Autosomal Recessive Disorder

Presented By:Aamir Sharif

M.Phil Human Pathology

Autosomal Recessive Disorder

• Largest category of Mendelian disorder

• Usually does not affect the parent of the affected individual,

but sibling may show the disease.

• Complete penetrance is common.

• Onset is frequently early in life.

• NEW mutations rarely detected clinically

• Usually affect enzymatic proteins (show LOSS of

FUNCTION)

Pattern Of Inheritance:

Typical pattern is two heterozygous unaffected (carrier) parent.

• The triat does not usually affect the parent, but siblings may show the disease

• Siblings have one chance in four of being affected

Both sexes affected equally.

Autosomal Recessive Disorder

MetabolicCystic fibrosis , Phenylketonuria Galactosemia , HomocystinuriaGlycogen storage disease

Hematopoietic Sickle cell anaemiaThalassemias

Endocrine Congenital Adrenal hyperplasia

Skeletal Alkaptonuria

Nervous Friedrich ataxiaSpinal muscular atrophy

Cystic Fibrosis (Mucoviscidosis)

• Autosomal recessive• Most common lethal genetic disease affecting

Caucasians (1 in 3,200 live births in the USA)– 2-4% of population are carriers– Uncommon in Asians and African-Americans

• Widespread disorder in epithelial chloride transport affecting fluid secretion in

–exocrine glands– epithelial lining of the respiratory, gastrointestinal,

and reproductive tracts• Abnormally viscid mucus secretions

Pathogenesis

• Primary defects in CF is abnormal function of an epithelial choloride channel protein encoded by the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene at chromosome no 7.

CFTR Gene: Normal

• Cystic Fibrosis Transmembrane Conductance Regulator (CFTR)

• CTFR → epithelial chloride channel protein– agonist induced regulation of the chloride channel– interacts with epithelial sodium channels (ENaC)

• Sweat gland– CTFR activation increases luminal Cl− resorption– ENaC increases Na+ resorption– sweat is hypotonic

• Respiratory and Intestinal epithelium– CTFR activation increases active luminal secretion of chloride– ENaC is inhibited

CFTR Gene: Cystic Fibrosis

• Sweat gland– CTFR absence decreases luminal Cl− resorption– ENaC decreases Na+ resorption– sweat is hypertonic

• Respiratory and Intestinal epithelium– CTFR absence decreases active luminal secretion of chloride– lack of inhibition of ENaC is opens sodium channel with

active resorption of luminal sodium– secretions are decreased but isotonic

Chloride Channel Defect and Effects

• Chloride channel defect in the sweat duct (top) causes increased chloride and sodium concentration in sweat.

• In the airway (bottom), cystic fibrosis patients have decreased chloride secretion and increased sodium and water reabsorption leading to dehydration of the mucus layer coating epithelial cells, defective mucociliary action, and mucus plugging of airways predispose to recurrent pulmonry function.

CFTR Gene: Mutational Spectra

• More than 800 mutations are known• These are grouped into six classes

– mild to severe– In severe case there is complete loss of

CFTR– Occurs in 70% of CF chromosomes– 3 base pair deletion leading to absence of

phenylalanine at position 508 (DF508) of the CF transmembrane conductance regulator (CFTR)

Genetics of CF

• DF508 mutation leads to improper processing and intracellular degradation of the CFTR protein

• Other mutations in the CF gene produce fully processed CFTR proteins that are either non-functional or partially functional

Morphology

• Plugging of ducts with viscous mucus and loss of ciliary function of respiratory mucosa

• Pancreas– atrophy of exocrine pancreas with fibrosis– islets are not affected

• Liver– plugging of bile canaliculi with portal inflamation– biliary cirrhosis may develop

• Genitalia– Absence of vas deferens and azoospermia

• Sweat glands– normal histology

Pancreas in Cystic Fibrosis.

Note that the ducts contain inspisated material and the acini are atrophic and the stroma exhibits fibrosis and chronic inflamation. The islets are preserved. Normal pancreas for comparison

Lung Pathology in CF

• More than 95% of CF patients die of complications resulting from lung infection

• Viscous bronchial mucus with obstruction and secondary infection– S. aureus– Pseudomonas– Hemophilus

• Bronchiectasis– dilatation of bronchial lumina– scarring of bronchial wall

• Lungs of a patient dying of cystic fibrosis.

• There is extensive mucus plugging and dilation of the tracheobronchial tree. The pulmonary parenchyma is consolidated by a combination of both secretions and pneumonia—the green color associated with Pseudomonas infections.

Manifestations

• Common presentations– Chronic cough– Recurrent pulmonary infiltrates– Failure to thrive – Meconium ileus

Manifestations

• Respiratory tract– Chronic sinusitis

• Nasal obstruction• Rhinorrhea• Nasal polyps in 25%; often requires surgery

– Chronic cough• Persistent• Viscous, purulent, green sputum

CF Diagnosis

• Clinical criteria• One are more characteristic phenotypic features OR

history of cystic fibrosis in siblings or positive new born screening test result

• Or inc Sweat chloride concentration on two or more occasion

• DNA Analysis– gene sequencing

PHENYLKETONURIA (PKU)• Ethnic distribution

– common in persons of Scandinavian descent – uncommon in persons of African-American and Jewish descent

• Autosomal recessive• 98 % cases of PKU is due to Phenylalanine hydroxylase

deficiency and 2 % arise from abnormalities in synthesis or recycling of the cofactor tetrahydrobiopterin.

• Affected patients are normal at birth but within a week exhibt a rising plasma phenylalanine level.

• At 6 months less than 4 % untreated patients show mental retardation.

• About one 3rd are never able to walk and two 3rd talk.• Decreased pigmentation of hair and skin often accompany

metal retardation.

Pathogenesis & C/F

• Normally 50% of dietary intake of phenylalanine is necessary for protein synthesis and rest is converted into tyrosine by PAH.

• When phenylalanine metabolism is blocked because of a lack of PAH enzyme, minor shunt pathways come into play, yielding several intermediates that are excreted in urine and sweat that impart a strong musty or mousy odor to affected infants.

• Excess phenylalanine and its metabolites contribute to the brain damage in PKU.

• Concomitant lack of tyrosine, a precursor of melanin is responsible for the light color of skin and hair.

Diagnosis

• Approximately 500 mutant alleles of PAH gene have been identified, only some of which cause a severe deficiency of the enzyme.

• Those with 6% residual activity present with milder disease.

• Some mutation results in modest elevations of blood phenylalanine without associated neurological damage (benign hyper phenylalaninemia).

• Serum phenylalanine levels differentiate BHP from PKU.

GALACTOSEMIA• Autosomal recessive disorder affects 1 in 60,000 live-born

infants.• Normally Lactose → glucose + galactose• Galactose-1-phosphate uridyl transferase (GALT)

– GALT is involved in the first step in the transformation of galactose to glucose

– Lack of this enzyme due to homozygous mutation in the encoding gene GALT → galactosemia

– As a result of this transferase deficiency, glactose-1-phosphate,and galactitol accumulate in manny tissues,including liver, spleen, lens of eyes, kidney and cereberal cortex.

Pathogenesis

• Fatty changes Hepatomegally• Glactititol swells in lens Opacity of lens Cataract• CNS Loss of nerve cell, Gliosis, edema.• Infants affected by galactosemia typically present

with symptoms of lethargy, vomiting, diarrhea, failure to thrive, and within a few days of milk ingestion.

• Jaundice and hepatomegally in first week• Accumulation of metabolites in kidney impair A.acid

transport resulting in aminoacidurea.• E.coli septicemia occur frequently.

• Presences of reducing sugar other than glucose in urine

• Transferase dificiency in leukocyte and RBC• Antenal GALT activity assay in cultured amniotic fluid

cells• Or Galactilol level in amniotic fluid supernatant.

Lysomal storage disease

• Lysosome breakdown complex substrates like sphingoloipids , mucopolysaccharides into soluble end products.

• Inherited deficiency of lysosomal enzyme, catabolism of its substrate remain incomplete, leading to accumulation of partially degraded metabolites into lysosomes.

• Approximately 40 lysomal storage diseases are identified that shows certain common features.Autosomal recessive transmissionPt. population consisting of infants and young children.Storage of insoluble intermediates in the mononuclear

phagocyte system, giving rise to hepatospleenomegaly.Cellular dysfunction not only by undigested material but also by

activation of macrophages and release of cytokines.

Tay-Sachs Disease

• It is characterized by accumulation of gangliosides principally in the brain as a result of a mutation in and consequent deficiency of the β subunit of the hexosaminidase A.

• Based on gangliosides these disorders are sub classified into GM₁ and GM₂

• GM₂ is the most common gangliosides.• More than 100 mutations have been described; most affect protein

folding or intracellular transport.• The brain is principally affected because it is involved in gangliosides

metabolism.• GM₂ storage occur throughout the CNS.

• The molecular basis of neuronal injury is not fully understood.

• Because in many cases the mutant protein is misfolded, so called “unfolded protien” response.

• If such misfolded protein are not stabilized by chaperones they triggers appoptosis.

Morphology

• Affected cell appear swollen and sometimes foamy.• Electron microscopy reveals whorled configuration

within lysosomes composed of onion-skin layers of membranes throughout the CNS.

• In Retina there is pallor produced by swollen ganglion cells in the peripheral retina result in contrasting “cherry red” spot in the relatively unaffected central macula.

• In the most common acute infantile variant of Tay-Sach disease infants appear normal at birth but motor weakness begin at 3 to 6 month of age followed by neurological impairment, onset of blindness and progressively more severe neurological dysfunction.

• Death occur within 2 or 3 years.• Level of Hexosaminidase in the serum or DNA analysis of

Heterozygote carriers has a diagnostic importance

Niemann-Pick Disease Type A and B

• Niemann-Pick Disease Type A and B diseases are characterized by a primary deficiency of a sphingo-myelinase and the resultant accumulation of sphingomyelin.

• In type A severe deficiency of sphingo-myelinase, the break down of sphingomyelin into ceramide and phosporylcholine is impaired and excess sphingomyelin accumulates in all phagocytic cell and in the neuron.

• Macrophages become stuffed with droplets of the complex lipid, imparting a fine vacculation or foaminess to the cytoplasm.

Morphology

• Electron microscopy confirms that vacuoles are engorged secondary lysosomes that often contain cytoplasmic bodies resembling concentric lamellated myelin figures, sometimes called “zebra ” bodies.

• Because of their high content of phagocytic cells, the organ most severely affected are spleen, liver, bone marrow, Lymph nodes and lungs.

• That lead to visceromegaly and severe neurologic deterioration but in type 2 only visceromegaly.

• Death occur within first 3 year of life.• Estimation of Sphingomyelinase activity in the leukocytes or cultured

fibroblast can be used for diagnosis of suspected case and carriers.• Antenatal diagnosis is possible by enzyme assay or DNA probe

analysis.

Niemann-Pick Disease Type C

• It is more common than Type A and B.• Mutation in 2 related gene NPC1 and NPC2• NPC1 is being responsible for a majority of cases.• Niemann-Pick Disease Type C is due to primary defect

in lipid transport that lead to accumulation of cholesterol and gangliosides GM₁ and GM₂ in the affected cells.

• NPC is clinically hetrogeneouses• The most common manifestation are: ataxia, dystonia,

vertical supra nuclear gaze, dysarthria, and psychomotor regression.

Gaucher Disease

• Normally Glycolipids derived from the breakdown of senescent blood cell are degraded in liver , spleen, and bone marrow into ceramide and then into glucose by glucocerebrosidases .

• Deficient glucocerebrosidases lead to accumulation of glucocerebrosides in mononuclear phagocytic cells that result in enlargment of Phagocytes “Gaucher cells” upto 1micrometer.

• Accumulation of distended lysosmes gave cytoplasm a characterized appearance “wrinkled tissu paper”

• No distnict vacuolation is present.• This disease is caused not just by the burden of storage material

but also by activation of the macrophages and derived cytokines.

• 3 Autosomal recessive variants of Gaucher disease• Type 1 also called chronic non neuronopathic form account for

99% of cases and characterized by• Clinical or radiographic bone involvement 70-100%(osteopenia, focal lytic lesion and osteonecrosis)• Hepatospleenomegaly and abscence of CNS involvement while in

Type II and Type III neurologic manifestation (convulsion, progressive deterioration) appear during infancy and in adulthood

• Gaucher cells are found in the liver , spleen, lymph nodes and bone marrow

• Estimation of glucocerebrosides level in the leukocytes or cultured fibroblast can be used for diagnosis of suspected case and carriers.

Mucopolysaccharidoses

• Mucopolysaccharides form a part of ground substance and are synthesized by connective tissue fibroblasts.

• Certain fraction is degraded within lysosomes.• The lack of lysosomal enzymes leads to accumulation of

mucopolysaccharides in lysosomes.• Several clinical variants of MPS classified numerically from MPS I to

MPS VII.• Mucopolysaccharides that accumulate in tissues are Dermatan

sulphate, heparaan sulphate, keratan sulphate and Chondroitin sulphate.

• Hepatospleenomegaly, skeltal deformities, lesion of heart valves and sub endothelial arterial deposits and lesion in brain are common in all MPS.

Type I Huler syndrome

• It is caused by a deficiency of alpha-L-iduronidase• Children have a life expectancy of 6-10 yr and death is often

due to cardiac complications.• Accumulation of dermatan sulphate and heparan sulphate

is seen in cells of mononuclear phagocyte system, in fibroblast, within endothelium and smooth muscle cells of vascular wall.

• The affected cell are swollen and have clear cytoplasm, resulting from accumulation of material positive for periodic acid-Schiff staining within engorged, vaculated lysosomes.

• Lysosomal inclusions are also found in neurons accounting for mental retardation

MPS type II or Hunter Syndrome

• Differ from Hurler syndroms in its mode of of inheritance (X-linked), the absence of corneal clouding and often milder in clinical course.

• Accumulation of dermatan sulphate and heparan sulphate from deficiency of L-iduronate sulpatase.

Glycogen Storage Diseases (Glycogenosis)

Clinicopathological Catageory

Specific type Enzyme deficiency

Morphologic changes Clinical Features

Hepatic Type Hepatorenal( von Gierke disease)

Glucose-6-phosphatase

Hepato spleenomegally :intra cytoplasmic accumulation of glycogen small amount of lipidsRenomegally:intra cytoplasmic accumulation of glycogen in cortical tubules

In untreated pt.HepatomegalyHypoglycemiaHyperlipidemiaHyperuricemiaBleeding tedencyWith treatment:Patients surive and develop hepatic adenomas

Myopathic Type

McArdle Syndrome(type v)

Muscle phosphorylase

Glycogen in subsarcolemmal location in skelton muscle

Painfull cramps after sternous exercisesNo rise in lactate level in venous bloodMyoglobinurea in 50% cases

Miscellaneous Type

Generalized glycogenosis(pompe disease type II)

Lysosomal glucosidase (acid maltase)

Cardiac and hepatomegalyLacy cytoplasmic pattern(due to glycogen deposition)

Massive cardiomegallyMuscle hypotoniaCardiorespiratory failure before 2 yr

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ss

SICKLE CELL

ANAEMIA

Definition and Molecular Basis of Disease

• Sickle cell disease (SCD): a recessively inherited chronic hemolytic anemia

• Caused by a single nucleotide substitution in the β globin gene on chromosome 11 – Hemoglobin S (most common): GTG GAG results in

substitution of valine (hydrophobic) for glutamate (hydrophilic)

– Many other variant hemoglobins are described

• Mutant hemoglobin polymerizes under low oxygen conditions and form bundles that distort red cells into the classic sickle shape

Prognosis

• Over the past 30 years in US/Europe, median survival has increased from 14yrs to 45-55yrs for SS disease

• Figures not available for Africa but estimated 50% of affected die before 5yrs

• WHO estimates that SCD complicates up to 9% of under 5 deaths in West Africa

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Normally, humans have

Of these, Haemoglobin A makes up around 96-97% of the normal haemoglobin in humans.

Haemoglobin

A two alpha two beta

A2 two alpha two delta

F two alpha two gamma

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Common types of Sickle Cell DisordersType of anaemia

Hemoglobin variation

comment

Sickle Cell Anemia

Sickle haemoglobin (HbS) + Sickle haemoglobin

Most Severe – No HbA

Hemoglobin S-Beta thalassemia

Sickle haemoglobin (HbS) + reduced HbA

Mild form of Sickle Cell Disorder

Hemoglobin S-C disease

Sickle haemoglobin (HbS) + (HbC)

Mild form of Sickle Cell Disorder

Sickle Cell Trait

Sickle haemoglobin (S) + Normal haemoglobin (A)

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Pathophysiology

Is caused by • point mutation in the β-globin chain of Hb glutamic (hydrophobic amino acid)

valine( hydrophilic )

• at the 6th position

Life span • RBC 90–120 days• sickle cells 10–20 days.

• exposure to P O2 < 40 mmHg for 2 to 4 minutes • polymerization of Hb• The initiation of polymerization may be -

incomplete and -reversible , if re-oxygenation occurs early in the

process. • Repetitive exposure to alternating de-oxygenated and

oxygenated states lead to -membrane distortion,

-irreversible sickling.

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Pathophysiology

Deoxygenation

polymerization of hemoglobin

sickling of red cells

endothelial damage/activation

RBC and leukocyte adhesion to endothelium, vasoconstriction

vascular occlusion, organ ischemia and end-organ damage

OXY-STATE DEOXY-STATE

Initial Clinical Presentation

• Typically presents in infancy after 6 months of age, when Hb F is waning

– Birth hemoglobin F: α2γ2– Hemoglobin A: α2β2– Hemoglobin S: α2S2

• Pain and anemia are hallmarks of disease

Initial Clinical Presentation

• Suggestive historical findings:

– Family history of known SCD– Family history of sudden death in young child– Frequent pain– Frequent chest infections– Failure to gain weight despite good nutrition– Persistent jaundice– Classic sequelae (hand/foot syndrome, priapism)

Initial Clinical Presentation

• Physical Exam findings are nonspecific:

– scleral icterus– pale mucous membranes– systolic murmur throughout precordium– splenomegaly

Initial Clinical Presentation

• Predictors of adverse outcome at presentation:

– Dactylitis (iinflammation of an entire digits) in infant <1y/o

– Hb<7g/dL

– leukocytosis absent infection

– priapism

Sequelae: Vaso-occlusive/pain crisis

• Occurs in 60% of SS patients when vaso-occlusion tissue ischemia

• May be triggered by infection, temperature extremes, dehydration or stress but usually w/o identifiable cause.

• Characterized by severe pain often in extremities, involving the long bones, or the abdomen. May last hours to days.

• Number of pain crises/year varies widely between individuals with some patients w/ constant low level pain.

Sequelae: Infection

• By age 1 30% of Hb SS pts are asplenic, by age 6, 90% are asplenic due to microinfarcts.

• This makes children especially vulnerable to infection/sepsis with encapsulated organisms, esp. Strep pneumoniae (400x higher risk vs. general population)

• Sickle Cell patients are also more susceptible to osteomyelitis (Salmonella and Staph spp.)

Sequelae: Acute Chest Syndrome

• Characterized by new respiratory distress, CXR infiltrate, hypoxia(O2<95%) and/or chest pain

• Occurs in 40% of patients with SS disease• Can progress rapidly to ARDS• May be caused by viral or bacterial infection,

fat embolism(2/2 bone marrow infarction), cause unknown in most cases

Sequelae: Stroke

• 11% of SS patients have a stroke by age 20 with peak incidence between 2 and 10

• Presents as focal neurologic deficit or seizure

Sequelae:Acute Splenic Sequestration

• Sudden enlargement of the spleen accompanied by a >2g/dL decrease in Hb from baseline, often w/ thrombocytopenia

• Occurs in children <3y/o• Can cause sudden circulatory collapse

Sequelae: Aplastic Crisis

• Caused by infection with Parvovirus B19 (fifth disease) which invades young erythroblasts in bone marrow

• Often presents with fever, URI sx and drop in Hb• RBC life expectancy in SS disease is 10-20 days, thus

decrease in RBC production has profound effect.• Bone marrow recovery typically within 7-10 days

Sequelae: Anemia

• Compensated anemia at baseline• Baseline Hb normally 8-9 g/dl• Overtransfusion can predispose to transfusion

transmitted infections and iron overload

Other Sequelae

• Priapism• Dactylitis (hand-foot syndrome): painful swelling of

hands and feet which occurs in infants.• Avascular necrosis of the humeral/femoral head• Cholelithiasis• Retinopathy• Chronic leg ulcers

Laboratory investigations

Complete blood count

Level of Hb -: 6–8 g/dL (Normal range-: Male=13.5-17.5g/dl Female=11.5-15.5g/dl) High reticulocyte count (10–20%).

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Blood film The blood film is microcytic and hypocromic

Sickled cell anaemia Normal

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Sickle solubility test

Mixture of Hb S in a reducing solution

Gives a turbid appearance

(Precipitation of Hb S)

Normal Hb gives a clear solution

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Hb electrophoresis • To confirm the diagnosis. • There is, no Hb A 80–95% Hb SS 2–20% Hb F

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Results of laboratory examination in sickle cell anaemiaLaboratory examinations

results Values in this disease

Values in health

WBC count increased 10000 -30000 5000 -10000

RBC count decreased 1 -4 million/mm3 4 -6 million/mm3

Hb count decreased 6 -8g/100ml Male=13.5-17.5g/dlFemale=11.5-15.5g/dl

Haematocrit reading decreased 10 -30% 45%

Reticulocyte count increased 10-40% 1-2%

Hb electrophoresis positive HbS & HbF HbA

Urine analysis Albumin casts positive negative

Serum bilirubin increased 1-3 mg/100ml 0.2-0.8mg/100ml

Platelet count increased 40000-50000/mm3 150000-400000/mm3

Bone marrow exanination

Increased red cells 40-70% 8-30%

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Tests to detect sickle cell genes before birth

Diagnosed in an unborn baby

Sampling amniotic fluid

Look for the sickle cell gene

Fluorescent in situ hybridization

Fluorescent in situ hybridization

• Fluorescent in situ hybridization, also known as fluorescence in situ hybridization, is more commonly referred to as FISH.

• It is a technique used to detect the presence or absence and location of specific gene sequences

• Can visualize specific cytogenetic abnormalities (copy number aberrations) – chromosomal deletion, amplification,

translocation

• FISH is fast and accurate at finding it.• It can also be performed even if cells are not actively

dividing, and it provides more specific information about abnormalities in chromosomes.

• More conventional techniques, such as karyotyping, simply tell investigators the number and size of chromosomes within a cell.

• Such analysis can be performed on prenatal samples (amniocentesis, chronic villus biopsy, umbilical cord blood ) , peripheral blood lymphocytes.

How does FISH work?

• FISH identify where a particular gene falls within an individual's chromosomes.

• The first step is to prepare short sequences of single-stranded DNA that match a portion of the gene the researcher is looking for. These are called probes.

• The next step is to label these probes by attaching one of a number of colors of fluorescent dye.

• DNA is composed of two strands of complementary molecules that bind to each other like chemical magnets.

• Since the researchers' probes are single-stranded, they are able to bind to the complementary strand of DNA, wherever it may reside on a person's chromosomes.

• When a probe binds to a chromosome, its fluorescent tag provides a way for researchers to see its location.

Interpretation of FISH

• Each fluorescently labeled probe that hybridizes to a cell nucleus in the tissue of interest will appear as a distinct fluorescent dot– Diploid nuclei will have two dots– If there is duplication in the region of interest, the gain

will result in more than two dots– If there is a loss in the region of interest, one or zero dot

will result