NRF2 Gene Polymorphism (RS6721961) Pak Armed Forces Med J 2022; 72 (2): 398 Association of NRF2 Gene Polymorphism (RS6721961) with Semen Parameters of Infertile Males in Pakistani Population Bushra Rehman, Amena Rahim*, Muhammad Afzal*, Aqsa Tazarrat**, Humaira Kashif, Sadia Qayyum*** Foundation University Islamabad, Pakistan, *Islamic International Medical College Islamabad, Pakistan, **Mohi-Ud-Din Islamic Medical College Azad Jammu & Kashmir, Pakistan, ***Hazara University, Mansehra Pakistan ABSTRACT Objective: To determine an association of single nucleotide polymorphism of the NRF2 gene (RS6721961) with semen parameters of primary male infertility patients in the Pakistani population. Study Design: Comparative cross-sectional study. Place and Duration of Study: Department of Biochemistry, Islamic International Medical College, Rawalpindi, in collaboration with two private infertility clinics (American Infertility Center, Rawalpindi and Mother and Child International Hospital and Research Institute, Mirpur, AJK) from Oct 2020 to Sep 2021. Methodology: A total of 288 participants were included in this study. There were 144 diagnosed cases of primary male infertility and 144 healthy fertile males, age and ethnicity matched controls. Blood samples were collected from participants after obtaining written informed consent. DNA was extracted by the Chelex TM Methodoogy. Multiplex PCR was done to determine the respective allelic frequencies of NRF2 (RS6721961) genotypes using specific primers. Results: There was no significant association between NRF2 genotypes and semen volume and semn colour in infertile males (p-value 0.32 and 0.84, respectively). Out of 144 cases, 111 (77%) had normal sperm count, 29 (20.1%) had oligospermia, while 4 (2.7%) patients had azoospermia. A significant association was observed between NRF2 genotypes and sperm concentration of infertile males (p-value <0.001). Out of 144 cases, 80 (55.5%) had normal motility, 57 (39.5%) had asthenospermia while 7 (8.4%) patients had necrospermia. A strong association was observed between NRF2 genotypes and sperm motility in infertile males (p-value <0.001). Conclusion: Single nucleotide polymorphism (RS6721961) of the NRF2 gene is significantly associated with sperm concentration and sperm motility in infertile males in the studied Pakistani population. Keywords: Male infertility, Nuclear factor erythroid 2-related factor 2 (NRF2), Polymerase chain reaction, Single nucleotide polymorphism. How to Cite This Article: Rehman B, Rahim A, Afzal M, Tazarrat A, Kashif H, Qayyum S. Association of NRF2 Gene Polymorphism (RS6721961) with Semen Parameters of Infertile Males in Pakistani Population. Pak Armed Forces Med J 2022; 72(2): 398-402. DOI: https://doi.org/10.51253/pafmj.v72i2.7565 INTRODUCTION Infertility affects approximately 70 million coup- les worldwide, with male factors accounting for over half of the cases. 4 In Pakistan, the pooled prevalence of primary male infertility was 76.19%, reported in 2015. 1,2 Gene polymorphism refers to multiple forms of a single gene. 3 Single nucleotide polymorphism in a genome is a single nucleotide change that aids in determining the link between gene variations and prevalent disorders. 4,5 Male infertility mainly accounts for sperm abnormalities, and many of them are caused by genetic defects. Genetic factors are found in about 15% of infertile males, including chromosomal abnor- malities or single-gene mutations. 6 The nuclear trans- criptional protein, erythroid two related factors 2 (NRF2), induces antioxidant enzymes via the signal- ling pathway of the antioxidant response element (ARE). 7 NRF2 protects the male reproductive tract from oxidative stress by playing a vital role in cellular antioxidant defence during spermatogenesis and fer- tilisation. - Numerous genes are involved in normal spermatogenesis and sperm functions. Oxidative stress (OS) is a significant contributor to a decline in male fertility by producing irreparable damage to DNA, lipids, proteins and enzymatic systems, ultimately lea- ding to cell death and a reduction in semen parameters linked to male infertility. 9 Many studies have reported that impairment of the NRF2-ARE signalling pathway and functional polymorphisms of the NRF2 promoter region is associated with many human diseases, including male infertility. 10 There is limited information and statistical data available regarding the genetic basis of infertility in Pakistani males. Hence, it is important to carry out genetic studies for male infertility to decrease the This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by-nc/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Correspondence: Dr Bushra Rehman, Lecturer Biochemistry, Foundation University, Islamabad Pakistan Received: 25 Aug 2021; revision received: 10 Apr 2022; accepted: 19 Apr 2022 mzahid689@gmail.com Original Article Open Access