Influence of flaxseed-derived diglyceride-based high internal phase Pickering emulsions on the rheological and physicochemical properties of myofibrillar protein gels.

The study aimed to investigate the influence of flaxseed-derived diglyceride-based high internal phase Pickering emulsions (HIPPE) at different levels (0%, 10%, 20%, 30%, 40%, and 50%) on the rheological and physicochemical properties of myofibrillar protein (MPs) gels. The study indicated that with increasing HIPPE levels, there was a significant increase in whiteness while a decrease in water-holding capacity. The gels with 10% HIPPE levels had higher ionic bonds, while those with 40% and 50% HIPPE levels showed higher hydrogen bonds. By increasing HIPPE levels in the formation of MP gels, the T2 relaxation time was found to decrease. Additionally, in all MP gels, G' values were significantly higher than G" values over time. Adding lower contents of HIPPE levels resulted in a more compact microstructure. These findings indicate that flaxseed-derived diglyceride-based HIPPEs could be utilized as fat substitutes in meat products to enhance their nutritional quality.

Gene Expression, Morphology, and Electrophysiology During the Dynamic Development of Human Induced Pluripotent Stem Cell-Derived Atrial- and Ventricular-Like Cardiomyocytes.

Background and Objectives: Gene expression, morphology, and electrophysiological combination are essential for assessing the dynamic development of human induced pluripotent stem cell-derived atrial- and ventricular-like cardiomyocytes (iPS-AM and iPS-VM, respectively). Methods: For iPS-AM/VM differentiation, we performed the small molecule-based temporal modulation of the retinoic acid and bone morphogenetic protein signaling pathways. We investigated the gene expression and morphology using immunofluorescence, quantitative real-time polymerase chain reaction, flow cytometry, and transmission electron microscopy as well as registered electrophysiological functions using a whole-cell patch clamp on days 20, 30, and 60 post-differentiations. Results: Pan-cardiomyocyte marker, including troponin T2 (TNNT2) and alpha-actinin-2 (ACTN2), expressions increased both in iPS-AMs and iPS-VMs. Similarly, the mRNA expression of both iPS-AM-specific markers, ie, natriuretic peptide A (NPPA), myosin light chain 7 (MYL7), and K+ channel Kir3.4 (KCNJ5), and iPS-VM-specific markers, ie, gap junction α-1 (GJA1), myosin light chain 2 (MYL2), and alpha-1-subunit of a voltage-dependent L-type calcium channel (CACNA1C), increased from 0 to 20 days, and then decreased from 30 to 60 days. Concerning morphology, cardiac troponin-T (cTnT) arrangement was progressively organized and developed from a disorderly myofibrillar distribution to an organized sarcomere pattern both in iPS-AMs and iPS-VMs. Mitochondrial numbers gradually increased and those of lipid droplets decreased during dynamic development. Regarding physiological function, the resting and action potential amplitudes remained statistically indifferent in both cell types, and the action potential duration was prolonged during the development. Conclusion: IPS-AMs/VMs displayed dynamic development concerning their gene expression, morphology, and electrophysiological function. The discoveries of this study could provide novel insights into heart development and encourage further research.

Development and validation of a clinical prediction model for ischemic stroke recurrence after successful stent implantation in symptomatic intracranial atherosclerotic stenosis.

OBJECTIVE: This study aimed to analyze the risk factors for recurrent ischemic stroke in patients with symptomatic intracranial atherosclerotic stenosis (ICAS) who underwent successful stent placement and to establish a nomogram prediction model. METHODS: We utilized data from a prospective collection of 430 consecutive patients at Jining NO.1 People's Hospital from November 2021 to November 2022, conducting further analysis on the subset of 400 patients who met the inclusion criteria. They were further divided into training (n=321) and validation (n=79) groups. In the training group, we used univariate and multivariate COX regression to find independent risk factors for recurrent stroke and then created a nomogram. The assessment of the nomogram's discrimination and calibration was performed through the examination of various measures including the Consistency index (C-index), the area under the receiver operating characteristic (ROC) curves (AUC), and the calibration plots. Decision curve analysis (DCA) was used to evaluate the clinical utility of the nomogram by quantifying the net benefit to the patient under different threshold probabilities. RESULTS: The nomogram for predicting recurrent ischemic stroke in symptomatic ICAS patients after stent placement utilizes six variables: coronary heart disease (CHD), smoking, multiple ICAS, systolic blood pressure (SBP), in-stent restenosis (ISR), and fasting plasma glucose. The C-index (0.884 for the training cohort and 0.87 for the validation cohort) and the time-dependent AUC (>0.7) indicated satisfactory discriminative ability of the nomogram. Furthermore, DCA indicated a clinical net benefit from the nomogram. CONCLUSIONS: The predictive model constructed includes six predictive factors: CHD, smoking, multiple ICAS, SBP, ISR and fasting blood glucose. The model demonstrates good predictive ability and can be utilized to predict ischemic stroke recurrence in patients with symptomatic ICAS after successful stent placement.

Effect of flaxseed-derived diglyceride-based high internal phase Pickering emulsion on the quality characteristics of reformulated beef burgers.

The study aimed to fabricate healthier beef burgers using high internal phase Pickering emulsion (HIPPE) as animal fat substitute. In this context, HIPPE stabilized by modified soy protein isolates was produced with flaxseed-derived diglycerides (DAGs). Beef burgers were prepared by substituting beef backfat with HIPPE at varying levels (0%, 25%, 50%, 75%, and 100%). Reformulated burgers showed a significant decrease in WHC (from 89.75 to 77.38%), pH (from 5.73 to 5.58), L* values (from 53.5 to 44.5), and b* values (22.9 to 21.8), while a significant increase in a* values (from 24.4 to 6.7), cooking loss (from 20.25 to 30.62), and cooking shrinkage (from 11.27 to 13.05). Texture attributes, including hardness, chewiness, and gumminess, decreased up to 50% fat substitution and increased with increasing levels of fat substitution. Moreover, the rheological properties (G' and G'') and T2 relaxation time were increased with increasing fat replacement. The reformulation with HIPPE resulted in a decrease in SFA (from 3896 to 1712 mg/100 g), ω-6/ω-3 ratio (from 5.29 to 0.47), atherogenic index (from 0.57 to 0.13), and thrombogenic index (from 1.46 to 0.15) and increase in PUFA/SFA ratio (from 0.20 to 2.79). Notably, burgers with 50% fat substitution were more preferred regarding tenderness, while those with 100% fat substitution obtained higher scores for color and flavor than all other treatments. In conclusion, 50% fat replacement using flaxseed-derived diglyceride-based HIPPE improved beef burgers' textural profile and fatty acid composition without compromising the sensory characteristics.

Mitochondrial dysfunction is associated with hypertrophic cardiomyopathy in Pompe disease-specific induced pluripotent stem cell-derived cardiomyocytes.

Pompe disease (PD) is a rare autosomal recessive disorder that presents with progressive hypertrophic cardiomyopathy. However, the detailed mechanism remains clarified. Herein, PD patient-specific induced pluripotent stem cells were differentiated into cardiomyocytes (PD-iCMs) that exhibited cardiomyopathic features of PD, including decreased acid alpha-glucosidase activity, lysosomal glycogen accumulation and hypertrophy. The defective mitochondria were involved in the cardiac pathology as shown by the significantly decreased number of mitochondria and impaired respiratory function and ATP production in PD-iCMs, which was partially due to elevated levels of intracellular reactive oxygen species produced from depolarized mitochondria. Further analysis showed that impaired fusion and autophagy of mitochondria and declined expression of mitochondrial complexes underlies the mechanism of dysfunctional mitochondria. This was alleviated by supplementation with recombinant human acid alpha-glucosidase that improved the mitochondrial function and concomitantly mitigated the cardiac pathology. Therefore, this study suggests that defective mitochondria underlie the pathogenesis of cardiomyopathy in patients with PD.

Establishment of a novel human induced pluripotent stem cell line (SIPDi001-A) with compound heterozygous mutations in the UBR7 gene from a Li-Campeau syndrome patient.

Li-Campeau syndrome (LICAS) is a syndromic neurodevelopmental disorder characterized by autosomal recessive inheritance and global developmental delay. In this study, we reported the generation of a novel induced pluripotent stem cell (iPSC) line derived from peripheral blood mononuclear cells (PBMCs) obtained from a 7-year-old male patient with Li-Campeau syndrome. The patient carries compound heterozygous variants in the UBR7 gene (c.35_54dup, p.S19Rfs*42; c.863 T > C, p.L288P). The iPSC line showed typical cell morphology, robust expression of pluripotent and self-renewal markers, normal karyotype, and trilineage differentiation potential. This iPSC cell line could be valuable for investigating the underlying pathological mechanisms of neurodevelopmental disorders caused by UBR7 mutations.

Patient-specific induced pluripotent stem cell properties implicate Ca2+-homeostasis in clinical arrhythmia associated with combined heterozygous RYR2 and SCN10A variants.

We illustrate use of induced pluripotent stem cells (iPSCs) as platforms for investigating cardiomyocyte phenotypes in a human family pedigree exemplified by novel heterozygous RYR2-A1855D and SCN10A-Q1362H variants occurring alone and in combination. The proband, a four-month-old boy, presented with polymorphic ventricular tachycardia. Genetic tests revealed double novel heterozygous RYR2-A1855D and SCN10A-Q1362H variants inherited from his father (F) and mother (M), respectively. His father showed ventricular premature beats; his mother was asymptomatic. Molecular biological characterizations demonstrated greater TNNT2 messenger RNA (mRNA) expression in the iPSCs-induced cardiomyocytes (iPS-CMs) than in the iPSCs. Cardiac troponin Ts became progressively organized but cytoplasmic RYR2 and SCN10A aggregations occurred in the iPS-CMs. Proband-specific iPS-CMs showed decreased RYR2 and SCN10A mRNA expression. The RYR2-A1855D variant resulted in premature spontaneous sarcoplasmic reticular Ca2+ transients, Ca2+ oscillations and increased action potential durations. SCN10A-Q1362H did not confer any specific phenotype. However, the combined heterozygous RYR2-A1855D and SCN10A-Q1362H variants in the proband iPS-CMs resulted in accentuated Ca2+ homeostasis disorders, action potential prolongation and susceptibility to early afterdepolarizations at high stimulus frequencies. These findings attribute the clinical phenotype in the proband to effects of the heterozygous RYR2 variant exacerbated by heterozygous SCN10A modification. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Cardiac deficiency of P21-activated kinase 1 promotes atrial arrhythmogenesis in mice following adrenergic challenge.

P21-activated kinase 1 (Pak1) signalling plays a vital and overall protective role in the heart. However, the phenotypes of Pak1 deficiency in the cardiac atria have not been well explored. In this study, Pak1 cardiac-conditional knock-out (cKO) mice were studied under baseline and adrenergic challenge conditions. Pak1 cKO mice show atrial arrhythmias including atrial fibrillation (AF) in vivo, detected during anaesthetized electrocardiography without evidence of interstitial fibrosis upon Masson's trichrome staining. Optical mapping of left atrial preparations from Pak1 cKO mice revealed a higher incidence of Ca2+ and action potential alternans under isoprenaline challenge and differences in baseline action potential and calcium transient characteristics. Type-2 ryanodine receptor (RyR2) channels from Pak1 cKO hearts had a higher open probability than those from wild-type. Reverse transcription-quantitative polymerase chain reaction and Western blotting indicated that pCamkIIδ and RyR2 are highly phosphorylated at baseline in the atria of Pak1 cKO mice, while the expression of Slc8a2 and Slc8a3 as a Na+-Ca2+ exchanger, controlling the influx of Ca2+ from outside of the cell and efflux of Na+ from the cytoplasm, are augmented. Chromatin immunoprecipitation study showed that pCreb1 interacts with Slc8a2 and Slc8a3. Our study thus demonstrates that deficiency of Pak1 promotes atrial arrhythmogenesis under adrenergic stress, probably through post-translational and transcriptional modifications of key molecules that are critical to Ca2+ homeostasis. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Generation of cardiomyocytes from human-induced pluripotent stem cells resembling atrial cells with ability to respond to adrenoceptor agonists.

Atrial fibrillation (AF) is the most common chronic arrhythmia presenting a heavy disease burden. We report a new approach for generating cardiomyocytes (CMs) resembling atrial cells from human-induced pluripotent stem cells (hiPSCs) using a combination of Gremlin 2 and retinoic acid treatment. More than 40% of myocytes showed rod-shaped morphology, expression of CM proteins (including ryanodine receptor 2, α-actinin-2 and F-actin) and striated appearance, all of which were broadly similar to the characteristics of adult atrial myocytes (AMs). Isolated myocytes were electrically quiescent until stimulated to fire action potentials with an AM profile and an amplitude of approximately 100 mV, arising from a resting potential of approximately -70 mV. Single-cell RNA sequence analysis showed a high level of expression of several atrial-specific transcripts including NPPA, MYL7, HOXA3, SLN, KCNJ4, KCNJ5 and KCNA5. Amplitudes of calcium transients recorded from spontaneously beating cultures were increased by the stimulation of α-adrenoceptors (activated by phenylephrine and blocked by prazosin) or ß-adrenoceptors (activated by isoproterenol and blocked by CGP20712A). Our new approach provides human AMs with mature characteristics from hiPSCs which will facilitate drug discovery by enabling the study of human atrial cell signalling pathways and AF. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Small-conductance calcium-activated potassium channels in the heart: expression, regulation and pathological implications.

Ca2+-activated K+ channels are critical to cellular Ca2+ homeostasis and excitability; they couple intracellular Ca2+ and membrane voltage change. Of these, the small, 4-14 pS, conductance SK channels include three, KCNN1-3 encoded, SK1/KCa2.1, SK2/KCa2.2 and SK3/KCa2.3, channel subtypes with characteristic, EC50 ∼ 10 nM, 40 pM, 1 nM, apamin sensitivities. All SK channels, particularly SK2 channels, are expressed in atrial, ventricular and conducting system cardiomyocytes. Pharmacological and genetic modification results have suggested that SK channel block or knockout prolonged action potential durations (APDs) and effective refractory periods (ERPs) particularly in atrial, but also in ventricular, and sinoatrial, atrioventricular node and Purkinje myocytes, correspondingly affect arrhythmic tendency. Additionally, mitochondrial SK channels may decrease mitochondrial Ca2+ overload and reactive oxygen species generation. SK channels show low voltage but marked Ca2+ dependences (EC50 ∼ 300-500 nM) reflecting their α-subunit calmodulin (CaM) binding domains, through which they may be activated by voltage-gated or ryanodine-receptor Ca2+ channel activity. SK function also depends upon complex trafficking and expression processes and associations with other ion channels or subunits from different SK subtypes. Atrial and ventricular clinical arrhythmogenesis may follow both increased or decreased SK expression through decreased or increased APD correspondingly accelerating and stabilizing re-entrant rotors or increasing incidences of triggered activity. This article is part of the theme issue 'The heartbeat: its molecular basis and physiological mechanisms'.

Case report: Catecholamine cardiomyopathy in children with neuroblastoma.

Introduction: Many endocrine diseases, such as neuroblastoma (NB), can be linked with acquired cardiomyopathy and heart failure. Neuroblastoma's cardiovascular manifestations are typically hypertension, electrocardiogram (ECG) changes, and conduction disturbances. Case Presentation: A 5-year-old 8-month-old girl was admitted to the hospital with ventricular hypertrophy and hypertension (HT) and heart failure. She had no previous history of HT. On color doppler echocardiography, the left atrium and left ventricle were enlarged. The left ventricular ejection fraction (EF) was as low as 40%, and the ventricular septum and left ventricular free wall were thickened. The internal diameters of both coronary arteries were widened. Abdominal computed tomography scan (CT) demonstrated an 8.7 cm × 7.1 cm × 9.5 cm tumor behind the left peritoneum. In urine catecholamines analysis, free-norepinephrine (f-NE), free-dopamine (f-DA), free-normetanephrine (f-NMN), free-3-methoxytyramine (f-3MT), vanillylmandelic acid (VMA), and homovanillic acid (HVA) levels were all greater than the normal range for 24 h except free-metanephrine (f-MN) and free-epinephrine (f-E). Based on these findings, we diagnosed her as NB complicated by catecholamine cardiomyopathy manifested by hypertrophic cardiomyopathy (HCM). Oral metoprolol, spironolactone, captopril and amlodipine furosemide, and intravenously injected sodium nitroprusside and phentolamine were employed for treating HT. After the tumor resection, the blood pressure (BP) and urinary catecholamine levels were all restored. After a follow-up of 7 months, echocardiography indicated normalization of ventricular hypertrophy and function. Conclusion: This is a rare report showing catecholamine cardiomyopathy in NB children. Tumor resection leads to a return to normal of the catecholamine cardiomyopathy manifested as HCM.

A Clinical Prediction Model for Patients with Acute Large Vessel Occlusion Due to Underlying Intracranial Atherosclerotic Stenosis.

BACKGROUND: Acute large vessel occlusion due to underlying intracranial atherosclerotic stenosis (ICAS-LVO) increases the difficulty of revascularization, resulting in frequent re-occlusion. The establishment of its pathogenesis before endovascular treatment (EVT) is beneficial for patients. We aimed at developing and validating a clinical prediction model for ICAS-LVO patients before EVT. METHODS: Patients with acute large vessel occlusion at Jining No. 1 People's Hospital from January 2019 to September 2021 were retrospectively included as the training cohort. The 70 patients who met the inclusion and exclusion criteria were included in the validation cohort (October 2021 to May 2022). Demographics, onset form, medical history, digital subtraction angiography (DSA) imaging data, and laboratory test data were collected. Preprocedural parameters for the ICAS-LVO risk prediction model were established by stepwise logistic regression controlling for the confounding effects. Then, we constructed a nomogram model and evaluated its performance via the Hosmer-Lemeshow goodness-of-fit test, area under the ROC curve (AUC) analysis. RESULTS: The 231 acute LVO patients were included in the final analysis, 74 (32.3%) patients were ICAS-LVO. A preoperative diagnosis prediction model consisting of five predictors for ICAS-LVO, including fluctuating symptoms, NIHSS < 16, atrial fibrillation, tapered sign, and ASITN/SIR score ≥ 2. The model depicted an acceptable calibration (Hosmer-Lemeshow test, p = 0.451) and good discrimination (AUC, 0.941; 95% confidence interval, 0.910-0.971). The optimal cut-off value for the ICAS-LVO scale was 2 points, with 86.5% sensitivity, 91.1% specificity, and 90.5% accuracy. In the validation cohort, the discriminative ability was promising with an AUC value of 0.897, implying a good predictive performance. CONCLUSION: The established ICAS-LVO scale, which is composed of five predictors: fluctuating symptoms, NIHSS < 16, atrial fibrillation, tapered sign, and ASITN/SIR score ≥ 2, has a good predictive value for ICAS-LVO in Chinese populations.

Generation of one human induced pluripotent stem cell line (XACHi004-A) with heterozygous mutation of RYR2 gene from an atrial fibrillation patient.

As the most frequently diagnosed arrhythmia, atrial fibrillation (AF) has been recently reported to be closely related to ryanodine receptor (RyR2) dysfunction and calcium leak. Here, using non-integrating sendai viral method, we generated one iPSC line from peripheral blood mononuclear cells (PBMC) isolated a 10-year-old boy with simple atrial fibrillation which carries the heterozygous mutation in RYR2 gene (c.14638G > A, p.V4880I). The generated iPSC line was identified by the typical cell morphology, highly expressed pluripotent markers, normal karyotype, and in-vitro trilineage differentiation potential. It will provide a useful model for studying the pathophysiological consequences of RYR2 mutation on the AF pathogenesis.

Generation of two heterozygous GAA mutation-carrying human induced pluripotent stem cell lines (XACHi005-A, XACHi006-A) from parents of an infant with Pompe disease.

Pompe disease results from GAA mutations that leads to lysosomal glycogen accumulation and cardiac and skeletal muscle pathology. We have previously generated an infantile-onset Pompe disease patient-derived human-induced pluripotent stem cells (iPSCs) line carrying compound GAA mutations (R608X and E888X). Using his parents' peripheral blood mononuclear cells (PBMCs), we here generated two iPSCs lines which carry mutations of R608X E888X respectively. Both lines show typical cell morphology, high expressed pluripotent and self-renewal markers, normal karyotype, and trilineage differentiation potential. These two lines are valuable re-sources for studying the pathological mechanisms of GAA mutation-caused Pompe disease.

The Biphasic Effect of Retinoic Acid Signaling Pathway on the Biased Differentiation of Atrial-like and Sinoatrial Node-like Cells from hiPSC.

Background and Objectives: Although human-induced pluripotent stem cells (hiPSC) can be efficiently differentiated into cardiomyocytes (CMs), the heterogeneity of the hiPSC-CMs hampers their applications in research and regenerative medicine. Retinoic acid (RA)-mediated signaling pathway has been proved indispensable in cardiac development and differentiation of hiPSC toward atrial CMs. This study was aimed to test whether RA signaling pathway can be manipulated to direct the differentiation into sinoatrial node (SAN) CMs. Methods and Results: Using the well-characterized GiWi protocol that cardiomyocytes are generated from hiPSC via temporal modulation of Wnt signaling pathway by small molecules, RA signaling pathway was manipulated during the differentiation of hiPSC-CMs on day 5 post-differentiation, a crucial time point equivalent to the transition from cardiac mesoderm to cardiac progenitor cells in cardiac development. The resultant CMs were characterized at mRNA, protein and electrophysiology levels by a combination of qPCR, immunofluorescence, flow cytometry, and whole-cell patch clamp. The results showed that activation of the RA signaling pathway biased the differentiation of atrial CMs, whereas inhibition of the signaling pathway biased the differentiation of sinoatrial node-like cells (SANLCs). Conclusions: Our study not only provides a novel and simple strategy to enrich SANLCs but also improves our understanding of the importance of RA signaling in the differentiation of hiPSC-CMs.

The safety and efficacy of the Neuroform EZ stent for the treatment of symptomatic atherosclerotic stenosis in the middle cerebral artery.

OBJECTIVE: To evaluate the safety and efficacy of Neuroform EZ stent placement for patients with symptomatic atherosclerotic stenosis of the middle cerebral artery (MCA). METHOD: We retrospectively reviewed the clinical data of 70 patients (36 males and 34 females; mean age: 62.5 ± 1.25 years) with symptomatic atherosclerotic stenosis of the MCA who underwent Neuroform EZ stent insertion between January 2018 and June 2020. We reviewed the clinical data of each patient so that we could evaluate outcomes and angiographic findings at follow-up. RESULTS: The technical success rate for the 70 patients was 100%. The mean rate of stenosis improved from 82.6 ± 6.5% (pre-stenting) to 21.11 ± 2.6% (post-stenting). During the post-stenting follow-up period, the 1-year frequencies of stroke, transient ischemic attack, and death, were all 0%. 42 patients were reviewed in hospital by DSA or CTA. Of the patients undergoing DSA or CTA review, four patients were found to have in-stent restenosis. CONCLUSION: Our analysis indicates that the Neuroform EZ stent represents a valuable endovascular treatment option for patients with severe stenosis of the MCA. Additional studies are now needed to evaluate the long-term outcomes arising from the use of this technique.

Generation of induced pluripotent stem cells (iPSCs) from a Chinese infant (XACHi015-A) with type 2 Long QT syndrome carrying the heterozygous mutation c.1814C>T(p.P605L) in KCNH2.

Induced pluripotent stem cell lines (iPSCs) were generated from peripheral blood mononuclear cells (PBMCs) isolated from the peripheral blood of a ten years old boy with the type 2 Long QT syndrome carrying the heterozygous mutation c.1814C>T(p.P605L) in KCNH2. PBMCs were reprogrammed using non-integrative Sendai viral vectors containing reprogramming factors OCT4, SOX2, KLF4 and C-MYC. The iPSCs were shown to express pluripotent markers, have trilineage differentiation potential, carry c.1814C>T(p.P605L) mutation in KCNH2 and have a normal karyotype. Thuse the iPSC line we established will be useful for studying the pathogenesis of the type 2 long QT syndrome and drug testing.

Establishment of iPSC line from a Chinese infant (XACHi012-A) with Jervell and Lange-Nielsen syndrome carrying combined KCNQ1 frameshift c.431delC(p.I145Sfs*92) and nonsense c.1175G > A (p.W392X) variants and two iPSC lines from the parents (XACHi013-A, XACHi014-A).

Induced pluripotent stem cell lines (iPSCs) were generated from peripheral blood mononuclear cells (PBMCs) isolated from the peripheral blood of a two month-old boy and the parents. Jervell and Lange-Nielsen syndrome (JLNS) was diagnosed in the boy carrying combined KCNQ1 frameshift c.431delC (p.I145Sfs*92) and nonsense c.1175G > A(p.W392X) variants inherited from his mother and father respectively. PBMCs were reprogrammed using non-integrative Sendai viral vectors containing reprogramming factors OCT4, SOX2, KLF4 and C-MYC. IPSCs were shown to express pluripotent markers, have trilineage differentiation potential, carrying identified KCNQ1 variants with corresponding PBMC, and have a normal karyotype. Thus we established three iPSC lines as useful tools for studying the pathophysiological mechanism of JLNS and drug testing.

Downregulation of microRNA­221 facilitates H1N1 influenza A virus replication through suppression of type­IFN response by targeting the SOCS1/NF­κB pathway.

Accumulating data has indicated that host microRNAs (miRNAs/miRs) play essential roles in innate immune responses to viral infection; however, the roles and the underlying mechanisms of miRNAs in influenza A virus (IAV) replication remain unclear. The present study examined on the effects of miRNAs on hemagglutinin (H)1 neuraminidase (N)1 replication and antiviral innate immunity. Using a microarray assay, the expression profiles of miRNA molecules in IAV­infected A549 cells were analyzed. The results indicated that miR­221 was significantly downregulated in IAV­infected A549 cells. It was also observed that IAV infection decreased the expression levels of miR­221 in A549 cells in a dose­ and time­dependent manner. Functionally, upregulation of miR­221 repressed IAV replication, whereas knockdown of miR­221 had an opposite effect. Subsequently, it was demonstrated that miR­221 overexpression could enhance IAV­triggered IFN­α and IFN­ß production and IFN­stimulated gene expression levels, while miR­221­knockdown had the opposite effect. Target prediction and dual luciferase assays indicated that suppressor of cytokine signaling 1 (SOCS1) was a direct target of miR­221 in A549 cells. Furthermore, knockdown of SOCS1 efficiently abrogated the influences caused by miR­221 inhibition on IAV replication and the type­I IFN response. It was also found that the miR­221 positively regulated NF­κB activation in IAV­infected A549 cells. Taken together, these data suggested that miR­221­downregulation promotes IAV replication by suppressing type­I IFN response through targeting SOCS1/NF­κB pathway. These findings suggest that miR­221 may serve as a novel potential therapeutic target for IAV treatment.