The Therapeutic Effects Of Pentapeptide Compound PLNPK On Adjuvant Arthritis And Its Mechanisms

Objeclive:To investigate the therapeutic effect of pentapeptide compound PLNPK on adjuvant arthritis and its possible mechanisms.Methods:1. Freund's Complete adjuvant(10mg/ml) was injected 0.1ml intradermally in the right hind footpad of Wistar rats to induce AA. Intracutaneous injection with given drugs was commenced daily from the seventh day after immunization. The fight and left ankle perimeter of AA rats were measured every four days. The effect of PLNPK on the pathology damage of ankle joints was assayed by HE colorization. The effect of PLNPK on the proliferation of fibroblast-like synoviocytes(FLS), the infiltration and activation of macrophages were assayed by Vimentin, CD68 and CD86 immunohistochemistry in the synovial tissue of AA rats.2. Type II collagenase and trypsin digested synovial tissue of AA rats and establish the cultivate system of FLS in AA rats in vitro. The inhibiting effect of PLNPK on the proliferation of FLS was assayed by MTS method. The sterile Fluid Thioglycollate Medium was injected to the abdominal cavity of AA rats, then we extracted peritoneal macrophages(PEM_ψ). PEM_ψ were stimulated by PLNPK in vitro and we extracted the supernatant liquid. FLS and the supernatant liquid were incubated together. The inhibiting effect of PLNPK on FLS was assayed by MT method.3. The inhibiting effect of PLNPK on the proliferation of RAW264.7 was assayed by MTS method. The inhibiting effect of PLNPK on the proliferation and activation degree of PEM_ψ in normal rats was assayed by MTS method and CD86 immunocytochemistry. The inhibiting effect of PLNPK on the proliferation and activation degree of PEM_ψ in AA rats was assayed by MTS method and CD86 immunocytochemistry. The inhibiting effect of PLNPK on the function of AA rats ??secreting TNF-αand IL-1βwas assayed by ELISA.Results:1. PLNPK can relieve the primary and secondary ankle mortise swelling at the dose of 200ug/kg/d and 100ug/kg/d (P＜0.05). PLNPK can ameliorate the ankle pathological damage of AA rats in vivo. PLNPK can inhibit the proliferation of FLS, the infiltration and the activation degree of macrophages in vivo.2. PLNPK(0.4-3.2mg/ml) can not significantly directly inhibit the proliferation of FLS in vitro. PLNPK(0.8-3.2mg/ml) can significantly indirectly inhibit the proliferation of FLS in vitro and the inhibiting rate is between 17.89% and 35.49% (P＜0.05).3. PLNPK(0.8-3.2mg/ml) can significantly inhibit the proliferation of RAW264.7 in vitro and the inhibiting rate is between 21.24% and 81.68% (P＜0.05). PLNPK(1.6-3.2mg/ml) can significantly inhibit the proliferation of PEM_ψ in normar rats in vitro and the inhibiting rate is between 21.32% and 77.80% (P＜0.05). PLNPK(1.6-3.2mg/ml) can decrease the number of CD68⁺ PEM_ψ in normal rats in vitro. PLNPK(0.8-3.2mg/ml) can significantly inhibit the proliferation of PEM_ψ in AA rats in vitro and the inhibiting rate is between 36.41% and 82.85% (P＜0.05). PLNPK(0.4-3.2mg/ml) can decrease the number of CD68+ PEM_ψ in AA rats in vitro. PLNPK(0.4-3.2mg/ml) can significantly reduce the amount of IL-1βsecreted by PEM_ψ and the inhibiting rate is between 39.23% and 94.72% in vitro (P＜0.05). PLNPK(0.8-3.2mg/ml) can significantly reduce the amount of TNF-αsecreted by PEM_ψ and the inhibiting rate is between 40.70% and 79.32% in vitro (P＜0.05).Conclnsions:Pentapeptide compound PLNPK has the therapeutic effect on AA rats. PLNPK can relieve the primary and secondary ankle swelling, and ameliorate the pathology damage. PLNPK may exert its effect by inhibiting macrophage proliferation,activation and secreting TNF-αand IL-1β, therefore inhibiting the proliferation and ??activation of FLS.