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ABSTRACT. Alterations in the activities of enzymes related to energy metabolism in canine lymphoma cells were investigated. Cytosolic pyruvate kinase (PK) and mitochondrial malate dehydrogenase (MDH) activities in lymphoma cells were significantly higher than those in lymphocytes obtained from lymph nodes of healthy dogs, whereas cytosolic lactate dehydrogenase (LDH) activity was significantly lower in lymphoma cells. The cytosolic M/L ratio (MDH activity/LDH activity), which is considered to be a good indicator of energy metabolism related to glucose utilization in animal tissues, was significantly higher in lymphoma cells than in the normal lymphocytes. KEY WORDS: canine, lymphoma, malate dehydrogenase.J. Vet. Med. Sci. 67(6): 615-616, 2005 Lymphoma is one of the most common neoplasms observed in the dog and primarily affects middle-aged to older dogs [14]. The most common form of canine lymphoma is the multicentric form, which usually presents with superficial lymphadenopathy. Morphology, immunophenotypes, responsiveness to chemotherapy, and correlations between morphology and survival time of canine lymphoma have been reported [8,9,17]. Several reports have discussed the metabolic alterations of dogs with lymphoma [15,16,20], however, the enzyme activities of lymphoma cells per se have not been investigated. In the present study, lymphoma cells and normal lymphocytes from the lymph nodes of healthy dogs were obtained, and the activities of the enzymes related to energy metabolism including hexokinase (HK), piruvate kinase (PK), lactate dehydrogenase (LDH), malate dehydrogenase (MDH) and aspartate aminotransferase (AST) were measured.Five healthy mixed breed dogs (2 males and 3 females; 3-6 years old) maintained for research in our laboratory and fed on a commercial diet (Hill's Pet Products, Topeka, KS, U.S.A.) were used as control dogs. Their hematological and serum chemistry values were within the normal ranges. In addition, ten dogs referred to our Veterinary Medical Teaching Hospital and diagnosed as multicentric, stage III lymphoma were used. Signalment of the ten dogs are shown in Table 1.Popliteal lymph nodes of normal dogs were obtained surgically under general anesthesia with isoflurane (Rhodia, England). Lymphoma cells were collected by fine needle aspiration biopsy from enlarged lymph nodes of the dogs with lymphoma before the initiation of chemotherapy and put in RPMI 1640 medium. Lymphocytes and lymphoma cells were further purified by gradient centrifugation with a commercially available mononuclear cell isolating solution (Histopaque-1077, Sigma-Aldrich Co., St. Louis, MO, U.S.A.), followed by three washes with cold phosphatebuffered saline (0.15 M NaCl, pH 7.2).Washed lympyocytes were resuspended in 2 ml ice-cold STE solution (0.25 M sucrose, 10 mM Tris-Hcl buffer, pH 7.5, containing 2 mM EGTA), followed by homogenization with an ultrasonic processor (VP-5, Taitek, Koshigaya, Japan) for 5 sec. The homogenate was centrifuged at 100 × g for 1 min to remove nuclei and cell debris, and the supe...

contrasting

confidence: 41%

Changes in Activities of Enzymes Related to Energy Metabolism in Canine Lymphoma Cells

Washizu

Azakami

Bonkobara

et al. 2005

“…High ML ratio in leukocytes is considered to reflect elevation of energy metabolism, including ATP production, in various animal tissues [3,24]. Significant increase in the ML ratio was observed in leukocytes of race horse with continuous hard training [3] and tumor tissues with hyperplasia [4]. On the other hand, the ML ratio is significantly decreased in leukocytes of diabetic dogs with insulin deficiency [5].…”

mentioning

confidence: 99%

Comparison of Enzyme Activities in Plasma and Leukocytes in Dairy and Beef Cattle

Arai

Inoue

Takeguchi

et al. 2003

ABSTRACT. Concentrations of plasma glucose, immunoreactive insulin (IRI) and free fatty acid (FFA) and activities of enzymes related to energy metabolism and lactate dehydrogenase (LDH) isoenzyme pattern in plasma and leukocytes were investigated in lactating Hols tein cows (dairy cattle) and fattening Japanese Black Wagyu x Holstein steers (beef cattle). IRI concentrations and LDH and malate dehydrogenase (MDH) activities in the plasma of beef cattle were significantly higher than those in dairy cattle. The cytosolic rat io of MDH/ LDH activity in the leukocytes of beef cattle was significantly higher than that of dairy cattle. These findings might be associated with the different energy metabolism between dairy and beef cattle. KEY WORDS: beef cattle, insulin, malate dehydrogenase.J. Vet. Med. Sci. 65(11): 1241-1243, 2003 It has been reported that activities of enzymes related to energy metabolism including ATP production in peripheral leukocytes are very variable in animal species [23,24]. In particular, activities of enzymes in the malate-aspartate shuttle, which has a very important role in the transfer from cytosolic NADH to mitochondrial NADH [13], change remarkably according to variation in energy metabolism of animals [2]. Lactate dehydrogenase (LDH) is a tetrameric molecule made up of four subunits of the two parent molecules, the H and M subunits, and each tissue has a characteristic composition of isoenzymes [8]. The isoenzyme patterns of LDH are variable in plasma and leukocytes of each animal species [24]. Activities of enzymes related to energy metabolism in leukocytes are considered to reflect the metabolic states in animal tissues [2,3,24]. On the other hand, plasma glucose and insulin concentrations are different between dairy and beef cattle [12,15,20,22]. It has been reported that Japanese Black heifers have higher plasma insulin level at 18 mo of age than at or before 12 mo of age, whilst Holstein heifers have very similar values at all stages [19].In the present study, activities of enzymes in the malateaspartate shuttle and LDH and LDH isoenzyme pattern were determined in plasma and peripheral leukocytes of the lactating dairy cattle and the fattening beef cattle to clarify one aspect of the differences in energy metabolism between the dairy and beef cattle.Eight lactating dairy Holstein cows (5 to 8 years old) maintained with good quality hay and concentrates in the Fuji Animal Farm of our university and four fattening Japanese Black Wagyu × Holstein steers (2 years old) kept with high concentrate feed at a farm in Shimane Prefecture were used. Blood was withdrawn from the jugular vein into heparinized tubes, with the animals at rest and unfasted. The plasma was recovered by centrifugation at 4°C. Leukocytes were isolated by centrifugation with a lymphocyte-isolating solution (ICN Biochemical Inc., Aurora, OH, U.S.A.). Cytosolic and mitochondrial fractions of leukocytes were prepared by the method described previously [23]. The activities of the enzymes were measured by using reported met...

“…Sci. 64(11): 1065-1067, 2002 A transgenic mouse strain carrying human prototype cHa ras gene, Tg-rasH2 mice, was established by Saitoh et al. [20].…”

mentioning

confidence: 99%

“…On the other hand, it has been reported that activities of enzymes in the glycolysis and the pentose phosphate pathway are significantly increased in developed tumor cells [1,7,17]. Activities and mRNA expression of malate dehydrogenase (MDH) in the malate-aspartate shuttle are significantly induced in the early stage of testicular tumor of dogs [3]. In the present study, activities of several enzymes related to energy metabolism were measured in livers of the Tg-rasH2 mice treated with DEN to investigate availability of certain enzymes as a diagnostic marker for energy metabolism at the early stage in tumor cells.…”

mentioning

confidence: 99%

“…Increasing in MDH activities is induced by some kinds of factors such as excess of glucose utilization [8] and intensive training [2] in animal tissues. Although activities of enzymes in glycolysis and pentose phosphate pathway were significantly increased in well developed tumor cells [1,7], these enzymes activities were not so high at the early neoplastic stage [3]. MDH and AST may be one of important factors to lead the cytosolic energy metabolism to more ATP production before activation of glycolysis and pentose phosphate pathway at the early stage of tumor.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Changes in Hepatic Enzyme Activities in Transgenic Mice Carrying Human Prototype c-Ha-ras Gene Treated with Diethylnitrosamine.

Arai

Ogawa

Nakamura

et al. 2002

ABSTRACT. Activities of malate dehydrogenase (MDH) and aspartate aminotransferase in the malate-aspartate shuttle were significantly increased in the cytosolic fractions of livers with early neoplastic symptoms such as swelling and discoloration in transgenic mice carrying the prototype human c-Ha-ras gene, Tg-rasH2 mice, which were administered with diethylnitrosamine (DEN) as a carcinogenic chemical at a dose of 200 mg/kg body weight. Cytosolic MDH/LDH (lactate dehydrogenase) (ML) ratio increased significantly and w as considered to be a useful marker to characterize the energy metabolism at early neoplastic stage in livers of the Tg-rasH2 mice. KEY WORDS: aspartate aminotransferase, malate dehydrogenase, transgenic mouse.J. Vet. Med. Sci. 64(11): 1065-1067, 2002 A transgenic mouse strain carrying human prototype cHa ras gene, Tg-rasH2 mice, was established by Saitoh et al. [20]. These mice show high and rapid susceptibility to various carcinogens and are considered to be promising as an animal model for an alternative rapid carcinogenicity test [25]. Spontaneous hepatocellular carcinomas, forestomach papillomas, skin papillomas and lymphomas were observed in 20 to 30% of Tg-rasH2 mice by 82 weeks and these incidences were increased by treatments of some kinds of carcinogens including N,N'-diethylnitrosamine (DEN) known as Salmonella mutagenesis assay-positive trans-species carcinogen [24]. The major target organs of DEN are the liver, lung, forestomach and hematopoietic system [12]. In our previous reports, Tg-rasH2 mice were characterized by stable activities of some enzymes related to drug detoxication and energy metabolism in their livers and kidneys compared to those in the non-transgenic control [18]. On the other hand, it has been reported that activities of enzymes in the glycolysis and the pentose phosphate pathway are significantly increased in developed tumor cells [1,7,17]. Activities and mRNA expression of malate dehydrogenase (MDH) in the malate-aspartate shuttle are significantly induced in the early stage of testicular tumor of dogs [3]. In the present study, activities of several enzymes related to energy metabolism were measured in livers of the Tg-rasH2 mice treated with DEN to investigate availability of certain enzymes as a diagnostic marker for energy metabolism at the early stage in tumor cells.F1 hybrids of a transgenic male C57BL/6J, Tg-rasH2, mice and normal female BALB/cByJ mice were used. The F1 offsprings were screened by polymerase chain reaction (PCR), and divided into transgenic (-/Tg) and non-transgenic mice (-/-). Six-week-old male transgenic mice, which showed high expression of ras gene, were used in the present study. Their mean (± SD) body weights were 16.8 (± 0.4) g. Mice were housed in an air-conditioned room (23 ± 2°C, 55 ± 10%) with a light period of 12 hr. Food and water were available ad libitum. All procedures involving laboratory animals were performed in accordance with the care and use guidelines of the Central Institute of Experimental Animals (Kawasaki, Japan)...