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Title of Thesis

Muhammad Arshad Rafiq
Institute/University/Department Details
Department of Biological Sciences/ Quaid-i-Azam University, Islamabad
Number of Pages
Keywords (Extracted from title, table of contents and abstract of thesis)
inherited ectodermal dysplasias, alopecias, ed, ed-malformation syndrome, ed syndrome, ectodermal dysplasias, ectodermal appendages, edpl, x-linked hypohidrotic ectodermal dysplasia, autosomal recessive ectodermal dysplasias, nail dysplasia, , onycholysis, anonychia, homozygosity, envoplakin gene, desmoglein genes, desmocolin genes, glycoproteins, desmosomes

One of the most exciting areas of cutaneous research has focused on the biology of skin appendages and particularly of the hair follicle because of its involvement in a variety of skin disorders and its increasingly recognized role in epidermal homeostasis, wound healing, and tumorigenesis. Discovery of genes directly implicated in the pathogenesis of skin diseases have far-reaching implications for basic science and, importantly, for affected individuals. The study presented here include genetic mapping of two of the hereditary skin disorders: Ectodermal Dysplasia and Alopecia.

Ectodermal Dysplasia (ED) represents a heterogeneous group of genetic disorders that are distinguished by the impaired development of two or more ectodermal appendages, hair, teeth, nails and sweat glands. A condition characterized by only ectodermal signs is called a pure ED; if it combines ectodermal signs and malformations, it is termed an ED/malformation syndrome or an ED syndrome. More than 170 different pathological, clinical, and molecular conditions have been recognized and defined as ectodermal dysplasias (ED). Some complex phenotypes are now explained in the light of the molecular causative defects involving widely expressed proteins with essential roles in control of cell cycle regulation, in replication and/or DNA repair.

There are several forms of inherited alopecia or hair loss, showing extensive variation in age of onset and severity. However, only a small number of causative genes for alopecias have been identified and/or localized. Genetic conditions affecting hair structure or hair growth cycling may be isolated or they may be a component of complex syndromes with associated abnormalities of other ectodermal appendages.

For the present study, three families with hereditary ectodermal dysplasias and three with alopecias were ascertained from different parts of Pakistan. One of the families EDPl showed X-linked hypohidrotic ectodermal dysplasia (XLHED) phenotype. Two families EDP2 & EDP3 with autosomal recessive ectodermal dysplasias have been identified, on the basis of the presence of multiple affected members including females in highly consanguineous families. Affected persons from three other families LAP 1, LAP2 and LAP3 showed clinical features of hypotrichosis, an autosomal recessive form of alopecia. In family EDP 1, clinical features of the affected individuals resembled those of the other families with XLHED in which mutation in the EDl gene were reported earlier. Therefore, DNA from two affected individuals and one obligate carrier was analyzed for a mutation in the ED 1 gene. Sequence analysis of exon 9 from two affected males revealed a Cā†’T transition at nucleotide position 1375 of EDl gene, resulting in a threonine to methionine amino acid substitution at codon 378 (T378M).

In family EDP2, the clinical features of the affected individuals include highly dystrophic nails and thin hair on scalp, fine eyebrows and eyelashes, and thin body hair. Analysis of the family pedigree suggests autosomal recessive inheritance of the disease. Genome wide linkage analysis of 390 microsatellite markers mapped the ectodermal dysplasia gene to the 3.92 cM interval flanked by markers D1OS1710 and D1OS1741 on chromosome 10q24.32-q25.1. A maximum two-point LOD score of 2.97 was obtained for marker D10S1267. Multipoint linkage analysis generated a maximum lod score of 4.79 in the interval D10S1239-D10S1264. The genetic interval identified in this family corresponds to 6.35 Mb.

In family EDP3 with autosomal recessive form of hereditary nail dysplasia, affected individuals presented with onycholysis of fingernails and anonychia of toenails. Associated abnormalities of ectodermal appendages were not observed in any of the affected individuals. Linkage has been established to chromosome 17q. A maximum multipoint analysis LOD score of 4.85 was obtained at marker D17S1301. Due to the consanguineous nature of the kindred, the gene for nail dysplasia is probably contained within a 5.0 cM (3 MB on the sequence-based physical map) region of homozygosity flanked by markers D17S1807 and D17S937. Envoplakin gene, located in the candidate region, was searched for mutation in two affected individuals and an unaffected control. The PCR amplified 22 exons and splice junctions of the gene were purified and sequenced on an ABI Prism 310 DNA sequencer. However, no disease-causing mutation was detected.

In three consanguineous families (LAP 1, LAP2 & LAP 3 ), with hypotrichosis segregating as a single abnormality without associated defects, a novel hypotrichosis locus was mapped to a 5.5 cM region on chromosome 18q12.1. Affected individuals in all the three families showed similar clinical features. At birth, hair was present on the scalp, but regrew sparsely after ritual shaving, which is usually performed a week after birth. Affected persons are nearly devoid of eyebrows and eyelashes; however, axillary and pubic hairs are normal. Affected male individuals have normal beard hair. A candidate gene approach was used to localize the hypotrichosis locus segregating in these families. Significant evidence of linkage was obtained for marker D18S36 that localizes to the candidate 18q12 region. For the three kindreds, a maximum two-point LOD score of 5.25 was obtained at marker D18S36. In family LAP3, recombination events observed in the affected individuals localized the disease locus to 5.57 cM flanked by markers D18S847 and D18S536. The candidate region contains desmoglein and desmocolin genes, which are the glycoproteins of desmosomes. One of the desmoglein genes DSG4 was screened for pathogenic mutation. PCR amplification of all the exons and splice junctions of DSG4 gene, followed by sequencing in ABI Prism 310 sequencer, identified an identical 5 kb deletion mutation, encompassing exon 5 to 8 (EX5 _ 8del), in affected individuals from all the three families.

Download Full Thesis
3730.92 KB
S. No. Chapter Title of the Chapters Page Size (KB)
1 0 Contents 0
403.25 KB
2 1 Introduction I
529.4 KB
  1.1 Human Skin 2
  1.2 Ectodermal Dysplasia 4
  1.3 Genes Involved In Generation Of Ectodermal Dysplasias 7
  1.4 Mutations In EDs Genes Generates Diverse Phenotypes 8
  1.5 Alopecia 17
3 2 Materials And Methods 35
1177.56 KB
  2.1 Families Studied 35
  2.2 Genotyping 38
  2.3 Linkage Studies 38
  2.4 Mutations Screening 40
4 3 Ectodermal Dysplasia 68
854.07 KB
  3.1 Families Studied 70
  3.2 Molecular Genetic Studies 73
  3.3 Discussion 76
5 4 Alopecia 110
1005.09 KB
  4.1 Families Studied 111
  4.2 Molecular Genetic Studies 113
  4.3 Discussion 115
  4.4 References 145
  4.5 Electronic Database Information 177