OBJECTIVE: To construct a transgene expressing human endomorphin-2 by linking the signal peptide of mouse nerve growth factor (PN) to a human endomorphin-2 DNA sequence containing a short linker recognized by the protease FURIN and test the analgesic effect of endomorphin-2 on neuropathic pain. METHODS: The transgene was inserted into the cosmid pAxCAwt to generate PN-EM-2-pAxCAwt. The recombinant adenovirus Ad-PNEM2 was packaged and propagated in HEK293 cells. After the Ad-PNEM2-infected NIH3T3 cells had been cultured, protein expression was examined by immunofluorescence and ELISA. A CCI rat model was constructed and the Ad-PNEM2 was administered intrathecally. The rats' pain thresholds (PWL) were measured and the presence of endomorphin-2 in the cerebrospinal fluid was confirmed through ELISA. RESULTS: The Ad-PNEM2 expressed endomorphin-2 smoothly and abundantly in NIH3T3 cells at a significantly higher rate than the viral control (P<0.01) or blank control (P<0.01). The expressed endomorphin-2 was mainly observed in the cytoplasm. The concentration of endomorphin-2 in the cerebrospinal fluid increased 1 day after injection and peaked between 7 and 14 days after injection. After injection, PWL approached normal levels in the operated study group. No significant change was observed in the control groups. There was a significant correlation between PWL and endomorphin-2 level (r = 0.944, P<0.001). CONCLUSION: The constructed human endomorphin-2 transgene was expressed effectively, and endomorphin-2 expressed by the recombinant adenovirus altered the threshold to thermal stimulus and showed good analgesic effect.
OBJECTIVE: To construct a transgene expressing human endomorphin-2 by linking the signal peptide of mouse nerve growth factor (PN) to a human endomorphin-2 DNA sequence containing a short linker recognized by the protease FURIN and test the analgesic effect of endomorphin-2 on neuropathic pain. METHODS: The transgene was inserted into the cosmid pAxCAwt to generate PN-EM-2-pAxCAwt. The recombinant adenovirus Ad-PNEM2 was packaged and propagated in HEK293 cells. After the Ad-PNEM2-infected NIH3T3 cells had been cultured, protein expression was examined by immunofluorescence and ELISA. A CCI rat model was constructed and the Ad-PNEM2 was administered intrathecally. The rats' pain thresholds (PWL) were measured and the presence of endomorphin-2 in the cerebrospinal fluid was confirmed through ELISA. RESULTS: The Ad-PNEM2 expressed endomorphin-2 smoothly and abundantly in NIH3T3 cells at a significantly higher rate than the viral control (P<0.01) or blank control (P<0.01). The expressed endomorphin-2 was mainly observed in the cytoplasm. The concentration of endomorphin-2 in the cerebrospinal fluid increased 1 day after injection and peaked between 7 and 14 days after injection. After injection, PWL approached normal levels in the operated study group. No significant change was observed in the control groups. There was a significant correlation between PWL and endomorphin-2 level (r = 0.944, P<0.001). CONCLUSION: The constructed human endomorphin-2 transgene was expressed effectively, and endomorphin-2 expressed by the recombinant adenovirus altered the threshold to thermal stimulus and showed good analgesic effect.
Authors: L F Tseng; M Narita; C Suganuma; H Mizoguchi; M Ohsawa; H Nagase; J P Kampine Journal: J Pharmacol Exp Ther Date: 2000-02 Impact factor: 4.030
Authors: M Cavazzana-Calvo; S Hacein-Bey; G de Saint Basile; F Gross; E Yvon; P Nusbaum; F Selz; C Hue; S Certain; J L Casanova; P Bousso; F L Deist; A Fischer Journal: Science Date: 2000-04-28 Impact factor: 47.728