Preliminary study on the role and mechanism of Wnt non-canonical pathway in mouse Alzheimer's disease model ã€Abstract】 Objective To investigate the role and mechanism of Wnt non-canonical pathway in mouse Alzheimer's disease model. Methods Ninety SD male mice of 4 months old were randomly divided into normal control group, sham operation group, model group, low dose group, medium dose group and high dose group, with 15 rats in each group. The normal control group was not treated. The sham operation group injected a certain amount of normal saline. The mice in the model group, the low-dose group, the middle-dose group and the high-dose group were injected with kainic acid to establish the Alzheimer's disease model. After successful modeling, lithium chloride was continuously administered, and the number of passive avoidances was measured by a mouse platform test. After the mice were sacrificed, interleukin-1β (IL-1β) and interleukin-6 (IL-6) were measured in the brain tissue. Changes in the concentration of tumor necrosis factor (TNF-α). Results The number of passive avoidances in the model group and the low-dose group was significantly higher than that in the sham-operated group (P < 0.05). The number of passive avoidances in the high- and medium-dose groups was significantly lower than that in the model group (P < 0.05). Compared with the model group, the cytokines IL-1β, IL-6 and TNF-α in the low, medium and high dose groups were significantly decreased (P < 0.05), and the decrease was more significant in the middle and high dose groups (P <0). 05) . Conclusion High- and medium-dose doses of lithium chloride can significantly improve the learning and memory scores of mice, and the curative effect is better than low-dose lithium chloride. The lithium chloride Wnt non-canonical pathway plays a positive role in the mouse Alzheimer's disease model and contributes to the improvement of Alzheimer's disease. [Key words] mouse Alzheimer's disease Wnt non-classical pathway lithium chloride mechanism Wnt is a secreted protein, and the receptor for Wnt is Frizzled or low-density lipoprotein-related protein (LRP), which, when combined, activates downstream signaling pathways. These signaling pathways are a series of two, mainly divided into two groups: the classical pathway and the non-canonical pathway. The non-canonical pathway includes the PCP pathway and the Wnt / Ca2+ pathway. Frizzled receptors include Friz-zled-5, Frizzled-1, and the like, and Wnt ligands include Wnt8b, Wnt7a, Wnt7b, Wnt5a, Wnt3a, Wnt2b, and the like, which activate different corresponding receptors. The ligand or receptor for the Wnt non-canonical pathway is essentially Wnt5a or Friz-zled-5, which activates the downstream pathway when the Wnt5a ligand binds to its Friz-zled-5 receptor. When the pathway is activated, the resulting Wnt-Frizzled complex will then activate the protein dishevelle (DVL). For the classical pathway of Wnt, it is a pathway that controls cell fate. Activated DVL inhibits glycogen synthase kinase 3β, which is an increase in p-glycogen synthase kinase 3β, which ultimately leads to enhanced stability of β-catenin. After β- catenin has strong stability, it will transduce into the nucleus. There is a nuclear transcription factor TCF/LEF family in the nucleus, and β-catenin will bind to the nuclear transcription factor TCF/LEF family. To regulate the transcription of the gene of interest. There are two types of Wnt non-canonical pathways. In the PCP pathway that controls tissue polarity and cell movement, activated DVL cascades Rho A, JNK, and NLK signals; regulates cells during cardiac development of the gastrula. In the Wnt/Ca2+ pathway, the Wnt ligand binds to the Frizzled receptor and activates PKC and Ca MKII. The Wnt signaling pathway is more involved in cancer and tumors. The Wnt signaling pathway is also closely related to neuropsychiatric diseases. The main role of Wnt in the nervous system is to promote hippocampal nerve development, dendritic development, axon guidance and Release of neurotransmitters, etc. This study investigated the role and mechanism of the Wnt non-canonical pathway in the mouse model of Alzheimer's disease. 1 Materials and methods 1. 1 Materials and modeling In this study, 90 male SD rats of 4 months old were selected, with a body mass of 24 to 28 g, all purchased from the Experimental Animal Research Center of Hunan Province. After purchase, they were fed in a rat cage and fed for 1 week, allowing them to freely feed and drink. The patients were randomly divided into normal control group, sham operation group, model group, low dose group, middle dose group and high dose group, with 15 rats in each group. Next, model mice with Alzheimer's disease were established. The mice in the model group, low-dose group, middle-dose group and high-dose group were anesthetized with 1% sodium pentobarbital (20 mg / kg). The locator (supplied by Shanghai Xinsoft Co., Ltd.) fixed it, cut the skin at the skull line to determine the position of the Meynert basal ganglia (nb M), and then drilled a diameter of about 1. For small holes of 0 mm, inject 1 μl of kainic acid into the sides of the nb M with a micro syringe. In order to destroy the cholinergic cells in the area, slowly inject the alginic acid into the sea, preferably for 5 minutes. Postoperative postoperative treatment, not only suture the skin, but also to give penicillin to the mice on time for 3 days. Fifteen normal mice were not treated as normal controls, and the sham operation group only injected the same amount of normal saline according to the same method. 1. 2 methods 1. 2. 1 Experimental method Lithium chloride solution was prepared from distilled water, and a high concentration solution of lithium chloride (20 mg / ml), a medium concentration solution (15 mg / ml), and a low concentration solution (10 mg / ml) were prepared. 2nd after success Except for the distilled water of normal control group, sham operation group and model group, the other groups were given the corresponding drugs according to the above dosage, and were administered according to the daily volume of 5 ml/kg, once a day for 15 days. The mice were weighed once every 3 days throughout the experiment and the dose was adjusted in turn. During the intragastric administration, 3 mice died in each group, and 12 mice in each group were finally tested. 1. 2. 2 Mouse learning and memory test The mouse jumping test method (provided by Shanghai Xinsoft Co., Ltd.) was designed to investigate the effect of lithium chloride on learning and memory of model mice. The platform test is an experiment of one-time stimulation avoidance response. On the 10th day after surgery, the mouse platform test was carried out. The mouse was first placed on the grid. When the grid was energized, there was an electric shock. When the electric shock was hit by the mouse, the mouse jumped onto the platform and was controlled by the platform software. The mouse jumped again. After the platform, it will be shocked, then jump back to the platform, and record the number of times each mouse is shocked. The mouse is shocked and jumped onto the platform to call it a passive avoidance response. After the test was repeated 1 day, the mice were first placed on the platform, and the time of the first jump (ie, the incubation period) was recorded. The mice passively evaded the response and used as the learning and memory score for 5 days. The memory retention of the mice was observed. 1. 2. 3 Determination of IL-1β, IL-6 and TNF-α in hippocampus of mice The mice were sacrificed on the 2nd day after operation, and the brain was decapitated. 2 ~ 1. 0 g, rinsed in physiological saline to remove blood stains, and made a 10% homogenate with physiological saline. After centrifuging the homogenate at 3 000 rpm for 5 min, the supernatant was removed and frozen. Strictly follow the radioimmunoassay kit (purchased from Beijing Furui Runkang Biotechnology Co., Ltd.) to explain the calculation of IL-1β, IL-6 and TNF-α in hippocampus per milligram of tissue. 1. 3 Statistical methods obtained using SPSS 13. 0 Software performs statistical processing and comparative analysis. Data are expressed as mean ± standard deviation ( x ç‹ Â± s ), measurement data using t test, and count data using χ 2 test. P < 0. 05 is statistically significant. 3 Discussion Alzheimer's disease (AD) is a degenerative disease of the nervous system. It is more common in the elderly over 70 years old, and there are more women than men. Before the age of 65, the disease is called Alzheimer's disease, and those over 65 years old are called the elderly. Sexual dementia. Mainly manifested as memory impairment, aphasia, visual space skills damage, executive dysfunction and other characteristics, there is no clear cause, possible factors and hypotheses as many as 30 kinds. Family history is one of the risk factors for this disease. The family members of some AD patients have the same rate of disease as the general population, and may also have the risk of congenital disease. It can be seen that dementia is related to heredity. Some physical diseases such as thyroid diseases and immune system diseases have been studied as risk factors for the disease. Head trauma with disturbance of consciousness is also considered a risk factor for the disease, and there are many reports. In addition, research speculation, progressive failure of the immune system, and other social and psychological factors such as living alone and life bumps may also be the cause of the disease. Because the incidence factors involve many aspects, it must not be treated with drugs alone. According to cognitive ability and deterioration of physical function, it can be divided into three stages: mild dementia, moderate dementia, and severe dementia. The first period is generally characterized by memory loss, decreased judgment ability, inadvertent work, indifferent emotions, and often suspicious symptoms. The second period is generally characterized by severe damage to the memory of the near and far, the visual space ability of the simple structure is reduced, and the outdoor space cannot be independently performed. Activities, various neurological symptoms, frequent movements and other symptoms; in the third period, patients have completely relied on caregivers, and they cannot take care of themselves in their daily lives. The patients are mainly examined by neuropsychological tests, hematology, neuroimaging, electroencephalography (EGG), cerebrospinal fluid testing, and genetic testing. At present, the clinical treatment of Alzheimer's disease is to control the associated psychopathological symptoms, mainly including psychotic and pathological symptoms such as anxiety, depression, and psychosis, and the corresponding drugs can be selected for treatment. In addition, it is necessary to use nootropics or drugs that improve cognitive function to delay the progression of the disease. Drugs acting on neurotransmitters can enhance central cholinergic activity and improve learning and memory in older adults. The cerebral metabolic activating drug can dilate the cerebral blood vessels, increase the utilization of oxygen and amino acids by the cerebral cortical cells, improve the function of brain cells, and improve memory. Numerous studies have confirmed that the pathological process of Alzheimer's disease is closely related to the dysfunction of Wnt signaling pathway. At present, it is mainly through the transplantation of stem cells and activation of its intrinsic Wnt signaling pathway to promote the production of newborn hippocampal neurons to compensate for the loss of hippocampal neurons to treat Alzheimer's disease. The cytokines IL-1β, IL-6 and TNF-α play an important role in the central nervous system, contributing to the growth and development of the body and the stability of the internal environment. Under pathological conditions, these cytokines are abnormally expressed and inflammation occurs. Reaction, damage to tissue cells, lithium chloride (Li Cl), through the action of drugs, mainly through the Wnt non-classical signaling pathway, has a significant inhibitory effect on the expression and activity of some abnormal cytokines, and protects nerve cells and cardiomyocytes. Function to improve the patient's memory and judgment. As a highly selective inhibitor, Li Cl is used in research work for the activation of Wnt non-canonical signaling pathways and plays an important role in cardiomyocyte differentiation by inhibiting the cytokines IL-1β, IL-6, TNF- The activity of alpha improves myocardium cells. This study used basal ganglia injection of kainic acid to establish a model of Alzheimer's disease, resulting in the destruction of nb M cholinergic neurons, thereby damaging the choline projection system from nb M to the neocortex, cerebral cortex and hippocampal choline. Reduced acetyltransferase activity and acetylcholinesterase activity ultimately lead to significant changes in learning and memory function. One of the most important animal models in memory research is the suppression of imitative activities or learning habits. Passive avoidance experiments evade certain annoying things by the animal learning to remove a particular behavior. The mouse jumping platform experiment is simple and easy to operate. It can observe multiple animals at the same time, which can objectively reflect the animal's memory after a single stimulation, and can analyze the mechanism of action of the puzzle drug. The results of this experiment showed that there was no significant difference in behavior between the normal control group and the sham operation group (P > 0.05). However, compared with the sham operation group, the passive avoidance times of the model group and the low dose group were within 5 days of training. There was a significant increase (P < 0.05), but there was no significant difference between the high dose group and the middle dose group (P > 0.05). Compared with the model group, the number of passive avoidances in the high-dose group and the middle-dose group was significantly decreased within 5 days of training (P < 0.01), and the number of passive avoidances in the low-dose group was also significantly reduced from the 15th day of training. (P < 0.05). The results showed that the high-dose group and the middle-dose group can significantly improve the learning and memory scores of mice, and the curative effect is better than the low-dose group. After drug intervention, the cytokines IL-1β, IL-6 and TNF-α decreased significantly ( P < 0 . 05), the drop in the middle dose group and the high dose group was more significant (P < 0.05). 4 Conclusion In conclusion, high and medium doses of lithium chloride can significantly improve the learning and memory performance of mice, and the efficacy is better than low-dose lithium chloride. The non-classical pathway of lithium chloride Wnt plays an active role in the mouse Alzheimer's disease model. Helps to improve Alzheimer's disease. Being a reliable supplier of above products, Medton works under the following main principles: quality, credit, innovation, and close partnership with customers. Marine Lifeguard Rescue Medton Medical , https://www.medtonmedical.com
Preliminary study on the role and mechanism of Wnt non-canonical pathway in mouse Alzheimer's disease model