Application of artemisia selengensis extract in preparation of medicine for improving neurodevelopmental disorder and medicine

Abstract

The invention belongs to the technical field of food nutrition, and discloses application of an artemisia selengensis extract in preparation of a medicine for improving neurodevelopmental disorder and the medicine. The artemisia selengensis extract has the following advantages and effects: the research finds that the artemisia selengensis extract has the effect of improving neurodevelopmental disorders, which is specifically embodied in that the artemisia selengensis extract can effectively increase the activity of acetylcholin esterase, lower the content of ROS (reactive oxygen species), up-regulate antioxidant indexes (GSH (glutathione) content and sod1 and cat gene expression) and relieve oxidative stress injury of zebrafish caused by NP (natriuretic peptide); meanwhile, the artemisia selengensis extract can also up-regulate the expression of a neural development gene (alpha 1-tubulin, mbp) so as to improve the neural development disorder of the zebrafish; besides, the mechanism of improving the zebrafish neurodevelopmental disorder by the artemisia selengensis extract is mainly realized through a p38 / 5-HT / Wnt / beta-catenin signal channel, expression of p38a is down-regulated (network pharmacology finds that p38 in MAPHK is a main mechanism channel), and expression of htr1aa, maoa, wnt4a and beta-catenin is down-regulated.

Description

Technical Field

[0001] The invention relates to the technical field of food nutrition, and in particular to application of an Artemisia selengensis extract in preparing a drug for improving neurodevelopmental disorders and the drug. Background Art

[0002] Neurodevelopmental disorders (NDDs) are a group of conditions characterized by abnormal brain development that affects mood, learning, cognition, and memory. NDDs have a profound impact on society at large, reducing quality of life, lowering academic performance, and disrupting behavior. Despite worldwide efforts to understand the root causes of NDDs, there is no cure for these disorders. Only a few FDA-approved drugs such as aripiprazole, risperidone, and lurasidone are used to relieve symptoms. The development of the central nervous system is a complex process that is determined by both genetic and external factors, but genetic factors appear to account for 30-40% of all cases of neurodevelopmental disorders. The developing brain is particularly vulnerable to toxic chemicals such as lead, methylmercury, bisphenol A, phthalates, polybrominated diphenyl ethers, polycyclic aromatic hydrocarbons, perfluoroalkyl acids, NPs, octylphenol (OP), etc. There are a wide variety of chemicals used for commercial purposes, and only a small number of them have been studied for neurotoxicity. Therefore, it is necessary to identify risk factors for neurodevelopmental disorders and to research and develop a wider variety of drugs to improve neurodevelopmental disorders. Summary of the invention

[0003] The purpose of the present invention is to provide an application of an Artemisia selengensis extract in preparing a drug for improving neurodevelopmental disorders and the drug.

[0004] The above technical purpose of the present invention is achieved through the following technical solution: use of Artemisia selengensis in the preparation of drugs for improving neurodevelopmental disorders.

[0005] The present invention is further provided as follows: use of the Artemisia selengensis extract in the preparation of a medicament for improving neurodevelopmental disorders.

[0006] The present invention is further configured as follows: the preparation method of the Artemisia selengensis extract comprises the following steps: taking Artemisia selengensis stems, placing them in an organic solvent for ultrasonic extraction, removing impurities from the extract, concentrating, re-dissolving and freeze-drying, to obtain the Artemisia selengensis extract.

[0007] The present invention is further configured as follows: in the preparation method of the Artemisia selengensis extract, the Artemisia selengensis stems are baked before ultrasonic extraction, the baking conditions are 60°C, 3h, the solvent is 50% ethanol, the mass ratio of the solvent to the Artemisia selengensis is 20:1, the conditions for ultrasonic extraction are 40°C, the power is 100W-110W, the extraction time is 80-100min, the number of extractions is 3 times, the impurity removal is centrifugal impurity removal, the centrifugal speed is 3000rpm / min, the centrifugal time is 25min, the concentration is reduced pressure concentration at 45°C, and after concentration, an appropriate amount of ultrapure water is added for freeze drying.

[0008] The present invention is further provided as follows: use of the Artemisia selengensis extract in preparing a medicine for increasing the activity of acetylcholinesterase.

[0009] The present invention is further provided as follows: use of the Artemisia selengensis extract in preparing a drug for improving the expression of neural developmental genes.

[0010] The present invention is further configured as follows: the neural developmental genes include mbp Gene, α1-tubulin Gene.

[0011] The present invention is further configured as follows: the Artemisia selengensis extract is used in the preparation of a drug for lowering the content of ROS and raising the antioxidant index.

[0012] By adopting the above technical solution, the antioxidant indicators are increased, including increasing the GSH content, upregulating the expression of sod1 and cat genes The present invention is further configured as follows: use of the Artemisia selengensis extract in preparing a drug for up-regulating genes related to the p38 / 5-HT / Wnt / β-catenin signaling pathway.

[0013] The present invention is further provided as follows: a drug for improving neurodevelopmental disorders, comprising an Artemisia selengensis extract.

[0014] The beneficial effects of the present invention are: 1. Artemisia selengensis, also known as water wormwood, Artemisia selengensis, Artemisia selengensis, Artemisia selengensis, etc., is a perennial herbaceous plant of the genus Artemisia in the Asteraceae family. It is a specialty vegetable crop in southern my country. Artemisia selengensis is rich in nutrition and has a unique flavor. Both the above-ground tender stems and underground creeping stems are edible. It is rich in protein, carbohydrates, crude fiber, vitamins and trace elements. In addition to its edible value, the various active ingredients contained in Artemisia selengensis have rich biological activities and are widely used in the pharmaceutical industry. The main chemical components include polyphenols, flavonoids, volatile oils, polysaccharides and terpenes. Artemisia selengensis has been proven to have medical activity, mainly focusing on anti-inflammatory and antioxidant effects. At the same time, studies have also found that Artemisia selengensis extracts have anti-tumor, blood pressure lowering, prevention and alleviation of diabetes and its complications, and liver protection. However, research on neuroprotection is still in its initial stage, especially in improving neurodevelopmental disorders. There is no report yet. The results have good practical significance for the implementation of toxicity intervention programs based on edible plants or their active ingredients and neurodevelopmental disorder improvement programs in the population in the future.

[0015] 2. The present invention found that the extract of Artemisia selengensis has the effect of improving neurodevelopmental disorders, which is specifically reflected in: Artemisia selengensis extract can effectively increase the activity of acetylcholinesterase, downregulate the content of ROS, upregulate antioxidant indicators (GSH content, sod1 and cat gene expression), and reduce the oxidative stress damage of zebrafish caused by NP; at the same time, Artemisia selengensis extract can also upregulate neurodevelopmental genes ( α1-tubulin, MBP) expression to improve zebrafish neurodevelopmental disorders; in addition, the mechanism by which Artemisia selengensis extract improves zebrafish neurodevelopmental disorders is mainly achieved through the p38 / 5-HT / Wnt / β-catenin signaling pathway, among which downregulation p38a expression (network pharmacology found that p38 in MAPHK is the main mechanism pathway), downregulating htr1aa , maoa , wnt4a , β-catenin expression. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The effect of Artemisia selengensis extract on mortality, hatching rate and acetylcholinesterase; A: mortality; B: hatching rate; C: acetylcholinesterase activity; * P <0.05 means significant difference between the two comparisons, ** P <0.01 indicated extremely significant differences in pairwise comparisons.

[0017] Figure 2 Effects of Artemisia selengensis extract on neurodevelopmental genes;* P <0.05 means significant difference between the two comparisons, ** P <0.01 indicated extremely significant differences in pairwise comparisons.

[0018] Figure 3 Effects of Artemisia selengensis extract on oxidative stress. A: ROS content; B: GSH content; C: sod1 and cat Gene expression;* P <0.05 means significant difference between the two comparisons, ** P <0.01 indicated extremely significant differences in pairwise comparisons.

[0019] Figure 4 This is a network pharmacology analysis; A: Genes shared by Artemisia selengensis extract and neurodevelopmental disorders; B: Shared gene protein interaction map; C: GO analysis; D: KEGG analysis; E: p38a Expression;* P <0.05 means significant difference between the two comparisons, ** P <0.01 indicated extremely significant differences in pairwise comparisons.

[0020] Figure 5 Effects of Artemisia selengensis extract on 5-HT / Wnt / β-catenin signaling pathway;* P <0.05 means significant difference between the two comparisons, ** P <0.01 indicated extremely significant differences in pairwise comparisons. DETAILED DESCRIPTION Example

[0021] Application of Artemisia selengensis extract in preparing medicine for improving neurodevelopmental disorders.

[0022] The preparation method of Artemisia selengensis extract comprises the following steps: Using fresh Artemisia selengensis stems as raw materials, the washed fresh Artemisia selengensis stems were placed in an oven, treated at 60°C for 3 hours, and crushed into powder using an ultrafine grinder; Using 50% ethanol as solvent, ultrasonic extraction was performed at 40°C and power of 100 W-110 W for 80-100 min. The mass ratio of extraction solvent to Artemisia selengensis was 20:1, and the extraction times were 3 times. The extract was centrifuged at a centrifugal speed of 3000 rpm / min for 25 min to remove impurities, and then concentrated under reduced pressure at 45°C. An appropriate amount of ultrapure water was added and freeze-dried to obtain the Artemisia selengensis extract.

[0023] Pharmacodynamic evaluation of Artemisia selengensis extract in improving neurodevelopmental disorders in zebrafish (1) Experimental animals and groups: Zebrafish embryos 4 h after fertilization (4 hpf) were collected and placed in 6-well cell culture plates, with 30 embryos placed in each well. The experiment was divided into a blank group, a NP-exposed group, and a low / high dose ASTE intervention group.

[0024] (2) Preparation and administration of neurodevelopmental models 4-120 hpf: NP-treated group and ASTE low / high dose group were given NP (0.8 mg / L); blank control group was given fresh aerated water. 48-120 hpf: ASTE low / high dose group were given ASTE (0.11 and 0.33 mg / L). At 120 hpf, the drug solution was removed, the larvae were washed three times with embryo water, and the larvae were transferred to 1.5 mL EP tubes and stored at −80°C.

[0025] (3) Determination of mortality, hatching rate and acetylcholinesterase After exposure to 120 hpf, the mortality and hatching rate of zebrafish were calculated. The larvae in each well were then collected and added to 10 volumes of ice-cold saline. After being fully ground in a tissue grinder, the supernatant was obtained by centrifugation at 3000 rpm for 15 min at 4°C. The activity of acetylcholinesterase (Nanjing Jiancheng Biological, China) in the supernatant was determined by a kit. The experimental results are shown in Figure 1 . The results showed that the mortality rate increased significantly after NP treatment, while the mortality rate decreased significantly after Artemisia selengensis extract treatment. There was no significant change in the hatching rate. NP treatment significantly reduced the activity of acetylcholinesterase, while the activity of acetylcholinesterase increased after Artemisia selengensis extract treatment. This indicates that Artemisia selengensis extract has a certain regulatory effect on the nerve damage caused by NP. Therefore, Artemisia selengensis extract can be used to prepare drugs that increase the activity of acetylcholinesterase.

[0026] (4) Determination of neurodevelopmental genes RT-qPCR analysis of neurodevelopmental genes α1-tubulin, gap43, mbp The expression of Figure 2 The results showed that NP treatment significantly downregulated α1-tubulin and mbp The expression of NP in the rats was significantly upregulated by Artemisia selengensis extract compared with the NP group. α1-tubulin and mbp The expression of 147 genes in the 147 genes was significantly decreased, indicating that the extract of Artemisia selengensis can improve the neurodevelopmental disorders caused by NP. The extract of Artemisia selengensis can be used to prepare a method for improving the expression of neurodevelopmental genes. α1-tubulin and mbp of the expression of drugs.

[0027] (5) Oxidative stress analysis The content of ROS and GSH was determined using the Nanjing Jiancheng kit; RT-qPCR was used to determine sod1 and cat The expression of Figure 3 The results showed that NP treatment significantly increased the content of ROS, decreased the content of GSH, and down-regulated sod1 and cat The expression of NP in zebrafish was shown to be induced by NP. Compared with the NP group, the extract of Artemisia selengensis significantly reduced the content of ROS, increased the content of GSH, and up-regulated sod1 and cat The expression of 147 amino acids in zebrafish with neurodevelopmental disorders was significantly decreased, indicating that the extract of Artemisia selengensis can improve the oxidative stress level of zebrafish with neurodevelopmental disorders. Therefore, the extract of Artemisia selengensis can be used to prepare drugs that downregulate the content of ROS and upregulate antioxidant indicators.

[0028] (6) Network pharmacology analysis Through network pharmacology methods, potential targets of Artemisia selengensis extract for improving neurodevelopmental disorders were screened and its mechanism for improving neurodevelopmental disorders was analyzed. Databases such as Swiss, PharmMapper, GeneCards, and DisGeNET were used to mine Artemisia selengensis extract and neurodevelopmental disorder targets, and the common targets of Artemisia selengensis extract and neurodevelopmental disorders were determined; then cytoscape 3.7.1 was used to construct a protein-protein interaction (PPI) network; finally, the DAVID database was used to perform GO function and KEGG pathway enrichment analysis, key signaling pathways were screened, and RT-qPCR was used to verify the key pathways. The results are shown in Figure 4 The results showed that Artemisia selengensis extract may improve neurodevelopmental disorders through the MAPK pathway. Further RT-qPCR found that Artemisia selengensis extract significantly reduced NP-induced p38a The results showed that Artemisia selengensis extract increased the expression of MAPK pathway genes. p38a Improve neurodevelopmental disorders 。

[0029] (7) Analysis of 5-HT / Wnt / β-catenin signaling pathway 5-HT / Wnt / β-catenin pathway genes were determined by RT-qPCR htr1aa , maoa , wnt4a , β-catenin The expression of Figure 5 The results showed that NP treatment significantly downregulated htr1aa , maoa , wnt4a , β-catenin This indicates that NP causes neurodevelopmental disorders through the 5-HT / Wnt / β-catenin pathway; compared with the NP group, Artemisia selengensis extract significantly upregulated htr1aa , maoa , wnt4a , β-catenin The expression of Artemisia selengensis extract can improve the neurodevelopmental disorders caused by NP through the 5-HT / Wnt / β-catenin pathway. MAPK can regulate the 5-HT / Wnt / β-catenin pathway. In summary, Artemisia selengensis extract can improve neurodevelopmental disorders through the p38 / 5-HT / Wnt / β-catenin signaling pathway. Therefore, Artemisia selengensis extract can be used to prepare drugs that upregulate genes related to the p38 / 5-HT / Wnt / β-catenin signaling pathway. In summary, the above experimental results show that the Artemisia selengensis extract has the effect of improving neurodevelopmental disorders. The drug including the Artemisia selengensis extract can be used to improve neurodevelopmental disorders.

Claims

1. Application of Artemisia selengensis in the preparation of medicines for improving neurodevelopmental disorders.

2. Application of Artemisia selengensis extract in the preparation of drugs for improving neurodevelopmental disorders.

3. The use of the Artemisia selengensis extract according to claim 2 in the preparation of a medicament for improving neurodevelopmental disorders, characterized in that: The preparation method of the Artemisia selengensis extract comprises the following steps: taking Artemisia selengensis stems, placing them in an organic solvent for ultrasonic extraction, removing impurities from the extract, concentrating, re-dissolving and freeze-drying the extract to obtain the Artemisia selengensis extract.

4. The use of the Artemisia selengensis extract according to claim 3 in the preparation of a medicament for improving neurodevelopmental disorders, characterized in that: In the preparation method of the Artemisia selengensis extract, the Artemisia selengensis stems are baked before ultrasonic extraction, the baking conditions are 60°C and 3h, the organic solvent is 50% ethanol, the mass ratio of the solvent to the Artemisia selengensis is 20:1, the conditions for ultrasonic extraction are 40°C, the power is 100W-110W, the extraction time is 80-100min, the number of extractions is 3 times, the impurity removal is centrifugal removal, the centrifugal speed is 3000rpm / min, the centrifugal time is 25min, the concentration is reduced pressure concentration at 45°C, and after concentration, an appropriate amount of ultrapure water is added for freeze drying.

5. Use of Artemisia selengensis extract in the preparation of a drug for increasing the activity of acetylcholinesterase.

6. Use of Artemisia selengensis extract in the preparation of drugs for increasing the expression of neural developmental genes.

7. Use of the Artemisia selengensis extract according to claim 6 in the preparation of a drug for increasing the expression of neural development genes, characterized in that: The neurodevelopmental genes include mbp Gene, α1-tubulin Gene.

8. Application of Artemisia selengensis extract in the preparation of drugs for downregulating ROS content and upregulating antioxidant indicators.

9. Application of Artemisia selengensis extract in the preparation of drugs for upregulating genes related to the p38 / 5-HT / Wnt / β-catenin signaling pathway.

10. A drug for improving neurodevelopmental disorders, characterized in that: Includes Artemisia selengensis extract.

Images