Journal of Human Reproductive Science
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Year : 2022  |  Volume : 15  |  Issue : 2  |  Page : 187-190

Cytogenetic studies in primary amenorrhoea cases

1 Clinical Research Development Unit of Tabriz Valiasr Hospital, Tabriz University of Medical, Sciences, Tabriz, Iran
2 Food and Drug Administrator, University of Medical Sciences, Isfahan, Iran
3 Department of Cell and Molecular Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran

Date of Submission29-Jan-2022
Date of Decision31-May-2022
Date of Acceptance04-Jun-2022
Date of Web Publication30-Jun-2022

Correspondence Address:
Elham Ghadirkhomi
Clinical Research Development Unit of Tabriz Valiasr Hospital, Tabriz University of Medical, Sciences, Tabriz
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/jhrs.jhrs_13_22

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Background: Amenorrhoea is considered a kind of menstrual disorder in a woman of reproductive age. It is a symptom with many potential causes such as an abnormality in the hypothalamic–pituitary–ovarian axis, anatomical abnormalities of the genital tract or functional causes. Aims: In this study, we aimed to investigate chromosomal abnormalities in patients presenting with primary amenorrhoea. Setting and Design: This study was conducted in the medical genetic laboratory. Materials and Methods: Chromosomal analysis was carried out in 134 cases that were referred to the human genetic laboratory from 2010 to 2017, employing (GTG) Giemsa banding. Statistical Analysis Used: Statistical analyses were carried out by Microsoft Office Excel (2019). Results: The karyotype results revealed 77.6% (n = 104) with normal chromosome composition while 22.38% (n = 30) showed chromosomal abnormalities. Among the patients with abnormal chromosome constituents, 53.54% exhibited numerical aberration and 46.66% showed structural abnormalities. Conclusion: The present study has emphasised that karyotyping is one of the fundamental investigations in the evaluation of primary amenorrhoea.

Keywords: Chromosomal abnormality, cytogenetic study, karyotyping, primary amenorrhoea

How to cite this article:
Ghadirkhomi E, Ghdirkhomi A, Angaji SA. Cytogenetic studies in primary amenorrhoea cases. J Hum Reprod Sci 2022;15:187-90

How to cite this URL:
Ghadirkhomi E, Ghdirkhomi A, Angaji SA. Cytogenetic studies in primary amenorrhoea cases. J Hum Reprod Sci [serial online] 2022 [cited 2022 Sep 30];15:187-90. Available from:

   Introduction Top

Most girls begin menstruating between the ages of 9 and 18, with an average of around 12 years. If other pubertal changes occur naturally, the absence of menstruation until the age of 15 is not considered primary amenorrhoea. Primary amenorrhoea may occur with or without other signs of puberty.[1],[2]

Although there are many reasons for primary amenorrhoea including constitutional delay, chronic systemic disease, hypothalamic-pituitary dysfunction and absent ovarian function, genetic or chromosomal causes are the most important as their presence affects subsequent management.[3],[4]

Primary amenorrhoea can be caused by chromosomal disorders such as Turner syndrome, gonadal disorders, structural problems with the uterus and structural abnormalities of the Fallopian tubes which could prevent or inhibit the normal development or the release of eggs into the uterus.[5],[6] According to previous studies, the percentage of chromosomal abnormalities varies from 15.9% to 63.3% in patients with primary amenorrhoea.[3],[6] This study was undertaken to determine the frequency and type of chromosomal abnormalities that result in primary amenorrhoea in the local population.

   Materials and Methods Top

In this study, we used the data obtained from patients (n = 134) referred with primary amenorrhoea for chromosomal analysis. The cases were referred from different parts of Iran between 2010 and 2017. The age group of the participants ranged from 17 to 30 years. For chromosomal abnormality investigation, routine lymphocyte cell culture and the G-banding methods were used.

About 1 ml of blood was added to 6 ml of Roswell Park Memorial Institute medium supplemented with 10% of foetal bovine serum and 0.1 μg/ml of phytohaemagglutinin and was incubated at 37°C. Cells were treated with 0.1 μg/ml of colcemid after a 72-h incubation period, after which metaphase chromosomes of each patient were spread on a slide and stained using the standard G-banding technique.[7],[8] A minimum of 25 metaphase cells per slide were analysed to find any numerical or structural chromosomal abnormalities using a microscope. If an abnormal karyotype was found in this first analysis, further cells (up to 60) could be examined.

This research and methods were performed under the ethical principles, national norms, relevant guidelines and regulations for conducting medical research in Iran. Written informed consent was obtained from all eligible subjects. The project was approved by the science and research branch of Tehran's Islamic azad university's ethical committee (Tehran, Iran, IR.IAU.SRB.REC.1398.001).

For this cross-sectional study, the sample size was calculated based on Cochran's formula using an online sample size calculator website ( Obtained desired sample size for this study was 130 individuals.

   Results Top

Among the 134 patients, 30 had abnormal karyotypes. The chromosomal analysis and the obtained karyotype for the patients are summarised in [Table 1]. Chromosomal abnormalities can be classified into five main types with or without mosaicism [Figure 1]:
Table 1: Chromosomal abnormalities in primary amenorrhea

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Figure 1: Chromosome abnormalities in primary amenorrhoea

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  1. The most frequent karyotype was X chromosome aneuploidies (n = 14) that include Turner syndrome 45, X (n = 6), mosaic Turner 45, X/46, XX (n = 7) and 48, XXXX (n = 1).
  2. A male karyotype of 46, XY was present in four patients.
  3. Structural anomalies of the X chromosome were detected in 10 cases. Four patients were found to have an isochromosome of the long arm of the X chromosome (×46, i[Xq]), one patient had an isochromosome of the short arm of the X chromosome (×46, i[Xp]) and five patients had a partial deletion of the X chromosome.
  4. Mosaicism of male chromosome constitution and X chromosome aneuploidy was present in one case (45, X [39]/46, XY [21]).
  5. Mosaicism of X chromosome aneuploidy and structural anomalies of the X chromosome were found in one case 45, X [12]/46, i (Xq) [27]/47, X, i (Xq), i (Xq).[17] Partial karyotype images of the chromosome abnormalities are shown in [Figure 2].
Figure 2: Partial karyotype images of the chromosome abnormalities (only sex chromosome images of the karyotypes are shown). (a) Normal female. (b) Male karyotype: XY. (c) Monosomy X. (d) Iso-chromosome of long arm of X: 46,X,i(Xq). (e) 48,XXXX. (f) 47,X,i(Xq),i(Xq). (g) Iso-chromosome of short arm of X: 46,X,i(Xp)

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   Discussion Top

Primary amenorrhoea is the complete absence of menstruation. Genetic factors have a causal effect in addition to endocrine disturbances and constitutional and environmental factors in primary amenorrhoea.[9] Several cytogenetic studies have been performed aimed at understanding the frequency and nature of chromosomal abnormalities in primary amenorrhoea.[10],[11] In the present study, the incidence of chromosomal abnormalities in amenorrhoea was found to be 22.38%. The reported incidence in different kinds of literature ranges from 13% to 32% and numerical X chromosomal abnormality is frequent.[12]

Among the different types of chromosomal abnormalities, either complete monosomy of X (n = 6) or mosaicism (n = 7) was observed in 43.3% of cases. The obtained results were in line with the previous reports that indicate Turner's syndrome as the leading cause of primary amenorrhoea. The obtained results further strengthened the role of gene composition on the X chromosome in normal female physiology and reproduction. The presence of extra X chromosomes causes ovarian failure due to hormonal imbalance.[14] Complete or partial deletion of chromosome X and consequent gene insufficiency in deleted regions also leads to gonadal dysgenesis.[13] It is known that deletion of the long arm of the X chromosome leads to ovarian failure.[14] The ZFX gene is another critical gene for the normal development of female gonads that is located on the short arm of the X chromosome.[15] The lack of Short Stature Homeobox (SHOX), a vital growth development gene located on the short arm of the X chromosome, has also been reported to be a major cause of Turner syndrome phenotype and primary amenorrhoea.[14]

Sex-reversed female, individuals with female phenotype but 46, XY karyotype, occurs in the embryos with a mutation or deletion on testis determining factor, the sex determination region of the chromosome Y (SRY gene).[14],[16] These subjects often represent normally developed Müllerian ducts, gonadal dysgenesis, poorly developed breast, absence of the uterus or ovaries and primary amenorrhoea.[14],[16] The chromosomal abnormalities in primary amenorrhoea could be grouped as the X numerical; X structural, structural and numerical abnormalities together, 46, XY female and sex reversal condition [Figure 3]. Their frequencies from different reported articles are: 45, X (40%–50%), X mosaicism (25%–36%), X structural (8%), 46, XY female (16%) and mosaicism of male chromosome constitution (45, X/46, XY) (1.36%).[5],[6],[15],[17] The frequency of chromosomal anomalies among different populations is summarised in [Table 2]. In the present study, the respective frequency is 20% (45, X), 23.3% (X mosaicism), 32% (X structural), 13.3% (46, XY) and 3.3% (45, X/46, XY). A rare type of aneuploidy together with structural anomalies of the X chromosome (45, X/46, X, i[Xq]/47, X, i[Xq], i[Xq]) was observed in one of the studied cases. Two reports of this anomaly have previously been published by Melaragno MI, et al.

, (1993) and Gorukmez O, et al. (2015).[18],[19] Although the results of this study emphasise the role of chromosomal abnormalities as a causative factor of primary amenorrhoea, as a limitation of the study, other possible factors such as molecular markers have not been included in this study, which need further investigation.
Figure 3: Chromosomal abnormalities in primary amenorrhoea

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Table 2: Frequency of chromosomal abnormalities among different populations

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   Conclusion Top

Our study confirms that chromosomal abnormalities are significant factors in gonadal dysgenesis resulting in primary amenorrhoea. Furthermore, an appreciable proportion of these individuals with sex chromosome abnormalities is mosaics. Genetic counselling should include the risk of premature menopause for patients with Turner syndrome, use of hormonal replacement therapy, risk of gonadal malignancy in patients with XY gonadal dysgenesis and possibility of infertility in the future children of patients with mosaic Turner syndrome.

Availability of data and materials

All data obtained and analysed during this study are included in this article. Details are available from the corresponding author, on reasonable request.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.

   References Top

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[PUBMED]  [Full text]  
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Gorukmez O, Sag SO, Gulten T, Ture M, Yakut T. TURNER SYNDROME WITH 45, X/46, X, I (Xq)/47, X, I (Xq), I (Xq) KARYOTYPE. Genetic Counseling. 2015;26(2):267.  Back to cited text no. 19
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  [Figure 1], [Figure 2], [Figure 3]

  [Table 1], [Table 2]


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