Belinda Baquero Perez

Publications (2)4.59 Total impact

• Article: A simplified but robust method for the isolation of avian and mammalian muscle satellite cells.

  Belinda Baquero-Perez, Suresh V Kuchipudi, Rahul K Nelli, Kin-Chow Chang
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  ABSTRACT: Current methods of isolation of muscle satellite cells from different animal species are highly variable making inter-species comparisons problematic. This variation mainly stems from the use of different proteolytic enzymes to release the satellite cells from the muscle tissue (sometimes a single enzyme is used but often a combination of enzymes is preferred) and the different extracellular matrix proteins used to coat culture ware. In addition, isolation of satellite cells is frequently laborious and sometimes may require pre-plating of the cell preparation on uncoated flasks or Percoll centrifugation to remove contaminating fibroblasts. The methodology employed to isolate and culture satellite cells in vitro can critically determine the fusion of myoblasts into multi-nucleated myotubes. These terminally differentiated myotubes resemble mature myofibres in the muscle tissue in vivo, therefore optimal fusion is a keystone of in vitro muscle culture. Hence, a simple method of muscle satellite cell isolation and culture of different vertebrate species that can result in a high fusion rate is highly desirable. We demonstrate here a relatively simple and rapid method of isolating highly enriched muscle satellite cells from different avian and mammalian species. In brief, muscle tissue was mechanically dissociated, digested with a single enzyme (pronase), triturated with a 10-ml pipette, filtered and directly plated onto collagen coated flasks. Following this method and after optimization of the cell culture conditions, excellent fusion rates were achieved in the duck, chicken, horse and cow (with more than 50% cell fusion), and to a lesser extent pig, pointing to pronase as a highly suitable enzyme to release satellite cells from muscle tissue. Our simplified method presents a quick and simple alternative to isolating highly enriched muscle satellite cell cultures which can subsequently rapidly differentiate into well developed primary myotubes. The use of the same isolation protocol allows better inter-species comparisons of muscle satellite cells. Of all the farm animal species investigated, harvested chicken muscle cells showed the highest percentage of muscle satellite cells, and equine muscle cells presented the highest fusion index, an impressive ≈ 77%. Porcine cells displayed the lowest amount of satellite cells but still achieved a modest fusion rate of ≈ 41%.
  BMC Cell Biology 06/2012; 13:16. · 2.59 Impact Factor
• Source

  Available from: Suresh Kuchipudi

  Article: Comparative distribution of human and avian type sialic acid influenza receptors in the pig.

  Rahul K Nelli, Suresh V Kuchipudi, Gavin A White, Belinda Baquero Perez, Stephen P Dunham, Kin-Chow Chang
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  ABSTRACT: A major determinant of influenza infection is the presence of virus receptors on susceptible host cells to which the viral haemagglutinin is able to bind. Avian viruses preferentially bind to sialic acid alpha2,3-galactose (SAalpha2,3-Gal) linked receptors, whereas human strains bind to sialic acid alpha2,6-galactose (SAalpha2,6-Gal) linked receptors. To date, there has been no detailed account published on the distribution of SA receptors in the pig, a model host that is susceptible to avian and human influenza subtypes, thus with potential for virus reassortment. We examined the relative expression and spatial distribution of SAalpha2,3-GalG(1-3)GalNAc and SAalpha2,6-Gal receptors in the major organs from normal post-weaned pigs by binding with lectins Maackia amurensis agglutinins (MAA II) and Sambucus nigra agglutinin (SNA) respectively. Both SAalpha2,3-Gal and SAalpha2,6-Gal receptors were extensively detected in the major porcine organs examined (trachea, lung, liver, kidney, spleen, heart, skeletal muscle, cerebrum, small intestine and colon). Furthermore, distribution of both SA receptors in the pig respiratory tract closely resembled the published data of the human tract. Similar expression patterns of SA receptors between pig and human in other major organs were found, with exception of the intestinal tract. Unlike the limited reports on the scarcity of influenza receptors in human intestines, we found increasing presence of SAalpha2,3-Gal and SAalpha2,6-Gal receptors from duodenum to colon in the pig. The extensive presence of SAalpha2,3-Gal and SAalpha2,6-Gal receptors in the major organs examined suggests that each major organ may be permissive to influenza virus entry or infection. The high similarity of SA expression patterns between pig and human, in particular in the respiratory tract, suggests that pigs are not more likely to be potential hosts for virus reassortment than humans. Our finding of relative abundance of SA receptors in the pig intestines highlights a need for clarification on the presence of SA receptors in the human intestinal tract.
  BMC Veterinary Research 01/2010; 6:4. · 2.00 Impact Factor