Effect Of Pyrroloquinoline Quinone(PQQ) On Exercise Capacity And Its Mechanism In Mice Exposed To Acute High Altitude

Plateau is an important part of our territory,accounting for 26%of the land area of our country.Low pressure and low oxygen are its prominent environmental factors.As the altitude rises,the partial pressure of oxygen becomes lower,especially at altitudes above 3500m,the exercise capacity and labor efficiency of the human organism decrease significantly.However,the potential molecular mechanism underlying the decrease in exercise capacity in the plateau environment is still unclear,and the interventions for preventing acute plateau reaction and improving physical work capacity are still in a scarce stage.It is of great practical and military significance to find small-molecule drugs to improve the exercise capacity in the face of the increasingly frequent plateau emergencies.Pyrroloquinoline quinone(PQQ)is an aromatic tricyclic orthoquinone that cannot be synthesized in mammals and is mainly obtained through diet.Experiments have confirmed that PQQ,as a redox cofactor,can stimulate mitochondrial genesis,protect mitochondria from free radical oxidative damage,improve energy utilization,and regulate basal energy metabolism.The effect of PQQ on mitochondrial function under hypoxic conditions has not been reported;therefore,the present study focused on the improvement of PQQ on the exercise capacity of mice under simulated plateau hypoxic conditions and the study of its mitochondrial mechanism.PurposeTo investigate the effect of PQQ on the improvement of exercise ability in mice under acute low-pressure hypoxic environment and its molecular mechanism.Methods1.Effects of different duration of hypoxic exposure on blood biochemical indexes.Male Kunming mice were exposed to acute plateau hypoxia for 12 h,7 d and 28 d using a low-pressure oxygen chamber simulating a low-pressure hypoxic environment at 6000m above sea level.The serum glucose(Glu),triglycerides(TG),creatine kinase(CK),creatine kinase isoenzyme(CK-MB),α-hydroxybutyrate dehydrogenase(α-HBDH),CO2,homocysteine(HCY)and lactate dehydrogenase(LDH)were measured using a fully automated dry biochemical analyzer..2.Effect of PQQ on the anti-hypoxic capacity of mice and the locomotor capacity of mice exposed to acute low-pressure hypoxia.Male Kunming mice were given different doses of PQQ(5,10 mg/kg.bid/d)and CoQ10(20 mg/kg.qd/d)by gavage for 7 d.The normobaric hypoxia tolerance and decompression hypoxia tolerance of mice were evaluated by the normobaric confinement hypoxia test and 10,000 m decompression hypoxia test,respectively.The exercise capacity of mice was evaluated by simulated hypoxic weight-bearing swimming test at 4000m above sea level.The EKF portable lactate meter was also used to detect the blood lactate in the mice with hypoxic swimming;the biochemical kits and biochemical analysers were used to detect the changes of liver glycogen and serum superoxide dismutase,serum malondialdehyde and serum urea.3.Study on the molecular mechanism of PQQ to improve anti-hypoxia and exercise capacity in mice exposed to acute low pressure hypoxia.Male Kunming mice were given different doses of PQQ(5 and 10 mg/kg.bid/d)and CoQ10(20 mg/kg.qd/d)by gavage,and after 7 d,the exposed mice were placed in a low-pressure oxygen chamber at a simulated altitude of 6000m for 12 h.Changes in the relative copy number of cardiac mitochondrial DNA were measured by real-time fluorescence quantitative PCR,and protein immunoblotting was used to detect The expression levels of mitochondrial-associated protein mitochondrial transcription factor A(Tfam),mitochondrial fusion protein 2(Mfn2),optic nerve atrophy-associated protein 1(Opal),and mitochondrial splitting protein 1(Fis1)were detected using real-time fluorescence quantitative PCR.Rat cardiomyocytes H9C2 were cultured using a three-gas incubator at 1%oxygen partial pressure while administered with different concentrations of PQQ(5 μmol/L、2 μmol/L、1 μmol/L、0.5 μmol/L、0.1 μmol/L、0.01 μmol/L)and incubated for 12 h and 24 h,respectively,and cell viability was detected by CCK8 to determine the intervention concentration of PQQ.The PQQ concentrations of 0.1 mol/L and 1 mol/L were selected to intervene for 12 h and 24 h,respectively,and the cellular ROS level was detected by luminescence enzyme standardization;the cellular ATP level was detected by chemiluminescence.Result1.Compared with the normoxic control group,GLU was significantly elevated in the 12 h and 7 d exposure groups(P<0.05),while GLU was significantly lower in the 28 d exposure group(P<0.001);CO2 was significantly decreased in the exposure group of 12 h,7 d and 28 d(P<0.01);HCY and TG were significantly higher in the 12 h,7 d and 28 d exposure groups(P<0.05).CK,CK-MB,α-HBDH and LDH were not significantly changed in the 12 h and 7 d exposure groups,but were significantly increased in the 28 d exposure group(P<0.05).These results suggest that the molecular regulation mechanisms of acute hypoxia and chronic hypoxia are different,causing different changes in serum biochemical parameters in mice.2.The PQQ intervention group significantly improved the survival time(P<0.05)of the mice in the normal pressure airtight hypoxia experiment and the survival rate of 10000m acute decompression hypoxia.The experimental results suggest that PQQ has a significant anti-hypoxia effect.Compared with the hypoxic control group,the swimming exhaustion time of the PQQ intervention group was significantly increased(P<0.05),the liver glycogen(P<0.05)and serum SOD levels(P<0.01)were up-regulated,and the blood lactic acid(P<0.05),blood urea(P<0.05)and MDA levels were down-regulated.The experimental results suggest that PQQ can improve the exercise capacity of mice exposed to acute hypoxia and reduce oxidative stress damage.3.Compared with the normoxic control group,the heart mitochondrial copy number,Tfam,Mfn2,and Opal protein expression of the hypoxia control group decreased,and the Fis1 protein expression increased.Compared with the hypoxia control group,PQQ intervention significantly up-regulated the expression of mitochondrial copy number,Tfam,Mfn2,and Opal,and down-regulated the expression of Fis1 protein.The viability of H9C2 cells in the low-dose PQQ intervention group increased significantly,but did not show a significant dose-dependence.Meanwhile,the ATP levels of H9C2 cells in hypoxic culture were significantly increased and ROS levels were significantly decreased after 0.1 μmol/L and 1 μmol/L PQQ interventions.The experimental results suggest that the molecular mechanism of anti-hypoxia and exercise capacity of PQQ may be related to its regulation of cardiac mitochondrial biogenesis,mitochondrial fusion-division dynamics,mitochondrial oxidative stress and energy metabolism.Conclusion1.Acute and chronic low-pressure hypoxia exposure have different molecular regulatory mechanisms and different physiological and biochemical effects.2.PQQ has the effect of improving anti-hypoxia and exercise capacity under acute low-pressure hypoxic conditions,and reducing oxidative stress damage under low-pressure hypoxia.3.The molecular mechanism of PQQ improving anti-hypoxia and exercise ability under acute altitude hypoxia may be related to its regulation of cardiac mitochondrial biogenesis,mitochondrial fusion-division dynamics,mitochondrial oxidative stress and energy metabolism.