Membrane-type 5-matrix metalloproteinase (MT5-MMP) is a proteinase mainly expressed in the nervous system with emerging roles in brain pathophysiology. The implication of MT5-MMP in Alzheimer’s disease (AD), notably its interplay with the amyloidogenic process, remains elusive. Accordingly, we crossed the genetically engineered 5xFAD mouse model of AD with MT5-MMP-deficient mice and examined the impact of MT5-MMP deficiency in bigenic 5xFAD/MT5-MMP−/− mice. At early stages (4 months) of the pathology, the levels of amyloid beta peptide (Aβ) and its amyloid precursor protein (APP) C-terminal fragment C99 were largely reduced in the cortex and hippocampus of 5xFAD/MT5-MMP−/−, compared to 5xFAD mice. Reduced amyloidosis in bigenic mice was concomitant with decreased glial reactivity and interleukin-1β (IL-1β) levels, and the preservation of long-term potentiation (LTP) and spatial learning, without changes in the activity of α-, β- and γ-secretases. The positive impact of MT5-MMP deficiency was still noticeable at 16 months of age, as illustrated by reduced amyloid burden and gliosis, and a better preservation of the cortical neuronal network and synaptophysin levels in bigenic mice. MT5-MMP expressed in HEKswe cells colocalized and co-immunoprecipitated with APP and significantly increased the levels of Aβ and C99. MT5-MMP also promoted the release of a soluble APP fragment of 95 kDa (sAPP95) in HEKswe cells. sAPP95 levels were significantly reduced in brain homogenates of 5xFAD/MT5-MMP−/− mice, supporting altogether the idea that MT5-MMP influences APP processing. MT5-MMP emerges as a new pro-amyloidogenic regulator of APP metabolism, whose deficiency alleviates amyloid pathology, neuroinflammation and cognitive decline.Electronic supplementary materialThe online version of this article (doi:10.1007/s00018-015-1992-1) contains supplementary material, which is available to authorized users.

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Astrocytes play an active role in the central nervous system and are critically involved in astrogliosis, a homotypic response of these cells to disease, injury, and associated neuroinflammation. Among the numerous molecules involved in these processes are the matrix metalloproteinases (MMPs), a family of zinc-dependent endopeptidases, secreted or membrane-bound, that regulate by proteolytic cleavage the extracellular matrix, cytokines, chemokines, cell adhesion molecules, and plasma membrane receptors. MMP activity is tightly regulated by the tissue inhibitors of MMPs (TIMPs), a family of secreted multifunctional proteins. Astrogliosis in vivo and astrocyte reactivity induced in vitro by proinflammatory cues are associated with modulation of expression and/or activity of members of the MMP/TIMP system. However, nothing is known concerning the intracellular distribution and secretory pathways of MMPs and TIMPs in astrocytes. Using a combination of cell biology, biochemistry, fluorescence and electron microscopy approaches, we investigated in cultured reactive astrocytes the intracellular distribution, transport, and secretion of MMP-2, MMP-9, TIMP-1, and TIMP-2. MMP-2 and MMP-9 demonstrate nuclear localization, differential intracellular vesicular distribution relative to the myosin V and kinesin molecular motors, and LAMP-2-labeled lysosomal compartment, and we show vesicular secretion for MMP-2, MMP-9, and their inhibitors. Our results suggest that these proteinases and their inhibitors use different pathways for trafficking and secretion for distinct astrocytic functions.

Matrix metalloproteinases (MMPs) are pleiotropic endopeptidases involved in a variety of neurodegenerative/neuroinflammatory processes through their interactions with a large number of substrates. Among those, the amyloid precursor protein (APP) and the beta amyloid peptide (Aβ) are largely associated with the development of Alzheimer’s disease (AD). However, the regulation and potential contribution of MMPs to AD remains unclear. In this study, we investigated the evolution of the expression of MMP-2, MMP-9, and membrane-type 1-MMP (MT1-MMP) in the hippocampus at different stages of the pathology (asymptomatic, prodromal-like and symptomatic) in the 5xFAD transgenic mouse AD model. In parallel we also followed the expression of functionally associated factors. Overall, the expression of MMP-2, MMP-9, and MT1-MMP was upregulated concomitantly with the tissue inhibitor of MMPs-1 (TIMP-1) and several markers of inflammatory/glial response. The three MMPs exhibited age- and cell-dependent upregulation of their expression, with MMP-2 and MMP-9 being primarily located to astrocytes, and MT1-MMP to neurons. MMP-9 and MT1-MMP were also prominently present in amyloid plaques. The levels of active MT1-MMP were highly upregulated in membrane-enriched fractions of hippocampus at 6 months of age (symptomatic phase), when the levels of APP, its metabolites APP C-terminal fragments (CTFs), and Aβ trimers were the highest. Overexpression of MT1-MMP in HEK cells carrying the human APP Swedish mutation (HEKswe) strongly increased β-secretase derived C-terminal APP fragment (C99) and Aβ levels, whereas MMP-2 overexpression nearly abolished Aβ production without affecting C99. Our data consolidate the emerging idea of a regulatory interplay between MMPs and the APP/Aβ system, and demonstrate for the first time the pro-amyloidogenic features of MT1-MMP. Further investigation will be justified to evaluate this MMP as a novel potential therapeutic target in AD.

Matrix Metalloproteinases (MMPs) play a role in migration of many cell types outside the central nervous system (CNS). Among neural cells, astrocytes are one of the main sources of MMPs in physiological and postlesional conditions. However, no data are available on the possible role of MMPs in astrocyte motility. Using an in vitro model of 2D migration and broad spectrum and selective MMP inhibitors, the authors demonstrated that MMP-2, but not MMP-9, is a key enzyme for astrocyte migration. In support of these data, the authors found constitutive expression of MMP-2 in astrocytes, while MMP-9 was nearly undetectable by gel zymography and immunocytochemical methods. The inhibition of migration by MMP inhibitors correlated with changes in cell morphology and in the organization of the actin cytoskeleton. In parallel, the characteristic focalized distribution of MMP-2 at the migration front observed in control cells became more diffuse and internalized by treatments that inhibited migration. The disruption of actin by cytochalasin D caused the partial recruitment of MMP-2 and gelatinolytic activity into actin aggregates, indicating a connection between the proteinase and the actin cytoskeleton. Finally, the authors found a co-localization of beta1-integrin with MMP-2 at the leading edge of migrating astrocytes. Altogether, these data provide the first evidence for the implication of MMP-2 in astrocyte motility, probably through the interaction of the proteinase with beta1-integrin that could act as a linker between pericellular proteolysis and the actin cytoskeleton.

The tissue inhibitor of metalloproteinases-1 (TIMP-1) belongs to a family of multifunctional proteins that inhibit matrix metalloproteinases (MMPs), but also regulate cell growth, proliferation, migration and apoptosis in non-nervous tissues. We had previously reported that kainate (KA)-mediated excitotoxic seizures induce the expression of TIMP-1 in resistant neurons and reactive astrocytes of the rat CNS, but the functional implications of these changes had not been elucidated. In the present work we used a targeted gene null mutation in mice to investigate in vivo the involvement of TIMP-1 in neuronal death and axonal sprouting following KA. We found no differences in seizure behaviour between the wild-type (WT) and the TIMP-1 knock-out (KO) mice, without any compensation by other TIMPs, at least at the mRNA level. However, the TIMP-1 KO mice were resistant to excitotoxicity and did not undergo the typical mossy fibre sprouting observed in WT mice. The lack of TIMP-1 paradoxically hampered the increase in the activity of MMPs observed in the seizing WT mice. In addition, we demonstrate that learning and memory are impaired in untreated KO mice. In conclusion, this study provides the first in vivo evidence for the implication of TIMP-1 in neuronal death and axonal sprouting in a pathological situation, but also suggests the involvement of TIMP-1 in the synaptic mechanisms underlying learning and memory in physiological conditions. More generally, these data support the idea that the control of proteolysis is instrumental for pathological and physiological processes in the brain.

We previously reported that deficiency of membrane-type five matrix metalloproteinase (MT5-MMP) prevents amyloid pathology in the cortex and hippocampus of 5xFAD mice, and ameliorates the functional outcome. We have now investigated whether the integrity of another important area affected in Alzheimer’s disease (AD), the frontal cortex, was also preserved upon MT5-MMP deficiency in 4-month old mice at prodromal stages of the pathology. We used the olfactory H-maze (OHM) to show that learning impairment associated with dysfunctions of the frontal cortex in 5xFAD was prevented in bigenic 5xFAD/MT5-MMP−/− mice. The latter exhibited concomitant drastic reductions of amyloid beta peptide (Aβ) assemblies (soluble, oligomeric and fibrillary) and its immediate precursor, C99. Simultaneously, astrocyte reactivity and tumor necrosis factor alpha (TNF-α) levels were also lowered. Moreover, MT5-MMP deficiency induced a decrease in N-terminal soluble fragments of amyloid precursor protein (APP), including soluble APPα (sAPPα), sAPPβ and the MT5-MMP-linked fragment of 95 kDa, sAPP95. However, the lack of MT5-MMP did not affect the activity of β- and γ-secretases. In cultured HEKswe cells, transiently expressed MT5-MMP localized to early endosomes and increased the content of APP and Aβ40 in these organelles, as well as Aβ levels in cell supernatants. This is the first evidence that the pro-amyloidogenic features of MT5-MMP lie, at least in part, on the ability of the proteinase to promote trafficking into one of the amyloidogenic subcellular loci. Together, our data further support the pathogenic role of MT5-MMP in AD and that its inhibition improves the functional and pathological outcomes, in this case in the frontal cortex. These data also support the idea that MT5-MMP could become a novel therapeutic target in AD.

Altogether, these data strongly suggest that obesity may interfere with the orosensory system responsible for the detection of free long-chain fatty acids in humans. This trial was registered at clinicaltrials.gov as NCT02028975.