Aldo-keto reductases in the eye

Aldose reductase (AKR1B1) is an NADPH-dependent aldo-keto reductase best known as the rate-limiting enzyme of the polyol pathway. Accelerated glucose metabolism through this pathway has been implicated in diabetic cataract and retinopathy. Some human tissues contain AKR1B1 as well as AKR1B10, a closely related member of the aldo-keto reductase gene superfamily. This opens the possibility that AKR1B10 may also contribute to diabetic complications. The goal of the current study was to characterize the expression profiles of AKR1B1 and AKR1B10 in the human eye. Using quantitative reverse transcriptase-PCR and immunohistochemical staining, we observed expression of both AKR genes in cornea, iris, ciliary body, lens, and retina. Expression of AKR1B1 was the highest in lens and retina, whereas AKR1B10 was the highest in cornea. Lenses from transgenic mice designed for overexpression of AKR1B10 were not significantly different from nontransgenic controls, although a significant number developed a focal defect in the anterior lens epithelium following 6 months of experimentally induced diabetes. However, lenses from AKR1B10 mice remained largely transparent following longterm diabetes. These results indicate that AKR1B1 and AKR1B10 may have different functional properties in the lens and suggest that AKR1B10 does not contribute to the pathogenesis of diabetic cataract in humans

Zoledronic acid blocks overactive Kir6.1/SUR2-dependent KATP channels in skeletal muscle and osteoblasts in a murine model of Cantú syndrome

Cantú syndrome (CS) is caused by the gain of function mutations in th

Consequences of SUR2[A478V] mutation in skeletal muscle of murine model of Cantu syndrome

(1) Background: Cantu syndrome (CS) arises from gain-of-function (GOF) mutations in th

Consequences of SUR2[A478V] mutation in skeletal muscle of murine model of Cantu syndrome

(1) Background: Cantu syndrome (CS) arises from gain-of-function (GOF) mutations in th

Glibenclamide reverses cardiovascular abnormalities of Cantu syndrome driven by KATP channel overactivity

Cantu syndrome (CS) is a complex disorder caused by gain-of-function (GoF) mutations in ABCC9 and KCNJ8, which encode the SUR2 and Kir6.1 subunits, respectively, of vascular smooth muscle (VSM) KATP channels. CS includes dilated vasculature, marked cardiac hypertrophy, and other cardiovascular abnormalities. There is currently no targeted therapy, and it is unknown whether cardiovascular features can be reversed once manifest. Using combined transgenic and pharmacological approaches in a knockin mouse model of CS, we have shown that reversal of vascular and cardiac phenotypes can be achieved by genetic downregulation of KATP channel activity specifically in VSM, and by chronic administration of the clinically used KATP channel inhibitor, glibenclamide. These findings demonstrate that VSM KATP channel GoF underlies CS cardiac enlargement and that CS-associated abnormalities are reversible, and provide evidence of in vivo efficacy of glibenclamide as a therapeutic agent in CS

Cardiovascular consequences of KATP overactivity in Cantu syndrome

Cantu syndrome (CS) is characterized by multiple vascular and cardiac abnormalities including vascular dilation and tortuosity, systemic hypotension, and cardiomegaly. The disorder is caused by gain-of-function (GOF) mutations in genes encoding pore-forming (Kir6.1, KCNJ8) and accessory (SUR2, ABCC9) ATP-sensitive potassium (KATP) channel subunits. However, there is little understanding of the link between molecular dysfunction and the complex pathophysiology observed, and there is no known treatment, in large part due to the lack of appropriate preclinical disease models in which to test therapies. Notably, expression of Kir6.1 and SUR2 does not fully overlap, and the relative contribution of KATP GOF in various cardiovascular tissues remains to be elucidated. To investigate pathophysiologic mechanisms in CS we have used CRISPR/Cas9 engineering to introduce CS-associated SUR2[A478V] and Kir6.1[V65M] mutations to the equivalent endogenous loci in mice. Mirroring human CS, both of these animals exhibit low systemic blood pressure and dilated, compliant blood vessels, as well dramatic cardiac enlargement, the effects being more severe in V65M animals than in A478V animals. In both animals, whole-cell patch-clamp recordings reveal enhanced basal KATP conductance in vascular smooth muscle, explaining vasodilation and lower blood pressure, and demonstrating a cardinal role for smooth muscle KATP dysfunction in CS etiology. Echocardiography confirms in situ cardiac enlargement and increased cardiac output in both animals. Patch-clamp recordings reveal reduced ATP sensitivity of ventricular myocyte KATP channels in A478V, but normal ATP sensitivity in V65M, suggesting that cardiac remodeling occurs secondary to KATP overactivity outside of the heart. These SUR2[A478V] and Kir6.1[V65M] animals thus reiterate the key cardiovascular features seen in human CS. They establish the molecular basis of the pathophysiological consequences of reduced smooth muscle excitability resulting from SUR2/Kir6.1-dependent KATP GOF, and provide a validated animal model in which to examine potential therapeutic approaches to treating CS

Hypotension due to Kir6.1 gain‐of‐function in vascular smooth muscle

BACKGROUND: K(ATP) channels, assembled from pore‐forming (Kir6.1 or Kir6.2) and regulatory (SUR1 or SUR2) subunits, link metabolism to excitability. Loss of Kir6.2 results in hypoglycemia and hyperinsulinemia, whereas loss of Kir6.1 causes Prinzmetal angina–like symptoms in mice. Conversely, overactivity of Kir6.2 induces neonatal diabetes in mice and humans, but consequences of Kir6.1 overactivity are unknown. METHODS AND RESULTS: We generated transgenic mice expressing wild‐type (WT), ATP‐insensitive Kir6.1 [Gly343Asp] (GD), and ATP‐insensitive Kir6.1 [Gly343Asp,Gln53Arg] (GD‐QR) subunits, under Cre‐recombinase control. Expression was induced in smooth muscle cells by crossing with smooth muscle myosin heavy chain promoter–driven tamoxifen‐inducible Cre‐recombinase (SMMHC‐Cre‐ER) mice. Three weeks after tamoxifen induction, we assessed blood pressure in anesthetized and conscious animals, as well as contractility of mesenteric artery smooth muscle and K(ATP) currents in isolated mesenteric artery myocytes. Both systolic and diastolic blood pressures were significantly reduced in GD and GD‐QR mice but normal in mice expressing the WT transgene and elevated in Kir6.1 knockout mice as well as in mice expressing dominant‐negative Kir6.1 [AAA] in smooth muscle. Contractile response of isolated GD‐QR mesenteric arteries was blunted relative to WT controls, but nitroprusside relaxation was unaffected. Basal K(ATP) conductance and pinacidil‐activated conductance were elevated in GD but not in WT myocytes. CONCLUSIONS: K(ATP) overactivity in vascular muscle can lead directly to reduced vascular contractility and lower blood pressure. We predict that gain of vascular K(ATP) function in humans would lead to a chronic vasodilatory phenotype, as indeed has recently been demonstrated in Cantu syndrome

Complex consequences of Cantu syndrome SUR2 variant R1154Q in genetically modified mice

Cantu syndrome (CS) is caused by gain-of-function (GOF) mutations in pore-forming (Kir6.1, KCNJ8) and accessory (SUR2, ABCC9) ATP-sensitive potassium (KATP) channel subunits, the most common mutations being SUR2[R1154Q] and SUR2[R1154W], carried by approximately 30% of patients. We used CRISPR/Cas9 genome engineering to introduce the equivalent of the human SUR2[R1154Q] mutation into the mouse ABCC9 gene. Along with minimal CS disease features, R1154Q cardiomyocytes and vascular smooth muscle showed much lower KATP current density and pinacidil activation than WT cells. Almost complete loss of SUR2-dependent protein and KATP in homozygous R1154Q ventricles revealed underlying diazoxide-sensitive SUR1-dependent KATP channel activity. Surprisingly, sequencing of SUR2 cDNA revealed 2 distinct transcripts, one encoding full-length SUR2 protein; and the other with an in-frame deletion of 93 bases (corresponding to 31 amino acids encoded by exon 28) that was present in approximately 40% and approximately 90% of transcripts from hetero- and homozygous R1154Q tissues, respectively. Recombinant expression of SUR2A protein lacking exon 28 resulted in nonfunctional channels. CS tissue from SUR2[R1154Q] mice and human induced pluripotent stem cell-derived (hiPSC-derived) cardiomyocytes showed only full-length SUR2 transcripts, although further studies will be required in order to fully test whether SUR2[R1154Q] or other CS mutations might result in aberrant splicing and variable expressivity of disease features in human CS

Resultant Effects of Head Impacts on Vestibular Function Among NCAA Division 1 Male Lacrosse Players

Context: Few studies have evaluated the cumulative effects of the number and magnitude of head impacts on vestibular function among collegiate men\u27s lacrosse players. Objective: To determine theinfluence of the total number of head impacts (THI), peak linear acceleration, and Head Injury Criteria (HIC) score on vestibular ocular reflex (VOR) function after one competitive collegiate lacrosse season. Design: Prospective cohort. Setting: Combined field-based and laboratory study. Patients or Other Participants: 41 male NCAA Division I lacrosse players (age = 20.6 ± 1.4 yrs, height = 181.9 ± 6.6 cm). Interventions: Head impact measures were collected with the GForce Tracker(TM) system, and included total number of head impacts greater than 20 g (THI), magnitude of head impacts as assessed by peak linear acceleration, and Head Injury Criteria (HIC) score. Paired samples t-tests were used to compare thepretest and posttest VOR results. A multiple regression approach was used to explore the contributions of accelerometer variables, participant-reported number of previous concussions, and symptom severity during VOR testing to the post-season VOR measurements (α = 0.05). Main Outcome Measures: VOR parameters, as assessed by the inVision(TM) system included: maximum gaze velocity of the gaze stabilization test (GST) yaw, visual acuity difference of the dynamic visual acuity test (DVA) for pitch and yaw head movements, and absolute change in directional bias for each test. Results: A total of 8,648 impacts were recorded during the 2015 NCAA lacrosse season among the 32 athletes who participated in the post-season VOR assessments. Multiple regression analyses indicated that the total number (mean, 270.3 + 158.2), peak linear acceleration (mean, 50.9 + 158.2 g), HIC score (mean, 33.9 + 38.3), and symptom severity (mean, 2.35 + 2.97) were not significantly related to the changes observed in vestibular function (R = 0.201 to 0.535, P \u3e 0.05). However, participant-reported number of previous concussions was significantly related to the absolute change in directional bias during the GST (P = .036). Theabsolute change in directional bias of participants\u27 vestibular system was 6.64 + 19.57 for GST, 2.25 + 10.73 for DVA pitch, and 2.90 + 8.03 for DVA yaw (P \u3e 0.05). Conclusions: Participant reported concussion history had the greatest influence on changes in VOR directional bias measures and vestibular asymmetry in collegiate male lacrosse players over the course of one competitive season. These findings support the inclusion of vestibular assessment and treatment in concussion management protocols to improve patient care and health related quality of life among lacrosse participants with previous medical histories of concussion. Supported by a Southwest Athletic Trainers\u27 Association Graduate Student Research Grant