Objective To investigate the protective effects of purified effective component group in extract from Xiaoshuan Tongluo(CGXT)formula on chronic brain ischemia in rats.Methods CGXT 75,150,and 300 mg/kg or vehicle were ig administered daily for four weeks to rats with bilateral common carotid arteries ligation(BCCAL).From the day 24 to 28 after BCCAL,Morris water maze was performed to assess the learning and memory impairment of rats.Four weeks after BCCAL,brain gray and white matter damage were assessed.Results In Morris test,the mean escape latency of rats in the CGXT(150 and 300 mg/kg)groups was significantly shorter than that in the vehicle group.CGXT also attenuated the neuronal damage in hippocampus and cortex and reduced the pathological damage in the optic tract and corpus callosum.Conclusion CGXT could improve learning and memory impairment resulted from BCCAL in rats.These results provide the experimental basis for the clinical use of CGXT in stroke treatment and may help in investigation of multimodal therapy strategies in ischemic cerebrovascular diseases including stroke.

Objective To compare the biological characteristics of several different anxiety rat models established by different methods of stress at different time points and provide experimental basis for the most appropriate modeling methods .Methods 60 rats were randomly divided into normal , empty bottle stress , chronic emotional stress ( CES ) group, restraint stress for 3h, 6h, and modeling respectively .In the experimental 7 d, 14 d, 21 d, elevated plus maze and fear condition system was used to test anxiety-like behavior in rats , open field test to study anxiety or depression-like behavior , forced swimming test was used to detect depression-like behavior in rats , and using the Elisa test kit to detect the contents of 5-HT, DA in the hippocampus in rats .Results Anxiety-like behavioral test results showed that rats in empty bottle stress, CES, 6 h restraint stress group started to have anxiety-like behavior since 14 d, then anxiety-like behavior was becoming increasingly apparent .Forced swimming test results showed that immobility time in 6 h restraint rats was significantly increased in the first 7 d(P <0.05).Meanwhile, compared with control group, hippocampal 5-HT, DA contents in empty bottle stress and CES rats increased significantly since 14 d.Conclusions Among several stress methods established anxiety model , anxiety-like behavior in 3 h restraint stress was not obvious; 6 h restraint stress exhibited a depression-like behavior in the forced swimming test might be due to prolonged stress .Empty bottle stress and CES can successfully establish the anxiety rat model , and the anxiety behavior of the rats have some differences . Corresponding model methods can be selected according to different experimental purposes .

Objective To observe the effects of Zuogui Jiangtang Jieyu Formula (ZJJF) on the ability of learning and memory and the expressions of JNK, Bcl-2 and Bax in hippocampus in diabetic rats with depression; To explore the protective mechanism of hippocampal damage in diabetic rats with depression. Methods High-fat gavage combined with intravenous injection of STZ was used to establish the model of diabetic rats. 28 days of chronic stress was given continuously and diabetic rats complicated with depression were built successfully. Then rats were randomly divided into 6 groups, including normal, model, positive medicine, high-, medium-, and low-dose of ZJJF groups. After the last administration, Morris water maze was used to detect escape latency time;Western blot was used to disclose the protein expressions of JNK, Bcl-2 and Bax in rat hippocampaus;RT-PCR was used to test the gene expressions of JNK and Bcl-2 and Bax. Results Compared with the normal group, escape latency time in model rats was significant longer (P<0.01), the protein and gene expression of JNK and Bax in rat hippocampaus significantly increased, Bcl-2 was markedly decreased (P<0.05, P<0.01);Compared with the model group, escape latency time in positive medicine group and high-dose of ZJJF group was significant shorter (P<0.01), the protein and gene expressions of JNK and Bax significantly decreased, and Bcl-2 markedly increased (P<0.05, P<0.01). Conclusion ZJJF can significantly improve the ability of learning and memory in diabetic rats with depression, which might be associated with preventing neuronal apoptosis in hippocampus.

Objective To study the content of monoamine neurotransmitters and neurotrophic factor in the hippocampus, amygdala and prefrontal cortex in anxious depression rats, and explore the possible pathogenesis.Methods 60 SD rats were randomly divided into normal group, vehicle group, anxiety group, depression group, and anxious depression group, 12 rats in each group.Chronic restraint stress combined with corticosterone injection was used to establish anxiety and depression model, the modeling time was 21 d.After modeling, elevated plus maze test, open field test, and forced swimming test were used to evaluate the anxiety and depression-like behavior, HPLC-ECD was used to detect the content of 5-HT, NE, and DA in the hippocampus, amygdala, and prefrontal cortex of rats.Western-blotting was used to detect the expression of BDNF and NT-3 in rats.Results Rats in anxious depression model group were comparable to the anxiety group in time and frequency entering open arm time, and number of locomotor activity in open field, and it had a significant difference when compared with the control and depression groups (P<0.01 or P<0.05).Immobile time in anxious depression model rats was increased significantly when compared with the control and anxiety groups (P<0.01).Meanwhile, compared with the control group, 5-HT in hippocampus and 5-HT, NE in amygdala or prefrontal cortex were significantly decreased in the depressive rats with anxiety (P<0.01 or P<0.05).Moreover, the content of BDNF and NT-3 was significantly decreased in each brain regions compared with the control group (P<0.01 or P<0.05), and BDNF levels were obviously decreased compared with the anxiety group (P<0.05).Conclusions Rats of anxious depression have significant anxiety and depression-like behaviors.Its mechanism may be associated with the down-regulation of monoamine neurotransmitters and neurotrophic factors BDNF and NT-3 in hippocampus, amygdala, and prefrontal cortex region.

Objective To investigate the effects of dizocilpine(MK801) on depressive-like behaviors and damaged hippocampus in rats with diabetes-related depression.MethodsThe animal models of diabetes-related depression were established and they were randomly divided into two groups based on random number table: model group and MK801 group,while 6 rats were included in each group.And another six health rats were regarded as control group.The Open-field test was used to detect the activities.The damage of hippocampus was valued by HE staining,Nissl staining,and Tunel staining.The protein expressions of Bax,Bcl-2 in hippocampus were detected by Western blot.ResultsThe number of activities was significant decreased in Open-field test in model group when compared with control.Hippocampal neurons vacuoles,Nissl bodies were decreased and apoptotic cells were increased in hippocampus in model group as well.Furthermore,the expression of Bax was significant up-regulated(94.57±7.97,P<0.01),while the Bcl-2 was declined(24.65±5.26,P<0.01).Compared with the model group,the animals in MK801 group exhibited increased activities(12.50±4.42,P<0.01),which accompany with an increased Nissl body(133.55±16.74,P<0.01) and a decreased apoptosis(22.50±6.35,P<0.01).Moreover,the expression of Bax was decreased and the Bcl-2 was increased in MK801 group when they were contrasted to model(33.00±4.57,P<0.01).Conclusion MK801 is a significant element to regulate the expression of apoptosis protein including Bax,Bcl-2,and to protecte the hippocampal neuron in rats with diabetes-related depression effectively.

Objective To investigate the effects of Zuogui Jiangtang Jieyu Prescription on neurotrophic effects of astrocytes in diabetes mellitus rats with depression. Methods The diabetes mellitus with depression rat models were established by composite method, and then were randomly divided into 5 groups: model group, positive group, Zuogui Jiangtang Jieyu Prescription high-, medium-, low-dose groups, with 12 rats in each group. 12 normal rats were set as control group. Each administration group was given relevenat medicine for gavage for continuous 4 weeks. Open-field test was used to evaluate the depression-like behavior. The expression of GFAP in astrocyte and MAP2 in neuron were tested by immunohistochemistry. The expression of protein and mRNA of BDNF, GDNF, and NGF were tested by Western blot and RT-PCR. Results Compared with the control group, the activities of rats in model group were significantly reduced. The expression of GFAP increased, while the expressions of MAP2, BDNF, GDNF and NGF decreased. Compared with the model group, the depression-like behavior of rats in model group were significantly improved. The expression of GFAP decreased, while the expressions of MAP2, BDNF, GDNF and NGF increased, and GFAP decrseaed significantly. Conclusion The secretion function of astrocyte can be improved by Zuogui Jiangtang Jieyu Prescription. Its anti-depression and neuron-protection function might be correlated with the enhancement of neurotrophic effects of astrocytes.

In this study, about 350 bp cDNA fragment was amplified by PCR. After being sequenced, the AE1-c-end gene fragment was cloned into EcoR I-Pst I site of pGADT7 to form AD ends in the yeast two-hybrid system. The recombinant plasmid was transformed into yeast AH109, and the expression in the yeast was observed. The results demonstrate that AE1-c-end was obtained. pGADT7-AE1-c-end has no toxic effect on the yeast. It can serve as a target gene of yeast two-hybrid system.
Cloning, Molecular ; DNA, Complementary ; Plasmids ; Polymerase Chain Reaction ; Two-Hybrid System Techniques ; Yeasts

OBJECTIVETo study the chemical constituents in roots of Saussurea lappa.

METHODIsolation and purification were carried out by silica gel, Sephadex LH-20 and RP-18 column chromatography. The chemical structures of constituents were elucidated on the basis of spectral data.

RESULTEleven compounds were isolated and identified as: 5,7-dihydroxy-2-methylchromone (1), p-hydroxybenzaldehyde (2), 3,5-dimethoxy-4-hydroxy-benzaldehyde (3), 3,5-dimethoxy-4-hydroxy-acetophenone (4), ethyl 2-pyrrolidinone-5(s)-carboxylate (5), 5-hydroxymethyl-furaldehyde (6), palmitic acid (7), succinic acid (8), glucose (9), daucosterol (10), beta-sitosterol (11).

CONCLUSIONCompounds 1, 2, 4, 5, 7, 9 were isolated from the genus Saussurea for the first time.

Benzaldehydes ; chemistry ; isolation & purification ; Palmitic Acid ; chemistry ; isolation & purification ; Plant Roots ; chemistry ; Pyrrolidinones ; chemistry ; isolation & purification ; Saussurea ; chemistry ; Sitosterols ; chemistry ; isolation & purification

CRISPR/Cas9, emerged as an efficient and powerful gene editing technology, has become the mainstream genome editing technology. Constructing mutants using CRISPR/Cas9 system is of great significance to the functional study and breeding application of useful genes. As the basis of the technology, a method for identification of mutation with efficiency and lower cost is needed. In this report, we studied the factors influencing mutation detected by CEL Ⅰ crude extracts, such as the amount of protein, enzyme incubation time, PCR buffers. Under the optimized conditions, we can integrate the mutation detection steps into one-tube reaction. We used this system to examine the mutation types and frequency of rice stn1 mediated by CRISPR/Cas9. We also used this method to identify different mutation types including homozygous, heterozygous and bi-allelic mutations. The accuracy of this method reached 100% verified by sequencing. Altogether, our results showed that using CELⅠ crude extracts was an efficient and low cost method for identification of CRISPR/Cas9 mediated mutation.