Method of diagnosis or treatment of neurological disorders with p75ECD and/or p75
A kind of technology of p75ecd-fc, use, apply in the field of neurotrophin receptor p75 and/or its extracellular domain
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Embodiment 1
[0133] Embodiment 1 Diagnosis and treatment of neurological diseases
[0134] Materials and methods
[0135] Reagents and Antibodies
[0136] The vectors of AAV8-CBA-p75NTR-ECD-Fc (AAV-ECD-Fc) and AAV8-CBA-GFP (AAV-GFP) were produced and purified by Virovek (Hayward, CA, USA). pcDNA3-human full-length p75NTR was from Dr. Rainer Niedenthal (Institute for Physiological Chemistry / Biochemistry, Hannover Medical School, Hannover, Germany). Synthetic Aβ42 (Cat. No. 62080) and Promiscuous Aβ42 (Cat. No. 62046) were purchased from American Peptide (Sunnyvale, CA, USA).
[0137] Human p75ECD-Fc and Fc recombinant proteins were prepared as follows. Human p75ECD and Fc nucleotide constructs were produced under standard conditions using primers (Table 2) and PCR, respectively, using human cDNA as the DNA source, and then for the p75ECD-Fc fusion protein by overlapping PCR using standard conditions. Fragments blend together.
[0138] Table 2 PCR primer pair sequences for human p75ECD-...
Embodiment 2
[0241] Example 2 Specificity of Effect of p75ECD
[0242] To investigate the specificity of p75ECD-Fc on amyloid load, 8-month-old AD mice were injected with recombinant p75ECD-Fc or one of two control proteins (human IgG or recombinant human adult fused to human IgG Fc) in the hippocampus of 8-month-old AD mice. Fibroblast growth factor receptor 4ECD (FGFR4-Fc)), additional experiments were performed. A single injection of recombinant p75ECD-Fc, but not human IgG or FGFR4-Fc, also significantly reduced the area fraction of amyloid plaques at the injection site one week after injection (data not shown). To investigate the underlying mechanism of how p75ECD-Fc affects amyloid load in the brain, the effect of p75ECD-Fc on Aβ fibril formation and depolymerization was examined using ThT assay and transmission electron microscopy (TEM) visualization. The data showed that p75ECD-Fc, but not the control proteins human IgG or FGFR4-Fc, suppressed A[beta] fluorescence in fibril format...
Embodiment 3
[0245] Example 3 Intramuscular delivery of p75ECD-Fc in AD mice alleviates AD pathology and improves cognitive function
[0246] AD mice (APP / PS1 transgenic mice) or wild type were injected intramuscularly with AAV-ECD-Fc as follows: At 3 months of age, female APP / PS1 transgenic mice were injected with 5 μl 2×10 in 9 An AAV-ECD-Fc or AAV-GFP vector genome was injected intramuscularly. At 12 months of age, all mice were sacrificed and brains were sampled. The control group received AAV-GFP, but was otherwise treated the same. After treatment, mice were tested using the Morris water maze test. In certain experiments, mice were injected once intraperitoneally with recombinant p75ECD-Fc peptide at 50 μg / 50 μl per mouse. Other experiments were performed as described above.
[0247] Intramuscular delivery of p75ECD ameliorates cognitive decline in AD mice
[0248] Mice treated with AAV-ECD-Fc performed better in the Morris water maze test compared to AAV-GFP-treated mice. This ...
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