Multiorgan fibrosis commonly occurs in ageing mammals and its causes are not apparent. We have previously established a colony of mice with endothelial deletion of sirtuin 1 (SIRT1; SIRT1endo-/- mice) and noticed that these mice spontaneously develop traces of fibrosis in the kidneys and the heart and mount an exaggerated fibrotic response after challenge with stressors. Taking into the account the fact that SIRT1 becomes depleted in ageing, these findings raised the possibility of its participation in fibrosis of ageing. To address this possibility, we characterized the animal model of SIRT1endo-/-. Data showed that endothelial and endothelial progenitor cells exhibit higher levels of premature senescence, apoptosis, and reduced resistance to stressors, along with the impaired angiogenesis and reduced endothelium-dependent vasorelaxation. Morphologic examination of the heart and kidneys from these mice did not reveal any pathological changes in heterozygote animals, but showed a spontaneously developing mild patchy interstitial fibrosis in SIRT1endo-/- mice already at the age of 12 weeks. Quantitative PCR data demonstrated significantly higher expression of collagen I and a slightly increased collagen III under basal conditions. Echocardiography studies showed development of a mild diastolic dysfunction in 30 week-old SIRT1endo-/- mice. Applying a selection pressure on angiogenic and regenerative processes, we next examined adriamycin-induced cardiomyopathy and folate-induced nephropathy models. Data showed that trichrome staining, collagen I and III mRNA expression, as well as the short/long ratio of endoglin isoforms, a marker of senescence, were all elevated in SIRT1endo-/- mice. Folic acid-treated SIRT1endo-/- mice exhibited microvascular rarefaction, which was undetectable under basal conditions. It was previously reported that SIRT1 inhibition reduces MMP-14 mRNA abundance in cultured endothelial cells. We were able to reproduce these findings and showed that SIRT1endo-/- mice exhibited a 5-fold decrease in MMP-14 staining intensity compared to control animals and SIRT1 inhibition reduced MMP-14 protein abundance and matrylitic activity. Next, we examined other putative targets of MMP-14, endoglin and tissue transglutaminase (both normally cleaved by MMP-14), over-expression of both potentially contributing to pro-fibrotic program via type II TGF-β receptor and matrix proteins cross-linking, respectively. Inhibition of SIRT1 in HUVEC resulted in accumulation of endoglin Similarly, expression of cell-associated tissue transglutaminase was elevated in cells treated with SIRT1 inhibitor. Collectively, these findings confirmed that MMP-14 represents a novel target for SIRT1 and that depletion of the latter reduces MMP-14 expression with the resulting loss of proteolytic matrix-degrading properties of endothelial cells and subsequently decreased cleavage of endoglin and tissue transglutaminase. ConA has been previously shown to induce MMP-14. We confirmed this activity of ConA in vitro and showed that ConA effect was independent on the expression of SIRT1. Therefore, we next treated wild-type B6;129 mice with highly specific SIRT1 inhibitor III together with weekly intravenous injections of ConA, and compared those with mice receiving SIRT1 inhibitor III treatment alone. To validate the efficacy of this model, we performed quantitative real-time PCR and immunoblot analysis, which showed suppression of renal MMP-14 in the inhibitor-treated mice and its induction in ConA co-treated animals. In these mice, ConA treatment resulted in the reduction of proteinuria and significant prevention of fibrosis, as demonstrated by Masson’s trichrome staining. The mRNA synthesis of collagen I and III did not differ between SIRT1 inhibitor III-treated and SIRT1 inhibitor III/ConA-treated mice, demonstrating that ConA does not directly affect synthesis of extracellular matrix (ECM), but rather stimulates the cleavage of ECM excess through induction of MMP-14. Moreover, the angiogenic sprouting and matrilytic activity of aortic rings cultured in 3D matrigel, both suppressed in mice receiving SIRT1 inhibitor alone, showed a dramatic increase in ConA-treated animals. This data confirms the pathogenetic role of suppressed MMP-14 in exaggerated fibrotic response of SIRT1endo-/- mice. It also suggests that SIRT1 depletion with ageing and propensity toward fibrosis are linked via this novel MMP-14 mechanism.