Identification of the nucleotide sequence of the lipoprotein lipase gene as well as its role in the development of hyperlipidemia and pancreatitis in the Miniature Schnauzer

Lipoprotein Lipase (LPL) is a key enzyme in lipid transport. It
catalyses the hydrolysis of the triacylglycerol component of
chylomicrons and very low-density lipoproteins (VLDL), providing
non-esterified fatty acids for tissue utilization. The gene
encoding for LPL has already been identified in several species
except the dog. Mutations of the human LPL-gene have been shown to
cause partial or complete malfunction of the enzyme, resulting in
accumulation of lipoproteins in the blood. This condition is called
familial LPL deficiency. LPL malfunction results in
hyperlipoproteinemia, recurrent acute pancreatitis, and ultimately
pancreatic insufficiency. Several authors have postulated a genetic
cause for pancreatitis in the Miniature Schnauzer. An idiopathic
increase in serum triglyceride concentration can also be found in
this breed. Based on these findings we were evaluating a possible
role of the lipoprotein lipase gene in the development of
pancreatitis and hyperlipidemia in the Miniature Schnauzer. First,
we identified the genetic sequence of the LPL gene in the dog. We
determined clones on the Trace Archive database for the canine
genome project that contain the genomic sequence of a particular
exon as well as its adjacent intronic regions. Based on these
findings we designed primers for each exon using the software
Netprimer (www.premierbiosoft.com/netprimer/index.html). Canine
subjects were chosen from a pool of 170 Miniature Schnauzers from
the database at the Gastrointestinal Laboratory at Texas A&M
University. Based on clinical history, serum cPLI concentrations,
and serum triglyceride concentrations 21 Miniature Schnauzers were
chosen and were selected into a clinically normal control group (9
dogs) and an affected group (12 dogs). DNA was then collected from
either white blood cells or mucosal cells of these dogs. After PCR
optimization, exon 1 through 9 including the adjacent intronic
regions were amplified in all dogs using MasterAmp Extra – Long PCR
Kit (Epicentre, WI, USA) and were sequenced in triplicates.
Differences in the nucleotide sequences were then compared among
the two groups. 10 exonic SNPs and 9 intronic SNPs were identified.
Upon analysis, none of these variations could be associated with
the disease status. We conclude that pancreatitis associated with
hyperlipidemia in the Miniature Schnauzer is not linked to
mutations of the lipoprotein lipase gene or its splicing regions.