In Vivo Imaging of Matrix Metalloproteinase 12 and Matrix Metalloproteinase 13 Activities in the Mouse Model of Collagen-Induced Arthritis
In Vivo Imaging of Matrix Metalloproteinase 12 and Matrix Metalloproteinase 13 Activities in the Mouse Model of Collagen-Induced Arthritis
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DOI：
10.1002/art.38295
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发表时间：
2014-03-01
影响因子：
13.3
通讯作者：
Nagase, Hideaki
中科院分区：
文献类型：
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作者：
Lim, Ngee Han;Meinjohanns, Ernst;Nagase, Hideaki
目标。目的：研制可激活的Forster共振能量转移(FRET)底物探针，检测炎性关节炎小鼠体内基质金属蛋白酶12(MMP12)和MMP13的活性。由基质金属蛋白酶-12和基质金属蛋白酶-13激活的多肽FRET探针是从从膦基肽抑制剂文库中选择的抑制剂中反向设计的。体外用基质金属蛋白酶-1、基质金属蛋白酶-2、基质金属蛋白酶-3、基质金属蛋白酶-12和基质金属蛋白酶-13证明了探针的选择性。在酵母多糖诱导的小鼠炎症模型中检测探针的体内激活，并通过基质金属蛋白酶抑制剂GM6001和特定的合成抑制剂基质金属蛋白酶-12和基质金属蛋白酶-13来评价探针的特异性。在活体胶原诱导性关节炎(CIA)模型中，这些探针被用于监测这些酶的活性。MMP12和MMP13活性探针(MMP12ap和MMP13ap)区分两种酶活性。GM6001和它们各自的特异性抑制剂抑制了这些探针的体内激活。在CIA模型中，MMP12ap的活性在发病后5天达到高峰，并在此期间与疾病严重程度呈显著相关(r=0.85P<；0.0001)。MMP13ap活性在发病后逐渐升高，且在较长时间内与病情严重程度呈正相关(r=0.58P&0.0001)。我们产生了两个选择性的FRET探针，可以用来监测活体动物中基质金属蛋白酶-12和基质金属蛋白酶-13的活性。MMP12ap跟踪CIA炎症的初始阶段，而MMP13ap跟踪疾病的进展。这些探针的特异性有助于监测基质金属蛋白酶抑制剂的疗效。
Objective. To develop enzyme-activatable Forster resonance energy transfer (FRET) substrate probes to detect matrix metalloproteinase 12 (MMP-12) and MMP-13 activities in vivo in mouse models of inflammatory arthritis.Methods. Peptidic FRET probes activated by MMP-12 and MMP-13 were reverse designed from inhibitors selected from a phosphinic peptide inhibitor library. Selectivity of the probes was demonstrated in vitro using MMP-1, MMP-2, MMP-3, MMP-12, and MMP-13. In vivo activation of the probes was tested in the zymosan-induced mouse model of inflammation, and probe specificity was evaluated by the MMP inhibitor GM6001 and specific synthetic inhibitors of MMP-12 and MMP-13. The probes were used to monitor these enzyme activities in the collagen-induced arthritis (CIA) model in vivo.Results. The MMP-12 and MMP-13 activity probes (MMP12ap and MMP13ap, respectively) discriminated between the activities of the 2 enzymes. The in vivo activation of these probes was inhibited by GM6001 and by their respective specific inhibitors. In the CIA model, MMP12ap activation peaked 5 days after disease onset and showed strong correlation with disease severity during this time (r = 0.85, P < 0.0001). MMP13ap activation increased gradually after disease onset and correlated with disease severity over a longer period of 15 days (r = 0.58, P < 0.0001).Conclusion. We generated two selective FRET probes that can be used to monitor MMP-12 and MMP-13 activities in live animals. MMP12ap follows the initial stage of inflammation in CIA, while MMP13ap follows the progression of the disease. The specificity of these probes is useful in monitoring the efficacy of MMP inhibitors.