title: Mitochondrial complex 1 gene analysis in keratoconus.
abstract: OBJECTIVE
  Keratoconus is characterized by the thinning of corneal stroma, resulting in reduced vision. The exact etiology of keratoconus (KC) is still unknown. The involvement of oxidative stress (OS) in this disease has been reported. However, the exact mechanism of OS in keratoconus is still unknown. Thus we planned this study to screen mitochondrial complex I genes for sequence changes in keratoconus patients and controls, as mitochondrial complex I is the chief source of reactive oxygen species (ROS) production.
  
  
  METHODS
  A total of 20 keratoconus cases and 20 healthy controls without any ocular disorder were enrolled in this study. Mitochondrial complex I genes (ND1, 2, 3, 4, 4L, 5, and 6) were amplified in all patients and controls using 12 pairs of primers by PCR. After sequencing, DNA sequences were analyzed against the mitochondrial reference sequence NC_012920. Haplogroup frequency based Principle Component Analysis (PCA) was constructed to determine whether the gene pool of keratoconus patients is closer to major populations in India.
  
  
  RESULTS
  DNA sequencing revealed a total 84 nucleotide variations in patients and 29 in controls. Of 84 nucleotide changes, 18 variations were non-synonymous and two novel frame-shift mutations were detected in cases. Non-synonymous mtDNA sequence variations may account for increased ROS and decreased ATP production. This ultimately leads to OS; which is a known cause for variety of corneal abnormalities. Haplotype analysis showed that most of the patients were clustered under the haplogroups: T, C4a2a, R2'TJ, M21'Q1a, M12'G2a2a, M8'CZ and M7a2a, which are present as negligible frequency in normal Indian population, whereas only few patients were found to be a part of the other haplogroups like U7 (Indo-European), R2 and R31, whose origin is contentious.
  
  
  CONCLUSIONS
  Mt complex I sequence variations are the main cause of elevated ROS production which leads oxidative stress. This oxidative stress then starts a cascade of events which ultimately can lead to keratoconus. Prompt antioxidant therapy should be initiated in keratoconus patients to minimize ROS related damage.
authors.affiliation: Laboratory For Molecular Reproduction and Genetics, Department of Anatomy, All India Institute of Medical Sciences, Ansari Nagar, New Delhi, India.
authors.forename: Dhananjay
authors.initials: D
authors.lastname: Pathak
authors.forename: Bhagabat
authors.initials: B
authors.lastname: Nayak
authors.forename: Manvendra
authors.initials: M
authors.lastname: Singh
authors.forename: Namrata
authors.initials: N
authors.lastname: Sharma
authors.forename: Radhika
authors.initials: R
authors.lastname: Tandon
authors.forename: Rajesh
authors.initials: R
authors.lastname: Sinha
authors.forename: Jeewan S
authors.initials: JS
authors.lastname: Titiyal
authors.forename: Rima
authors.initials: R
authors.lastname: Dada
mesh_terms: D000293:Adolescent; D019943:Amino Acid Substitution; D001483:Base Sequence; D016022:Case-Control Studies; D002648:Child; D004252:DNA Mutational Analysis; D017931:DNA Primers / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics; D042967:Electron Transport Complex I / Q000235:genetics* / Q000378:metabolism; D019143:Evolution, Molecular; D005260:Female; D016368:Frameshift Mutation; D050259:Genes, Mitochondrial; D014644:Genetic Variation; D006239:Haplotypes; D006801:Humans; D007194:India; D007640:Keratoconus / Q000235:genetics* / Q000378:metabolism; D008297:Male; D018384:Oxidative Stress; D020641:Polymorphism, Single Nucleotide; D017382:Reactive Oxygen Species / Q000378:metabolism; D017384:Sequence Deletion; D055815:Young Adult
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial; D017382:Reactive Oxygen Species; D042967:Electron Transport Complex I