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An idic(15) associated with POF (premature ovarian failure): Molecular cytogenetic definition of a case and review of the literature
Gene 503 (2012) 123–125
Contents lists available at SciVerse ScienceDirect
Gene journal homepage: www.elsevier.com/locate/gene
Short Communication
An idic(15) associated with POF (premature ovarian failure): Molecular cytogenetic definition of a case and review of the literature Veronica Bertini a, David Viola b, Paolo Vitti b, Paolo Simi a, Angelo Valetto a,⁎ a b
Cytogenetic and Molecular Genetic Unit, Children Department, A.O.U. Pisana, S. Chiara Hospital, 56100 Pisa, Italy Department of Endocrinology and Metabolism, University of Pisa, 56100 Pisa, Italy
a r t i c l e
i n f o
Article history: Accepted 22 April 2012 Available online 2 May 2012 Keywords: Lymphocytic karyotype Chromosome aberration Infertility Premature ovarian failure FISH CGH array
a b s t r a c t We report on a 36-year-old infertile woman, presenting a premature ovarian failure with an otherwise normal female phenotype. Cytogenetic analyses showed the presence of a supernumerary marker chromosome, that was characterized by FISH (fluorescent in situ hybridization) and array CGH (comparative genomic hybridization). This marker chromosome was derived from chromosome 15, and contained only heterochromatic material. The Prader Willi/Angelman region was not present. No duplications of the 15q regions were detected by array CGH. Supernumerary markers of chromosome 15 have been reported in cases of infertility and amenorrhea, that is also described in cases with marker derived by other acrocentric chromosomes. The case here presented constitutes a further example that etiology of POF is not always associated with a defective gene, but in some cases oocytes atresia can be the consequence of the abnormal meiotic pairing of chromosomes. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Premature ovarian failure (POF) is a disorder in which amenorrhea, associated to elevated levels of gonadotropins, occurs before the age of 40 years (Albright et al., 1942; Coulam, 1982). This condition differs from menopause because in 5–10% of cases the patients can conceive and deliver a child after the diagnosis (Van Kasteren and Schoemaker, 1999). About 90% of cases POF are idiopathic, whereas in the remaining 10% the etiology is quite heterogeneous, including environmental, metabolic, iatrogenic, autoimmune and genetic factors (Vujovic, 2009). A genetic basis for POF has been well established by the report of numerous familial cases (Van Kasteren et al., 1999) even if the molecular mechanisms underlying POF are far from being completely elucidated.
Abbreviations: 15q, long arm of chromosome 15; 17OHP, 17 hydroxyprogesterone; ALUI, restriction enzyme ALUI; CGH, comparative genomic hybridization; CR1, critical region 1; CR2, critical region 2; Cy3, cyanine 3; Cy5, cyanine 5; DA-DAPI, distamycin A-4′,6-diamidino-2-phenylindole; Idic(15), isodicentric chromosome 15; FISH, fluorescent in situ hybridization; FMR1, fragile X mental retardation 1; FRAXA, fragile X A; FSH, follicle stimulating hormone; FT3, triiodothyronine; FT4, tetraiodothyronine; LH, luteinizing hormone; POF, premature ovarian failure; RSAI, restriction enzyme RSAI; SMC, supernumerary marker chromosome; TSH, thyroid-stimulating hormone; UTR, untranslated region; XY, chromosomes X and Y. ⁎ Corresponding author at: Cytogenetic and Molecular Genetic Unit, Children Department, A.O.U. Pisana, Ospedale S. Chiara, Via Roma 57, 56100 Pisa, Italy. Tel.: + 39 050 992777; fax: + 39 050 993498. E-mail address: [email protected] (A. Valetto). 0378-1119/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.gene.2012.04.071
In the last years an extensive research of candidate genes for POF has been performed, but so far, the genes identified can be considered as a risk factor for this pathology; for example, it is known that expansion of CGG repeats at 5′UTR of FMR1, the gene responsible for the Fragile X syndrome, is associated with POF in female carriers (Bretherick et al., 2005). These data are consistent with the fact that POF is a multifactorial disease, where the phenotype is most probably the result of the interaction of more than one gene. The complex genetic etiology of POF can be exemplified by the studies on chromosome X. This chromosome plays a primary role in the pathology (Toniolo, 2006), and molecular analysis of X rearrangements (deletions, inversions, and X;autosome translocation) has suggested that 2 regions, called ‘critical region 1’ (CR1) and ‘critical region 2’ (CR2), are essential for normal ovarian function and normal reproductive lifespan (Rizzolio et al., 2006). It was initially hypothesized that genes for ovary development and/or function were clustered in CR1 e CR2, but a search for genes interrupted by the breakpoints in balanced-X autosome translocations identified only five genes, out of >40 translocations mapped. Moreover, none of them showed a clear function in ovarian physiology and their mutational analysis in patients with POF failed to confirm their causative role (Prueitt et al., 2002; Rizzolio et al., 2006). The reason why these X anomalies cause POF is still debated, but it is clear that this is one of the few pathologies where the chromosomal alteration does not cause the phenotypic effects through the silencing of a gene/s, disrupted in the rearrangement, but where the cytogenetic alteration ‘per se’ causes the phenotype.
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V. Bertini et al. / Gene 503 (2012) 123–125
It emerges from all these data that genetic etiology of POF is complex but challenging: even if there are genes predisposing to the disease, the classic paradigm one gene/one disease cannot explain this pathology. Here, we present a case of secondary amenorrhea, associated to an isodicentric chromosome 15 or idic(15). The object of this study was to perform a detailed molecular characterization of the marker chromosome, in order to get further sights on the POF etiology in our patient.
2.2. Cytogenetic, FISH and FRAXA analyses
2. Materials and methods
2.3. Array CGH
2.1. Patient
Genomic DNA of the patient was isolated from peripheral blood by standard methods; a normal control female DNA was obtained by Promega (Madison, Wisconsin, USA). 1 μg of genomic DNA from both the patient (test sample) and the control (reference sample) was digested with RSAI and ALUI restriction enzymes. Test and reference DNA were differentially labeled with Cy5-dCTP or with Cy3dCTP using random primer labeling according to manufacturer's protocol (Agilent, Santa Clara, California, USA). The labeling reactions were applied to the 44 K arrays and incubated in an oven for 24 h at 65 °C. Slides were washed and scanned using the Agilent scanner. Identification of individual spots on scanned arrays was performed with the Agilent dedicated software, as well as filtration, normalization and exclusion of spots with aberrant morphology or high background.
The proband, a 36-year-old woman, was born at term after an uneventful pregnancy and delivery from healthy non-consanguineous parents. The birth weight was 2.870 g, length 50 cm; growth and mental developmental milestones were normal. At the age of 12 years she presented menarche followed by regular menses; she had no miscarriage and had a successful pregnancy at the age of 27 years. At 35 years of age she experienced asymptomatic abrupt amenorrhea. The medical history was negative for environmental, infectious and iatrogenic causes of amenorrhea and also for autoimmune diseases. Physical examination was unremarkable with no evidence of major malformations as well as facial dysmorphisms. Intellectual development was normal. Cardiac, renal, retinal and limb malformations were absent. Hypertension was absent. Pelvic ultrasound examination was normal except for the absence of follicles in the ovaries. Hormonal profile was normal for FT3, FT4, TSH, prolactin, cortisol, 17OHP, androstenedione and testosterone; LH and FSH were in the menopausal range (55.0 and 166.4 mU/mL, respectively) and were confirmed over a period of one year. Serum thyroid (antithyroglobulin and antithyroperoxidase), adrenal, ovarian, pancreatic islets and gastric autoantibodies were undetectable. Other routine blood tests were normal.
G- and Q-banded chromosomes of cultured peripheral blood lymphocytes were analyzed. Chromosomal preparations were obtained according to standard techniques. Fluorescent in situ hybridisation (FISH) was performed using probes for the centromeric region of chromosome 15 and for the Prader-Willi/ Angelmann syndrome critical region (Vysis, Downers, Grove), following the manufacturer's protocol. FRAXA expansion was analyzed by standard methods.
3. Results 3.1. Cytogenetic, FISH and FRAXA Standard cytogenetic investigations in the proband showed the presence of a small supernumerary marker in all 50 metaphases analyzed. The marker appeared bi-satellited, DA-DAPI positive. FISH analysis with 15 centromeric alphoid probe showed two signals on the marker, suggesting that this was an isodicentric chromosome derived by chromosome 15 (Figs. 1A,B,C,D). FISH with Prader Willi/
Fig. 1. A—G metaphase spread showing the marker chromosome (circle). B—Q metaphase spread showing the marker chromosome (circle).C—DA-DAPI staining showing that the marker is positive (circle). D —FISH with alphoid centromeric chromosome 15 probe. The marker shows two positive signals (circle).
V. Bertini et al. / Gene 503 (2012) 123–125
Angelmann syndrome probe was negative. FRAXA alleles were in the normal range. 3.2. Array CGH Array-CGH using a 44 K oligo platform (Agilent Technologies, Santa Clara, California, USA) excluded DNA duplications of the chromosome 15 long arm regions; no duplications or deletions of the other chromosomes were shown at a resolution of about 100 kB.
125
that amenorrhea is frequently reported also in carriers of SMCs derived from other acrocentric chromosomes, consistent with this theory of meiotic disturb (Manvelyan et al., 2008). More data are available in male carriers of SMC(15), where the segregation ratio of the marker during spermatogenesis can be analyzed by techniques like fluorescent in situ hybridization (FISH) performed on sperm. Spermatogenetic failure occurs during prophase of meiosis I, where the marker chromosome preferentially associates with the XYbivalent, leading to an early disruption of spermatogenesis (Oracova et al., 2009).
4. Discussion 5. Conclusions Here, we report on a POF case whose karyotype presented a supernumerary marker chromosome (SMC) identified as an idic(15). In order to make a better comparison with idic(15) cases previously reported we performed a detailed molecular characterization of this marker by array CGH. This technique has been applied to very few POF cases so far, thus the present case is helpful to get further sights on the POF etiology. We did not detect 15q euchromatic material with protein-codinggenes in the marker and we excluded any major rearrangement of the other chromosomes that could be missed by conventional cytogenetics. These results are consistent with the otherwise normal phenotype of our patient whose only pathological aspect is POF. FMR1 gene was in the normal range, thus also the most common risk factor could be excluded. The marker does not contain protein-coding-genes, and it can be inferred that is probably the cytogenetic alteration ‘per se’ to cause POF. Relationship between SMCs and infertility is still debated. Several studies reported an increased incidence of SMC(15) in infertile males with sperm count anomalies, whereas females with SMC(15) usually show a normal fertility (Crolla et al., 1995, 2005; Liehr et al., 2004; Morel et al., 2004). However, in a recent paper (Manvelyan et al., 2008) cases with SMCs associated with fertility problems were collected: among 14 cases of female carriers of inv dup 15, half of them presented repeated abortions, as expected in cases of chromosomal aberration carriers, and 3 cases presented with primary or secondary amenorrhea. The reasons why SMCs can lead to POF are so far speculative, but two observations can be made. First, it is known that SMCs may interfere with the segregation of other chromosomes during meiosis and cause a ‘meiotic disturb’. Faulty meiotic pairing have been described in about one third of oocytes of chromosomally normal females and this phenomenon is considered the cause of the “physiological” atresia of the germ cells which occurs during the fetal life. Defective pairing caused by SMCs might lead to death of germ cells, perhaps through a checkpoint mechanism (Bhalla, 2010; Speed, 1988) that reduces the number of ovarian follicles. Second, it is well established that also constitutional chromosome aneuploidies (i.e. trisomies 13, 18, 21 and 45,X0) (Hall, et al., 2006) cause severe pairing failure that probably determines the degeneration of nearly all oocytes leading to gonad dysgenesis. Unfortunately, to the best of our knowledge, there are no studies on fetal oocytes in carriers of supernumerary marker chromosomes, but is worth noting
In conclusion, detection of a SMC(15) in a routine karyotyping of a woman with fertility problems reinforces the importance of detailed clinical and molecular-cytogenetic evaluation, in order to better elucidate its relationship with POF and to offer an exhaustive genetic counseling before reproductive options are considered. References Albright, F., Smith, P.H., Fraser, R., 1942. A syndrome characterized by primary ovarian insufficiency and decreased stature: report of 11 cases with a digression on hormonal control of axillary and pubic hair. Am. J. Med. Sci. 204, 625–648. Bhalla, N., 2010. Meiotic checkpoints: repair or removal? Curr. Biol. 20, 1014–1016. Bretherick, L.K., Fluker, M.R., Robinson, W.P., 2005. FMR1 repeat sizes in the gray zone and high end of the normal range are associated with premature ovarian failure. Hum. Genet. 117, 376–382. Coulam, C.B., 1982. Premature gonadal failure. Fertil. Steril. 38, 645–655. Crolla, A.J., Aharvey, J.F., Sitch, F.L., Dennis, N.R., 1995. Supernumerary marker 15 chromosomes: a clinical, molecular and FISH approach to diagnosis and prognosis. Hum. Genet. 95, 161–170. Crolla, A.J., Youings, S.A., Ennis, S., Jacobs, P.A., 2005. Supernumerary marker chromosomes in man: parental origin, mosaicism and maternal age revisited. Eur. J. Hum. Genet. 13, 154–160. Hall, H., Hunt, P., Hassold, T., 2006. Meiosis and sex chromosome aneuploidy: how meiotic errors cause aneuploidy; how aneuploidy causes meiotic errors. Curr. Opin. Genet. Dev. 16, 323–329. Liehr, T., Claussen, U., Starke, H., 2004. Small supernumerary marker chromosomes (sSMC) in humans. Cytogenet. Genome Res. 107, 55–67. Manvelyan, M., et al., 2008. Thirty-two new cases with small supernumerary marker chromosomes detected in connection with fertility problems: detailed molecular cytogenetic characterization and review of the literature. Int. J. Mol. Med. 21, 705–714. Morel, F., et al., 2004. Chromosomal abnormalities in couples undergoing intracytoplasmic sperm injection. A study of 370 couples and review of the literature. Int. J. Androl. 27, 178–182. Oracova, E., et al., 2009. Sperm and embryo analysis in a carrier of supernumerary inv dup(15) marker chromosome. J. Androl. 30, 233–239. Prueitt, R.L., Chen, H., Barnes, R.I., Zinn, A.R., 2002. Most X;autosome translocations associated with premature ovarian failure do not interrupt X-linked genes. Cytogenet. Genome Res. 97, 32–38. Rizzolio, F., et al., 2006. Chromosomal rearrangements in Xq and premature ovarian failure: mapping of 25 new cases and review of the literature. Hum. Reprod. 21, 1477–1483. Speed, R.M., 1988. The possible role of meiotic pairing anomalies in the atresia of human fetal oocytes. Hum. Genet. 78, 260–266. Toniolo, D., 2006. X-linked premature ovarian failure: a complex disease. Curr. Opin. Genet. Dev. 16, 293–300. Van Kasteren, Y.M., Schoemaker, J., 1999. Premature ovarian failure: a systematic review on therapeutic interventions to restore ovarian function and achieve pregnancy. Hum. Reprod. Update 5, 483–492. Van Kasteren, Y.M., Hundscheid, R.D., Smits, A.P., Cremers, F.P., van Zonneveld, P., Braat, D.D., 1999. Familial idiopathic premature ovarian failure: an overrated and underestimated genetic disease? Hum. Reprod. 14, 2455–2459. Vujovic, S., 2009. Aetiology of premature ovarian failure. Menopause. Int. 15, 72–75.
Contents lists available at SciVerse ScienceDirect
Gene journal homepage: www.elsevier.com/locate/gene
Short Communication
An idic(15) associated with POF (premature ovarian failure): Molecular cytogenetic definition of a case and review of the literature Veronica Bertini a, David Viola b, Paolo Vitti b, Paolo Simi a, Angelo Valetto a,⁎ a b
Cytogenetic and Molecular Genetic Unit, Children Department, A.O.U. Pisana, S. Chiara Hospital, 56100 Pisa, Italy Department of Endocrinology and Metabolism, University of Pisa, 56100 Pisa, Italy
a r t i c l e
i n f o
Article history: Accepted 22 April 2012 Available online 2 May 2012 Keywords: Lymphocytic karyotype Chromosome aberration Infertility Premature ovarian failure FISH CGH array
a b s t r a c t We report on a 36-year-old infertile woman, presenting a premature ovarian failure with an otherwise normal female phenotype. Cytogenetic analyses showed the presence of a supernumerary marker chromosome, that was characterized by FISH (fluorescent in situ hybridization) and array CGH (comparative genomic hybridization). This marker chromosome was derived from chromosome 15, and contained only heterochromatic material. The Prader Willi/Angelman region was not present. No duplications of the 15q regions were detected by array CGH. Supernumerary markers of chromosome 15 have been reported in cases of infertility and amenorrhea, that is also described in cases with marker derived by other acrocentric chromosomes. The case here presented constitutes a further example that etiology of POF is not always associated with a defective gene, but in some cases oocytes atresia can be the consequence of the abnormal meiotic pairing of chromosomes. © 2012 Elsevier B.V. All rights reserved.
1. Introduction Premature ovarian failure (POF) is a disorder in which amenorrhea, associated to elevated levels of gonadotropins, occurs before the age of 40 years (Albright et al., 1942; Coulam, 1982). This condition differs from menopause because in 5–10% of cases the patients can conceive and deliver a child after the diagnosis (Van Kasteren and Schoemaker, 1999). About 90% of cases POF are idiopathic, whereas in the remaining 10% the etiology is quite heterogeneous, including environmental, metabolic, iatrogenic, autoimmune and genetic factors (Vujovic, 2009). A genetic basis for POF has been well established by the report of numerous familial cases (Van Kasteren et al., 1999) even if the molecular mechanisms underlying POF are far from being completely elucidated.
Abbreviations: 15q, long arm of chromosome 15; 17OHP, 17 hydroxyprogesterone; ALUI, restriction enzyme ALUI; CGH, comparative genomic hybridization; CR1, critical region 1; CR2, critical region 2; Cy3, cyanine 3; Cy5, cyanine 5; DA-DAPI, distamycin A-4′,6-diamidino-2-phenylindole; Idic(15), isodicentric chromosome 15; FISH, fluorescent in situ hybridization; FMR1, fragile X mental retardation 1; FRAXA, fragile X A; FSH, follicle stimulating hormone; FT3, triiodothyronine; FT4, tetraiodothyronine; LH, luteinizing hormone; POF, premature ovarian failure; RSAI, restriction enzyme RSAI; SMC, supernumerary marker chromosome; TSH, thyroid-stimulating hormone; UTR, untranslated region; XY, chromosomes X and Y. ⁎ Corresponding author at: Cytogenetic and Molecular Genetic Unit, Children Department, A.O.U. Pisana, Ospedale S. Chiara, Via Roma 57, 56100 Pisa, Italy. Tel.: + 39 050 992777; fax: + 39 050 993498. E-mail address: [email protected] (A. Valetto). 0378-1119/$ – see front matter © 2012 Elsevier B.V. All rights reserved. doi:10.1016/j.gene.2012.04.071
In the last years an extensive research of candidate genes for POF has been performed, but so far, the genes identified can be considered as a risk factor for this pathology; for example, it is known that expansion of CGG repeats at 5′UTR of FMR1, the gene responsible for the Fragile X syndrome, is associated with POF in female carriers (Bretherick et al., 2005). These data are consistent with the fact that POF is a multifactorial disease, where the phenotype is most probably the result of the interaction of more than one gene. The complex genetic etiology of POF can be exemplified by the studies on chromosome X. This chromosome plays a primary role in the pathology (Toniolo, 2006), and molecular analysis of X rearrangements (deletions, inversions, and X;autosome translocation) has suggested that 2 regions, called ‘critical region 1’ (CR1) and ‘critical region 2’ (CR2), are essential for normal ovarian function and normal reproductive lifespan (Rizzolio et al., 2006). It was initially hypothesized that genes for ovary development and/or function were clustered in CR1 e CR2, but a search for genes interrupted by the breakpoints in balanced-X autosome translocations identified only five genes, out of >40 translocations mapped. Moreover, none of them showed a clear function in ovarian physiology and their mutational analysis in patients with POF failed to confirm their causative role (Prueitt et al., 2002; Rizzolio et al., 2006). The reason why these X anomalies cause POF is still debated, but it is clear that this is one of the few pathologies where the chromosomal alteration does not cause the phenotypic effects through the silencing of a gene/s, disrupted in the rearrangement, but where the cytogenetic alteration ‘per se’ causes the phenotype.
124
V. Bertini et al. / Gene 503 (2012) 123–125
It emerges from all these data that genetic etiology of POF is complex but challenging: even if there are genes predisposing to the disease, the classic paradigm one gene/one disease cannot explain this pathology. Here, we present a case of secondary amenorrhea, associated to an isodicentric chromosome 15 or idic(15). The object of this study was to perform a detailed molecular characterization of the marker chromosome, in order to get further sights on the POF etiology in our patient.
2.2. Cytogenetic, FISH and FRAXA analyses
2. Materials and methods
2.3. Array CGH
2.1. Patient
Genomic DNA of the patient was isolated from peripheral blood by standard methods; a normal control female DNA was obtained by Promega (Madison, Wisconsin, USA). 1 μg of genomic DNA from both the patient (test sample) and the control (reference sample) was digested with RSAI and ALUI restriction enzymes. Test and reference DNA were differentially labeled with Cy5-dCTP or with Cy3dCTP using random primer labeling according to manufacturer's protocol (Agilent, Santa Clara, California, USA). The labeling reactions were applied to the 44 K arrays and incubated in an oven for 24 h at 65 °C. Slides were washed and scanned using the Agilent scanner. Identification of individual spots on scanned arrays was performed with the Agilent dedicated software, as well as filtration, normalization and exclusion of spots with aberrant morphology or high background.
The proband, a 36-year-old woman, was born at term after an uneventful pregnancy and delivery from healthy non-consanguineous parents. The birth weight was 2.870 g, length 50 cm; growth and mental developmental milestones were normal. At the age of 12 years she presented menarche followed by regular menses; she had no miscarriage and had a successful pregnancy at the age of 27 years. At 35 years of age she experienced asymptomatic abrupt amenorrhea. The medical history was negative for environmental, infectious and iatrogenic causes of amenorrhea and also for autoimmune diseases. Physical examination was unremarkable with no evidence of major malformations as well as facial dysmorphisms. Intellectual development was normal. Cardiac, renal, retinal and limb malformations were absent. Hypertension was absent. Pelvic ultrasound examination was normal except for the absence of follicles in the ovaries. Hormonal profile was normal for FT3, FT4, TSH, prolactin, cortisol, 17OHP, androstenedione and testosterone; LH and FSH were in the menopausal range (55.0 and 166.4 mU/mL, respectively) and were confirmed over a period of one year. Serum thyroid (antithyroglobulin and antithyroperoxidase), adrenal, ovarian, pancreatic islets and gastric autoantibodies were undetectable. Other routine blood tests were normal.
G- and Q-banded chromosomes of cultured peripheral blood lymphocytes were analyzed. Chromosomal preparations were obtained according to standard techniques. Fluorescent in situ hybridisation (FISH) was performed using probes for the centromeric region of chromosome 15 and for the Prader-Willi/ Angelmann syndrome critical region (Vysis, Downers, Grove), following the manufacturer's protocol. FRAXA expansion was analyzed by standard methods.
3. Results 3.1. Cytogenetic, FISH and FRAXA Standard cytogenetic investigations in the proband showed the presence of a small supernumerary marker in all 50 metaphases analyzed. The marker appeared bi-satellited, DA-DAPI positive. FISH analysis with 15 centromeric alphoid probe showed two signals on the marker, suggesting that this was an isodicentric chromosome derived by chromosome 15 (Figs. 1A,B,C,D). FISH with Prader Willi/
Fig. 1. A—G metaphase spread showing the marker chromosome (circle). B—Q metaphase spread showing the marker chromosome (circle).C—DA-DAPI staining showing that the marker is positive (circle). D —FISH with alphoid centromeric chromosome 15 probe. The marker shows two positive signals (circle).
V. Bertini et al. / Gene 503 (2012) 123–125
Angelmann syndrome probe was negative. FRAXA alleles were in the normal range. 3.2. Array CGH Array-CGH using a 44 K oligo platform (Agilent Technologies, Santa Clara, California, USA) excluded DNA duplications of the chromosome 15 long arm regions; no duplications or deletions of the other chromosomes were shown at a resolution of about 100 kB.
125
that amenorrhea is frequently reported also in carriers of SMCs derived from other acrocentric chromosomes, consistent with this theory of meiotic disturb (Manvelyan et al., 2008). More data are available in male carriers of SMC(15), where the segregation ratio of the marker during spermatogenesis can be analyzed by techniques like fluorescent in situ hybridization (FISH) performed on sperm. Spermatogenetic failure occurs during prophase of meiosis I, where the marker chromosome preferentially associates with the XYbivalent, leading to an early disruption of spermatogenesis (Oracova et al., 2009).
4. Discussion 5. Conclusions Here, we report on a POF case whose karyotype presented a supernumerary marker chromosome (SMC) identified as an idic(15). In order to make a better comparison with idic(15) cases previously reported we performed a detailed molecular characterization of this marker by array CGH. This technique has been applied to very few POF cases so far, thus the present case is helpful to get further sights on the POF etiology. We did not detect 15q euchromatic material with protein-codinggenes in the marker and we excluded any major rearrangement of the other chromosomes that could be missed by conventional cytogenetics. These results are consistent with the otherwise normal phenotype of our patient whose only pathological aspect is POF. FMR1 gene was in the normal range, thus also the most common risk factor could be excluded. The marker does not contain protein-coding-genes, and it can be inferred that is probably the cytogenetic alteration ‘per se’ to cause POF. Relationship between SMCs and infertility is still debated. Several studies reported an increased incidence of SMC(15) in infertile males with sperm count anomalies, whereas females with SMC(15) usually show a normal fertility (Crolla et al., 1995, 2005; Liehr et al., 2004; Morel et al., 2004). However, in a recent paper (Manvelyan et al., 2008) cases with SMCs associated with fertility problems were collected: among 14 cases of female carriers of inv dup 15, half of them presented repeated abortions, as expected in cases of chromosomal aberration carriers, and 3 cases presented with primary or secondary amenorrhea. The reasons why SMCs can lead to POF are so far speculative, but two observations can be made. First, it is known that SMCs may interfere with the segregation of other chromosomes during meiosis and cause a ‘meiotic disturb’. Faulty meiotic pairing have been described in about one third of oocytes of chromosomally normal females and this phenomenon is considered the cause of the “physiological” atresia of the germ cells which occurs during the fetal life. Defective pairing caused by SMCs might lead to death of germ cells, perhaps through a checkpoint mechanism (Bhalla, 2010; Speed, 1988) that reduces the number of ovarian follicles. Second, it is well established that also constitutional chromosome aneuploidies (i.e. trisomies 13, 18, 21 and 45,X0) (Hall, et al., 2006) cause severe pairing failure that probably determines the degeneration of nearly all oocytes leading to gonad dysgenesis. Unfortunately, to the best of our knowledge, there are no studies on fetal oocytes in carriers of supernumerary marker chromosomes, but is worth noting
In conclusion, detection of a SMC(15) in a routine karyotyping of a woman with fertility problems reinforces the importance of detailed clinical and molecular-cytogenetic evaluation, in order to better elucidate its relationship with POF and to offer an exhaustive genetic counseling before reproductive options are considered. References Albright, F., Smith, P.H., Fraser, R., 1942. A syndrome characterized by primary ovarian insufficiency and decreased stature: report of 11 cases with a digression on hormonal control of axillary and pubic hair. Am. J. Med. Sci. 204, 625–648. Bhalla, N., 2010. Meiotic checkpoints: repair or removal? Curr. Biol. 20, 1014–1016. Bretherick, L.K., Fluker, M.R., Robinson, W.P., 2005. FMR1 repeat sizes in the gray zone and high end of the normal range are associated with premature ovarian failure. Hum. Genet. 117, 376–382. Coulam, C.B., 1982. Premature gonadal failure. Fertil. Steril. 38, 645–655. Crolla, A.J., Aharvey, J.F., Sitch, F.L., Dennis, N.R., 1995. Supernumerary marker 15 chromosomes: a clinical, molecular and FISH approach to diagnosis and prognosis. Hum. Genet. 95, 161–170. Crolla, A.J., Youings, S.A., Ennis, S., Jacobs, P.A., 2005. Supernumerary marker chromosomes in man: parental origin, mosaicism and maternal age revisited. Eur. J. Hum. Genet. 13, 154–160. Hall, H., Hunt, P., Hassold, T., 2006. Meiosis and sex chromosome aneuploidy: how meiotic errors cause aneuploidy; how aneuploidy causes meiotic errors. Curr. Opin. Genet. Dev. 16, 323–329. Liehr, T., Claussen, U., Starke, H., 2004. Small supernumerary marker chromosomes (sSMC) in humans. Cytogenet. Genome Res. 107, 55–67. Manvelyan, M., et al., 2008. Thirty-two new cases with small supernumerary marker chromosomes detected in connection with fertility problems: detailed molecular cytogenetic characterization and review of the literature. Int. J. Mol. Med. 21, 705–714. Morel, F., et al., 2004. Chromosomal abnormalities in couples undergoing intracytoplasmic sperm injection. A study of 370 couples and review of the literature. Int. J. Androl. 27, 178–182. Oracova, E., et al., 2009. Sperm and embryo analysis in a carrier of supernumerary inv dup(15) marker chromosome. J. Androl. 30, 233–239. Prueitt, R.L., Chen, H., Barnes, R.I., Zinn, A.R., 2002. Most X;autosome translocations associated with premature ovarian failure do not interrupt X-linked genes. Cytogenet. Genome Res. 97, 32–38. Rizzolio, F., et al., 2006. Chromosomal rearrangements in Xq and premature ovarian failure: mapping of 25 new cases and review of the literature. Hum. Reprod. 21, 1477–1483. Speed, R.M., 1988. The possible role of meiotic pairing anomalies in the atresia of human fetal oocytes. Hum. Genet. 78, 260–266. Toniolo, D., 2006. X-linked premature ovarian failure: a complex disease. Curr. Opin. Genet. Dev. 16, 293–300. Van Kasteren, Y.M., Schoemaker, J., 1999. Premature ovarian failure: a systematic review on therapeutic interventions to restore ovarian function and achieve pregnancy. Hum. Reprod. Update 5, 483–492. Van Kasteren, Y.M., Hundscheid, R.D., Smits, A.P., Cremers, F.P., van Zonneveld, P., Braat, D.D., 1999. Familial idiopathic premature ovarian failure: an overrated and underestimated genetic disease? Hum. Reprod. 14, 2455–2459. Vujovic, S., 2009. Aetiology of premature ovarian failure. Menopause. Int. 15, 72–75.