CARNITINE PALMITOYL-TRANSFERASE ENZYME-INHIBITION PROTECTS PROXIMAL TUBULES DURING HYPOXIA

The role of inhibition of the CPT enzymes responsible for accumulation of long chain acylcarnitines (LCAC) during hypoxia in the proximal tu bule has not been previously examined. We have characterized CPT enzym e activities in mitochondrial fractions of rabbit proximal tubules. Ma lonyl CoA-sensitive CPT I activity (1.1 +/- 0.3 nmol/min/mg protein): and detergent-solubilized, malonyl CoA-insensitive CPT II activity (2. 3 +/- 0.4 nmol/min/mg protein) were readily detected in proximal tubul e mitochondrial fractions. Subjecting rabbit proximal tubules to vario us periods of hypoxia did not significantly change mitochondrial CPT I or CPT II activities. Thirty minutes of hypoxia resulted in an increa se in lysophospholipid mass from 440 +/- 105 to 720 +/- 93 pmol/mg pro tein, N = 5, LCAC mass from 79 +/- 11 to 618 +/- 34 pmol/mg protein; N = 5, and lactate dehydrogenase (LDH) release from 9 +/- 1% to 46 +/- 3%, N = 8. Pretreatment of proximal tubules with two different CPT inh ibitors, glybenclamide (Glyb) 400 mu M and oxfenicine (Oxfe) 1 mM, res ulted in reduction in the magnitude of hypoxia-induced lysophospholipi d formation 490 +/- 160 (Glyb), 342 +/- 150 pmol/mg protein (Oxfe), N = 4, hypoxia-induced LCAC formation 295 +/- 27 (Glyb), 128 +/- 16 pmol /mg protein (Oxfe), N = 5, and LDH release 25 +/- 1% (Glyb) and 19 +/- 2% (Oxfe), N = 8. The protective effect of CPT inhibition was also as sociated with increased production of lactate suggesting the modulatio n of a substrate-mediated metabolic switch. Immunoblots demonstrated t hat hypoxia caused a time dependent hydrolysis of fodrin-alpha subunit and that CPT inhibition protected against hypoxia-induced fodrin prot eolysis. These data suggest a unifying hypothesis that links phospholi pase A(2) (PLA(2)) activation, and hypoxia-mediated fodrin proteolysis to the proximal tubule mitochondrial CPT system. I propose that CPT i nhibition may represent a novel mechanism to ameliorate proximal tubul e cell death during hypoxia.