Metformin Improves Cognitive And Memory Dysfunction In Mice With Hepatic Encephalopathy By Inhibiting Neuroinflammation Through Modulation Of The CGAS/STING Signaling Pathway

Objective: Hepatic encephalopathy(HE)is a syndrome of central nervous system dysfunction caused by severe liver disease and based on metabolic disorders,manifested by brain metabolic disorders,motor and neurocognitive deficits,typically characterized by fluttering tremor,etc.The onset of HE is insidious and not easily detected in the early stages,mainly manifested by neuropsychological or psychological test abnormalities,such as The main manifestations are neuropsychological or psychological test abnormalities,such as: cognitive impairment,memory loss,personality and behavioral changes,etc.In this study,we established a mouse model of Hyperammonia(HY)and used Metformin(MET)for prophylactic treatment to investigate the effect of MET on cognitive and memory impairment and the mechanism of neuroinflammation regulation of HE.The preliminary investigation of the effect of MET on HE cognitive and memory impairment and the modulation mechanism of neuroinflammation provides a new idea for effective prevention and treatment of HE,in order to contribute to HE prevention and treatment.Methods: Adult male ICR mice(6-8 weeks old)were randomly divided into control group(Control),model group(UR),metformin group(MET),and model group + metformin group(UR + MET),and drug intervention experiments were conducted after one week of adaptive feeding.mice in the Control group were given saline for 10 days continuously;mice in the UR group were given saline for 7 days first and then The mice in the MET group were given MET(300 mg/kg)for 7 days,followed by MET and saline for the next 3 days,and in the UR+MET group,MET was given prophylactically for 7days,followed by MET and UR for 3 days.Behavioral tests were performed on the next day: the Morris water maze test was performed to assess the behavioral performance of each group of mice.The expression of cyclic GMP-AMP synthase(c GAS)and stimulator of interferon gene(STING)in the prefrontal cortex of mice was measured by western blot(WB).The expression of the interferon gene(STING)was measured by real-time fluorescence PCR(q PCR).The expression of interferon regulator 7(Irf7)and the release of mitochondrial DNA(mt DNA)in the cytoplasm of the mice were observed.Results: RESULTS: First,we performed behavioral analysis to detect neurobehavioral parameters in mice.In the Morris water maze positioning navigation experiment,there was basically no difference in the time spent searching the platform between the mice in each group on day 1(P>0.05);from day 2 onwards,the time spent searching the platform was significantly increased in the UR group compared to the Control group(P<0.0001),and after treatment with MET intervention,the MET and UR+MET groups compared to the Control group,respectively There were no significant differences(P>0.05),and the latency time was significantly shorter in the UR+MET group compared to the UR group(P<0.0001).In the Morris water maze spatial exploration experimental period,mice in the UR group traversed the original platform less often(P<0.01)and spent less time in the original platform quadrant compared with the Control group(P<0.001),while the UR+MET group traversed the original platform more often(P<0.01)and spent more percentage of time in the original platform quadrant compared with the UR group(P<0.001).The number of neurons in the prefrontal cortex of the mice in the Control group was high,and more NICs were visible,and the neurons were dense and dark;compared with the Control group,the number of neurons in the UR group became less(P<0.001),and the NICs were lightly colored and had shrunken cytosomes.In contrast,the number and size of neurons in the prefrontal cortex of the mice in the MET and UR+MET groups were not statistically different from those in the Control group(P>0.05),and the number of cortical neurons was higher in the UR+MET group compared with the UR+MET group,with darker coloring and fuller cytosomes in the UR+MET group.Next,to detect the function of mitochondria,we quantified the intracytoplasmic mt DNA content.Compared with the Control group,mt DNA levels in the cytoplasm of the prefrontal cortex of the UR mice were increased(P<0.01),whereas mt DNA levels in the cytoplasm of the prefrontal cortex of the UR+MET mice were decreased after MET intervention(P<0.05).Subsequently,we explored the regulatory mechanism of neuroinflammation.Compared with the Control group,c GAS and STING protein expressions were significantly higher in the cerebral cortex of UR mice(P<0.01),and decreased in the UR+MET group after MET intervention(P<0.01).Finally,to detect whether the cGAS/STING signaling pathway was fully activated,we examined the expression of its downstream inflammatory factors.Compared with the Control group,the expression of inflammatory factors ISG15,PKR,and Irf7 in the prefrontal cortex of mice in the UR group was significantly upregulated(P<0.01),while the expression of inflammatory factors ISG15,PKR,and Irf7 in the prefrontal cortex of mice in the UR+MET group was significantly decreased after administration of MET intervention treatment(P<0.05).Conclusions: MET may improve learning and memory dysfunction in HE mice by inhibiting c GAS-STING signaling pathway,downregulating Irf7,ISG15,and PKR inflammatory factors,reducing neuroinflammation,and exerting a protective effect on neurons.To provide new ideas for effective prevention and treatment of cognitive dysfunction in HE,with a view to contribute to the discovery and treatment of HE.