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BASIC CYTOGENETICS AND CYTOGENETICS OF INFERTILITY Richard Hall BSc SRCS Cytogenetics Department, Guy's & St Thomas' NHS Foundation Trust Basic Genetics.

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Presentation on theme: "BASIC CYTOGENETICS AND CYTOGENETICS OF INFERTILITY Richard Hall BSc SRCS Cytogenetics Department, Guy's & St Thomas' NHS Foundation Trust Basic Genetics."— Presentation transcript:

1 BASIC CYTOGENETICS AND CYTOGENETICS OF INFERTILITY Richard Hall BSc SRCS Cytogenetics Department, Guy's & St Thomas' NHS Foundation Trust Basic Genetics for ART Practitioners

2 Chromosomes? The most important objects in the living world, for the genes they carry determine the existence and form of organisms.

3 G-banded karyotype

4 Cytogenetics? The study of the genetic constitution of cells through the visualisation and analysis of chromosomes. –G-banding (and other traditional techniques) –Fluorescence in situ hybridization (FISH) –Molecular techniques (QF-PCR, MLPA)

5 Chromosome analysis techniques QF-PCR 8p del 4p dup 10p del MLPA MICROARRAYS FISHFISH CGH

6 Preparation of metaphases CULTURE SYNCHRONISEHARVEST PREPARE SLIDES STAIN SLIDES ANALYSE CHROMOSOMES 72 hours to 14 days

7 Traditional microscopy METAPHASE Low power x100High power x1000

8 Traditional microscopy High power (1000x) view. Next stage of analysis involves locating each chromosome pair and comparing them band for band. Random distribution of chromosomes can hinder the accuracy and efficiency of the band comparison. Typically 1000 bands per cell.

9 Chromosome abnormalities AneuploidyAneuploidy –too many chromosomes –too few chromosomes RearrangementsRearrangements –translocations balancedbalanced unbalancedunbalanced –inversions

10 Chromosome abnormalities Chromosome abnormalities seen in adults referred for: infertilityinfertility mostly sex chromosome aneuploidy rearrangements involving sex chromosomes recurrent miscarriagerecurrent miscarriage balanced chromosome rearrangements e.g. translocations and inversions 6% 2.5% However, up to 50% of first trimester loss is due to foetal chromosome abnormality – mostly de novo

11 Spontaneous abortion products 46,N+1645,X Triploidy Other Other autosomal trisomy 15% of first trimester pregnancies are lost 50% normal 50% abnormal

12 Aneuploidy Mostly from meiotic non-disjunction. Meiosis is the specialised cell division that generates haploid gametes. Errors in meiotic segregation occur frequently in human females, especially in MI.

13 Chromosome abnormalities and maternal age

14 Meiosis I non-disjunction Meiosis I Meiosis II DisomicNullisomic

15 Mosaicism The presence of two or more cell lines that are genetically identical, except for the chromosomal difference between them, in a single zygote. Frequently seen in patients with sex chromosome aneuploidy. Abnormal cell line may be in the minority.

16 47,XXY 46,XY 47,XXY 46,XY Anaphase lag – loss of one X 47,XXY/46,XY Mosaicism

17 Turner syndrome High mortality in first trimester foetuses Oedema of extremities Oedema of extremities Coarctation of the aorta Coarctation of the aorta Webbed neck Webbed neck Classical karyotype = 45,X (45%)

18 Turner syndrome Phenotype very variable, often mild and dependant on karyotype Short stature Short stature Increased carrying angle Increased carrying angle Infertility Infertility 7% mosaic, eg 45,X/46,XX 45% structural abnormality, eg 46,X,i(X)(q10)

19 Structural abnormalities of the X-chromosome Monosomy for short arm is associated with features of Turner syndrome or primary ovarian failure Partial monosomy for, or balanced rearrangements with, breakpoint in long arm more likely to be associated with premature ovarian failure The location of the breakpoint in the X may influence gonadal function

20 Structure of the X chromosome Xp11.2-p22.1 –Ovarian failure (gonadal dysgenesis) Xq13 –X inactivation centre (XIST) Xq13-q26 –Critical region for ovarian function –Breakpoints within this region are associated with gonadal insufficiency –Except breakpoints in Xq22

21 Klinefelter syndrome Incidence = 1/1000 Usually taller than average Disproportionately long limbs 30–50% gynaecomastia Infertility/azoospermia IQ may be reduced relative to siblings Example karyotypes =47,XXY 47,XXY/46,XY

22 Klinefelter syndrome Phenotype very variable – some patients are not diagnosed until they try for a family. Mosaics 47,XXY/46,XY may have milder phenotype and may be fertile. Therefore always carry out mosaicism check as infertility is the main clinical problem.

23 Chromosome translocations Exchange of material between chromosomesExchange of material between chromosomes Two typesTwo types –Robertsonian –reciprocal

24 Acrocentric chromosomes Normal male karyotype: 46,XY

25 Robertsonian translocations der(14;21)(q10;q10)

26 Robertsonians and infertility Some male carriers are infertile as they have spermatogenic arrest. Thought to be due to failure of pairing of the translocation in meiosis, which allows it to interfere with the X-Y bivalent. The more often this occurs the greater the effect on the sperm count. Prevalence of 1 in 1000.Prevalence of 1 in 1000. 10x excess in infertile men.10x excess in infertile men.

27 Alternate segregation Adjacent segregation Robertsonians and miscarriage Behaviour at meiosis Female carriers of der(14;21) have 10% risk of Downs syndrome child

28

29 Robertsonian translocations Summary Result from fusion of two acrocentric chromosomes (13, 14, 15, 21, 22)Result from fusion of two acrocentric chromosomes (13, 14, 15, 21, 22) Prevalence of 1 in 1000Prevalence of 1 in 1000 Balanced carriers have reproductive risks present as:Balanced carriers have reproductive risks present as: –recurrent miscarriage –Patau syndrome –Downs syndrome –male infertility

30 Reciprocal translocations Exchange of material between two non- homologous chromosomesExchange of material between two non- homologous chromosomes Prevalence of 1 in 500Prevalence of 1 in 500 Balanced carriers are generally phenotypically normalBalanced carriers are generally phenotypically normal Reproductive consequences because of behaviour at meiosisReproductive consequences because of behaviour at meiosis

31 The homologous chromosomes cannot pair properly Instead they must form a quadrivalent Behaviour at meiosis

32 Balanced (like carrier parent) Normal Alternate segregation

33 Unbalanced monosomy and trisomy Unbalanced trisomy and monosomy Adjacent-1 segregation

34 Reciprocal translocations: reproductive risks For most translocations, ~50% of conceptions will have either normal chromosomes or the balanced translocation.For most translocations, ~50% of conceptions will have either normal chromosomes or the balanced translocation. Unbalanced products result in:Unbalanced products result in: –miscarriage (large segments) –dysmorphic delayed child (small segments).

35 Reciprocal translocations Summary Chromosome rearrangements are rare, but chromosome analysis is indicated if a couple have had three or more miscarriages of unknown aetiology. Essential that both partners are investigated as either the male or the female could carry a balanced rearrangement. Aneuploidy is the most common chromosomal cause of early miscarriage and requires no follow-up.

36 Suggested reading Gardner, RJM & Sutherland GR (2004). Chromosome abnormalities and genetic counseling. 3rd edn. Oxford University Press, New York.


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