Study On Anti-dementia Effects And Its Mechanisms Of Yulangsan Polysaccharides (YLSP)

Alzheimer’s disease is one kind of disease characterized by recent memory handicap, senile plaque(SP), neurofibrillary tangle(NFT), and neuron loss in pathology and gradually neuron’s functional deterioration in clinics. It has seriously affected the patient’s cognition function, memory function, language function and space cognition function, social activities ability, personal living ability and emotion personality etc. Its cause of disease and the outbreak mechanism are still not clear. Several theories now widely recognized are: neurotransmitter impairment theory; calcium homeostasis imbalance theory; free radicals injure theory; gene mutation theory; energy dysbolism theory; nerve cell apoptosis theory; excitatory transmitter toxicity;β-Amyloid deposition theory etc. YLS, the root of one plant named Millettia pulchra Kurz var laxior (Dunn) Z.Wei, is a commonly used Chinese herb in Guangxi province. It has the function of supplying QI and blood, increasing the human ability of immunity, anti-aging, anti-stress etc. It can also make the body strong and allay tiredness so as to be applied to treat poor memory in old people and children’s hypophrenia in clinics. Physically weak person, sickness person and postnatal weakness women can also be used. Our research is to extract polysaccharides from YLS and to observe the effects of YLS polysaccharides on anti-dementia and its mechanism. 1. Research on the extraction and assaying method of polysaccharides in YulangsanThe linear range of glucose concentration was15.12-65.52μg·mL-1, and the results showed good precision, the average recovery ratio was98.25%, RSD=1.72%; By the orthogonal design of extracting conditions we found that the conditions as three hours for water-soaking time, adding ten times amount of water, three hours for extract time and extract three times had the highest extraction rate. Through the optimized extraction conditions, the higher rates and higher purity of YLSP can be extracted. Phenol-sulfuric acid method has high accuracy and stability, it can be used for determination of polysaccharides in Yulangsan.2. Protective effects of YLSP on Aβ25-35-induced injury in PC12cellsPC12cells were treated with Aβ25-35to establish a cell model of Alzheimer’s disease. The cell morphology, cell viability and proliferation, persistence, neurite outgrowth and average length were observed. The activities of SOD, GSH-Px, NOS and the contents of GSH, NO, MDA and LDH in culture supernatant and (or) cells were examined by Spectrophotometry. ROS,[Ca2+]i and MMP were examined by FCM. The results showed that LYSP could protect PC12cells from Aβ25-35neurotoxicity:the cells morphology was at good condition and the cell viability, proliferation, persistence, neurite outgrowth and average length were significantly elevated. LYSP-treated group could enhance SOD, GSH-Px activity and GSH content, decrease NOS activity and MDA, NO, ROS, LDH contents significantly in culture supernatant and (or) cells. LYSP-treated group also could decrease [Ca2+]i and increase MMP. It suggests that YLSP can protect PC12cells from injury induced by Aβ25-35.3. Protective effects of YLSP on Aβ325-35-induced apoptosis in PC12cells We measured PC12cell apoptosis ratio by FCM and DAPI staining, and investigated whether there are any apoptotic gene expression changes involved in the apoptosis and the effects of YLSP by real-time PCR. The results showed that PC12cell apoptosis ratio of LYSP-treated group was decreased obviously, Decrease of Bcl-2mRNA and increase of p53, Bax and Caspase-3mRNA under the treatment of AP25-35were reversed by the treatment of YLSP to PC12cells.4. Effects of YLSP on Caspase-3expression, Caspase-3activity and Tau(Ser202) phosphorylation in the frontal lobe and hippocampus of SAMP8mouseThe SAMP8mouse was adopted as AD model group. Caspase-3expression, Caspase-3activity and Tau(Ser202) phosphorylation in the frontal lobe and hippocampus were respectively detected by immunohistochemical method. The results showed that the abnormal Caspase-3expression, Caspase-3activity and Tau(Ser202) phosphorylation were reduced by YLSP.Based on the above results, it is concluded that LYSP have markedly protective effects on Aβ25-35-induecd injury and apoptosis by scavenging free radicals, attenuating the MMP loss, preventing the influx of Ca2+and release of LDH, up-regulating the expressions of Bcl-2mRNA, down-regulating the expressions of p53, Bax and Caspase-3mRNA and reducing Tau(Ser202) phosphorylation.