Mesoporous　bioactive　glasses　(MBGs)　have　displayed　great　potential　in　dental　restorations　and　bone　regeneration　for　their　controllable　structure　and　great　bioactivity.　In　this　work,　we　have　systematically　investigated　the　effect　of　nitrogen　on　structure　evolution,　apatite-forming　ability　and　cytotoxicity　in　MBG　by　successfully　preparing　the　N-containing　MBG　nanospheres　by　a　modified　sol-gel　method.　It　is　interesting　to　demonstrate　that　all　obtained　samples　exhibit　uniform　interstitial　mesoporous　nanospheres　microstructure,　with　the　particles　size　increases　as　the　nitrogen　addition　increases　from　5　to　15　mol.%.　Notably,　a　large　amount　of　blooming　flower-like　hydroxyapatite　deposited　on　the　surface　of　15N-MBG　after　immersed　in　simulated　body　fluid　for　7　days,　exhibiting　excellent　apatite-forming　ability.　It　is　highlighted　that　the　incorporation　of　nitrogen　can　stimulate　cell　proliferation　at　early　incubation　time.　Moreover,　all　the　MBGs　have　no　obvious　inhibitory　impact　on　the　hPDLCs　growth　at　extracted　concentration.　Furthermore,　via　glass　network　connectivity　analysis　from　molecular　dynamics　simulations,　the　decrease　of　network　connectivity　caused　by　the　incorporation　of　nitrogen　could　provide　looser　glass　network　and　higher　bioactivity,　which　explains　the　experimental　results　well.