Introduction
Premature ovarian failure (POF) refers to the failure in ovarian function in women younger than 40 years in women, which accounts for about one in every 100 women younger than 40 years and one in every 1000 women younger than 40 years, according to which it is one of the causes of infertility in women [1, 2]. The clinical manifestations and etiology of POF are very heterogeneous and include primary or secondary amenorrhea for more than 4 months, incidence of hypogonadotropic hypogonadism for at least one month including menopause with the elevated levels of follicular stimulating hormone (FSH) in the serum (more than 40 mg/ml) and decreased levels of both estradiol and antimutrin hormone (AMH) in the patient's serum [2]. On the other hand, POF causes about 10 to 28% of primary amenorrhea and 4 to 18% of secondary amenorrhea. Due to the high heterogeneity of POF, several mechanisms are involved. In the embryonic period, the growing ovaries contain about 7 million primordial follicles, most of which are rapidly destroyed by the apoptotic mechanism. Only some germ cells continue to divide and a few enter the meiotic division and create primordial oocytes. Of these, half a million oocytes do not ovulate, and only about 400 oocytes are ovulated before physiological menopause. Several mechanisms are involved in the occurrence of POF, such as the presence of fewer primordial follicles, increased follicular atresia, and altered maturation of primordial follicles [3]. Many factors can be associated with the POF mechanisms, including genetic, immunological, metabolic factors and infection [4]. In addition, some POF cases are syndromic, such as turner syndrome. However, between 50% and 90% of cases are idiopathic, and genetic factors are possibly involved, as well.
The importance of genetics in the etiology of POF has shown, with about 10–20% of idiopathic POF cases also having first-degree relatives or women with affected mothers are six times more likely to develop POF [5-7]. About 70% of idiopathic cases are chromosomally normal and only about 10% of them have chromosomal abnormalities, most of which are due to X chromosome abnormalities and a smaller percentage have abnormalities in autosomal chromosomes. By comparing cytogenetic findings with disease manifestations, in 50% of adult primary amenorrhea and 13% of secondary amenorrhea in cases aged 30 years or younger, chromosomal abnormalities have been reported. The chromosomal abnormalities involved in POF include partial or complete deletions, duplications, X chromosome translocation, monosomy X, and triple X (XXX, 47) [5, 8, 9]. In addition, on the X chromosome, two critical regions involved in POF (Xq13.3-q21.1 and Xq21.3-q27) have been identified [3, 5]. The most common genetic finding in POF is the overlap in the FMR1 gene located in the Xq27.3 region. Complete mutation in the FMR1 gene is the cause of fragile X syndrome and is a common cause of mental retardation in boys [4, 10]. Moreover, other genes are also involved to develop POF. The aim of this study was to report chromosomal abnormalities, with X chromosome involvement, in two patients with POF, referred to Sarem Hospital in Tehran.
Patients and methods
Two patients diagnosed with POF were referred to the cytogenetic laboratory of Sarem Hospital in 2014-2015. According to the guidelines of the European Society of Reproduction and Human Embryology (ESHRE), the first step in the diagnosis of POF patients is chromosomal analysis using standard high resolution GTG-banding (Table 1) [11].
Table 1. Premature ovarian failure diagnostic guideline according to the European Society of Reproduction and Human Embryology (ESHRE) protocol [11]

 
Therefore, in both cases, chromosomal analysis of peripheral blood was performed. In this study, at least 20 metaphases with a band resolution between 450-band and 550-band resolution were analyzed. C-bonding was also evaluated in at least 10 metaphases if needed. The first case was a 16-year-old girl with diagnosed with POF and primary amenorrhea. Her chromosomal analysis revealed a deletion in most of the short arm of the X chromosome and her karyotype was 46, X, der (X) (Fig. 1).
Figure 1. 46, X, der (X)
The second patient was a 26-year-old female diagnosed with POF and secondary amenorrhea. She was found with 45, dic (X; 22) (q22; p12) karyotype. A portion of the long arm of the X chromosome was deleted at the q22 region to the end of the chromosome and replaced by chromosome 22. Therefore, a dysenteric chromosome was formed (Fig. 2).
Figure 2. 45, X, dic (X; 22) (q22; q12)
 
Discussion
POF is one of the causes of infertility worldwide and the first step in its genetic diagnosis is the high resolution chromosomes analysis, especially the X chromosome. The two studied cases were the diagnosed with POF had abnormalities on the X chromosome. The first case was a girl diagnosed with POF and primary amenorrhea with karyotype 46, X, der (X) and a large deletion of the short arm of the X chromosome from p11.2 to the telomeric end. Studies have shown that deletion in the Xp11.2 region accounts for 50% of primary amenorrhea and 45% of secondary amenorrhea and the studied case also had primary amenorrhea. In a study, 12 out of 27 people with deletion in Xp11.2, spontaneous menopause was reported. One of the important genes in the deleted region is SHOX, which lack of a copy of SHOX can result in short stature as Turner syndrome causes [13]. In two other studies, however, two cases with deletion in the same region had normal height and fertility [14].
The second case was a woman with POF and secondary amenorrhea reported with karyotype 45, dic (X; 22) (q22; p12). As previously mentioned, two critical regions on the X chromosome are associated with POF, Xq13.3-q21.1 and Xq21.3-q27. This patient also showed a deletion in the critical region of Xq21.3-q27, which was replaced by chromosome 22. Studies have shown that deletions are more common in the critical region of Xq21.3-q27, which was also observed in this patient [15]. There are many genes in the deleted region. The genes in the deleted region associated with POF are listed in Table 2, which can all explain the cause of POF in the patient [16].
Table 2. List of genes associated with premature ovarian failure in the Xq21 deleted region [16]

 
Conclusion
EOF is an important cause of infertility in couples, and chromosomal abnormalities are one of the most common genetic causes in EOF. Patients diagnosed with POF should preferably be subjected to precise chromosomal analysis according to ESHRE guidelines considering X chromosome and in the absence of chromosomal abnormalities, the presence of a fragment on Y chromosome and finally the presence of permutation in the FMR1 gene should be examined. Due to the effect of other factors to develop POF including infections and immune disorders, relevant tests should also be considered. However, accurate clinical examinations, history taking, and family history can help to some extent in determining the cause of EOF.
Acknowledgments: none declared.
Ethical permission: none declared.
Conflicts of interests: none declared.
Authors’ contribution: Akram Abdi (first author), writing introduction / main researcher / statistical analyst (40%); Iman Bagherizadeh (second author), assistant researcher (5%); Fateme Vand-Rajabpour, (third author), writing introduction / methodologist/ assistant researcher (10%); Yousef Shafaghati, (fourth author), assistant researcher (5%); Farkhondeh Behjati (fifth author), writing introduction / methodologist / main researcher/ statistical analyst/ writing discussion (40%)
Funding/Support: this study was supported by Sarem Cell Research Center & Medical Genetics and the Sarem Fertility and Infertility Research Center (SAFIR).