PARP-1 inhibitor alleviates cerebral ischemia/reperfusion injury by reducing PARylation of HK-1 and LDH in mice.

Poly (ADP-ribose) polymerase-1 (PARP-1) activity significantly increases during cerebral ischemia/reperfusion. PARP-1 is an NAD+-consumption enzyme. PARP-1 hyperactivity causes intracellular NAD+ deficiency and bioenergetic collapse, contributing to neuronal death. Besides, the powerful trigger of PARP-1 causes the catalyzation of poly (ADP-ribosyl)ation (PARylation), a posttranslational modification of proteins. Here, we found that PARP-1 was activated in the ischemic brain tissue during middle-cerebral-artery occlusion and reperfusion (MCAO/R) for 24 h, and PAR accumulated in the neurons in mice. Using immunoprecipitation, Western blotting, liquid chromatography-mass spectrometry, and 3D-modeling analysis, we revealed that the activation of PARP-1 caused PARylation of hexokinase-1 and lactate dehydrogenase-B, which, therefore, caused the inhibition of these enzyme activities and the resulting cell energy metabolism collapse. PARP-1 inhibition significantly reversed the activity of hexokinase and lactate dehydrogenase, decreased infarct volume, and improved neuronal deficiency. PARP-1 inhibitor combined with pyruvate further alleviated MCAO/R-induced ischemic brain injury in mice. As such, we conclude that PARP-1 inhibitor alleviates neuronal death partly by inhibiting the PARylation of metabolic-related enzymes and reversing metabolism reprogramming during cerebral ischemia/reperfusion injury in mice. PARP-1 inhibitor combined with pyruvate might be a promising therapeutic approach against brain ischemia/reperfusion injury.

The Depletion of NAMPT Disturbs Mitochondrial Homeostasis and Causes Neuronal Degeneration in Mouse Hippocampus.

Nicotinamide phosphoribosyltransferase (NAMPT) is the key enzyme in the salvaging synthesis pathway of the nicotinamide adenine dinucleotide (NAD). Both NAMPT and NAD progressively decline upon aging and neurodegenerative diseases. The depletion of NAMPT induces mitochondrial dysfunction in motor neurons and causes bioenergetic stress in neurons. However, the roles of NAMPT in hippocampus neurons need to be further studied. Using floxed Nampt (Namptflox/flox) mice, we knocked out Nampt specifically in the hippocampus CA1 neurons by injecting rAAV-hSyn-Cre-APRE-pA. The depletion of NAMPT in hippocampus neurons induced cognitive deficiency in mice. Nevertheless, no morphological change of hippocampus neurons was observed with immunofluorescent imaging. Under the transmission electron microscope, we observed mitochondrial swollen and mitochondrial number decreasing in the cell body and the neurites of hippocampus neurons. In addition, we found the intracellular Aß (6E10) increased in the hippocampus CA1 region. The intensity of Aß42 remained unchanged, but it tended to aggregate. The GFAP level, an astrocyte marker, and the Iba1 level, a microglia marker, significantly increased in the mouse hippocampus. In the primary cultured rat neurons, NAMPT inhibition by FK866 decreased the NAD level of neurons at > 10-9 M. FK866 dropped the mitochondrial membrane potential in the cell body of neurons at > 10-9 M and in the dendrite of neurons at > 10-8 M. FK866 decreased the number and shortened the length of branches of neurons at > 10-7 M. Together, likely due to the injury of mitochondria, the decline of NAMPT level can be a critical risk factor for neurodegeneration.

Nicotinamide phosphoribosyltransferase inhibitor ameliorates mouse aging-induced cognitive impairment.

Nicotinamide phosphoribosyltransferase (NAMPT) is the key enzyme for the synthesis of nicotinamide adenine dinucleotide (NAD) in the salvaging pathway. Though NAMPT inhibitors such as FK866 were originally developed as anti-cancer drugs, they also display neuroprotective effects. Here we show that the administration of FK866 at 0.5 mg/kg (ip, qod) for four weeks, i.e., ∼1% of the dose used for the treatment of cancer, significantly alleviates the aging-induced impairment of cognition and locomotor activity. Mechanistically, FK866 enhanced autophagy, reduced protein aggregation, and inhibited neuroinflammation indicated by decreasing TNFα, IL-6, GFAP, and Iba1 levels in the aged mouse brain. Though FK866 did not affect the total NAD and nicotinamide mononucleotide (NMN) levels in the mouse brain at the dose we used, FK866 increased nicotinamide (NAM) level in the young mouse brain and decreased NAM level in the aged mouse brain. On the other hand, FK866 did not affect the serum glucose, cholesterol, and triglyceride of young and aged mice and exhibited no effects on the various indices of young mice. Thus, the NAMPT inhibitor can be repurpose to counteract the cognitive impairment upon aging. We also envision that NAMPT inhibitor can be used for the treatment of age-related neurodegenerative diseases.