2Normal karyotype Study of chromosomeskaryotyping A karyotype is the standard arrangement of a photographed, stained chromosomes pairs which are arranged in order of decreasing length
3When chromosomes are preparing to divide, the DNA replicates itself into two strands called chromatidsReplicating chromosomeThe same chromosomeunder normal conditionsTelomereCentromereThe twochromatidsTelomere
4Chromosome nomenclature Two armsp (petite) small and q (follows p in alphabet)1-22 = autosome numbersX, Y = sex chromosomes
5Cytogenetic terminology Short arm p and long arm qEach Chromosome is divided into 2 or more regionsEach region is subdivided into bands and sub-bandsTotal no of chromosomes is given first followed by sex chromosome and finally description of abnormality in ascending order.eg:47,XY,+21 ,and Xp 21.246,XY,del(16)(p11.2 p13.1)
6The normal human karyotype Somatic cells: 22 pairs of autososmes & 1 pair of sex chromosomes (46,XX or 46,XY).The normal karyotype is diploid (2 copies of each chromosome).Sperm & eggs carry 23 chromosomes & are haploid (one copy of each chromosome).
7What is the difference between an Autosome and a Sex-chromosome? Autosomes are the first 22 homologous pairs of human chromosomes that do not influence the sex of an individual.Sex Chromosomes are the 23rd pair of chromosomes that determine the sex of an individual.
8Sperm determines genotypic sex by contributing either an X or a Y chromosome during fertilization.
10Three classes of chromosome Metacentric - centromere in middleSubmetacentric - centromere distant from middleAcrocentric - centromere at end
11Uses of karyotype analysis: Genotypic sex ( identification of X & Y chromosomes).Ploidy ( euploid, aneuploid or polyploid).Chromosomal structural defects (translocation, isochromosome, deletion etc..).
12Some definitionsHaploid (n)- refers to a single set of chromosomes (23 in humans).Sperm & eggs are haploid.Diploid (2n)- refers to a double set of chromosomes (46 in humans). Somatic cells are diploid.Euploid- refers to any multiple of the haploid set of chromosomes (from n-8n)
13Polyploid- refers to any multiple of the haploid set of chromosomes> diploid (2n). Aneuploid- refers to karyotypes that do not have multiples of the haploid set of chromosomes.Monosomy- refers to an aneuploid karyotype with one missing chromosome (XO in Turner’s syndrome).Trisomy- refers to an aneuploid karyotype with one extra chromosome (trisomy 21 in Down’s syndrome))
14Aneuploidy results from the failure of chromosomes to separate normally during cell division: Meiotic Nondisjunction
154N 2N N 2N NORMAL SEPARATION NORMAL ZYGOTE First meiotic division Second meioticdivisionNGametesFertilization2NZygotesNORMAL ZYGOTE
16NONDISJUNCTION TRISOMIC ZYGOTE MONOSOMIC ZYGOTE First meiotic division Second meioticdivisionGametesFertilizationZygotesTRISOMIC ZYGOTEMONOSOMIC ZYGOTE
17Aneuploidy usually results from non-disjunction Chromosomes or chromatids fails to separateAn error of mitotic or meiotic spindle attachment to centromereMay occur in either the maternal or the paternal germ cellsMore commonly arises in the motherFrequency of non-disjunction increases with maternal age
19Six main types Deletion Ring chromosome Duplication Isochromosome Inversionparacentric & pericentricTranslocationRobertsonian & reciprocal
20Deletion Involves loss of part of a chromosome Results in monosomy of that chromosomal segmentClinical effects due toInsufficient gene productsUnmasking of mutant alleles on normal chromosomeBeforedeletionAfterdeletion
22Ring chromosome Breaks occur in both arms of a chromosome Ring chromosome Breaks occur in both arms of a chromosome. The two broken ends anneal; the two acentric fragments are lost. Results in double deletion (in p and in q).
23Epilepsy, mental retardation and craniofacial abnormalities
24Isochromosome Mirror image chromosome Loss of one arm with duplication of otherLoss of p-armDuplication of q-arm
25Inversion Two breaks in one chromosome The fragment generated rotates 180o and reinserts into the chromosomePericentric - involves p and q armParacentric - involves only one arm
26Translocation - exchange of chromosomal material between two or more chromosomes ReciprocalRobertsonianIf no essential chromosome material lost or genes damaged then the individual is clinically normalHowever, there is an increased chance of chromosomally unbalanced offspring
27Reciprocal Translocation Involves two chromosomesOne break in each chromosomeThe two chromosomes exchange broken segmentsBefore translocationAfter translocation
28Robertsonian translocation Named after W. R. B. Robertson who first identified them in grasshoppers in 1916Most common structural chromosome abnormality in humansFrequency = 1/1000 livebirthsInvolves two acrocentric chromosomesTwo typesHomologous acrocentrics involvedNon-Homologous acrocentrics involved
29+ = + = Homologous acrocentric, i.e. chromosome 14 lost+=Non-homologous acrocentric, i.e. chromosomes 14 & 21lost+=
33What is mutation?A mutation may be defined as a permanent change in the DNA.These structural DNA changes affect protein expression & function.
34Mutations affect protein synthesis Transcription: Mutated DNA will produce faulty mRNA leading to the production of a faulty protein.
35Somatic & Germ cell mutations Mutations that occur in somatic cells such as skin cells or hair are termed Somatic.Germline mutations occur only in the gametes. These mutations are more threatening because they can be passed to offspring .
36Germline mutations can be transmitted to future generations. Those that occur in somatic cells may contribute to the pathogenesis of neoplasia.Drugs, chemical & physical agents that increase the rate of mutation act as carcinogens.
37Mutagens are agents that cause mutations. They include: 1. High Temperatures2. Toxic Chemicals (pesticides, etc)3. Radiation (nuclear and solar)
38Types of mutationsChromosomal mutation: affecting whole or a part of a chromosomeGene mutation: changes to the bases in the DNA of one gene
39Major types of genetic mutations Point mutations: Single base substitutions .Frameshift mutations: base pair insertions or deletions that change the codon reading frame.Large deletions: can result in loss of gene or juxtapose genes to create a hybrid that encodes a new “fusion” protein.Expansion of trinucleotide repeats: can arise in genes that have repeated sequences. Affected patients can have 100s or 1000s of repeats (normal:10-30).
41Gene Mutations: DNA base alterations Point mutation- eg:sickle cell anemiaInsertionDeletionInversionFrame Shifts
42Point mutation - when a base is replaced with a different base. CGG CCC AAT to CGG CGC AAT Guanine for CytosineInsertion - when a base is addedCGG CCC AAT to CGG CGC CAA T Guanine is addedDeletion - the loss of a baseCGG CCC AAT to CGG CCA A T loss of Cytosine
43Frame Shift mutationsA frame shift mutation results from a base deletion or insertion. Each of these changes the triplets that follow the mutation.CGG CCC AAT to CGG CGC CAA TFrame shift mutations have greater effects than a point mutation because they involve more triplets.This in turn changes the amino acids of the protein!
44Classification of genetic disorders 1.Gross chromosomal abnormalities 2.Diseases with multifactorial inheritance3.Disorders related to mutant genes of large effect
47Trisomy 21 - Down's Syndrome - karyotype 47, XX +21 or 47, XY+21- frequency about 1 in 600 births
48Trisomy 18 - Edward's Syndrome - karyotype 47, XX +18 or 47, XY+18- frequency about 1 in 8,000births
49Trisomy 13 - Patau's Syndrome - karyotype 47, XX +13 or 47, XY+13- frequency about 1 in 10,000 births
50Sex chromosome trisomies - 47, XXY (Klinefelter Syndrome), 47,XXX, 47,XYYTriploidies of other chromosomesRareusually incompatible with life
51- Polysomy X e.g. XXXX- Frequency about 1 in 1000
52Trisomy 21(Down’s syndrome) The most common malformationIncidence: 1 per 660 live births, closely related to maternal ageMother’s age<30 year risk:1 per 5000Mother’s age>35 year risk:1 per 250
53Clinical findings Flattened face Mental retardation Congenital heart disease:50%endocardial cushion,ASD,AV malformation,VSD10 to 20 fold increased risk of developing leukemiaInfection are commonPremature agingall patints older than 40 will have Alzheimer disease(degenerative disorder of brain)Musculoskeletal problems
67Cytogenetic diorders involving sex chromosomes They cause chronic problems relating to sexual development and fertilityThey are often difficult to diagnose at birth,and many are recognised at the time of pubertyHigher the number of x chromosomes, greater the likelihood of mental retardation
68Lyon hypothesis:In somatic cells of a female only one of the X chromosomes is activeX-inactivationOccurs early in embryonic lifeIs randomeither paternal or maternal XIs completeIs permanentIs clonally propagated through mitosisMary Lyon
69Y chromosomeRegardless of the number of X chromosomes, the presence of single Y determines male sexThe gene that indicates testicular development is sry gene (sex determining region Y gene)Located on distal arm of Y chromosome
70Turner syndrome Partial monosomy of X chromosome Hypogonadism in phenotypic femalesKaryotype:45,XMosaic patients with 45,X /46,XXCystic hygromas, Congenital heart disease (coarctation of aorta and bicuspid aortic valve), failure to develop secondary sexual charactersticsMental status is usually normal
74Klinefelter syndrome 47,XXY Results from meiotic nondisjunction The discovery of the karyotype of Klinefelter was the first demonstration that sex in humans is determined by the presence of the Y rather than the number of X chromosomesMale hypogonadism
76Klinefelter syndrome Lower IQ than sibs Tall stature Poor muscle tone Reduced secondarysexual characteristicsGynaecomastia(male breasts)Small testes/infertility
77Plasma gonadotropin levels( FSH) and estrodiol is elevated Testosterone levels are decreasedTesticular tubules are totally atrophiedSome shows primitive tubules
78HermaphroditismGenetic sex is determined by the presence or absence of Y chromosomeGonadal sex is based on histological characteristics of gonadsPhenotypic sex is based on the appearance of external genitalia
79True hermaphrodite implies the presence of both ovarian and testicular tissue Pseudohermaphrodite represents disagreement between the phenotypic and gonadal sex (eg:female pseudohermophrodite has ovaries but male external genitalia)
80Transmisson patterns of single gene disorders Autosomal dominantAutosomal recessiveX-linked
81Autosomal TraitsGenes located on Autosomes control Autosomal traits and disorders.2 Types of Traits:Autosomal DominantAutosomal Recessive
82Autosomal Dominant Traits If dominant allele is present on the autosome, then the individual will express the trait.A = dominant a = recessiveWhat would be the genotype of an individual with an autosomal dominant trait?AA and Aa (Heterozygotes are affected)
83Autosomal Dominant Inheritance Are manifested in heterozygous stateOne parent of an index case is usually affectedBoth males and females are affected and both can transmit the condition50% chance of affected heterozygote passing gene to childrenA new mutation in the gene resulting in the offspring being first affected and then may be inherited in a dominant fashionDominant genes may exhibit lack of penetrance, which is an all or none phenomenon; either the gene is expressed or not expressedMay show variable expressivity with different family members showing different manifestations of the trait
87Autosomal Recessive Traits If dominant allele is present on the autosome, then the individual will not express the trait.In order to express the trait, two recessive alleles must be present.
88A = dominant a = recessive What would be the genotype of an individual with an autosomal recessive trait?aaWhat would be the genotype of an individual without the autosomal recessive trait?AA or AaAa – called a Carrier because they carry the recessive allele and can pass it on to offspring, but they do not express the trait.
89Autosomal Recessive Traits Heterozygotes are Carriers with a normal phenotype.Most affected children have normal parents. (Aa x Aa)Two affected parents will always produce an affected child. (aa x aa)Close relatives who reproduce are more likely to have affected children.Both males and females are affected with equal frequency.Pedigrees show both male and female carriers.Complete penetrance is commonOnset is early in life
91X-Linked InheritanceInvolves particular genes located on the X chromosomeDisorders more commonly affect malesHeterozygote female will pass the gene to 50% of her sons who will express the trait, and to 50% of her daughters who will be carriers for the traitAffected males pass the gene to all of their daughters and none of their sonsHallmark is absence of male to male transmission
95Single-Gene “Mendelian” Disorders Structural proteins–Osteogenesis imperfecta and Ehlers-Danlos(collagens);Marfan syndrome (fibrillin);Duchenne and Becker muscular dystrophies (dystrophin)Enzymes and inhibitorsLysosomalstorage diseases;PKU (phenylalanine hydroxylase);Alpha-1 antitrypsin deficiencyReceptorsFamilial hypercholesterolemia (LDL receptor)Cell growth regulationNeurofibromatosis type I (neurofibromin);Hereditary retinoblastoma (Rb)TransportersCystic fibrosis (CFTR);Sickle cell disease (Hb);Thalassemias(Hb)
96Marfan syndrome (defect in the structural proteins) Is a disorder of connective tissues, manifested by changes in skeleton,eyes and cardiovascular systemAutosomal dominant
97PathogenesisMarfan syndrome results from inherited defect in extracellular glycoprotein –fibrillin-1Fibrillin is the major component microfibrilsThese fibrils form a basement on which tropoelastin is deposited to form elastic fibersMicrofibrils are abundant in aorta, ligaments,and ciliary zonules of lensMutations of FBN1 are mapped on the chromosome 15q21.
98MorphologyCardiovascular System: Dilatation of ascending aorta due to cystic medial necrosis, mitral vale insufficiency,aortic dissectionEyes: Dislocation of lens (usually outward and upward) called as ectopia lentis, severe myopia
99Musculoskeletal: exceptionally tall with long extremities and tapering fingers and toes The ratio of upper segment to the lower segment of the body is lower than normalJoint ligaments of hands and feet are lax;typically thumb can be hyperextended back to the wristThe head is dolicocephlic(long headed) with bossing of frontal eminencesPectus excavatum deformity, scoliosis
102Subluxation of the lens Marfan SyndromeSubluxation of the lens
103Ehlers-Danlos Syndrome A family of disorders with defect in synthesis and structure of fibrillar collagen characterized by hyperextensibility of skin, joint hypermobility, early bruisabilityMode of inheritence show all three types of Mendelian patternsOrthopaedic problems: joint instability, joint laxity, arthralgia and scoliosis
107Lysosomal storage disorders(defects in enzymes) Key component of intracellular “digestive” tractComposed of acid hydrolases that catalyse the breakdown of macromoleculesInherited deficiency-catabolism of macromolecules is incomplete accumulation of partially degraded macromoleculescell organelles become largelysosomal storage disease
109Cause of Tay-SachsThe absence of a vital enzyme called Hexosamindase A (Hex-A)Absence ofAccumulation of GM2 in neuronsHex-AThis syndrome is named for Warren Tay an ophthalmologist who first described a patient with symptoms and Bernard Sach, a neurologist, who described cellular changes caused by the disease a few years later. Tay-Sachs is caused by the absence of Hexosamindase A (Hex-A). Usually, this enzyme causes a fatty substance called GM2 ganglioside to accumulate in cells, especially in the nerve cells of the brain. However, the absence of Hex-A this does not happen leading several developmental problems. (1,2)Involvement of CNS, ANS and retina common
110Gene Location Chromosome 15 showing location of the syndrome The gene that controls the production of Hex-A is found on Chromosome 15. The picture above shows its location. (3)
111Characteristics Birth: Appear normal 6 months: Development slows 2 years: Seizures and deteriorating mental functions3 years: Blindness, mentally retardation, paralysis and non-responsiveness.Cherry red spot in the maculaCommon in JewsAt birth, babies affected with Tay-Sachs appear normal. After about six months, their development begins to slow. By two years of age, child are often affected by seizures and have fading mental functions. The syndrome continues to progress until blindness, mental retardation, paralysis and overall non-responsiveness result. Children usually die within the first four years of life. (1,2,7)
112Microscopy: neurons are ballooned with cytoplasmic vacuoles having lysosomes filled with gangliosidesEM: Whorled configuration with lysosomes composed of “onion skin” layer of membranes
114Detection Methods: Amniocentesis Chorionic villus sampling Blood samples to detect carriersAmniocentesis: Amniocentesis is done by removing and testing a small quantity of the fluid that surrounds the fetus in the uterus. This procedure is done at approximately the 16th week of pregnancy.Chorionic villus sampling: Choronic villus sampling is performed by the 10th week and usually provides a test answer much sooner than amniocentesis. The cell sample is obtained by withdrawing a small bit of the developing afterbirth.Blood samples to detect carriers: Bloods samples can be collected and tested for carriers by looking at the gene that is responsible for the production of Hex-A on chromosome 15. (7)
115In SummaryTay-Sachs is a genetic disorder that causes Hex-A, an enzyme important to the function of nerve cells, not to be produced.Babies with Tay-Sachs often appear normal at birth, but develop severe symptoms in the first few years of life.There is genetic counseling as well as support groups available for carriers of Tay-Sachs or parents with an affected child.
116Niemann –pick disease (type A and B) Deficiency of sphingomyelinase accumulation of sphingomyelin
117Type AMore severe infantile form with extensive neurological involvementMarked visceral accumulation of sphingomyelinProgressive wasting and early death within 3 yearsCherry red spot in the macula
118Type B Patients have organomegaly but no CNS involvement Survive to adulthood.
119Morphology Accumulation of sphingomyelin in mononuclear phagocytes Affected cell become largeInnumerable small vacuoles of uniform sizeimparting foaminess to the cytoplasmVacuoles stain for fat
120Phagocytic foam cells widely distributed in spleen ,liver, lymph node, bone marrow, tonsils, g.i.t,lungsBrain: Gyri shrunkened,sulci widened with vacuolation and ballooning of neuronsEM: zebra bodies
121Clinical Features: Evident by 6 months Protuberant abdomenFailure to thrive,vomiting,fever,Deterioration of psychomotor functionDeath by 2 yrs
122Gaucher disease Autosomal recessive Mutation in the gene encoding glucocerebrosidaseMost commonGlucocerebroside accumulates in phagocytic cells3 types
123Type I: Chronic non-neuronopathic form Storage limited to mononuclear phagocytes throughout the bodySplenic and skeletal involvement commonType II: acute neuronopathic ,dominated by CNS involvement,death by 2 yearsTypeIII: intermediate between I and II, progressive CNS involvement
124Morphology Glucocerebrosides accumulates in phagocytic cells Distended phagocytic cells (Gaucher)cells found in spleen liver,BM,LN,TONSILS,thymus and peyer patchesCells have fibrillary pattern instead of vacuolated (crumpled tissue paper )and have eccentrically placed nucleus.
125Gaucher cells (Phagocytic cells with a “crumpled tissue paper” appearance)
128Phenylketonuria Autosomal recessive disorder Deficiency of phenylalanine hydroxylase hyperphenylalaninemiaCommon in scandinavian peopleNormal at birthBy 6 months severe mental retardationSeizures,decreased pigmentation of hair and skinMental retardation can be avoided by restriction of phenylalanine intake early in life
129Galactosemia Autosomal recessive disorder Deficiency of galactose -1-phosphate uridyl transferaseGalactose -1-phosphate accumulates in liver, spleen, kidneys, lens of eye, cerebral corexAlternative metabolic pathways activated, leading to the production of galacitol
130Clinical features Failure to thrive Vomiting, diarrhea Hepatomegaly Opacification of lens (cataracts)Aminoaciduria
131Diagnosis can be suspected by demonstration in the urine of reducing sugars Many morphological changes can be prevented by early removal of galactose from diet
132Oochronosis (alkaptonuria) First human inborn error of metabolism to be discoveredAutosomal recessiveLack of homogentisic oxidase blocks metabolism of phenylalanine-tyrosine at the level of homogentisic acidHomogentisic acid accumulates in the bodyLarge amount is excreted,imparting a black color to the urine if allowed to stand
133MorphologyThe retained homogentisic acid selectively binds to collagen in connective tissues, tendons ,cartillage imparting blue black pigmentationMost evident in the ears, nose and cheeks.Wear and tear erosion of abnormal cartilage leads to denudation of subchondral bonedegenerative arthropathy
134Mucopolysaccharidoses Result from genetic deficiency of enzymes involved in the degradation of mucopolysaccharidesProgressive disorder chacterised by involvement of multiple organs like liver,spleen, heart and blood vesselsMost are associated with coarse facial features,joint stiffness and mental retardationThe accumulated mucopolysaccharides are grnerally found in mononuclear phagocytic cells,endothelial cells,smooth muscle cells and fibroblsts.
135Glycogen storage diseases Hereditary deficiency of one of the enzymes involved in the synthesis and breakdown of glycogen.Hepatic form: an inherited deficiency of hepatic enzymes involved in glycogen metabolism leads to storage of glycogen in liver and also hypoglycemia.eg :deficiency of glucose-6-phosphatase
136Myopathic form:in muscles glycogen is mainly used as a source of energy If the enzymes that fuel glycolytic pathway are deficient, glycogen storage occurs in muscles.eg: deficiency of muscle phosphofructokinase, muscle phosphorylase