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in Journal of the American Veterinary Medical Association

Abstract

Objective

To use canine glyceraldehyde-3-phosphate dehydrogenase complementary DNA (GAPDH cDNA) as a template in ribonuclease (RNase) protection assays to measure canine GAPDH mRNA expression.

Design and Procedure

Primers designed from the human GAPDH gene were used to amplify a 191 -base pair canine GAPDH cDNA by reverse-transcription polymerase chain reaction. The cDNA was sequenced, and used as a template for RNase protection assay.

Sample Population

Total RNA was isolated from a canine squamous carcinoma cell line.

Results

Canine GAPDH cDNA had a high degree of homology to human, rat, and mouse GAPDH. In vitro transcription of canine GAPDH cDNA was used to produce complementary RNA that detected canine GAPDH mRNA by RNase protection assay.

Conclusion

Canine GAPDH cDNA is a useful loading control to be used in RNase protection assays measuring mRNA expression in canine cells or tissues.

Free access
in American Journal of Veterinary Research

Abstract

Objective—To clone and sequence the cDNA for feline preproparathyroid hormone (preproPTH) and to compare that sequence with other known parathyroid hormone (PTH) sequences.

Sample Population—Parathyroid glands from 1 healthy cat.

Procedure—A cDNA library was constructed in λ phage from feline parathyroid gland mRNA and screened with a radiolabeled canine PTH probe. Positive clones were sequenced, and nucleic acid and deduced amino acid sequences were analyzed and compared with known preproPTH and PTH sequences.

Result—Screening of approximately 2 X 105 recombinant plaques revealed 3 that hybridized with the canine PTH probe; 2 clones comprised the complete sequence for feline preproPTH. Feline preproPTH cDNA consisted of a 63-base pair (bp) 5'-untranslated region (UTR), a 348-bp coding region, and a 326-bp 3'-UTR. The coding region encoded a 115-amino acid peptide. Mature feline PTH consisted of 84 amino acids. Amino acid sequence analysis revealed that feline PTH was > 83% identical to canine, bovine, swine, equine, human, and macaque PTH and 69, 71, and 44% identical to mouse, rat, and chicken PTH, respectively. Within the region responsible for hormonal activity (amino acids 1 to 34), feline PTH was > 79% identical to other mammalian PTH sequences and 64% identical to the chicken sequence.

Conclusions and Clinical Relevance—The amino acid sequence of PTH is conserved among mammalian species. Knowledge of the cDNA sequence for feline PTH may be useful to investigate disturbances of calcium metabolism and alterations in PTH expression in cats. (Am J Vet Res 2002;63:194–197)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To evaluate calcium balance and parathyroid gland function in healthy horses and horses with enterocolitis and compare results of an immunochemiluminometric assay (ICMA) with those of an immunoradiometric assay (IRMA) for determination of serum intact parathyroid hormone (PTH) concentrations in horses.

Animals—64 horses with enterocolitis and 62 healthy horses.

Procedures—Blood and urine samples were collected for determination of serum total calcium, ionized calcium (Ca2+) and magnesium (Mg2+), phosphorus, BUN, total protein, creatinine, albumin, and PTH concentrations, venous blood gases, and fractional urinary clearance of calcium (FCa) and phosphorus (FP). Serum concentrations of PTH were measured in 40 horses by use of both the IRMA and ICMA.

Results—Most (48/64; 75%) horses with enterocolitis had decreased serum total calcium, Ca2+, and Mg2+ concentrations and increased phosphorus concentrations, compared with healthy horses. Serum PTH concentration was increased in most (36/51; 70.6%) horses with hypocalcemia. In addition, FCa was significantly decreased and FP significantly increased in horses with enterocolitis, compared with healthy horses. Results of ICMA were in agreement with results of IRMA.

Conclusions and Clinical Relevance—Enterocolitis in horses is often associated with hypocalcemia; 79.7% of affected horses had ionized hypocalcemia. Because FCa was low, it is unlikely that renal calcium loss was the cause of hypocalcemia. Serum PTH concentrations varied in horses with enterocolitis and concomitant hypocalcemia. However, we believe low PTH concentration in some hypocalcemic horses may be the result of impaired parathyroid gland function. ( Am J Vet Res 2001;62:938–947)

Full access
in American Journal of Veterinary Research

Abstract

Objective—To determine effects of experimentally induced hypercalcemia on serum concentrations and urinary excretion of electrolytes, especially ionized magnesium (iMg), in healthy horses.

Animals—21 clinically normal mares.

Procedures—Horses were assigned to 5 experimental protocols (1, hypercalcemia induced with calcium gluconate; 2, hypercalcemia induced with calcium chloride; 3, infusion with dextrose solution; 4, infusion with sodium gluconate; and 5, infusion with saline [0.9% NaCl] solution). Hypercalcemia was induced for 2 hours. Dextrose, sodium gluconate, and saline solution were infused for 2 hours. Blood samples were collected to measure serum concentrations of electrolytes, creatinine, parathyroid hormone, and insulin. Urine samples were collected to determine the fractional excretion of ionized calcium (iCa), iMg, sodium, phosphate, potassium, and chloride.

Results—Hypercalcemia induced by administration of calcium gluconate or calcium chloride decreased serum iMg, potassium, and parathyroid hormone concentrations; increased phosphate concentration; and had no effect on sodium, chloride, and insulin concentrations. Hypercalcemia increased urinary excretion of iCa, iMg, sodium, phosphate, potassium, and chloride; increased urine output; and decreased urine osmolality and specific gravity. Dextrose administration increased serum insulin; decreased iMg, potassium, and phosphate concentrations; and decreased urinary excretion of iMg. Sodium gluconate increased the excretion of iCa, sodium, and potassium.

Conclusions and Clinical Relevance—Hypercalcemia resulted in hypomagnesemia, hypokalemia, and hyperphosphatemia; increased urinary excretion of calcium, magnesium, potassium, sodium, phosphate, and chloride; and induced diuresis. This study has clinical implications because hypercalcemia and excessive administration of calcium have the potential to increase urinary excretion of electrolytes, especially iMg, and induce volume depletion.

Full access
in American Journal of Veterinary Research

Summary

Serum lipoprotein concentrations, routine serum biochemical values, and morphologic changes of the liver were evaluated in cats undergoing weight loss. Food was witheld from 6 obese and 6 control cats for 3 days (days 0 to 2), followed by feeding 50% of previous food intake for 26 days (days 3 to 28). Percutaneous liver biopsy specimens were obtained from all cats on days 0, 7, 14, and 28. Blood samples for serum biochemical analysis and lipoprotein profiles were obtained on days 0, 3, 7, 14, and 28. All cats lost weight throughout the study, and none developed signs of clinical illness, including those of idiopathic hepatic lipidosis syndrome. Serum total cholesterol concentrations decreased initially in all cats, but rapidly returned to normal after day 3 in obese cats, suggesting altered cholesterol metabolism during dietary restriction. Low-density lipoprotein concentrations decreased throughout the study in control cats, but were unchanged in obese cats. Examination of liver biopsy specimens from each cat revealed minimal lipid accumulation in all specimens, although some specimens contained hydropic degeneration.

Free access
in American Journal of Veterinary Research