Application of Isoxaxifrage in the Preparation of Protective Drugs for Rat Pineal Gland Damage
A technology of iso-saxifrage and pineal gland, which is applied in the field of medicine, can solve the problems of not discovering iso-saxifrage in ultrastructure, tissue morphology, molecular level, etc.
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Embodiment 1
[0019] Example 1: Verification of the protective effect of IS on ultrastructural treatment of PCPA-induced pineal injury rat model
[0020] 1. Materials
[0021] 1. Drug: Isoxaxifrage monomer (IS): Shanghai Fusion Pharmaceutical Technology Co., Ltd., batch number: 150809, prepared into suspensions with different concentrations with 2% Tween-80 solution.
[0022] 2. Animals: rats, SD, SPF grade, male, weighing 160-180g, purchased from Speiford (Beijing) Biotechnology Co., Ltd., license number: SCXK (Beijing) 2016-0002; certificate number: No. 1103241911019500, raised in the central laboratory of Shanxi Academy of Traditional Chinese Medicine, temperature: 22-24°C, humidity: 40-50%.
[0023] 3. Reagents: p-chlorophenylalanine (PCPA): SIGMA Company of the United States, batch number: 10026000696; polysorbate-80 (Tween-80): chemical glass station of Hubei Pharmaceutical Company, batch number: 080308; chloral hydrate: Beijing Chaoyang District Chemical Pharmaceutical Factory, bat...
Embodiment 2
[0040] Example 2: Observation of the influence of IS on histopathological changes in rats with pineal gland injury:
[0041] A. Materials
[0042] Medicine: same as Example 1; Animal: same as Example 1; Reagent: same as Example 1.
[0043] Main instruments: universal microscope fluorescence imaging system: BX51+DPT2+IPE, Japan Olympus Company; automatic tissue dehydrator: EXCELSIOR ES, American Thermo SCIENTIFIC Company; rotary paraffin microtome: SHANDONFINESSE 325, American Thermo SCIENTIFIC Company; automatic sealing Machine: ClearVue, American Thermo Company; automatic dyeing machine: Gemini AS, American Thermo Company.
[0044] B. Method
[0045] Grouping and administration: same as in Example 1; pineal gland injury model: same as in Example 1.
[0046] C. Index detection: pineal gland histopathological observation: pineal gland was taken, fixed with 10% formaldehyde solution, dehydrated, paraffin embedded, sectioned, HE stained, observed under a microscope.
[0047] ...
Embodiment 3
[0048] Example 3: Effect of IS on the Regulation of Pineal Clock Genes in PCPA Injured Rats
[0049] 1. Pineal clock gene detection: real-time fluorescent quantitative (Real-Time PCR) was used to detect the expression of pineal clock genes Clock, Bmal1, Per1, Per2, Per3, Cry1, and Cry2 in rats with PCPA pineal injury.
[0050] (1) Extraction of total RNA: The extraction steps of total RNA were performed according to the instructions of the kit.
[0051] A. Take the pineal gland, fully grind the pineal gland in liquid nitrogen, add 1ml of Buffer RLT, pipette repeatedly, the sample is fully lysed, and stand at room temperature for 5 minutes.
[0052] B. Add 200 μl of chloroform to each 1ml of Buffer RLT and mix thoroughly, place at room temperature for 2 minutes, centrifuge at 12000 rpm for 5 minutes, and take the supernatant. Add the same volume of 70% ethanol, mix well, centrifuge at 12000rpm for 20s, and discard the waste liquid.
[0053] C. Add 350μl of Buffer RW1, centrif...
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