The localization of cholinephosphotransferase in the outer membrane of guinea-pig lung mitochondria

Sikpi, Matthew O.; Das, Salil K.

doi:10.1016/0005-2736(87)90236-7

Cited by 19 publications

(8 citation statements)

References 35 publications

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“…at incubation times of 1 h and 4 h. This decrease was more acute in microsomes than that in mitochondria ( Figure 2). We have previously demonstrated that in addition to its predominant localization in the microsomes, CPT exists also in mitochondria [38,39]. Thus, it is possible that during early stage of lung injury as observed in this study, cells try to repair the membrane damage by stimulating PC synthesis and therefore increased CPT activity, but with time lung cells lose their ability to repair membrane damage as evident from decreased CPT activity in both mitochondria and microsomes isolated from lung of CEES-treated animals (Figures 1 and 2).…”

Section: Discussionmentioning

confidence: 99%

Inhibition of cholinephosphotransferase activity in lung injury induced by 2-chloroethyl ethyl sulfide, a mustard analog

Roy

Mukherjee

Kabir

et al. 2005

J. Biochem. Mol. Toxicol.

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Exposure to mustard gas causes inflammatory lung diseases including acute respiratory distress syndrome (ARDS). A defect in the lung surfactant system has been implicated as a cause of ARDS. A major component of lung surfactant is dipalmitoyl phosphatidylcholine (DPPC) and the major pathway for its synthesis is the cytidine diphosphocholine (CDP-choline) pathway. It is not known whether the ARDS induced by mustard gas is mediated by its direct effects on some of the enzymes in the CDP-choline pathway. In the present study we investigated whether mustard gas exposure modulates the activity of cholinephosphotransferase (CPT) the terminal enzyme by CDP-choline pathway. Adult guinea pigs were intratracheally infused with single doses of 2-chloroethyl ethyl sulfide (CEES) (0.5 mg/kg b.wt. in ethanol). Control animals were injected with vehicles only. The animals were sacrificed at different time and the lungs were removed after perfusion with physiological saline. CPT activity increased steadily up to 4 h and then decreased at 6 h and stabilized at 7 days in both mitochondria and microsomes. To determine the dose-dependent effect of CEES on CPT activity we varied the doses of CEES (0.5-6.0 mg/kg b.wt.) and sacrificed the animals at 1 h and 4 h. CPT activity showed a dose-dependent increase of up to 2.0 mg/kg b.wt. of CEES in both mitochondria and microsomes then decreased at 4.0 mg/kg b.wt. For further studies we used a fixed single dose of CEES (2.0 mg/kg b.wt.) and fixed exposure time (7 days). Lung injury was determined by measuring the leakage of iodinated-bovine serum albumin into lung tissue and expressed as the permeability index. CEES exposure (2.0 mg/kg b.wt. for 7 days) caused a significant decrease of both CPT gene expression (approximately 1.7-fold) and activity (approximately 1.5-fold) in the lung. This decrease in CPT activity was not associated with any mutation of the CPT gene. Previously we reported that CEES infusion increased the production of ceramides which are known to modulate PC synthesis. To determine whether ceramides affect microsomal CPT activity the lung microsomal fraction was incubated with different concentrations of C(2)-ceramide prior to CPT assay. CPT activity decreased significantly with increasing dose and time. The present study indicates that CEES causes lung injury and significantly decreases CPT gene expression and activity. This decrease in CPT activity was not associated with any mutation of the CPT gene is probably mediated by accumulation of ceramides. CEES induced ceramide accumulation may thus play an important role in the development of ARDS by modulating CPT enzyme.

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Section: Discussionmentioning

confidence: 99%

Inhibition of cholinephosphotransferase activity in lung injury induced by 2-chloroethyl ethyl sulfide, a mustard analog

Roy

Mukherjee

Kabir

et al. 2005

J. Biochem. Mol. Toxicol.

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“…The fact that CPT antibody inhibited CPT activity of guinea pig liver mitochondria and microsomes in a dose dependent manner, it supports strongly the specificity of the antibody towards the CPT. We have used liver microsomes and mitochondrial fractions for our immunoblot analysis and immuno inhibition study as the existence of CPT in these two fractions were well characterized (1,8,10). This is the first demonstration of the antibody to cholinephosphotransferase.…”

Section: Discussionmentioning

confidence: 99%

“…The liver from animals were quickly excised, washed in ice cold saline (0.9%), blotted dry, weighed and homogenized in 10 vol of 0.25 M sucrose, 1 mM EDTA (pH 7.4) using a Potter Elvehjem homogenizer. After the removal of nuclei and cell debris by centrifugation at 600g for 10 min, mitochondria were sedimented at 10,000g for 10 min in a Sorvall SS-34 rotor at 4°C and further purified by sucrose density gradient centrifugation as described before (1,8). The 10,000g supernatant fluid was centrifuged at 105,000g for 30 min to sediment the microsomes.…”

Section: Dot Blot Analysismentioning

confidence: 99%

Development and Characterization of Cholinephosphotransferase Antibody

Chatterjee

Mukherjee

Das

2001

Biochemical and Biophysical Research Communications

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“…The liver from treated and control animals were quickly excised, washed in ice cold saline (0.9%), blotted dry, weighed, and homogenized in 10 vol of 0.25 M sucrose, 1 mM EDTA (pH 7.4) using a Potter Elvehjem homogenizer. After the removal of nuclei and cell debris by centrifugation at 600g for 10 min, mitochondria were sedimented at 10,000g for 10 min in a Sorvall SS-34 rotor at 4°C and further purified by sucrose density gradient centrifugation as described before (4,5). The 10,000g supernatant fluid was centrifuged at 105,000g for 30 min to sediment the microsomes.…”

Section: Methodsmentioning

confidence: 99%

“…The purity of the mitochondrial and microsomal fractions were monitored by assaying the marker enzymes: monoamino oxidase, succinic dehydrogenase, and rotenone sensitive NADPH cytochrome c reductase (4,5). Cholinephosphotransferase activity was assayed by measuring the incorporation of radioactivity from cytidine diphospho-[methyl-…”

Section: Methodsmentioning

confidence: 99%