ABSTRACT: A model of intracoronary stem cell delivery that enables transgenesis/gene targeting would be a powerful tool but is still
lacking. To address this gap, we compared intracoronary and intramyocardial delivery of lin−/c-kit+/GFP+ cardiac stem cells (CSCs) in a murine model of reperfused myocardial infarction (MI). Lin−/c-kit+/GFP+ CSCs were successfully expanded from GFP transgenic hearts and cultured with no detectable phenotypic change for up to ten
passages. Intracoronary delivery of CSCs 2days post-MI resulted in significant alleviation of adverse LV remodeling and dysfunction,
which was at least equivalent, if not superior, to that achieved with intramyocardial delivery. Compared with intramyocardial
injection, intracoronary infusion was associated with a more homogeneous distribution of CSCs in the infarcted region and
a greater increase in viable tissue in this region, suggesting greater formation of new cardiomyocytes. Intracoronary CSC
delivery resulted in improved function in the infarcted region, as well as in improved global LV systolic and diastolic function,
and in decreased LV dilation and LV expansion index; the magnitude of these effects was similar to that observed after intramyocardial
injection. We conclude that, in the murine model of reperfused MI, intracoronary CSC infusion is at least as effective as
intramyocardial injection in limiting LV remodeling and improving both regional and global LV function. The intracoronary
route appears to be superior in terms of uniformity of cell distribution, myocyte regeneration, and amount of viable tissue
in the risk region. To our knowledge, this is the first study to report that intracoronary infusion of stem cells in mice
is feasible and effective.
KeywordsIntracoronary administration–Cardiac stem cells–Regeneration–Mice–Myocardial infarction–Cardiac function
Archiv für Kreislaufforschung 04/2012; 106(5):849-864. · 7.35 Impact Factor
ABSTRACT: Agouti-related peptide (AgRP), a brain neuropeptide generated by AgRP/neuropeptide Y (NPY) neurons, plays a vital role in the hypothalamic regulation of energy homeostasis. RT-PCR and real-time PCR were carried out in various tissues to detect the AgRP expression pattern in pigs. Our RT-PCR results showed that the pig AgRP gene was ubiquitously expressed in all examined tissues including heart, liver, spleen, lung, kidney, stomach, bladder, m. longissimus, belly fat, brain, large intestine, lymph, back fat, skin, and hypothalamus. Real-time quantitative PCR experiments revealed that it is in the hypothalamus with the highest expression of AgRP both in adult Lantang and Landrace pigs compared to the back fat and m.longissimus muscle and the cDNA level of AgRP in the hypothalamus of adult Chinese indigenous Lantang pig (fat-type) is significantly higher than that of Landrace pig (lean-type). To understand the regulation of the pig AgRP gene, the 5'-flanking region was isolated from a pig bacterial artificial chromosome library and used in a luciferase reporter assay. A positive cis-acting element for efficient AgRP expression was identified at nucleotides -501 to -479, by 5'-serial deletion of the promoter. Electrophoretic mobility-shift assays (EMSA) with competing oligonucleotides revealed that the critical region contained a cis-acting element for Neurogenic Differentiation (NeuroD), which is a member of the NeuroD family of basic-helix-loop-helix transcription factors. This element has not been reported in human or mouse AgRP genes. Our results indicated that NeuroD might be an essential regulatory factor for transcription of pig AgRP, providing an important clue about energy homeostasis regulation in the porcine and human brain.
Molecular Biology Reports 04/2011; 38(4):2233-9. · 2.93 Impact Factor
ABSTRACT: The cocaine- and amphetamine-regulated transcript (CART) gene encodes an anorexigenic peptide. It has a key role in the hypothalamic regulation of energy balance through reducing food intake and enhancing lipid substrate utilization. To detect the CART expression pattern in pigs, reverse transcription (RT)-polymerase chain reaction (PCR) and real-time PCR were performed in various tissues. Our RT-PCR results revealed that the pig CART gene was ubiquitously expressed in all examined tissues including hypothalamus, m. longissimus, backfat, heart, liver, spleen, lung, kidney, stomach, bladder, belly fat, brain, large intestine, lymph, and skin. Real-time quantitative PCR experiments revealed that the cDNA level of CART in both the hypothalamus and backfat of adult Landrace pig (lean-type) was significantly higher than that of Chinese indigenous Lantang pig (fat-type), and it was in the hypothalamus where the highest expression of CART was observed for both adult Lantang and Landrace pigs, compared with backfat and m. longissimus muscle. To understand the regulation of the pig CART gene, the 5'-flanking region was isolated from a pig bacterial artificial chromosome library and used in a luciferase reporter assay. A positive cis-acting element for efficient CART expression was identified at nucleotides -73 to -53, using 5'-serial deletion of the promoter. Electrophoretic mobility shift assays with competing oligonucleotides revealed that the critical region contained a cis-acting element for the zinc-binding protein factor, a zinc-finger transcription factor of the Kruppel family. This element has not been reported in human or mouse CART genes. Our results indicated that zinc-binding protein factor might be an essential regulatory factor for transcription of pig CART, providing important insight into mechanisms involved in energy homeostasis regulation in the porcine and human brain.
DNA and cell biology 02/2011; 30(2):91-7. · 2.28 Impact Factor
ABSTRACT: Weed control is a substantial economic input for production of mint oils, the most commercially important of which are obtained from peppermint. The objective of this research is to obtain peppermint plants resistant to the broad-spectrum herbicide glufosinate, which can be used for development of economically efficacious weed control strategies and, perhaps, serve as a paradigm in perennial crops. The bar gene, which encodes phosphinothricin acetyltransferase (PAT) which inactivates glufosinate-ammonium or phosphinothricin (PPT), was constructed into Agrobacterium tumefaciens binary vectors under the nopaline synthase (NOS) or a chimeric promoter containing a trimer of the OCS-upstream-activating sequence (UAS) to a MAS promoter/activator region[(OCS)
MAS]. A total of 142 independent transgenic peppermint (cv. Black Mitcham) plants were obtained (107 and 35 were obtained with pGPTV (and pCAS1) and pATC940 vectors, respectively) and evaluated for herbicide resistance in the greenhouse after foliar application of glufosinate herbicide Liberty as the commercial product. All transgenic plants exhibited substantially less herbicide symptom development than non-transgenic Black Mitcham or untransformed tissue cultured-derived plants, albeit variation for herbicide resistance occurred amongst the transformed lines. Plants from 35 of the 142 lines were selected at random and all were PCR-positive for the presence of bar. Five lines, that were least affected, exhibited no injury symptoms to Liberty concentrations that are 4 times the standard level for control of weeds in peppermint fields. The most resistant transgenic plants had the greatest steady-state PAT mRNA levels and PAT activities. No experimental difference in herbicide resistance was evident between plant populations obtained with pGPTV (pCAS1)-bar or pATC940-bar vector. However, 4 of 35 lines transformed with (ocs)
MAS-bar exhibited maximal resistance while only 1 of 107 NOS-bar lines has comparable resistance. These herbicide resistant peppermint plants will facilitate development of post-emergent herbicide control strategies that use newer generation herbicides, like glufosinate, which have reduced environmental and product residual because of metabolism by microbes and the transgenic plants.
Molecular Breeding 08/2001; 8(2):109-118. · 2.85 Impact Factor