Mastromarino et al., 2019). FCER1G is engaged in a variety of immune responses as a constitutive component of the high-affinity immunoglobulin E (Ig E) receptor and interleukin-3 receptor complex (Liang et al., 2013). CCR1 is a member of the seven-transmembrane G-protein-coupled receptor family, which is widely expressed on myeloid cells and is involved in the activation and trafficking of immune cells (Schaller et al., 2008). Based on previous research, these nine hub genes have been shown to have key effects on immune response and inflammatory response. Moreover, it has been known that immune dysregulation and chronic inflammation are important factors in the development of PCOS and NAFLD (Anstee et al., 2013; Qin et al., 2016). Based on this, we speculate that: these genes may also contribute to the occurrence and development of PCOS and NAFLD, and may be potentially new treatment targets in the future.

MiRNAs are small non-coding RNA molecules (21–25 nucleotides long) that are partially or fully complementary to the 3′ URT of the target gene mRNA by inducing mRNA degradation or repressing mRNA translation (He and Hannon, 2004). In this study, we also constructed a miRNA-target gene network and selected the four miRNAs (miR-20a-5p, miR-129-2-3p, miR-124-3p, and miR-101-3p) that interacted the most with DEGs. miR-20a-5p was found to bind to pro-angiogenic genes and suppress their expression and thus exerting anti-angiogenic activity in PCOS (Patil et al., 2022). miR-20a-5p also played a protective role in lipid metabolism and IR of NAFLD (Wang et al., 2020; Zhang et al., 2021). Previous studies on miR‐129-2-3p mainly focused on cancer. miR-129-2-3p has not been reported in PCOS and NAFLD. A study has shown that miR-129-2-3p was involved in the functional regulation of neutrophils in chronic inflammatory processes. Thus, we speculate that miR-129-2-3 may be involved in the pathophysiology of PCOS through inflammatory processes. miR-124-3p was downregulated in PCOS patients (Ding et al., 2015) and was involved in lipid metabolism in NAFLD as a critical regulator in lipid homeostasis of the liver (Wang et al., 2020918). miR-101-3p was also downregulated in PCOS patients (Butler et al., 2020). Moreover, the downregulation of miR-101-3p was more strongly associated with hepatic injury in NAFLD (Meroni et al., 2019).

In conclusion, our study identified some hub genes and speculated that miRNA-gene regulatory networks contribute to the pathophysiological process of PCOS and NAFLD by bioinformatics analysis, which provided the potential diagnostic and therapeutic targets of PCOS and NAFLD. However, there are some limitations to our study. Firstly, only the top nine hub genes were involved in our current study. Secondly, a lack of research on detailed molecular mechanisms that the hub genes and miRNAs regulate in PCOS and NAFLD. Thirdly, the miRNA-gene interaction networks were only based on predictions from public databases. Thus, the molecular mechanism of these hub genes and miRNAs in the occurrence of PCOS and NAFLD need to be further studied.

Our study identified a miRNA-gene network potentially relevant for PCOS and NAFLD. The nine hub genes (including TREM1, S100A9, FPR1, NCF2, FCER1G, CCR1, S100A12, MMP9, and IL1RN) were significantly upregulated, which may have a critical influence on the pathophysiological mechanism of PCOS and NAFLD. Some potential target miRNAs (miR-20a-5p, miR-129-2-3p, miR-124-3p, and miR-101-3p) were also predicted and may participate in the pathophysiological process of PCOS and NAFLD through inflammation and immune response. These findings may contribute to the development of early diagnostic strategies, prognostic markers, and therapeutic targets.