May 2003
Volume 44, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2003
The Development of Transgenic Mice Over Expressing the Na/Myo-Inositol Cotransporter Using the BetaB2-Crystallin Gene Promoter and Their Use in Studying Adult Onset Osmotic Cataract
Author Affiliations & Notes
  • Z. Wang
    Department of Cell Biology and Genetics, University of North Texas Health Science Center at Fort Worth, Fort Worth, TX, United States
  • F.T. Nahi
    Department of Molecular and Human Genetics, Children's Research Institute, Columbus, OH, United States
  • M.L. Robinson
    Department of Molecular and Human Genetics, Children's Research Institute, Columbus, OH, United States
  • P.R. Cammarata
    Department of Molecular and Human Genetics, Children's Research Institute, Columbus, OH, United States
  • Footnotes
    Commercial Relationships  Z. Wang, None; F.T. Nahi, None; M.L. Robinson, None; P.R. Cammarata, None.
  • Footnotes
    Support  EY05570 (PRC)
Investigative Ophthalmology & Visual Science May 2003, Vol.44, 4263. doi:
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      Z. Wang, F.T. Nahi, M.L. Robinson, P.R. Cammarata; The Development of Transgenic Mice Over Expressing the Na/Myo-Inositol Cotransporter Using the BetaB2-Crystallin Gene Promoter and Their Use in Studying Adult Onset Osmotic Cataract . Invest. Ophthalmol. Vis. Sci. 2003;44(13):4263.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Abstract: : Purpose: Intracellular osmotic stress is alleged to be associated with the advancement of diabetic (i.e. osmotic) cataract. Prior studies from this laboratory described mouse lines which over expressed the bovine Na+/myo-inositol cotransporter (bSMIT) gene in lens fiber cells using the αA-crystallin promoter (Cammarata et al., Invest Ophthalmol Vis Sci. 1999; 40:1727-1737). Lens fiber swelling and related cataractous outgrowth positively correlated to the degree of lens bSMIT gene expression and intralenticular myo-inositol content. However, that model generated a nuclear cataract as early as E15.5 in the differentiating secondary fibers, consistent with the midgestation expression of the endogenous murine αA-crystallin gene. The ßB2-crystallin gene is active in the rat post-natal lens and its transcript reaches its maximal level by 6 months after birth (NH Lubsen, personal communication). Transgenic (tg+) mice were developed that over express the bSMIT gene in the lens using the rat ßB2-crystallin gene promoter to develop an adult onset osmotic cataract animal model. Methods: The rat ßB2-crystallin promoter, first exon and intron were cloned upstream of bSMIT, and this ßB2/bSMIT construct was used to generate tg+ mice. Tg+ mRNA expression was analyzed in adult tg+ mice by coupled RT-PCR. The tissue specificity of the promoter was assayed by RT-PCR from total RNA extracted from multiple tissues from a 4-week-old tg+ mouse and compared to RNA from the lens of an age-matched, tg- littermate. Results: Three independent tg+ lines were generated and no cataracts were evident in any of these lines at 4 weeks of age. Tg+ specific primers detected correctly spliced mRNA in the lenses of two tg+ lines using RT-PCR. Transgene transcript expression from each tg+ lens was equivalent as approximated from the densities of transgene-specific PCR product and ß-actin PCR product. No expression was observed with tg- lens or when the RT reaction was omitted with total RNA from tg+ lenses. The expected 517-bp band was authenticated by sequence analysis. ßB2/bSMIT predominated in tg+ lenses with trace tg+ expression in brain cortex and lung. Conclusions: These mice provide a unique diabetic animal model to study adult onset experimental osmotic cataractogenesis attributable to over expression of the Na+/myo-inositol cotransporter with corresponding alteration to lenticular phenotype.

Keywords: animal model • cataract • gene/expression 
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