BACKGROUND: The ADAMTS7 locus was genome-wide significantly associated with coronary artery disease. Lack of the ECM (extracellular matrix) protease ADAMTS-7 (A disintegrin and metalloproteinase-7) was shown to reduce atherosclerotic plaque formation. Here, we sought to identify molecular mechanisms and downstream targets of ADAMTS-7 mediating the risk of atherosclerosis. METHODS: Targets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe−/− and Apoe−/−Adamts7−/− mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, coimmunoprecipitation, and Förster resonance energy transfer–based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. RESULTS: In humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe−/− mice, atherosclerotic aortas of Apoe−/− mice lacking Adamts-7 (Apoe−/−Adamts7−/−) contained higher protein levels of Timp-1 (tissue inhibitor of metalloprotease-1). In coimmunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target MMP-9 (matrix metalloprotease-9). As a downstream mechanism, we investigated collagen content in plaques of Apoe−/− and Apoe−/−Adamts7−/− mice after a Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe−/− as compared to Apoe−/− Adamts7−/− mice. To facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim of decreasing TIMP-1 degradation, we designed a Förster resonance energy transfer–based assay targeting the ADAMTS-7 catalytic site. CONCLUSIONS: ADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise associated with coronary artery disease. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for the reduction of atherosclerosis-related events.

Background The ADAMTS7 locus was genome-wide significantly associated with coronary artery disease (CAD). Lack of the extracellular matrix (ECM) protease ADAMTS-7 was shown to reduce atherosclerotic plaque formation. Objective To identify molecular mechanisms and downstream targets of ADAMTS-7 mediating risk of atherosclerosis. Methods Targets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, co-immunoprecipitation, and Förster resonance energy transfer (FRET)-based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. Results In humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe-/- mice, atherosclerotic aortas of Apoe-/- mice lacking Adamts-7 (Apoe-/-Adamts7-/-) contained higher protein levels of tissue inhibitor of metalloproteases 1 (Timp-1). In co-immunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target matrix metalloprotease 9 (MMP-9) As a downstream mechanism, we investigated collagen content in plaques of Apoe-/- and Apoe-/-Adamts7-/- mice after Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. In order to facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim to decrease TIMP-1 degradation, we designed a FRET-based assay targeting the ADAMTS-7 catalytic site. Conclusion ADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise genome-wide significantly associated with CAD. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for reduction of atherosclerosis-related events.

Background: The gene encoding the extracellular matrix (ECM) protease ADAMTS-7 was associated with coronary artery disease (CAD) in genome-wide association studies. ADAMTS-7 is expressed at all stages in human plaques and mice lacking Adamts-7 displayed reduced atherosclerotic plaque formation. While these findings render ADAMTS-7 a promising therapeutic target, the underlying mechanisms remain unknown. Methods and Results: Here, we sought to identify downstream mechanisms of ADAMTS-7 in atherosclerotic plaque formation. Targets of Adamts-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques in Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. The endogenous inhibitor of matrix metalloproteinases (MMP) Timp-1 was identified as a novel target of Adamts-7. Adamts-7 and Timp-1 were found to be co-localized in atherosclerotic plaques and co-immunoprecipitation (Co-IP) studies revealed TIMP-1 as the first putative target to bind to the catalytic domain of ADAMTS-7. In vitro degradation assays demonstrated that ADAMTS-7 degrades TIMP-1. Co-IP furthermore revealed less binding of TIMP-1 to its canonical target MMP-9 when ADAMTS-7 was present. In line, scaffolding and degradation of TIMP-1 by ADAMTS-7 impaired TIMP-1-mediated inhibition of MMP-9 in vitro. As a downstream mechanism, we investigated collagen content in atherosclerotic plaques of Apoe-/- and Apoe-/- Adamts7-/- mice after Western diet. Collagen stainings of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. Since ADAMTS-7 might exert its pro-atherogenic effects through interaction with TIMP-1, we established a TIMP-1-ADAMTS-7-interaction assay based on Förster resonance energy transfer (FRET) to identify inhibitors of this protein-protein interaction. Conclusion and Outlook: TIMP-1 represents a novel downstream target of ADAMTS-7 that can explain the role of this novel risk factor in CAD. Our FRET-based protein-protein interaction assay may be used for high-throughput screening to identifiy inhibitors and prevent the initiation or slow the progression of atherosclerosis.

Introduction. Visual-motor integration (VMI) is defined as the degree to which visual perception (VP) and finger-hand movements are well coordinated. The VMI consists of two components: VP and motor coordination (MC). The main goal of our research was to determine whether there are differences in age and gender categories in VMI, VP and MC scores, as well as whether there is a correlation between VMI and school success of younger school-aged children. Methods. Out of 103 student respondents, 52 were female (50.5%), aged 6 to 11 years (8.05 ± 1.44 years), divided into two groups according to age: 6-8 years (first, second and third grade) and 9-11 years (fourth to fifth grade). Data on the level of VMI were obtained by applying the following tests: Beery-Buktenica Developmental Test of VMI, VP test and MC test. Results. In the older age group of subjects, a significant difference was observed in the mean values of the score on the VMI (12.67 ± 1.92), VP (23.69 ± 3.21) and MC (24.34 ± 3.23) tests comparing to the younger group of subjects (9.98 ± 2.12; 20.80 ± 3.2; 19.65 ± 3.82) (p < 0.001), while the difference in the mean values of scores in relation to gender was not observed. A significant, positive and strong correlation was observed between the scores on the VMI, VP and MC test with the success of second to fifth grade students (p < 0.050). Conclusion. Given such a strong correlation between VMI and the success of younger students, we conclude that it is important that VMI disabilities are identified in time, so that these students can be referred for further assessment and receive the necessary support.

Djeca sa smetnjama u razvoju manifestuju teškoće u razvoju i ne dosežu zadovoljavajući nivo razvoja, naročito ako im se ne pruži podrška u vidu njege i obrazovanja. Viši nivoi empatije su značajan faktor koji može uticati na pojavu pozitivnijih stavova učenika prema vršnjacima sa smetnjama u razvoju. Cilj istraživanja je bio da se procjeni da li empatija ima uticaj na stavove učenika prema vršnjacima sa smetnjama u razvoju. Studija je obuhvatila 120 učenika 4. i 5. razreda osnovnih škola, oba pola, uzrasta 10 i 11 godina. Istraživanje je rađeno u Banjaluci tokom 2020.godine. Ispitanici su podijeljeni na dvije grupe, one koji su pohađali nastavu sa učenicima sa smetnjama u razvoju i one koji nisu. Za ispitivanje stavova učenika prema učenicima sa smetnjama u razvoju korištena je Čedok Mekmaster skala, dok je za procjenu empatije korišten Indeks međuljudskog reagovanja. Korelacionom analizom utvrđena je značajna (r=0,18; p<0,05) pozitivna povezanost između stavova učenika prema vršnjacima sa smetnjama u razvoju i zauzimanja perspektive. Učenici koji pohađaju nastavu sa učenicima sa smetnjama u razvoju imaju značajno (t=-2,3; p=0,02) niže vrijednosti ukupnog skora stavova (3,05±0,28) u odnosu na učenike koji ne pohađaju (3,16±0,28). Naše istraživanje je pokazalo da empatija imaju značajan uticaj na stavove učenika prema vršnjacima sa smetnjama u razvoju i da učenici sa višim nivoem empatije imaju pozitivniji stav prema ovim vršnjacima. Takođe, učenici koji su u inkluziji imaju negativniji stav prema vršnjacima ometenim u razvoju u odnosu na učenike koji nisu u inkluziji.