Properties of microsomal and soluble diacylglycerol kinase in rat liver.
Explore the source record for details and available documents.
Biomedical subjects
Publications and source records attributed to H Kanoh.
Explore the source record for details and available documents.
The aim of this work is to describe the role of different phospholipases in hepatic phospholipid catabolism. Therefore isolated rat hepatocytes enriched in labeled dilinoleoyl-, dipalmitoyl- or dimyristoylglycerophosphocholine were prepared by pulse incubation with [3H]glycerol and 14C-labeled fatty acid. The labeled cells were chased up to 4 h in a tracer-free medium and the degradation of different phosphatidylcholines studied. After a 2-h chase about 40% of dilinoleoyl-, 70% of dipalmitoyl- and 30% of dimyristoylglycerophosphocholine were degraded. From the positional distribution of 14C-labeled fatty acid and the change in the doubly labeled molecular species of phospholipids, it was concluded that tb degradation of dilinoleoylglycerophosphocholine and that of phosphatidylethanolamine could be accounted for by the action of phospholipase A1, while the degradation of dipalmitoylglycerophosphocholine proceeded through the action of phospholipase A2. Dimyristoylglycerophosphocholine was probably cleaved by the combined action of both phospholipases A1 and A2. Up to 10 mM tetracain, added to the chase medium, effectively blocked the action of both phospholipase activities. A considerable part of 2-linoleoyl- and 1-palmitoylglycerophosphocholine liberated during the chase was reutilized for phosphatidylcholine synthesis without further degradation.
1. The procedure, which involved 2-step sonication of microsomes at pH 7.4 and then at pH 8.5 in the presence of sodium deoxycholate and subsequent dialysis, resulted in 4-5-fold purification of choline-phosphotransferase and ethanolaminephosphotransferase with the yield of 40-50%. 2. Ethanolaminephosphotransferase was further purified 8.5-fold over microsomes by sucrose density gradient centrifugation of the partially purified preparation, while cholinephosphotransferase activity was considerably lost during this procedure. No separation of the two transferases from each other was achieved at this step. 3. Cholinephosphotransferase required Mg2+ as cofactor, and microsomal phospholipids for its maximal activity. On the other hand, Mn2+ was more effective than Mg2+ as cofactor for ethanol aminephosphotransferase, and this enzyme was inhibited by microsomal phospholipids. 4. Both transferases were stimulated several-fold by sodium deoxycholate and also showed similar optimal pH ranging from pH 8.0 to 8.5. 5. Km values for 1,2-diacylglycerol emulsion were 81.0 muM for cholinephosphotransferase and 63.0 muM for ethanolaminephosphotransferase, respectively. CDP-choline and CDP-ethanolamine competitively inhibited, with the same Ki value (both 350 muM), ethanolaminephosphotransferase and cholinephosphotransferase, respectively. The Ki values obtained were much greater than the corresponding Km values for the cytidine substrates (36.4 muM for CDP-choline and 22.0 muM for CDP-ethanolamine). 6. The partially purified enzymes were further treated with Triton X-100. When enzyme activities were assayed with Mg2+, cholinephosphotransferase, although considerably inactivated, was partially separated from ethanolaminephosphotransferase by sucrose density gradient centrifugation of Triton-treated preparations. Furthermore, cholinephosphotransferase (but not ethanol-aminephosphotransferase) itself was partially separated into Mg2+ -requiring and Mn2+ -requiring components. In contrast, ethanolaminephosphotransferase assayed with either Mg2+ or Mn2+ formed a single peak together with Mn2+ -requiring cholinephosphotransferase.
Using the data reported previously by Moriya and Kanoh, the metabolic turnover of rat lung lecithins was calculated on the basis of the new assumption that the formation rate (Va) of lecithins from diacylglycerols did not equal the disappearance rate (Vb) of lecithins because of the presence of precursors other than diacylglycerols. The ratio of Va/Vb was almost the same in dienoic lecithin as that for total lecithins, whereas a very low ratio was found in saturated and hexaenoic species. The turnover time of lung lecithins estimated was markedly shorter than those determined in much later experimental periods after administration of labeled precursors. The reason for this discrepancy was discussed.
Explore the source record for details and available documents.
Rat liver microsomes containing 1,2-diacylglycerols formed from the endogenous phosphatidycholines by the action of 1,2-diacylglycerol: CDPcholine cholinephosphotransferase (E.C. 2.7.8.2.) were used as a source of enzymes and substrates. A marked selectivity of 1,2-diacylglycerol: CDPethanolamine ethanolaminephosphotransferase (E.C. 2.7.8.1.) was revealed for utilizing endogenous hexaenoic diaglycerol, while cholinephosphotransferase utilized without marked selectivity the endogenous 1,2-diacylglycerol species differing in the degree of unsaturation. By using microsomes prepared after injecting the animals with 1-acyl lysophosphatidylcholines labeled with radioactive myristic, palmitic, and stearic acids, the selectivity of the transferases towards the saturated fatty acids in the 1-position of the endogenous substrates was studied. In incubation of the labeled microsomes with CMP, cholinephosphotransferase appeared to utilize 1-myristyl phosphatidylcholine most rapidly and, in decreasing order, the 1-palmityl and 1-stearyl species. Ethanolaminephosphotransferase utilized the endogenous 1-stearyl diacylglycerol in preference to 1-palmityl type.
1. Mitochondrial and microsomal fractions of rat epididymal adipose tissue incorporated [1-(14)C]acetyl-CoA equally well into various fatty acids by a chain-elongation mechanism. C(18) and C(20) fatty acids were the two major products, and comprised about 80% of the total fatty acids synthesized in both particles. 2. When incubated in air, mitochondria synthesized stearic acid, octadecenoic acid and eicosamonoenoic acid in almost equal amounts (about 20% each), whereas in microsomal fractions, the synthesis of octadecenoic acid was more than fivefold the stearic acid formation. In both fractions, major components of synthesized monoenoic fatty acids were the Delta(11:12) isomers. Hexadecenoic acid and octadecenoic acid from whole adipose tissue contained approx. 11 and 14% of the Delta(11:12) isomer respectively. 3. When mitochondria or microsomal fractions were incubated in nitrogen, there was increased synthesis of stearic acid and palmitic acid and less of C(16) and C(18) monoenoic acids; synthesis of C(20) acids remained predominantly of the monoenoic acids. Determination of the position of the double bond in the monoenoic acids supported the view that the synthesis of hexadecenoic acid and octadecenoic acid involves a desaturase activity, whereas eicosamonoenoic acid and eicosadienoic acid are formed only by elongation of endogenous fatty acids. 4. Most of the radioactivity was found in free fatty acids (63%) and the phospholipid (26%) fraction. In phospholipids, phosphatidylcholine and phosphatidylethanolamine were the two major components. 5. Most of the fatty acids synthesized, including those not normally found in particle lipids (arachidic acid, eicosamonoenoic acid and eicosadienoic acid) were distributed fairly evenly in the phospholipid and free fatty acid fractions. However, stearic acid was found predominantly in the phospholipid fraction.
Explore the source record for details and available documents.
Explore the source record for details and available documents.
We describe a 67-year-old man with lichen planus-like keratosis associated with anti-230 kDa bullous pemphigoid antigen (BPAG1) autoantibody. The patient had noticed solitary dark brown macule more than 6 years previously on his left chest. Histological findings showed hypergranulosis, irregular acanthosis, liquefaction degeneration of basal cells, band-like infiltration of lymphocytes at the subepidermal portion, and a cleft at the basement membrane zone (BMZ), resulting in the formation of subepidermal blisters. Direct immunofluorescence findings of perilesional skin showed a linear deposition of IgG at BMZ. On indirect immunofluorescent study using normal human skin, circulating IgG autoantibody to BMZ was present in the patient's serum at a titer of 1:80. The antigen located on the epidermal site of normal skin split by 1M NaCl was reacted with the patient's serum. Immunoblot analysis using epidermal extracts demonstrated the presence of IgG antibody directed to BPAG1 in the patient's serum. These observations suggest that the presence of an antibody to BPAG1 could be caused by the damage of basal cells following lichen planus-like keratosis.
We describe a 50-year-old woman who noted acral hyperpigmentation, sclerodactyly and Raynaud's phenomenon with 1:160 of antinuclear antibody titer after treatment with a compound drug with tegafur and uracil. Histological findings of the finger and palm included hyperkeratosis, vacuolar degeneration of basal cells, thickened collagen fibers in the dermis, and dilatation of capillary vessels, perivascular mononuclear cell infiltration and melanophages in the upper dermis. IgG, IgA, IgM, C3 and C1q were not deposited in the skin by direct immunofluorescence study. After cessation of the drug, Raynaud's phenomenon and hyperpigmentation disappeared within 1 month and 4 months, respectively, and antinuclear antibody turned negative within 4 months. These observations suggest that tegafur may have caused not only hyperpigmentation in the palms and soles, but also sclerodactyly and Raynaud's phenomenon in the present case.
Four cases on non-A, non-B acute hepatitis have been observed in infants of the same family. No evidence of hepatitis A or B (HBs-Ag, anti-HBs antibodies, HBe-Ag, anti-HBe and anti-HBc antibodies) could be detected in sera. Virological investigations showed no evidence of cytomegalovirus, Epstein-Barr virus, infection, etc. The two younger infants developed disseminated intravascular coagulation and died. The other two recovered without sequelae.