ABSTRACT
ObjectiveTo study the effects of Wendantang on the expression of miRNA-219， N-methyl-D-aspartate receptor 2B （NR2B）， disrupted in schizophrenia 1 （DISC1）， and Ca2+/calmodulin-dependent protein kinase Ⅱγ （CaMKⅡγ） in the frontal lobe of rats with schizophrenia. MethodSixty rats were randomly divided into six groups， namely normal group， model group， high-， medium-， and low-dose Wendantang groups， and clozapine group， with 10 rats in each group. Rats in high-， medium-， and low-dose Wendantang groups were intragastric with 40， 20， and 10 g·kg-1 Wendantang， and the ones in clozapine group were intragastric with 0.02 g·kg-1 clozapine， those in normal and model group were intragastric with equal volume of normal saline， once a day. After 21 days of administration， rats in all groups except for the normal group were injected with 0.6 mg·kg-1 dizocilpine maleate （MK-801） into the left abdominal cavity for inducing acute schizophrenia. The stereotypic behavior and ataxia in rats were scored according to SAMS and HOFFMAN criteria. The morphological changes in the prefrontal cortex were observed by hematoxylin-eosin （HE） staining. The protein expression levels of NR2B, DISC1 and CaMKⅡγ in the frontal lobe was detected by Western blot. The mRNA expression levels of miRNA-219 was detected by real-time fluorescence quantitative polymerase chain reaction（Real-time PCR）. ResultCompared with normal group， the model group exhibited significantly increased stereotypic behavior and ataxia scores （P<0.01）， karyopyknosis and karyolysis in most neurons of the prefrontal cortex， and down-regulated NR2B， DISC1， and CaMKⅡγ protein expression （P<0.01） and miRNA-219， NR2B， DISC1， and CaMKⅡγ mRNA expression （P<0.01）. Compared with model group， Wendantang high-， medium-， and low-doses group lowered the scores of stereotypic behavior and ataxia at 50， 60 mmin（P<0.05，P<0.01）. In high- and medium-dose Wendantang groups， the neurons in the prefrontal cortex were densely arranged. The karyopyknosis and karyolysis were alleviated to varying degrees. The NR2B protein expression in the frontal lobe was up-regulated （P<0.01）. In the medium- and low-dose Wendantang groups， the DISC1 protein expression in the frontal lobe was up-regulated （P<0.05，P<0.01）. Wendantang at each dose significantly increased the CaMKⅡγ protein expression （P<0.05） and miRNA-219， NR2B， DISC1， and CaMKⅡγ mRNA expression in the frontal lobe （P<0.05，P<0.01）. ConclusionWendantang improves the scores of stereotypical behavior and ataxia， relieves the karyopyknosis and karyolysis of neurons in the prefrontal cortex， and increases the expression levels of miRNA-219， NR2B， DISC1， and CaMKⅡγ of rats with schizophrenia， so as to alleviate the schizophrenic-like symptoms and schizophrenia.
ABSTRACT
Objective:To investigate the protective effect of Wendantang-containing serum on astrocytes in glutamate environment and its effect on phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt)/glycogen synthetase kinase 3β (GSK3β) signal pathway. Method:Totally 60 rats were randomly divided into 5 groups, normal group (n=20), clozapine group, and high, medium and low-dose Wendantang groups, with 10 rats in each group. Normal group was given 20 mL·kg-1 normal saline, clozapine group was given 20 mg·kg-1 clozapine, Wendantang groups were given 40, 20, 10 g·kg-1 Wendantang, once a day. Eight days later, the rats were killed, their blood was taken, serum was centrifuged, inactivated, filtrated, sterilized and filled in separate centrifugal tubes. The astrocytes were divided into normal group, model group, clozapine group, and high, medium and low-dose Wendantang groups. The normal group and the model group were cultured with normal serum, and the rest groups were cultured with corresponding drug containing serum. The other groups were treated with 10 mmol·L-1 glutamic acid for 24 hours, and then the apoptosis of astrocytes was detected by flow cytometry. Western blot and Real-time fluorescent quantitative polymerase chain reaction (Real-time PCR) were used to determine protein, and mRNA expressions of PI3K, Akt, GSK3β in astrocytes. Result:Compared with the normal group, apoptosis in model group increased significantly (P<0.01). Compared with the model group, apoptosis in Wendantang groups decreased significantly (P<0.01). Compared with the normal group, protein and phosphorylation expressions of PI3K, Akt, GSK3β in model groups decreased significantly (P<0.05, P<0.01). Compared with the model group, protein and phosphorylation expressions of PI3K, Akt, GSK3β in Wendantang groups increased (P<0.05, P<0.01). Compared with the normal group, expressions of PI3K, Akt and GSK3β mRNA decreased in model group(P<0.01). Compared with the model group, expressions of PI3K, Akt and GSK3β mRNA increased significantly in Wendantang groups group (P<0.05，P<0.01). Conclusion:Wendantang-containing serum can effectively increase expressions of PI3K, Akt and GSK3β, so as to regulate PI3K/Akt/GSK3β signal pathway and protect nerve cells.
ABSTRACT
Objective:To investigate the effect of Wendantang on cyclic adenosine monophosphate （cAMP）-response element binding protein（CREB） gene silencing hippocampal cell activity， apoptosis and signal pathway of brain-derived neurotrophic factor/protomyosin related receptor kinase B/adenosine cyclophosphate effector binding protein （BDNF/TrkB/CREB）. Method:Wendantang-containing serum was prepared. Animal grouping： SD male rats were randomly divided into high， medium， low-dose groups， clozapine group and normal saline group， with 10 rats in each group， while 15 rats for the normal group. Dosage： 20 mL·kg-1 normal saline was given to normal group N， clozapine 0.02 g·kg-1 was given to dozapine group X， while high， medium and low-dose Wendantang groups were respectively given the same amount of Wendantang concentrated crude drug， with concentrations of 2， 1 and 0.5 g·mL-1 respectively once a day for 8 days continuously， and then blood was taken from femoral artery， and centrifuged for 15 min at 5 000 r·min-1. Supernatant was taken， inactivated， stored at -80 ℃ for standby. The CREB gene silenced hippocampal neuron cell line was constructed through transfection of liposomes into hippocampal cells， and Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to verify the effect of small interfering RNA （siRNA） transcription. The mRNA expressions of BDNF， TrkB， CREB and CaMKⅡ in normal hippocampal cells and CREB gene silenced hippocampal cells were measured. Result:Compared with normal group， the apoptosis of the normal gene silencing group was significantly increased （P<0.01）， compared with the normal gene silencing group， the apoptosis of each group was significantly reduced （P<0.01）. As for the mRNA expressions of BDNF， TrkB， CREB and CaMKⅡ， compared with the normal group， the mRNA expression of CREB， BDNF in the normal gene silencing group was significantly decreased （P<0.01）. Compared with the normal gene silencing group， the mRNA expression of BDNF in each administration group was highly increased （P<0.01）， but with no statistically significant difference between TrkB and CaMKⅡ groups. Conclusion:The Wendantang-containing serum could improve the mRNA expression of BDNF， protect hippocampal neurons and prevent cognitive impairment of schizophrenia by regulating BDNF/TrkB/CREB signal pathway.