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Table of Contents
ORIGINAL ARTICLE
Year : 2015  |  Volume : 2  |  Issue : 2  |  Page : 89-91

Evaluating the mRNA expression profile of APC in Pterygium


1 PhD Candidate, Departement of Biology, University of Sistan and Baluchestan, Zahedan, Iran
2 Associate Professor, Departement of Biology, University of Sistan and Baluchestan, Zahedan, Iran
3 Professor, Department of Ophthalmology, Al-Zahra Eye Hospital, Zahedan University of Medical Sciences, Zahedan, Iran

Date of Web Publication5-Jul-2017

Correspondence Address:
Maryam Najafi
Department of Biology, University of Sistan and Baluchestan, P. O. Box 98155.987, Zahedan
Iran
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Source of Support: None, Conflict of Interest: None


DOI: 10.5530/ami.2015.3.8

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  Abstract 


Introduction: The exact etiology of pterygium is unknown. Most of population based studies implicating that pterygium has a benign behavior rather than destructive condition. Changed expression of cell cycle-related genes may lead to the abnormal cell proliferation and finally to malignancy phenotype. The aim of this study was evaluation the expression profiles of APC in pterygium. Materials and Methods: The RNA extracted from the 23 pterygium tissues and 18 healthy tissues. After converting RNA to cDNA. The expression levels of these genes were assessed by real-time PCR. Results: The relative expression of APC gene in pterygium tissues was significantly different in comparison to conjunctiva tissues of healthy controls (mean ± SD was 1.82 ± 0.15 for cases versus 1.70  0.12 for controls, P value = 0.048). Conclusion: Detection of expression changes, leading to find molecular mechanism underlying the disease. As well as exploring pterygium markers paving the road for better therapy. We would like to propose further studies to identify exact molecular function of this gene in pterygium by using advanced molecular techniques such as RNAseq in various and larger genetic populations.

Keywords: APC, Gene, Expression, Pterygium


How to cite this article:
Najafi M, Kordi-Tamandani DM, Arish M. Evaluating the mRNA expression profile of APC in Pterygium. Acta Med Int 2015;2:89-91

How to cite this URL:
Najafi M, Kordi-Tamandani DM, Arish M. Evaluating the mRNA expression profile of APC in Pterygium. Acta Med Int [serial online] 2015 [cited 2019 Aug 17];2:89-91. Available from: http://www.actamedicainternational.com/text.asp?2015/2/2/89/209659




  Introduction Top


Pterygium is an external ocular mass that usually forms on the perilimbal conjunctiva and extends onto the corneal surface with prevalence rate between 0.7% and 33% globally.[1] Pterygia can differ from small, atrophic quiescent lesions to large, aggressive, rapidly growing fibrovascular lesions that can deform the corneal topography, and, in advanced cases, cover the optical center of the cornea.[2] [Figure 1] Traditionally regarded as a degenerative condition, but also display tumor-like features, such as a propensity to invade normal tissue and high recurrence rates following resection.[3] Pterygium are reported to occur in males twice as frequently as in females, as well as patients older than 40 years have the highest prevalence of pterygia.[4] Scientists demonstrate the role of different factors, such as viruses, oxidative stress, DNA methylation, apoptotic and oncogenic proteins, loss of heterozygosity, microsatellite instability, inflammatory mediators, extracellular matrix modulators, lymphangiogenesis, cell epithelial-mesenchymal transition, and alterations in cholesterol metabolism in pterygium development.[5] Internationally, there is a relationship between increased incidence of the pterygium and exposure to UV light.[6] One of the repair systems that is activated after ongoing UV exposure is base exision repair.[7] There is accumulating evidence supporting a role for the tumor suppressor APC (Adenomatous Polyposis Coli) which is located on the 5q21-q22 chromosome that blocks DNA polymeraseβ-dependent strand displacement synthesis during long patch base excision repair.[8] As well as over expression of APC leads to G1 cell cycle arrest by repressing transcription of cyclin D1 through Wnt signaling.[9] APC is transiently hyper-phosphorylated in the M phase of the cell cycle and associates with the kinetochore in dividing cells, so plays outstanding function in genomic stability and proper chromosome segregation.[10],[11],[12] APC induces G2/M cell cycle arrest in collaboration with topoisomerase IIα (topo IIα).[13] Another important role for APC is assigned in actin cytoskeletal integrity, cell-cell adhesion and cell migration.
Figure 1: Clinical photograph of a pterygium lesion on the ocular surface

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However, to date, little is known about the expression changes of APC in the pterygium. The aim of this study is the evaluation mRNA expression profile of APC in pterygium.


  Materials and Methods Top


Subject

This study was performed from 2010 to 2013 consisting of 23 primary pterygium tissues (45 males and 34 females) from patients with mean age of 52.44 + 20.611 and 18 normal conjunctiva tissues (41 males and 33 females) from healthy controls (a lembal conjunctiva of the patients who had gone under cataract surgery), with a mean age of 50.67 ± 23.318. These samples have collected from Alzahra eye hospital. All of the tissues were nasal, unilateral, inactive and primary. The biopsy tissue samples were frozen in -80°C until molecular analysis. All procedures in this study were approved by the Ethical Board at the Zahedan University of Medical Sciences. Informed consent was taken from all participants. Demographic table was described by Eye and Contact Lens journal previously [Table 1].
Table 1: The demographic characteristics of the case-control group

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Analysis of mRNA Expression

We extracted total RNA from 23 pterygium and 18 conjunctiva tissues using the RNX™- Plus solution (Cat No: MR7713C). The Revert Aid First Strand cDNA Synthesis Kit (Fermentas, Cat. no. K1621) was used to reverse-transcribe 1 mg of RNA in a final volume of 20 μL. An AB15700 sequence detection system (Applied Biosystems) was used to estimate the expression task.The normalization of data have been done by RNA18s as an internal standard. The sequence of primers was listed in [Table 2].
Table 2: Expression primer sequences and annealing temperatures

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Statistical Analysis

All statistical analyses have been done with SPSS version 20.0 (SPSS, Chicago, IL). Expression data have been assessed by compression of CT target/Ct housekeeping using the independent samples T- test between groups (healthy subjects and patients). The significance level was set at P ≤ 0. 05.


  Results and Discussion Top


The outcomes of expression analyses exposed a statistically significant variation between cases and healthy controls concerning the relative expression of APC (P value = 0.048). Increased expression in case group of candidate gene was detected. (Mean ± SD was 1.82 ± 0.15 for cases versus 1.70 ± 0.12 for controls) [Table 3].
Table 3: Comparison of relative gene expression APC gene between patients with pterygium and healthy controls

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As previously mentioned above the exact etiology of pterygium is yet unknown, but some characterizations of pterygium, such as angiogenesis, tissue structure and frequent recurrence, lead to compare its molecular mechanisms with a cancerous cell. APC is a multi-functional tumor suppressor that plays roles in several fundamental cellular processes. These include cell adhesion and migration, organization of the actin and microtubule networks, spindle formation and chromosome segregation.[14] Multi-tasking of APC might explain why its loss often leads to cancer.[15] The vast review literatures reported the expression level of cell cycle related genes in pterygium. In the line of our research, Lafontaine PO et al 2015, suggesting that increased expression of the trefoil protein might exert protective and beneficial roles during the pathogenesis of pterygia.[16] Xu YX et al 2014, reported the highly expressed of survivin in all pterygium tissues.[16] Detorakis et al 2010, have indicated that higher levels of FGF2 or VEGFA mRNA in pterygium may be involved in the pathogenesis or clinical behavior of the pterygium, including postoperative recurrence.[17] Tetsushi Nakagami et al 2000, found that stem cell factor is overexpressed in fibroblasts at the cap area of most pterygia by immune-histochemistry.[18] Di Girolamo N et al 2000, for the first time documented the increased cellular expression of MMPs and TIMP in pterygium.[19] Adiguzel U et al 2007, specified the COX-2 expression is increased in recurrent pterygium tissues and COX-2 expression may be a marker for the prediction of recurrence.[20] Yang SF et al 2009, have implicated the significantly increased expression of MMP-2 and MMP-9 expression by pterygium fibroblasts.[21] But on the other side, Schelline SA 2006, demonstrated that Matrix metalloproteinase expression showed no difference in normal Tenon's capsule and in primary or recurrent pterygia.[22] To the best our knowledge, analysis the expression of APC was done for the first time in this study. Detection of expression changes, leading to finding molecular mechanism underlying the disease. As well as exploring pterygium markers paving the road for better therapy. We would like to propose further studies to identify exact molecular function of APC gene in pterygium by using advanced molecular techniques such as RNA seq and HRM in various and larger genetic populations.


  Acknowledgments Top


We would like to express our gratitude to the department of Ophthalmology, Al-Zahra Eye Hospital, Zahedan University of Medical Sciences, and the department of Biology, University of Sistan and Baluchestan, for supporting this project financially.

Compliance with ethical standards


  Conflicts of Interest Top


The authors declare that they have no conflict of interest.

Research involving Human Participants

All procedures performed in studies involving human participants was in accordance with the ethical standards.

Informed Consent

Informed consent was obtained from all individual participants included in the study.



 
  References Top

1.
Jiao, W., et al., Prevalence and risk factors for pterygium in rural older adults in Shandong Province of China: a cross-sectional study. Biomed Res Int, 2014;pp 658648.  Back to cited text no. 1
    
2.
Zheng, K., et al., Comparison of pterygium recurrence rates after limbal conjunctival autograft transplantation and other techniques: meta-analysis. Cornea, 2012; 31(12): p. 1422–7.  Back to cited text no. 2
    
3.
Chui, J., et al., Ophthalmic pterygium: a stem cell disorder with premalignant features. Am J Pathol, 2011; 178(2): 817–27.  Back to cited text no. 3
    
4.
Tin, S.S. and V. Wiwanitkti, Risk factors for pterygium in the high-altitude communities. Nepal J Ophthalmol, 2014; 6 (12): pp 244.  Back to cited text no. 4
    
5.
Cardenas-Cantu, E., et al., Molecular Basis of Pterygium Development. Semin Ophthalmol, 2014; 1–17.  Back to cited text no. 5
    
6.
Achigbu, E. and U.F. Ezepue, Prevalence and Severity of Pterygium among Commercial Motorcycle Riders in South Eastern Nigeria. Ghana Med J, 2014;48(3): 153–7.  Back to cited text no. 6
    
7.
Rastogi, R.P., et al., Molecular Mechanisms of Ultraviolet Radiation-Induced DNA Damage and Repair. Journal of Nucleic Acids, 2010.; 592980.  Back to cited text no. 7
    
8.
Jaiswal, A.S., et al., Mechanism of adenomatous polyposis coli (APC)-mediated blockage of long-patch base excision repair. Biochemistry, 2006; 45(51): 15903–14.  Back to cited text no. 8
    
9.
Ishidate, T., et al., The APC-hDLG complex negatively regulates cell cycle progression from the G0/G1 to S phase. Oncogene, 2000; 19(3): p. 365–72.  Back to cited text no. 9
    
10.
Bhattacharjee, R.N., et al., The tumor suppressor gene product APC is hyperphosphorylated during the M phase. Biochem Biophys Res Commun, 1996; 220(1):. 192–5.  Back to cited text no. 10
    
11.
Fodde, R., et al., Mutations in the APC tumour suppressor gene cause chromosomal instability. Nat Cell Biol, 2001;3(4): 433–8.  Back to cited text no. 11
    
12.
Kaplan, K.B., et al., A role for the Adenomatous Polyposis Coli protein in chromosome segregation. Nat Cell Biol, 2001; 3(4): 429–32.  Back to cited text no. 12
    
13.
Wang, Y., et al., Interaction between Tumor Suppressor Adenomatous Polyposis Coli and Topoisomerase IIa: Implication for the G2/M Transition. Molecular Biology of the Cell, 2008;19(10): 4076–4085.  Back to cited text no. 13
    
14.
Aoki, K. and M.M. Taketo, Adenomatous polyposis coli (APC): a multi-functional tumor suppressor gene. J Cell Sci, 2007; 120(19): 3327–35.  Back to cited text no. 14
    
15.
Bienz, M., The subcellular destinations of APC proteins. Nat Rev Mol Cell Biol, 2002. 3(5): 328–38.  Back to cited text no. 15
    
16.
Xu, Y.X., et al., Differential expression and function of survivin during the progress of pterygium. Invest Ophthalmol Vis Sci, 2014; 55(12): 8480–7.  Back to cited text no. 16
    
17.
Detorakis, E.T., A. Zaravinos, and D.A. Spandidos, Growth factor expression in ophthalmic pterygia and normal conjunctiva. Int J Mol Med, 2010; 25(4): 513–6.  Back to cited text no. 17
    
18.
Nakagami, T., et al., Expression of stem cell factor in pterygium. Jpn J Ophthalmol, 2000; 44(3): 193–7.  Back to cited text no. 18
    
19.
Di Girolamo, N., et al., Expression of MMPs and TIMPs in human pterygia and cultured pterygium epithelial cells. Invest Ophthalmol Vis Sci, 2000; 41(3): 671–9.  Back to cited text no. 19
    
20.
Adiguzel, U., et al., Cyclooxygenase-2 expression in primary and recurrent pterygium. Eur J Ophthalmol, 2007; 17(6): 879–84.  Back to cited text no. 20
    
21.
Yang, S.F., et al., Increased expression of gelatinase (MMP-2 and MMP-9) in pterygia and pterygium fibroblasts with disease progression and activation of protein kinase C. Invest Ophthalmol Vis Sci, 2009; 50(10): 4588–96.  Back to cited text no. 21
    
22.
Schellini, S.A., et al., Matrix metalloproteinase-9 expression in pterygium. Arq Bras Oftalmol, 2006; 69(2): 161–4.  Back to cited text no. 22
    


    Figures

  [Figure 1]
 
 
    Tables

  [Table 1], [Table 2], [Table 3]



 

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