Application of tripeptide compound and copper ion chelate thereof in prevention and treatment of silicosis
A technology of copper ions and chelates, applied in tripeptide components, drug combinations, respiratory diseases, etc., to achieve the effects of inhibiting the formation of silicon nodules and collagen deposition, inhibiting inflammatory reactions, and inhibiting the development of
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Embodiment 1
[0023] Example 1 Establishment of silicon-induced mouse silicosis model, experimental grouping and administration method.
[0024] The C57BL / 6 mice were randomly divided into 5 groups: control group, silicon-induced mouse lung fibrosis model group (Si group), Si+0.2μg / g GHK-Cu group, Si+2μg / g GHK-Cu group, Si+ 20 μg / g GHK-Cu. Except for the control group, the other groups used oral perfusion of silica suspension to establish the mouse silicosis model. Dissolving 5mg of silica in 50ul of PBS to prepare the silica suspension, the mice were anesthetized by inhaling isoflurane Afterwards, inhale the silica suspension through the oropharynx. The GHK-Cu intervention group was intraperitoneally injected with the corresponding dose of GHK-Cu every day, and the control group was given the corresponding dose of normal saline. The mice were euthanized on the 28th day, and the left lung was perfused with 4% paraformaldehyde for 48 hours and embedded in paraffin. For other mice, 0.5 ml ...
Embodiment 2
[0025] Example 2 HE staining was used to detect lung inflammation and the formation of silicon nodules.
[0026] 1. HE staining.
[0027] Hematoxylin and eosin staining (HE staining for short) is the most widely used staining method for nuclear histology. The basic principle is: the chromatin in the nucleus is mainly DNA. In the double-helix structure of DNA, the phosphate groups on both sides of the chain are outward, negatively charged, and acidic. bonded or hydrogen bonded. Hematoxylin is blue in alkaline solution, so cell nuclei are stained blue. The main component in the cytoplasm is protein, which is an amphoteric compound, and the staining of the cytoplasm is closely related to the pH value. Eosin is a chemically synthesized acid dye that dissociates into negatively charged anions in water and combines with positively charged amino groups (cations) of proteins to stain cytoplasm, cytoplasm, red blood cells, muscles, connective tissues, and eosinophilic granules etc....
Embodiment 3
[0039] Example 3 The expression of inflammatory factors IL-6 and TNFα in BALF was detected by ELISA.
[0040] Take out the reagents and equilibrate to room temperature. Mix the reagents and samples thoroughly and avoid air bubbles as much as possible.
[0041] 1. Adding samples: set up blank holes, standard holes, and sample holes to be tested respectively. Add 10 μl of sample diluent to the blank well, add 100 μl of standard substance or test sample diluted in different proportions to the remaining wells, cover the membrane, and incubate at 37°C for 2 hours.
[0042]2. Discard the liquid, shake dry, add 100 μl of biotin antibody (1X) to each well, cover the membrane, and incubate at 37°C for 1 hour.
[0043] 3. Discard the liquid in the well, shake dry, wash the plate 3 times, soak for 1-2min each time, shake dry and pat dry the liquid in the well on absorbent paper.
[0044] 4. Add 100 μl of HRP-avidin (1X) to each well, cover the membrane, and incubate at 37°C for 1 hour....
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