RESUMO
Objective:To investigate the effect of methylene blue (MB) on motor dysfunction and its mechanism in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) mouse models.Methods:Forty healthy male C57BL/6 mice were randomly divided into 4 groups: control group, model group, low-dose treatment group and medium-dose treatment group ( n=10); PD mouse models were established by intraperitoneal injection of 25 mg/kg/d MPTP for a consecutive 7 d; low-dose treatment group and medium-dose treatment group were pretreated intraperitoneally with MB 2 mg/kg/d or MB 10 mg/kg/d for a consecutive 3 d, respectively; and then, MPTP 25 mg/kg/d+MB 2 mg/kg/d or MPTP 25 mg/kg/d+MB 10 mg/kg/d were injected intraperitoneally into the low-dose treatment group or medium-dose treatment group for a consecutive 7 d (MPTP and MB were given at 12 h of interval). Eight d after modeling, open field experiment, pole climbing experiment and rod rotating experiment were carried out to evaluate the spontaneous movement, coordination, endurance and motor ability. And then, the mice were sacrificed; immunofluorescent staining was used to observe tyrosine hydroxylase (TH) expression in the substantia nigra; Western blotting was used to detect the expressions of TH, α-synuclein, nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3), apoptosis-associated speck-like protein containing a CARD (ASC), cleaved-Caspase-1 and Gasdermin D (GSDMD) in the striatum and substantia nigra of mice. Contents of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-18 in the substantia nigra and striatum of mice were detected by ELISA. Results:Compared with the control group, the model group had shortened residence time in rod rotating, prolonged descent time in rod climbing, reduced total movement distance in open field, decreased number of TH-positive cells in the substania nigra, decreased TH protein levels in the substania nigra and striatum, and increased NLRP3, ASC, cleaved-Caspase-1, GSDMD and GSDMD-N protein levels in the substania nigra and striatum, and increased TNF-α, IL-1β and IL-18 contents in the substania nigra and striatum, with significant differences ( P<0.05). Compared with the model group, low-dose treatment group and medium-dose treatment group had prolonged residence time in rod rotating, shortened descent time in rod climbing, increased total movement distance in open field, increased number of TH-positive cells in the substania nigra, and increased TH protein levels in the substania nigra and striatum, decreased NLRP3, ASC, and cleaved-Caspase-1 levels in the substania nigra and striatum, and decreased TNF-α, IL-1β and IL-18 contents in the substania nigra and striatum, with significant differences ( P<0.05). No statistical differences in the above indexes were noted between the low-dose treatment group and medium-dose treatment group ( P>0.05). Conclusion:Low-/medium-dose MB can ameliorate motor dysfunction in PD mouse models, whose mechanism may be related to downregulate NLRP3 inflammasome and inhibit neuroinflammatory response to reduce dopaminergic neuron pyroptosis.
RESUMO
Objective @#To investigate the effects of D ⁃allose on the restoration of neurological function , Galectin⁃3 (Gal⁃3) , adenosine monophosphate⁃activated protein kinase/mammalian target of rapamycin (AMPK/mTOR) and the expression of some inflammatory factors in ischemia⁃reperfusion injury ( CIRI) mice . @*Methods @#A total of 50 male mice were randomly divided into control group (Con group) , sham group (Sham group) , cerebral ischemia⁃reperfusion injury group (MCAO group) , cerebral ischemia⁃reperfusion injury + D ⁃alolose group (MCAO + D ⁃allose group) and cerebral ischemia⁃reperfusion injury + modified citrus pectin group (MCAO + MCP group) . The middle cerebral artery occlusion/reperfusion (MCAO/R) model (reperfusion after 2 hours of MCA ischemia) was established by thread embolism . After successful modeling , the neurological function of mice was evaluated Longa score and rotated rod walking . TTC staining was used to observe the volume of cerebral infarction foci . The expression levels of Gal⁃3 and autophagy⁃related molecules were detected by Western blot and RT⁃PCR . Immunofluorescence was applied to detect the distribution of Gal⁃3 in brain tissue , and TNF⁃α , IL⁃8 secretion was detected with ELISA KIT . @*Results @#Compared with Con group and Sham group , the MCAO model represented significant increase in the Longa neurofunction score (P < 0. 01) , cerebral infarction volume ( P < 0. 01) , Gal⁃3 expression and manifasted enhanced autophagy (P < 0. 01) . After treatment with D ⁃allose , it could significantly improve neurological dysfunction , reduce cerebral infarction volume (P < 0. 01) , reduce the expression of Gal⁃3 ( P < 0. 01) , inhibit AMPK phosphorylation , promote mTOR phosphorylation , and inhibit autophagy (P < 0. 01) . The use of the Gal⁃3 inhibitor MCP alone could also achieve the effect of inhibiting autophagy . @*Conclusion @# D ⁃allose can effectively promote the recovery of neurological function and reduce the volume of infarct foci in CIRI mice . The mechanism may involve inhibiting excessive cell autophagy by downregulating the expression of Gal⁃3 , and reducing the release of inflammatory factors such as TNF⁃α and IL⁃8 , thereby exerting neuroprotective effects .