Identification of a novel interplaying loop of PPARγ and respective lncRNAs are involved in colorectal cancer progress

https://doi.org/10.1016/j.ijbiomac.2022.07.247Get rights and content

Abstract

Long noncoding RNAs (lncRNAs) as regulatory molecules play important roles in early treatment and diagnosis of cancers. Considering the role of PPARγ in colorectal cancer (CRC) as a tumor suppressor, the GEO database was used to identify candidate genes that affect the activation of PPARγ protein in CRC cell lines. Then were selected 5 genes containing PPARγ response element (PPRE) in up to 4000 bp upstream and were affected by PPARγ protein activation in HT-29 colon cancer cell line using UCSC database. Expression meta-analysis was applied to map the expression network between candidate genes and all known lncRNAs through expression correlation and lncRNAs that correlated with a greater number of candidate genes (R > 0.5, P.value < 0.001). Moreover, were selected 3 lncRNAs as lncRNAs affected by PPARγ protein activation. Next, the expression levels of candidate genes and lncRNAs were evaluated using RT-qPCR in HT-29 cell line. Results showed a significant increase (FDR <0.05) in the expression level of 5 candidate genes and lncRNAs LINC01133, MBNL1-AS, LOC100288911 after treatment with pioglitazone as PPARγ ligand compared to the untreated group in HT-29 cells. Although additional tests are needed to confirm bioinformatics predictions, it can be concluded that increased expression of PPARγ may increase genes and lncRNAs expression. In summary, this study could be suggested identifying lncRNAs affected by PPARγ activation could be a new strategy in understanding the function and activity of PPARγ in colon cancer.

Introduction

Colorectal cancer (CRC) is one of the most deadly malignant cancers in the world [1], accounting for >50 % of all cancers in developed countries, with a high mortality rate per year [2], [3]. The 5-year survival rate for people with CRC in the early stages usually has better results [4]. In addition, the highest metastatic CRC has been observed in patients under 50 years of age [5], [6]. Despite much progress in understanding the molecular mechanisms and treatment of CRC, the overall survival rate of patients with CRC has not improved significantly [7], [8].

In these days, targeted treatment based on tumor suppressor genes could to establish a new strategy in the treatment of cancers, including CRC [7]. Data from advanced studies have shown a reduction in the expression level of many tumor suppressor genes. Meanwhile, PPARγ plays an important role in the normalization of normal tissue hemostasis and the development of CRC pathology [9], [10], indicating that this receptor can be suitable as a drug target [11]. Most of the available data suggest that PPARγ has an anti-tumor effect in CRC [12], and its activation is associated with inhibition of cell growth in the laboratory and reduction of tumorigenesis in 90 x-ray mice [13]. Decreased or eliminated expression of PPARγ is associated with an increase in tumor formation in the small and large intestines and increases the tendency for CRC to progress [14].

Acquired information from experimental studies has revealed that PPARγ is one of the main tumor inhibitor genes in CRC. By activating PPARγ through its ligands like pioglitazone which is the most well-known synthetic ligand. PPARγ can affect genes that In CRC, they are reduced in expression and have tumor inhibitory activity [15]. Activation of these genes by activating PPARγ with its ligand is important because it results in the reduction of malignancy in CRC. Treatment of gene-based CRC with current methods may be associated with nonspecific effects as well as side effects [14].

Molecular biology of human tumors, in a series of studies, has highlighted the role of non-coding proteins RNAs, especially lncRNAs, in a variety of cancers [16]. These lncRNAs can be involved in tumorigenesis and important pathways in cancer that are associated with patient survival [17], [18]. However, the association between lncRNAs and PPARs in the progression of CRC has been less studied.

It seems that the study of PPARγ-related lncRNAs can help us to understand the function of this protein and identify tumor-inhibiting lncRNAs [19]. PPARγ plays a significant act as a nuclear transcription factor in CRC, so the target genes for this transcription factor may be valuable. We used microarray data as well as The Cancer Genome Atlas (TCGA) data to identify PPARγ-affected genes and lncRNAs using expression networks and responsive sequences. Also, the expression of desired genes was evaluated in CRC cell line under agonist treatment as in vitro study.

Section snippets

TCGA and GEO data analysis

The selected microarray data was downloaded from the GEO database in raw format, and the initial preprocessing of the data was performed, including the removal of backlight normalization and logarithmic data with base 2 using the AFFY and limma packages. The resulting expression matrix was used for gene set enrichment analysis (GSEA). RNASeq data was downloaded from the TCGA database in raw format (HTseq-Counts) using the TCGAbiolinks package. These data included 480 tumor and 41 normal

Identification of PPARγ target genes in CRC

Induction of cell cycle arrest, apoptosis and cell differentiation, as well as inhibition of angiogenesis, are all possible mechanisms to explain the antitumor activity of PPARγ in colon cancer. Therefore, PPARγ can affect the expression of many genes. In this regard, the expression of genes and lncRNAs affected by this protein were evaluated. For this purpose, a study with GSE40706 was selected using the GEO database. In this study, after treating T84 cells with rosiglitazone, the expression

Discussion

Regardless of great advances in understanding the molecular mechanisms and treatment of CRC during the last years, the overall survival of patients with CRC has not improved significantly [20], [21]. It has been revealed that a variety of factors and sophisticated processes are involved in this disease. Mutations in PPAR genes, especially PPARγ are among the genetic factors that increase the risk of this type of cancer [14]. Loss of function mutations in PPARγ is found in approximately 10 % of

Conclusion

Studies indicate that PPARγ plays a role in colon cancer malignancy, on the other hand, PPARγ can not only affect protein-coding genes, but also non-protein-coding genes, including lncRNAs. also affect its activation, which until now, the identification of lncRNAs under the influence of PPARγ has remained unknown. lncRNAs have been proposed in molecular biology today as molecules with diverse functions, play a role in transcriptional settings, post-transcriptional settings, translational

Funding

No funding was received.

Availability of data and materials

Data sharing not applicable to this article as no data-sets were generated or analyzed during the current study.

Ethics approval and consent to participate

We obtained the signed informed consent from all patients. This study was approved by the Biomedical Ethics Committee of ROYAN institute and also complies with the Ethics Code of IR.ACECR.ROYAN.REC.1398.089.

Consent for publication

Not applicable.

CRediT authorship contribution statement

The design and conceptualization of study and methodology were done by M.H., M.E., and M.M. Data mining, formal analysis and investigation were performed by M.H., and Kh.M. Supervision, validation and visualization were done by M.P., M.N. and K.G. Interpretation of the obtained information was done by M.P., M.M., and M.E. The manuscript was written by M.H. and reviewed, edited and approved by M.P., K.G, and M.N. All authors read and approved the final manuscript.

Declaration of competing interest

All of the authors have declared that no competing interests exist.

Acknowledgements

We thank our colleagues for their association and helpful discussions in this study.

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