DOI: 10.5061/dryad.6djh9w19c
title: Data for: Mitochondrial Ca2+-coupled generation of reactive oxygen species, peroxynitrite formation, and endothelial dysfunction in Cantú syndrome

description: Cantú syndrome is a multisystem disorder caused by gain-of-function (GOF)
 mutations in KCNJ8 and ABCC9, the genes
 encoding the pore-forming inward rectifier Kir6.1 and regulatory
 sulfonylurea receptor SUR2B subunits, respectively, of vascular
 ATP-sensitive K+ channels (KATP). In this study, we investigated
 changes in the vascular endothelium in mice in which Cantú syndrome
 -associated Kcnj8 or Abcc9 mutations were
 knocked-in to the endogenous loci. We found that endothelium-dependent
 dilation was impaired in small mesenteric arteries from Cantú mice. Loss
 of endothelium-dependent vasodilation led to increased vasoconstriction in
 response to intraluminal pressure or treatment with the adrenergic
 receptor agonist phenylephrine. We also found that either
 KATP GOF or acute activation of KATP channels with
 pinacidil increased the amplitude and frequency of wave-like
 Ca2+ events generated in the endothelium in response to the
 vasodilator agonist carbachol. Increased cytosolic Ca2+ signaling
 activity in arterial endothelial cells from Cantú mice was associated with
 elevated mitochondrial [Ca2+] and enhanced reactive oxygen species (ROS)
 and peroxynitrite levels. Scavenging intracellular or mitochondrial ROS
 restored endothelium-dependent vasodilation in the arteries of mice with
 KATP GOF mutations. We conclude that mitochondrial Ca2+ overload
 and ROS generation, which subsequently leads to nitric oxide consumption
 and peroxynitrite formation, cause endothelial dysfunction in mice with
 Cantú syndrome.
description: GraphPad Prism software (Version 9.4.1) was used to conduct
 statistical analyses and generate graphs. The data presented are expressed
 as means ± SEM, with "n" indicating the number of vessels or
 cells analyzed. One-way analysis of variance (ANOVA) with Tukey's
 multiple comparisons test and two-way ANOVA with Tukey's or
 Šídák's multiple comparisons test were used for statistical analyses.
 <em>P</em>-values &lt; 0.05 were considered statistically
 significant for all analyses.
description: # Data for: Mitochondrial Ca2+-coupled generation of reactive oxygen
 species, peroxynitrite formation, and endothelial dysfunction in Cantú
 syndrome --- ## Description of the data and file structure ###
 Figure\_1\_Processed\_Data 1D: Summary data of whole-cell current
 densities from voltage-clamp recordings of WT and Kir6.1wt/VM SMCs with
 indicated treatments\ 1F: Summary data of whole-cell current densities
 from voltage-clamp recordings of vascular endothelial cells from WT and
 Kir6.1wt/VM with indicated treatments 1H: Summary of current-clamp
 recordings of vascular endothelial cells from WT and Kir6.1wt/VM mice. ###
 Figure\_2\_Processed\_Data 2A: Representative recordings data showing
 endothelium-dependent vasodilation evoked by the muscarinic receptor
 agonist carbachol (CCh) in mesenteric arterioles from WT and Kir6.1wt/VM
 mice 2B: Summary data showing endothelium-dependent vasodilation evoked by
 the muscarinic receptor agonist carbachol (CCh) in mesenteric arterioles
 from WT and Kir6.1wt/VM mice 2C: Representative traces data showing
 vasodilatory responses to the nitric oxide donor sodium nitroprusside
 (SNP) in isolated mesenteric arteries from WT and Kir6.1wt/VM mice 2D:
 Summary data showing vasodilatory responses to the nitric oxide donor
 sodium nitroprusside (SNP) in isolated mesenteric arteries from WT and
 Kir6.1wt/VM mice ### Figure\_3\_Processed\_Data 3A: Representative traces
 data of the luminal diameter of isolated mesenteric arteries from WT and
 Kir6.1wt/VM mice showing changes in lumen diameter in response to
 increases in intraluminal pressure under active and passive (Ca2+-free)
 conditions 3B: Summary data of myogenic tone from WT and Kir6.1wt/VM mice
 3C: Representative traces data showing constriction of mesenteric arteries
 from WT and Kir6.1wt/VM mice in response to increasing concentrations of
 the α1-adrenoceptor agonist phenylephrine (PE) 3D: Summary data showing
 constriction of mesenteric arteries from WT and Kir6.1wt/VM mice in
 response to increasing concentrations of the α1-adrenoceptor agonist
 phenylephrine (PE) 3E: Representative recording data of the lumen diameter
 of myogenic tone showing changes in the lumen diameter of
 endothelium-denuded mesenteric arteries from WT and Kir6.1wt/VM mice 3F:
 Summary data of myogenic tone showing changes in the lumen diameter of
 endothelium-denuded mesenteric arteries from WT and Kir6.1wt/VM mice ###
 Figure\_4\_Processed\_Data 4B: Representative ΔF/F0 vs. time plots data of
 Ca2+ events from multiple Ca2+ event sites in indicated treatments 4C:
 Summary data showing the effects of pinacidil and glibenclamide on the
 amplitude (ΔF/F0), frequency (Hz), and the number of active sites ###
 Figure\_5\_Processed\_Data 5B: Representative ΔF/F0 vs. time plots data of
 Ca2+ events from multiple ROIs in WT and Kir6.1wt/VM 5C: Summary data
 showing the effects of pinacidil and glibenclamide on the amplitude
 (ΔF/F0), frequency (Hz), and the number of active sites 5D: Summary data
 showing the effects of pinacidil and glibenclamide on the frequency (Hz)
 5E: Summary data showing the effects of pinacidil and glibenclamide on the
 number of active sites ### Figure\_6\_Processed\_Data 6B: Representative
 ΔF/F0 vs. time plots data of X-Rhod-1 fluorescence for each condition 6C:
 Summary data showing changes in X-Rhod-1 fluorescence intensity 6E:
 Representative ΔF/F0 vs. time plots data of the change in fluorescence
 intensity under each condition 6F: Summary data showing the changes in
 CellRox fluorescence intensity ### Figure\_7\_Processed\_Data 7B: Summary
 data showing the change in Fl-B fluorescence intensity 7D: Summary data
 showing mean fluorescent intensity for nitrated tyrosine signal ###
 Figure\_8\_Processed\_Data 8A: Representative traces showing data
 CCh-evoked endothelium-dependent vasodilation in mesenteric arteries from
 Kir6.1wt/VM mice before and after treatment with the membrane-permeable
 intracellular ROS scavenger PEG-SOD 8B: Summary data showing CCh-evoked
 endothelium-dependent vasodilation in mesenteric arteries from Kir6.1wt/VM
 mice before and after treatment with the membrane-permeable intracellular
 ROS scavenger PEG-SOD 8C: Representative traces data showing CCh-evoked
 endothelium-dependent vasodilation in mesenteric arteries from Kir6.1wt/VM
 mice before and after treatment with the mitochondria-targeted-MitoTEMPO
 8D: Summary data showing CCh-evoked endothelium-dependent vasodilation in
 mesenteric arteries from Kir6.1wt/VM mice before and after treatment with
 the mitochondria-targeted-MitoTEMPO ### Supplemental Figure *1*
 Processed\_Data S1A: Representative traces data showing
 endothelium-dependent vasodilation induced by the muscarinic receptor
 agonist carbachol (CCh) in mesenteric arteries from WT and SUR2AV/AV mice
 S1B: Summary data showing endothelium-dependent vasodilation induced by
 the muscarinic receptor agonist carbachol (CCh) in mesenteric arteries
 from WT and SUR2AV/AV mice S1C: Representative recording data showing
 vasodilatory responses to the nitric oxide donor sodium nitroprusside
 (SNP) in isolated mesenteric arteries from WT and SUR2AV/AV mice S1D:
 Summary data showing vasodilatory responses to the nitric oxide donor
 sodium nitroprusside (SNP) in isolated mesenteric arteries from WT and
 SUR2AV/AV mice ### Supplemental Figure *2* Processed\_Data S2A: Typical
 recording data of the change in lumen diameter of third-order mesenteric
 arteries from WT and SUR2AV/AV mice in response to increases in
 intraluminal pressure under active and passive (Ca2+-free) conditions S2B:
 Summary data of the change in lumen diameter of third-order mesenteric
 arteries from WT and SUR2AV/AV mice in response to increases in
 intraluminal pressure under active and passive (Ca2+-free) conditions S2C:
 Representative recording data showing constriction of mesenteric arteries
 from WT and SUR2AV/AV mice in response to increasing concentrations of the
 α1-adrenoceptor agonist phenylephrine (PE) S2D: Summary data showing
 constriction of mesenteric arteries from WT and SUR2AV/AV mice in response
 to increasing concentrations of the α1-adrenoceptor agonist phenylephrine
 (PE) ### Supplemental Figure *3* Processed\_Data S3A: Representative
 recording data of changes in lumen diameter in response to increases in
 intraluminal pressure under active and passive (Ca2+-free) conditions in
 endothelium-denuded mesenteric arteries from SUR2AV/AV mice S3B: Summary
 data of changes in lumen diameter in response to increases in intraluminal
 pressure under active and passive (Ca2+-free) conditions in
 endothelium-denuded mesenteric arteries from SUR2AV/AV mice ###
 Supplemental Figure *4* Processed\_Data S4A: Representative traces of the
 luminal diameter of isolated pial arteries from WT and Kir6.1wt/VM S4B:
 Summary data of myogenic tone from WT and Kir6.1wt/VM mice S4C:
 Representative traces of the luminal diameter of isolated pial arteries
 from WT and SURAV/AV mice S4D: Summary data of myogenic tone from WT and
 SURAV/AV mice ### Supplemental Figure *5* Processed\_Data S5A: Summary
 data showing vasoconstriction of isolated mesenteric arteries from
 Kir6.1wt/VM compared to WT controls in response to 60 mM KCl S5B: Summary
 data showing vasoconstriction of isolated mesenteric arteries from
 SUR2AV/AV compared to WT controls in response to 60 mM KCl ###
 Supplemental Figure *6* Processed\_Data S6A: Summary data showing the
 passive diameter of isolated mesenteric arteries from Kir6.1wt/VM mice
 compared with WT controls S6B: Summary data showing the passive diameter
 of isolated mesenteric arteries from SURAV/AV mice compared with WT
 controls ### Supplemental Figure *7* Processed\_Data S7B: Representative
 ΔF/F0 vs. time plots data of Ca2+ events from multiple sites of Ca2+
 events S7C: Summary data showing the effects of pinacidil and
 glibenclamide on amplitude (ΔF/F0), frequency (Hz), and the number of
 active sites upon Tg treatment ### Supplemental Figure *8* Processed\_Data
 S8B: Representative ΔF/F0 vs. time plots of Ca2+ events from multiple Ca2+
 event sites. S8C: Summary data showing the effects of pinacidil on the
 amplitude (ΔF/F0) of Ca2+ signals in Ca2+-free solution or in a solution
 containing 2 mM extracellular Ca2+ S8E: Representative ΔF/F0 vs. time
 plots of Ca2+ events from multiple sites of Ca2+ events S8F: Summary data
 showing the effects of pinacidil at 0 mM or 2 mM extracellular Ca2+ in the
 presence of CCh ### Supplemental Figure *10* Processed\_Data S10B:
 Representative ΔF/F0 vs. time plots of the change in fluorescence
 intensity of CM-H2DCFDA under each condition S10C: Summary data showing
 the changes in CM-H2DCFDA fluorescence intensity. ### Supplemental Figure
 *11* Processed\_Data S11A: Representative traces data showing
 endothelium-dependent vasodilation evoked by carbachol (CCh) in mesenteric
 arteries from Kir6.1wt/VM mice before and after treatment with the
 extracellular ROS scavengers superoxide dismutase S11B: Summary data
 showing endothelium-dependent vasodilation evoked by carbachol (CCh) in
 mesenteric arteries from Kir6.1wt/VM mice before and after treatment with
 the extracellular ROS scavengers superoxide dismutase ### Supplemental
 Figure *12* Processed\_Data S12A: Representative traces of the lumen
 diameter of isolated mesenteric arteries from Kir6.1wt/VM mice showing the
 contraction response to increases in intraluminal pressure under active
 conditions and dilation under passive (Ca2+-free) conditions S12B: Summary
 data showing myogenic tone evoked in mesenteric arterioles from
 Kir6.1wt/VM mice before and after treatment with MitoTEMPO ##
 Sharing/Access information N/A ## Code/Software N/A