Characterization of the zebrafish as a model of ATP-sensitive potassium channel hyperinsulinism.

INTRODUCTION: Congenital hyperinsulinism (HI) is the leading cause of persistent hypoglycemia in infants. Current models to study the most common and severe form of HI resulting from inactivating mutations in the ATP-sensitive potassium channel (KATP) are limited to primary islets from patients and the Sur1 -/- mouse model. Zebrafish exhibit potential as a novel KATPHI model since they express canonical insulin secretion pathway genes and those with identified causative HI mutations. Moreover, zebrafish larvae transparency provides a unique opportunity for in vivo visualization of pancreatic islets. RESEARCH DESIGN AND METHODS: We evaluated zebrafish as a model for KATPHI using a genetically encoded Ca2+ sensor (ins:gCaMP6s) expressed under control of the insulin promoter in beta cells of an abcc8 -/- zebrafish line. RESULTS: We observed significantly higher islet cytosolic Ca2+ in vivo in abcc8 -/- compared with abcc8 +/+ zebrafish larvae. Additionally, abcc8 -/- larval zebrafish had significantly lower whole body glucose and higher whole body insulin levels compared with abcc8 +/+ controls. However, adult abcc8 -/- zebrafish do not show differences in plasma glucose, plasma insulin, or glucose tolerance when compared with abcc8 +/+ zebrafish. CONCLUSIONS: Our results identify that zebrafish larvae, but not adult fish, are a demonstrable novel model for advancement of HI research.

Cloning, functional expression, and pharmacological characterization of inwardly rectifying potassium channels (Kir) from Apis mellifera.

Potassium channels belong to the super family of ion channels and play a fundamental role in cell excitability. Kir channels are potassium channels with an inwardly rectifying property. They play a role in setting the resting membrane potential of many excitable cells including neurons. Although putative Kir channel family genes can be found in the Apis mellifera genome, their functional expression, biophysical properties, and sensitivity to small molecules with insecticidal activity remain to be investigated. We cloned six Kir channel isoforms from Apis mellifera that derive from two Kir genes, AmKir1 and AmKir2, which are present in the Apis mellifera genome. We studied the tissue distribution, the electrophysiological and pharmacological characteristics of three isoforms that expressed functional currents (AmKir1.1, AmKir2.2, and AmKir2.3). AmKir1.1, AmKir2.2, and AmKir2.3 isoforms exhibited distinct characteristics when expressed in Xenopus oocytes. AmKir1.1 exhibited the largest potassium currents and was impermeable to cesium whereas AmKir2.2 and AmKir2.3 exhibited smaller currents but allowed cesium to permeate. AmKir1 exhibited faster opening kinetics than AmKir2. Pharmacological experiments revealed that both AmKir1.1 and AmKir2.2 are blocked by the divalent ion barium, with IC50 values of 10-5 and 10-6 M, respectively. The concentrations of VU041, a small molecule with insecticidal properties required to achieve a 50% current blockade for all three channels were higher than those needed to block Kir channels in other arthropods, such as the aphid Aphis gossypii and the mosquito Aedes aegypti. From this, we conclude that Apis mellifera AmKir channels exhibit lower sensitivity to VU041.

The Identification of Goat KCNJ15 Gene Copy Number Variation and Its Association with Growth Traits.

(1) Background: Copy number variation (CNV) is a critical component of genome structural variation and has garnered significant attention. High-throughput screening of the KCNJ15 gene has revealed a correlation between the CNV region and the growth traits of goats. We aimed to identify the CNV of the KCNJ15 gene in five goat breeds and analyze its association with growth characteristics. (2) Methods: We utilized 706 goats from five breeds: Guizhou black goat (GZB), Guizhou white goat (GZW), Bohuai goat (BH), Huai goat (HH), and Taihang goat (TH). To evaluate the number of copies of the KCNJ15 gene using qPCR, we analyzed the correlation between the CNV and growth characteristics and then used a universal linear model. The findings revealed variations in the distribution of different copy number types among the different goat breeds. (3) Results: Association analysis revealed a positive influence of the CNV in the KCNJ15 gene on goat growth. In GZB, individuals with duplication types exhibited superior performance in terms of cannon bone circumference (p < 0.05). In HH, individuals with duplication types exhibited superior performance in terms of body slanting length (p < 0.05). Conversely, normal TH demonstrated better body height and body weight (p < 0.05), while in GZW, when CN = 3, it performed better than other types in terms of body weight and chest circumference (p < 0.05). However, in BH, it had no significant effect on growth traits. (4) Conclusions: We confirmed that the CNV in the KCNJ15 gene significantly influences the growth characteristics of four distinct goat breeds. The correlation between KCNJ15 gene CNVs and goat growth traits offers valuable insights to breeders, enabling them to employ precise and efficient breeding methods that enhance livestock welfare, productivity, and overall economic benefits in the industry.

A loss-of-function mutation in KCNJ11 causing sulfonylurea-sensitive diabetes in early adult life.

AIMS/HYPOTHESIS: The ATP-sensitive potassium (KATP) channel couples beta cell electrical activity to glucose-stimulated insulin secretion. Loss-of-function mutations in either the pore-forming (inwardly rectifying potassium channel 6.2 [Kir6.2], encoded by KCNJ11) or regulatory (sulfonylurea receptor 1, encoded by ABCC8) subunits result in congenital hyperinsulinism, whereas gain-of-function mutations cause neonatal diabetes. Here, we report a novel loss-of-function mutation (Ser118Leu) in the pore helix of Kir6.2 paradoxically associated with sulfonylurea-sensitive diabetes that presents in early adult life. METHODS: A 31-year-old woman was diagnosed with mild hyperglycaemia during an employee screen. After three pregnancies, during which she was diagnosed with gestational diabetes, the patient continued to show elevated blood glucose and was treated with glibenclamide (known as glyburide in the USA and Canada) and metformin. Genetic testing identified a heterozygous mutation (S118L) in the KCNJ11 gene. Neither parent was known to have diabetes. We investigated the functional properties and membrane trafficking of mutant and wild-type KATP channels in Xenopus oocytes and in HEK-293T cells, using patch-clamp, two-electrode voltage-clamp and surface expression assays. RESULTS: Functional analysis showed no changes in the ATP sensitivity or metabolic regulation of the mutant channel. However, the Kir6.2-S118L mutation impaired surface expression of the KATP channel by 40%, categorising this as a loss-of-function mutation. CONCLUSIONS/INTERPRETATION: Our data support the increasing evidence that individuals with mild loss-of-function KATP channel mutations may develop insulin deficiency in early adulthood and even frank diabetes in middle age. In this case, the patient may have had hyperinsulinism that escaped detection in early life. Our results support the importance of functional analysis of KATP channel mutations in cases of atypical diabetes.

An analysis of the relationship between ABCC8 and KCNJ11 gene polymorphisms and diabetic retinopathy in Turkish population.

BACKGROUND: Diabetic retinopathy (DR) occurs due to high blood glucose damage to the retina and leads to blindness if left untreated. KATP and related genes (KCNJ11 and ABCC8) play an important role in insulin secretion by glucose-stimulated pancreatic beta cells and the regulation of insulin secretion. KCNJ11 E23K (rs5219), ABCC8-3 C/T (rs1799854), Thr759Thr (rs1801261) and Arg1273Arg (rs1799859) are among the possible related single nucleotide polymorphisms (SNPs). The aim of this study is to find out how DR and these SNPs are associated with one another in the Turkish population. MATERIALS AND METHODS: This study included 176 patients with type 2 diabetes mellitus without retinopathy (T2DM-rp), 177 DR patients, and 204 controls. Genomic DNA was extracted from whole blood, and genotypes were determined by the PCR-RFLP method. RESULTS: In the present study, a significant difference was not found between all the groups in terms of Arg1273Arg polymorphism located in the ABCC8 gene. The T allele and the TT genotype in the -3 C/T polymorphism in this gene may have a protective effect in the development of DR (p = 0.036 for the TT genotype; p = 0.034 for T allele) and PDR (p = 0.042 and 0.025 for the TT genotype). The AA genotype showed a significant increase in the DR group compared to T2DM-rp in the KCNJ11 E23K polymorphism (p = 0.046). CONCLUSIONS: Consequently, the T allele and TT genotype in the -3 C/T polymorphism of the ABCC8 gene may have a protective marker on the development of DR and PDR, while the AA genotype in the E23K polymorphism of the KCNJ11 gene may be effective in the development of DR in the Turkish population.

Functional characterization of inactivating ABCC8 variants causing congenital hyperinsulinism.

Congenital hyperinsulinism (CHI; OMIM: 256450) is characterized by persistent insulin secretion despite severe hypoglycemia. The most common causes are variants in the ATP-binding cassette subfamily C member 8(ABCC8) and potassium inwardly-rectifying channel subfamily J member 11(KCNJ11) genes. These encode ATP-sensitive potassium (KATP) channel subunit sulfonylurea receptor 1 (SUR1) and inwardly rectifying potassium channel (Kir6.2) proteins. A 7-day-old male infant presented with frequent hypoglycemic episodes and was clinically diagnosed with CHI, underwent trio-whole-exome sequencing, revealing compound heterozygous ABCC8 variants (c.307C>T, p.His103Tyr; and c.3313_3315del, p.Ile1105del) were identified. In human embryonic kidney 293 (HEK293) and rat insulinoma cells (INS-1) transfected with wild-type and variant plasmids, KATP channels formed by p.His103Tyr were delivered to the plasma membrane, whereas p.Ile1105del or double variants (p.His103Tyr coupled with p.Ile1105del) failed to be transported to the plasma membrane. Compared to wild-type channels, the channels formed by the variants (p.His103Tyr; p.Ile1105del) had elevated basal [Ca2+]i, but did not respond to stimulation by glucose. Our results provide evidence that the two ABCC8 variants may be related to CHI owing to defective trafficking and dysfunction of KATP channels.

Role of Angiotensin II Type 1a Receptor (AT1aR) of Renal Tubules in Regulating Inwardly Rectifying Potassium Channels 4.2 (Kir4.2), Kir4.1, and Epithelial Na+ Channel (ENaC).

BACKGROUND: Kir4.2 and Kir4.1 play a role in regulating membrane transport in the proximal tubule (PT) and in the distal-convoluted-tubule (DCT), respectively. METHODS: We generated kidney-tubule-specific-AT1aR-knockout (Ks-AT1aR-KO) mice to examine whether renal AT1aR regulates Kir4.2 and Kir4.1. RESULTS: Ks-AT1aR-KO mice had a lower systolic blood pressure than Agtr1aflox/flox (control) mice. Ks-AT1aR-KO mice had a lower expression of NHE3 (Na+/H+-exchanger 3) and Kir4.2, a major Kir-channel in PT, than Agtr1aflox/flox mice. Whole-cell recording also demonstrated that the membrane potential in PT of Ks-AT1aR-KO mice was lesser negative than Agtr1aflox/flox mice. The expression of Kir4.1 and Kir5.1, Kir4.1/Kir5.1-mediated K+ currents of DCT and DCT membrane potential in Ks-AT1aR-KO mice, were similar to Agtr1aflox/flox mice. However, angiotensin II perfusion for 7 days hyperpolarized the membrane potential in PT and DCT of the control mice but not in Ks-AT1aR-KO mice, while angiotensin II perfusion did not change the expression of Kir4.1, Kir4.2, and Kir5.1. Deletion of AT1aR did not significantly affect the expression of αENaC (epithelial Na+ channel) and ßENaC but increased cleaved γENaC expression. Patch-clamp experiments demonstrated that deletion of AT1aR increased amiloride-sensitive Na+-currents in the cortical-collecting duct but not in late-DCT. However, tertiapin-Q sensitive renal outer medullary potassium channel currents were similar in both genotypes. CONCLUSIONS: AT1aR determines the baseline membrane potential of PT by controlling Kir4.2 expression/activity but AT1aR is not required for determining the baseline membrane potential of the DCT and Kir4.1/Kir5.1 activity/expression. However, AT1aR is required for angiotensin II-induced hyperpolarization of basolateral membrane of PT and DCT. Deletion of AT1aR had no effect on baseline renal outer medullary potassium channel activity but increased ENaC activity in the CCD.

Maternal Embryo Effect Arrest 31 (MEE31) is a moonlighting protein involved in GDP-D-mannose biosynthesis and KAT1 potassium channel regulation.

Due to anthropogenic global warming, droughts are expected to increase and water availability to decrease in the coming decades. For this reason, research is increasingly focused on developing plant varieties and crop cultivars with reduced water consumption. Transpiration occurs through stomatal pores, resulting in water loss. Potassium plays a significant role in stomatal regulation. KAT1 is an inward-rectifying potassium channel that contributes to stomatal opening. Using a yeast high-throughput screening of an Arabidopsis cDNA library, MEE31 was found to physically interact with KAT1. MEE31 was initially identified in a screen for mutants with delayed embryonic development. The gene encodes a conserved phosphomannose isomerase (PMI). We report here that MEE31 interacts with and increases KAT1 activity in yeast and this interaction was also confirmed in plants. In addition, MEE31 complements the function of the yeast homologue, whereas the truncated version recovered in the screening does not, thus uncoupling the enzymatic activity from KAT1 regulation. We show that MEE31 overexpression leads to increased stomatal opening in Arabidopsis transgenic lines. Our data suggest that MEE31 is a moonlighting protein involved in both GDP-D-mannose biosynthesis and KAT1 regulation.

Functional dissection of KATP channel structures reveals the importance of a conserved interface.

ATP-sensitive potassium channels (KATP) are inhibited by ATP but activated by Mg-ADP, coupling the intracellular ATP/ADP ratio to the potassium conductance of the plasma membrane. Although there has been progress in determining the structure of KATP, the functional significance of the domain-domain interface in the gating properties of KATP channels remains incompletely understood. In this study, we define the structure of KATP as two modules: KATPcore and SURABC. Based on this model, we identified two functionally important interfaces between these two modules, namely interface I and interface II. Further structure-guided mutagenesis experiments indicate that destabilizing interface II by deleting ECL3 on the SUR1 subunit impairs KNtp-independent Mg-ADP activation, demonstrating the essential role of intramolecular interactions between KATPcore and SURABC in Mg-ADP activation. Additionally, interface II is functionally conserved between SUR1 and SUR2, and the hydrophobic residue F351 on ECL3 of SUR1 is crucial for maintaining the stability of this interface.

Nedd4-2-dependent regulation of astrocytic Kir4.1 and Connexin43 controls neuronal network activity.

Nedd4-2 is an E3 ubiquitin ligase in which missense mutation is related to familial epilepsy, indicating its critical role in regulating neuronal network activity. However, Nedd4-2 substrates involved in neuronal network function have yet to be identified. Using mouse lines lacking Nedd4-1 and Nedd4-2, we identified astrocytic channel proteins inwardly rectifying K+ channel 4.1 (Kir4.1) and Connexin43 as Nedd4-2 substrates. We found that the expression of Kir4.1 and Connexin43 is increased upon conditional deletion of Nedd4-2 in astrocytes, leading to an elevation of astrocytic membrane ion permeability and gap junction activity, with a consequent reduction of γ-oscillatory neuronal network activity. Interestingly, our biochemical data demonstrate that missense mutations found in familial epileptic patients produce gain-of-function of the Nedd4-2 gene product. Our data reveal a process of coordinated astrocytic ion channel proteostasis that controls astrocyte function and astrocyte-dependent neuronal network activity and elucidate a potential mechanism by which aberrant Nedd4-2 function leads to epilepsy.

Genome mining yields putative disease-associated ROMK variants with distinct defects.

Bartter syndrome is a group of rare genetic disorders that compromise kidney function by impairing electrolyte reabsorption. Left untreated, the resulting hyponatremia, hypokalemia, and dehydration can be fatal, and there is currently no cure. Bartter syndrome type II specifically arises from mutations in KCNJ1, which encodes the renal outer medullary potassium channel, ROMK. Over 40 Bartter syndrome-associated mutations in KCNJ1 have been identified, yet their molecular defects are mostly uncharacterized. Nevertheless, a subset of disease-linked mutations compromise ROMK folding in the endoplasmic reticulum (ER), which in turn results in premature degradation via the ER associated degradation (ERAD) pathway. To identify uncharacterized human variants that might similarly lead to premature degradation and thus disease, we mined three genomic databases. First, phenotypic data in the UK Biobank were analyzed using a recently developed computational platform to identify individuals carrying KCNJ1 variants with clinical features consistent with Bartter syndrome type II. In parallel, we examined genomic data in both the NIH TOPMed and ClinVar databases with the aid of Rhapsody, a verified computational algorithm that predicts mutation pathogenicity and disease severity. Subsequent phenotypic studies using a yeast screen to assess ROMK function-and analyses of ROMK biogenesis in yeast and human cells-identified four previously uncharacterized mutations. Among these, one mutation uncovered from the two parallel approaches (G228E) destabilized ROMK and targeted it for ERAD, resulting in reduced cell surface expression. Another mutation (T300R) was ERAD-resistant, but defects in channel activity were apparent based on two-electrode voltage clamp measurements in X. laevis oocytes. Together, our results outline a new computational and experimental pipeline that can be applied to identify disease-associated alleles linked to a range of other potassium channels, and further our understanding of the ROMK structure-function relationship that may aid future therapeutic strategies to advance precision medicine.

Genotype-phenotype correlation in Taiwanese children with diazoxide-unresponsive congenital hyperinsulinism.

Objective: Congenital hyperinsulinism (CHI) is a group of clinically and genetically heterogeneous disorders characterized by dysregulated insulin secretion. The aim of the study was to elucidate genetic etiologies of Taiwanese children with the most severe diazoxide-unresponsive CHI and analyze their genotype-phenotype correlations. Methods: We combined Sanger with whole exome sequencing (WES) to analyze CHI-related genes. The allele frequency of the most common variant was estimated by single-nucleotide polymorphism haplotype analysis. The functional effects of the ATP-sensitive potassium (KATP) channel variants were assessed using patch clamp recording and Western blot. Results: Nine of 13 (69%) patients with ten different pathogenic variants (7 in ABCC8, 2 in KCNJ11 and 1 in GCK) were identified by the combined sequencing. The variant ABCC8 p.T1042QfsX75 identified in three probands was located in a specific haplotype. Functional study revealed the human SUR1 (hSUR1)-L366F KATP channels failed to respond to intracellular MgADP and diazoxide while hSUR1-R797Q and hSUR1-R1393C KATP channels were defective in trafficking. One patient had a de novo dominant mutation in the GCK gene (p.I211F), and WES revealed mosaicism of this variant from another patient. Conclusion: Pathogenic variants in KATP channels are the most common underlying cause of diazoxide-unresponsive CHI in the Taiwanese cohort. The p.T1042QfsX75 variant in the ABCC8 gene is highly suggestive of a founder effect. The I211F mutation in the GCK gene and three rare SUR1 variants associated with defective gating (p.L366F) or traffic (p.R797Q and p.R1393C) KATP channels are also associated with the diazoxide-unresponsive phenotype.

A beneficial role of GLP-1 receptor agonist therapy in ABCC8-MODY (MODY 12).

Maturity-onset diabetes of the young (MODY) is an inherited form of diabetes resulting from a mutation in a single gene. ABCC8-MODY is caused by mutations in the ABCC8 gene, which encodes sulfonylurea receptor 1 (SUR1), a regulatory component of the ATP-sensitive potassium (KATP) channel found in beta cells. In ABCC8-MODY, mutations in the ABCC8 gene interfere with insulin secretion in response to glucose. Recent evidence suggests that therapy with GLP-1 receptor agonists (GLP-1 RAs) may be beneficial in ABCC8-MODY. This report presents a successful treatment of a 49-year-old woman diagnosed with ABCC8-MODY using the GLP-1 RA semaglutide. The patient, who had been previously receiving insulin therapy, experienced significant improvements in glycemic control and weight loss after transitioning to semaglutide. GLP-1 RAs potentially enhance insulin secretion in ABCC8-MODY by activating multiple signaling pathways involved in insulin secretion. The report highlights the potential of GLP-1 RA therapy as an alternative to sulfonylureas and insulin for individuals with ABCC8-MODY. GLP-1 RAs have previously demonstrated benefits in other forms of MODY. Understanding the molecular mechanisms through which GLP-1 RAs promote insulin secretion, including their effects on KATP channels and activation of PKA and Epac signaling, offers valuable insights into their therapeutic effects.

Identification of a new Kir6 potassium channel and comparison of properties of Kir6 subtypes by structural modelling and molecular dynamics.

ATP-sensitive potassium ion channels (KATP) are transmembrane proteins that modulate insulin release and muscle contraction. KATP channels are composed of two types of subunit, Kir6 and SUR, which exist in two and three isoforms respectively with different tissue distribution. In this work, we identify a previously undescribed ancestral vertebrate gene encoding a Kir6-related protein that we have named Kir6.3, which may not have a SUR binding partner, unlike the other two Kir6 proteins. Whereas Kir6.3 was lost in amniotes including mammals, it is still present in several early-diverging vertebrate lineages such as frogs, coelacanth, and rayfinned fishes. Molecular dynamics (MD) simulations using homology models of Kir6.1, Kir6.2, and Kir6.3 from the coelacanth Latimeria chalumnae showed that the three proteins exhibit subtle differences in their dynamics. Steered MD simulations of Kir6-SUR pairs suggest that Kir6.3 has a lower binding affinity for the SUR proteins than either Kir6.1 or Kir6.2. As we found no additional SUR gene in the genomes of the species that have Kir6.3, it most likely forms a lone tetramer. These findings invite studies of the tissue distribution of Kir6.3 in relation to the other Kir6 as well as SUR proteins to determine the functional roles of Kir6.3.

Nutrigenomics of inward rectifier potassium channels.

Inwardly rectifying potassium (Kir) channels play a key role in maintaining the resting membrane potential and supporting potassium homeostasis. There are many variants of Kir channels, which are usually tetramers in which the main subunit has two trans-membrane helices attached to two N- and C-terminal cytoplasmic tails with a pore-forming loop in between that contains the selectivity filter. These channels have domains that are strongly modulated by molecules present in nutrients found in different diets, such as phosphoinositols, polyamines and Mg2+. These molecules can impact these channels directly or indirectly, either allosterically by modulation of enzymes or via the regulation of channel expression. A particular type of these channels is coupled to cell metabolism and inhibited by ATP (KATP channels, essential for insulin release and for the pathogenesis of metabolic diseases like diabetes mellitus). Genomic changes in Kir channels have a significant impact on metabolism, such as conditioning the nutrients and electrolytes that an individual can take. Thus, the nutrigenomics of ion channels is an important emerging field in which we are attempting to understand how nutrients and diets can affect the activity and expression of ion channels and how genomic changes in such channels may be the basis for pathological conditions that limit nutrition and electrolyte intake. In this contribution we briefly review Kir channels, discuss their nutrigenomics, characterize how different components in the diet affect their function and expression, and suggest how their genomic changes lead to pathological phenotypes that affect diet and electrolyte intake.

Kir7.1 knockdown and inhibition alter renal electrolyte handling but not the development of hypertension in Dahl salt-sensitive rats.

High K+ supplementation is correlated with a lower risk of the composite of death, major cardiovascular events, and ameliorated blood pressure, but the exact mechanisms have not been established. Inwardly rectifying K+ (Kir) channels expressed in the basolateral membrane of the distal nephron play an essential role in maintaining electrolyte homeostasis. Mutations in this channel family have been shown to result in strong disturbances in electrolyte homeostasis, among other symptoms. Kir7.1 is a member of the ATP-regulated subfamily of Kir channels. However, its role in renal ion transport and its effect on blood pressure have yet to be established. Our results indicate the localization of Kir7.1 to the basolateral membrane of aldosterone-sensitive distal nephron cells. To examine the physiological implications of Kir7.1, we generated a knockout of Kir7.1 (Kcnj13) in Dahl salt-sensitive (SS) rats and deployed chronic infusion of a specific Kir7.1 inhibitor, ML418, in the wild-type Dahl SS strain. Knockout of Kcnj13 (Kcnj13-/-) resulted in embryonic lethality. Heterozygous Kcnj13+/- rats revealed an increase in K+ excretion on a normal-salt diet but did not exhibit a difference in blood pressure development or plasma electrolytes after 3 wk of a high-salt diet. Wild-type Dahl SS rats exhibited increased renal Kir7.1 expression when dietary K+ was increased. K+ supplementation also demonstrated that Kcnj13+/- rats excreted more K+ on normal salt. The development of hypertension was not different when rats were challenged with high salt for 3 wk, although Kcnj13+/- rats excrete less Na+. Interestingly, chronic infusion of ML418 significantly increased Na+ and Cl- excretion after 14 days of high salt but did not alter salt-induced hypertension development. Here, we found that reduction of Kir7.1 function, either through genetic ablation or pharmacological inhibition, can influence renal electrolyte excretion but not to a sufficient degree to impact the development of SS hypertension.NEW & NOTEWORTHY To investigate the role of the Kir7.1 channel in salt-sensitive hypertension, its function was examined using complementary genetic and pharmacological approaches. The results revealed that although reducing Kir7.1 expression had some impact on maintaining K+ and Na+ balance, it did not lead to a significant change in the development or magnitude of salt-induced hypertension. Hence, it is probable that Kir7.1 works in conjunction with other basolateral K+ channels to fine-tune membrane potential.

Skeletal muscle delimited myopathy and verapamil toxicity in SUR2 mutant mouse models of AIMS.

ABCC9-related intellectual disability and myopathy syndrome (AIMS) arises from loss-of-function (LoF) mutations in the ABCC9 gene, which encodes the SUR2 subunit of ATP-sensitive potassium (KATP ) channels. KATP channels are found throughout the cardiovascular system and skeletal muscle and couple cellular metabolism to excitability. AIMS individuals show fatigability, muscle spasms, and cardiac dysfunction. We found reduced exercise performance in mouse models of AIMS harboring premature stop codons in ABCC9. Given the roles of KATP channels in all muscles, we sought to determine how myopathy arises using tissue-selective suppression of KATP and found that LoF in skeletal muscle, specifically, underlies myopathy. In isolated muscle, SUR2 LoF results in abnormal generation of unstimulated forces, potentially explaining painful spasms in AIMS. We sought to determine whether excessive Ca2+ influx through CaV 1.1 channels was responsible for myopathology but found that the Ca2+ channel blocker verapamil unexpectedly resulted in premature death of AIMS mice and that rendering CaV 1.1 channels nonpermeable by mutation failed to reverse pathology; results which caution against the use of calcium channel blockers in AIMS.

Case report: Better late than never, but sooner is better: switch from CSII to sulfonylureas in two patients with neonatal diabetes due to KCNJ11 variants.

Neonatal diabetes mellitus (NDM) is a rare genetic disease characterized by severe hyperglycemia requiring insulin therapy with onset mostly within the first 6 months and rarely between 6-12 months of age. The disease can be classified into transient (TNDM) or permanent neonatal diabetes mellitus (PNDM), or it can be a component of a syndrome. The most frequent genetic causes are abnormalities of the 6q24 chromosomal region and mutations of the ABCC8 or KCNJ11 genes coding for the pancreatic beta cell's potassium channel (KATP). After the acute phase, patients with ABCC8 or KCNJ11 mutations treated with insulin therapy can switch to hypoglycemic sulfonylureas (SU). These drugs close the KATP channel binding the SUR1 subunit of the potassium channel and restoring insulin secretion after a meal. The timing of this switch can be different and could affect long-term complications. We describe the different management and clinical outcome over the time of two male patients with NDM due to KCNJ11 pathogenetic variants. In both cases, continuous subcutaneous insulin infusion pumps (CSII) were used to switch therapy from insulin to SU, but at different times after the onset. The two patients kept adequate metabolic control after the introduction of glibenclamide; during the treatment, insulin secretion was evaluated with c-peptide, fructosamine, and glycated hemoglobin (HbA1c), which were within the normal range. In neonates or infants with diabetes mellitus, genetic testing is an indispensable diagnostic tool and KCNJ11 variants should be considered. A trial of oral glibenclamide must be considered, switching from insulin, the first line of NDM treatment. This therapy can improve neurological and neuropsychological outcomes, in particular in the case of earlier treatment initiation. A new modified protocol with glibenclamide administered several times daily according to continuous glucose monitoring profile indications, was used. Patients treated with glibenclamide maintain good metabolic control and prevent hypoglycemia, neurological damage, and apoptosis of beta cells during long-term administration.

Targeting the inward rectifier potassium channel 5.1 in thyroid cancer: artificial intelligence-facilitated molecular docking for drug discovery.

BACKGROUND: Recurrent and metastatic thyroid cancer is more invasive and can transform to dedifferentiated thyroid cancer, thus leading to a severe decline in the 10-year survival. The thyroid-stimulating hormone receptor (TSHR) plays an important role in differentiation process. We aim to find a therapeutic target in redifferentiation strategies for thyroid cancer. METHODS: Our study integrated the differentially expressed genes acquired from the Gene Expression Omnibus database by comparing TSHR expression levels in the Cancer Genome Atlas database. We conducted functional enrichment analysis and verified the expression of these genes by RT-PCR in 68 pairs of thyroid tumor and paratumor tissues. Artificial intelligence-enabled virtual screening was combined with the VirtualFlow platform for deep docking. RESULTS: We identified five genes (KCNJ16, SLC26A4, TG, TPO, and SYT1) as potential cancer treatment targets. TSHR and KCNJ16 were downregulated in the thyroid tumor tissues, compared with paired normal tissues. In addition, KCNJ16 was lower in the vascular/capsular invasion group. Enrichment analyses revealed that KCNJ16 may play a significant role in cell growth and differentiation. The inward rectifier potassium channel 5.1 (Kir5.1, encoded by KCNJ16) emerged as an interesting target in thyroid cancer. Artificial intelligence-facilitated molecular docking identified Z2087256678_2, Z2211139111_1, Z2211139111_2, and PV-000592319198_1 (-7.3 kcal/mol) as the most potent commercially available molecular targeting Kir5.1. CONCLUSION: This study may provide greater insights into the differentiation features associated with TSHR expression in thyroid cancer, and Kir5.1 may be a potential therapeutic target in the redifferentiation strategies for recurrent and metastatic thyroid cancer.

Association of polymorphisms of calcium reabsorption genes SLC12A1, KCNJ1 and SLC8A1 with colorectal adenoma.

BACKGROUND: In recent years, morbidity and mortality from colorectal cancer have increased. Colorectal adenoma is the main precancerous lesion. Understanding the pathogenesis of colorectal adenoma will help to improve the early diagnosis rate of colorectal cancer. METHODS: In this case-control study, we focused on three single nucleotide polymorphisms (SNPs) in genes SLC8A1 (rs4952490), KCNJ1 (rs2855798), and SLC12A1 (rs1531916). We analyzed 207 colorectal adenoma patients (112 high-risk cases and 95 low-risk cases) and 212 control subjects by Sanger sequencing. A food frequency questionnaire (FFQ) was used to survey demographic characteristics and dietary nutrition. RESULTS: In the overall analysis, the results suggested that the AA+AG and AG genotype carriers of rs4952490 had a 73.1% and 78% lower risk of colorectal adenoma compared to GG genotype carriers, respectively. However rs2855798 and rs1531916 were not associated with the incidence of colorectal adenoma. Additionally, stratified analysis showed that rs4952490 AA+AG and AG genotypes had a protective effect against low-risk colorectal adenoma in patients aged ≤ 60 years old who were non-smokers. We also observed that when calcium intake was higher than 616 mg/d and patients carried at least one gene with variant alleles there was a protective effect against low-risk colorectal adenoma. CONCLUSIONS: Interactions between dietary calcium intake and calcium reabsorption genes may affect the occurrence and development of colorectal adenoma.