doi: 10.1016/j.gendis.2024.101231.
eCollection 2024 Sep.
MMP-9 deficiency accelerates the progress of periodontitis
Affiliations
- PMID: 38882016
- PMCID: PMC11176640
- DOI: 10.1016/j.gendis.2024.101231
Item in Clipboard
MMP-9 deficiency accelerates the progress of periodontitis
Genes Dis.
.
No abstract available
Conflict of interest statement
The authors declared no conflict of interests.
Figures
MMP-9 deficiency impairs bone metabolism and cell proliferation as well as causes inflammation in the periodontium. (A) Representative radiographs show an overall reduction of the mineralization of the alveolar bone and loss of the alveolar bone in the root furcation regions (arrows) of the MMP-9 KO mice at 6, 12, and 29 months old, and the alveolar bone was porous in the apical space in the furcation regions. Molar cusps (arrowheads) were severely attrited in MMP-9 KO mice at these ages. (B) μCT shows that BMD of the alveolar bone of MMP-9 KO mice was decreased compared with the control groups. With aging, the alveolar bone resorption in the alveolar bone of MMP-9 KO mice was more severe. A porous region (arrows) in the apical areas was seen. Hyper-mineralized spots were deposited on the alveolar bones in 29-month-old MMP-9 KO mice. (C) Graph representation of BMD in the wild-type and MMP-9 KO mice acts as mean ± standard error of the mean. (D) Resin infiltration and acid etching SEM analysis revealed noteworthy findings in the alveolar bone of 2-, 6-, 12-, and 29-month-old MMP-9 knockout (KO) mice. Within the apical regions of the alveolar bone, trabecular bones displayed clear evidence of bone resorption, leading to porous spaces in the mutant mice. Additionally, calcium deposition was observed in the trabecular bones of 29-month-old MMP-9 KO mice. In 2-month-old wild-type mice, osteocytes within the trabecular bones exhibited an even distribution, whereas osteocytes in the alveolar bone of the same age MMP-9 KO mice displayed less organization. As aging progressed, the number of osteocytes in the trabecular bones of MMP-9 KO mice decreased, accompanied by an increase in matrix presence. Conversely, osteocytes and lacunae structures in the alveolar bones of age-matched control groups remained highly organized and uniformly distributed. (E) μCT analyzed alveolar bone microstructure parameters in the wild-type and MMP-9 KO mice. TV and BV on the alveolar bone were from the wild-type and MMP-9 KO mice at 2-, 6-, 12-, and 29-month-old. Quantitative trabecular bone significantly reduced BV, BV/TV, and Tb. Th. in MMP-9 KO mice at 6-, 12- and 29-month-old. BV, bone volume; TV, tissue volume; BV/TV, bone volume/tissue volume; Tb. Th., trabecular thickness. NS, no significance. (F) Alveolar bone loss by the linear CEJ-ABC distance was measured by a stereomicroscope. The samples were stained with methylene blue to reveal the CEJ. Teeth were worn in the MMP-9 KO mice (arrows). CEJ, cemental-enamel junction; ABC, alveolar bone crest. (G) Distance from the CEJ to the ABC in the wild-type and MMP-9 KO mice was measured by μCT. (H) Graphical representations of CEJ to ABC were measured as mean ± standard error of the mean (n = 4). (I) Apoptotic cells in the primary BMSCs. A fluorescent terminal deoxynucleotidyl transferase (TdT) assay was used for detection of apoptotic cells (green) in cultured BMSCs from 1- and 3-month-old wild-type and MMP-9 KO mice was performed. Hoechst was used for nucleus staining (Blue). Bar = 20 μm. Relative apoptotic cells in the BMSCs from the wild-type and null mice were shown. The expression of apoptotic cells from the wild-type mice acted as a 1.0. (J) Cell proliferation of the BMSCs from the 1- and 3-month-old wild-type and MMP-9 KO mice was observed. Cell proliferation was identified by BrdU incorporation. Cells were transferred into four-well glass slides and incubated with 30 mM BrdU in a culture medium for 6 h. The cells were treated with an anti-BrdU antibody, followed by the secondary antibody with Alexa Fluo® 488 green. For nucleus staining, the cells were incubated with a 1:5000 dilution of Hoechst. Images were obtained with a Nikon inverted microscope. Proliferative cells were expressed as a percentage of the number of BrdU-positive cells relative to the total number of Hoechst-positive nuclei. (K) Analysis of cell migration from wild-type and MMP-9 null mice. Migration of the BMSCs from wild-type and MMP-9 KO mice was measured using BD BiocoatTM Matrigel invasion chambers. Cell migration was quantified by counting the number of cell migrants passing through the membranes after 12 h. The wild-type cell migration was set as 100 %. (L) Functional role of the wild-type and MMP-9 mutant cells by gelatin zymography. Increases in MMP-9 levels from the wild-type cells catalyze gelatin with increasing MMP-9 concentrations whereas in MMP-9 KO cells failed to process gelatin. (M)In situ zymography of MMP-9 activity on its substrate in vivo. The BMSCs isolated from the wild-type and MMP-9 KO mice were grown on DQ-fluorescent Col4-coated slides for 12 h. The cells were fixed and images were observed under an immunofluorescent inverted microscope. Data showed that the number and intensity of Col4 degradation in the wild-type cells were higher than those of the MMP-9 null cells. The percentage of positive fluorescent spots in the BMSCs from the wild-type and MMP-9 KO mice was obtained. “n” indicates the number of animals used in each group. (N) Quantitative reverse transcription PCR analysis revealed that mRNA expression levels of ALP, BSP, Col1α1, DMP1, OPN, and OSN in BMSCs from 3-month-old MMP-9 null mice were significantly reduced when compared with age-matched wild-type mice. However, there was no significant difference in the expression of Col4α1 in BMSCs between the wild-type and MMP-9 KO mice. The data were presented as the mean ± standard deviation (n = 4). ∗P < 0.05 and ∗∗P < 0.01. Details of the primers used in this study are shown in Table S1. (O) Hematoxylin and eosin staining of periodontal tissues in the wild-type and MMP-9 KO mice. MMP-9 KO mice had progressive periodontal diseases with aging. (c, d) Higher magnification views from the boxed area in 2-month-old MMP-9 KO mice. Gingival epithelium and PDL had torn away from the cementum. There was no remarkable difference in the alveolar bone of the wild-type versus MMP-9 KO mice. (a, b) Higher magnification views from the boxed area in 2-month-old wild-type mice. (h–j, n–p) Higher magnification from the boxed areas in 6-, and 12-month-old MMP-9 null mice. The gingival membrane and PDL were severely detached from the cementum (arrows). Inflammation was seen in the periodontal tissue and alveolar bone resorption was present in MMP-9 KO mice (arrowheads). (e–g, k–m) Higher magnification views of the alveolar bone were from the boxed areas in 6- and 12-month-old wild-type mice. (t–v) Higher magnification views of the boxed area in 29-month-old MMP-9 KO mice showed periodontal tissue destruction and detachment of gingival epithelium and PDL from the cementum. Inflammation was noted in the periodontal tissue. Cement structure was abnormal, and bone resorption was observed in the alveolar bones in MMP-9 KO mice. (q–s) Higher magnification views from the boxed area in 29-month-old wild-type samples. AB, alveolar bone; C, cementum; G, gingival membrane; If, inflammatory cells; PDL, periodontal ligament. Bars, 50 μm. Bars in boxes, 200 μm. (P) Immunohistochemistry analysis of caspase 3, IL-1β, Ki67, osteoclast (OC), osteoprotegerin (OPG), and RANKL in the periodontal tissues of the wild-type and MMP-9 KO mice. A higher magnification image of the boxed areas shows the expression of caspase-3, IL-1β, Ki67, OC, OPG, and RANKL proteins in the wild-type and MMP-9 KO teeth. (Q) Graph representation demonstrated protein density as mean ± standard error of the mean in the alveolar bone of the wild-type and MMP-9 KO mice. The quantitative analysis of caspase-3, IL-1β, Ki67, OC, OPG, and RANKL protein detection in the periodontal tissues of the wild-type and MMP-9 KO mice was shown. (R) The RANKL/OPG ratio in the alveolar bone and PDL of the wild-type and MMP-9 KO mice was determined. BMD, bone mineral density; BMSCs, bone marrow stromal cells; Cas3, caspase-3; μCT, micro-computed tomography; M, month; n, number; KO, knockout; WT, wild type; ∗P < 0.05; ∗∗P < 0.01.
References
-
- Sharony R., Borochowitz Z., Cohen L., et al. Prenatal course of metaphyseal anadysplasia associated with homozygous mutation in MMP9 identified by exome sequencing. Clin Genet. 2017;92(6):645–648. - PubMed
-
- Wan C., Yuan G., Yang J., et al. MMP9 deficiency increased the size of experimentally induced apical periodontitis. J Endod. 2014;40(5):658–664. - PubMed
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