👤 Taeho Jo

Alzheimer's disease prediction using genomic data remains challenging due to the high dimensionality of whole-genome sequencing data and the complex relationships between genetic variants. We developed DuAL-Net (Dual Approach Local-global Network), a hybrid framework that integrates local genomic window analysis with global annotation-based modeling to prioritize disease-associated single-nucleotide polymorphisms (SNPs). As a proof of concept, we applied DuAL-Net to 14,094 SNPs within the Show less

Alzheimer's disease (AD) is marked by amyloid-β (Aβ) accumulation, tau pathology, and neuroinflammation. The β-site APP cleaving enzyme 1 (BACE1) is a key driver of Aβ production, while the NLRP3 inflammasome mediates microglial inflammatory responses. Histone deacetylase 6 (HDAC6), a cytoplasmic deacetylase, is upregulated in AD, yet its role in disease mechanisms remains unclear. Here, we show that HDAC6 promotes BACE1 protein stability through direct deacetylation of its C-terminal lysine (K501), thereby increasing Aβ production. HDAC6 also facilitated NLRP3 inflammasome activation in microglia, increasing IL-1β production in a catalytic domain-dependent manner. HDAC6 deficiency in 5xFAD mice reduced BACE1 accumulation, Aβ deposition, ASC speck formation, and IL-1β levels, accompanied by improved cognitive performance. Transcriptomic profiling further revealed downregulation of disease-associated microglial and neurotoxic astrocyte signatures alongside enrichment of synaptic pathways. These findings establish HDAC6 as a dual regulator of Aβ production and neuroinflammation, highlighting it as a promising therapeutic target in AD. Show less

Diabetes is a major risk factor for diabetic encephalopathy (DE), which is closely associated with sporadic Alzheimer's disease. Folic acid (FA) receptor signaling can suppress generation of neuropathogenic amyloid-beta (Aβ) induced by high extracellular glucose, suggesting that enhanced activation of this pathway could be a therapeutic strategy against DE-associated dementia, but the precise molecular signaling mechanisms are unclear. We report that high glucose levels increased the expression of amyloid precursor protein (APP) and β-secretase (BACE1) in cultured neurons and concomitantly induced amyloidogenesis, while FA treatment suppressed high glucose-stimulated expression of APP and BACE1, Aβ release, and accumulation of mitochondrial reactive oxygen species. Expression of nuclear factor erythroid 2-related factor 2 (Nrf2) was minimal under high glucose conditions, but was significantly upregulated together with downstream antioxidant enzymes following FA co-treatment. High glucose stimulation also increased folate receptor 1 (FOLR1) mRNA expression, suggesting a compensatory protective response. While treatment with 5-methyltetrahydrofolate (5-MTHF), the activated form of folate, did not significantly alter high glucose-induced upregulation of APP and BACE1, knockdown of FOLR1 mRNA reduced high glucose-stimulated Nrf2 expression and further augmented APP and BACE1 expression under high glucose conditions. Treatment with the STAT3 inhibitor 5'15-DPP also abolished high glucose-stimulated Nrf2 expression and increased APP and BACE1 expression levels. These findings indicate that FA/FOLR1 activation suppresses high glucose-induced amyloidogenesis by mitigating mitochondrial oxidative stress via STAT3/Nrf2 pathway signaling. In conclusion, present study suggests that the FA/FOLR1/STAT3/Nrf2 pathway is an effective therapeutic target for DE. Show less

Aberrant deposition of β-amyloid (Aβ) and hyperphosphorylated tau, along with neuroinflammation, are key drivers of Alzheimer's disease (AD) pathology. Here, we identify ramalin, a natural antioxidant, as a promising therapeutic agent that alleviates AD pathology by modulating β-site APP cleaving enzyme 1 (BACE1), histone deacetylase 6 (HDAC6), and the mitogen-activated protein kinases (MAPK) pathway. Ramalin reduced BACE1 protein levels, independently of its transcription, translation, or enzymatic activity, an effect mediated by inhibition of HDAC6. Consistently, HDAC6 knockout similarly decreased BACE1 levels, highlighting HDAC6 as a key regulator of BACE1. Ramalin further suppressed neuroinflammatory responses by downregulating inducible nitric oxide synthase (iNOS) and the NLR family pyrin domain containing 3 (NLRP3) inflammasome. In AD mouse models, ramalin treatment significantly attenuated neuroinflammation, Aβ plaque burden, and tau hyperphosphorylation, while improving cognitive performance. Notably, ramalin reversed Aβ oligomer-induced synaptic transmission impairment and restored synaptic vesicle recycling in hippocampal neurons. Transcriptomic analysis identified modulation of the MAPK pathway, with reduced phosphorylation of c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) implicated in tau pathology. These findings establish ramalin as a disease-modifying intervention that provides neuroprotection through concurrent regulation of BACE1, HDAC6, and MAPK signaling pathway. Collectively, our findings highlight ramalin as a compelling disease-modifying candidate with the potential to drive a breakthrough approach targeting AD pathology. Show less

The role of FGF is the least understood of the morphogens driving mammalian gastrulation. Here, we have investigated FGF function in a 2D gastruloid model for human gastrulation. We observed a ring of FGF-dependent ERK activity that closely follows the emergence of primitive streak (PS)-like cells but expands further inward. This ERK activity pattern depends on localized activation of basolateral FGF receptor 1 (FGFR1) by endogenous FGF gradients and is required for PS-like differentiation, with loss of PS-like cells upon FGF receptor inhibition rescued by direct ERK activation. Single cell transcriptome analysis confirmed that, among the ligands, FGF2 is broadly expressed, FGF8 is transiently expressed during PS-like differentiation and FGF4/17 are specifically expressed in PS-like cells - similar to the human and monkey embryo but different from the mouse. FGF4 knockdown greatly reduced PS-like differentiation, while FGF17 knockdown primarily affected subsequent mesoderm differentiation. FGF8 expression was spatially and temporally displaced from PS markers and FGF4 expression, while knockdown expanded PS-like cells, suggesting FGF8 may limit PS-like differentiation. Thus, we have identified a previously unreported role for FGF-dependent ERK signaling in 2D gastruloids and possibly the human embryo, where FGF4 and FGF17 signal through basolateral FGFR1 to induce PS-like cells and derivatives, potentially restricted by FGF8. Show less

Dysregulated extracellular matrix (ECM) deposition and epithelial-mesenchymal transition (EMT) in the trabecular meshwork (TM) contribute to glaucoma-associated fibrotic remodeling, and lysophosphatidic acid (LPA) potently induces these profibrotic responses in human trabecular meshwork (HTM) cells. We investigated whether an ethanolic extract of Show less

Alzheimer's disease (AD) is characterized by progressive cognitive decline, amyloid plaque accumulation, synaptic dysfunction, and neuroinflammation. This study reports the therapeutic potential of (S)-4-amino-5,5-difluoro-N'-methyl-N'-phenylpentanehydrazide hydrochloride (RA-058HM), a novel compound, in ameliorating these pathological features of AD in the 5xFAD mouse model. RA-058HM was administered orally for 8 weeks, and its multi-target effects - including relief from neuroinflammation, normalization of synaptic transmission, reduction of amyloidogenesis (plaque and soluble oligomers, as well as BACE1 levels), and rescue of cognitive function-were evaluated. To our knowledge, RA-058HM is the first compound to demonstrate simultaneous modulation of these key pathways in the 5xFAD model, highlighting its potential as a comprehensive disease-modifying therapy for AD. Behavioural tests revealed marked improvements in spatial and recognition memory in RA-058HM-treated 5xFAD mice, suggesting a reversal of cognitive deficits. At the molecular level, RA-058HM treatment reduced amyloidogenesis, as evidenced by decreased levels of amyloid precursor protein (APP) and β-secretase (BACE1) in the hippocampus, accompanied by reduced plaque formation, as detected by Thioflavin-S staining. Furthermore, synaptic transmission was restored to near-normal levels in RA-058HM-treated neurons, indicating that RA-058HM effectively rescues synaptic deficits without altering synaptic protein levels of PSD95 and synaptophysin. In addition, treatment of RA-058HM downregulated hippocampal levels of the NLRP3 inflammasome, TNF-α, and GFAP, suggesting a decrease in neuroinflammatory signaling and a modulation of glial activity. Restoration of mitochondrial motility in hippocampal neurons further suggests that RA-058HM may improve cellular energy dynamics. Collectively, these findings indicate that RA-058HM has multifaceted effects on AD pathology, targeting amyloid accumulation, synaptic transmission, neuroinflammation, and mitochondrial function. This study highlights RA-058HM as a promising candidate for AD therapy and underscores the potential of multi-targeted approaches in addressing the complex mechanisms underlying AD progression. Show less

Alzheimer's disease (AD) is a complex neurodegenerative disorder characterized by cognitive decline, oxidative stress, neuroinflammation, amyloid-beta (Aβ) accumulation, and tau protein hyperphosphorylation. In this study, we synthesized novel Ramalin derivatives and evaluated their therapeutic potential against AD, focusing on antioxidant, anti-inflammatory, and neuroprotective activities. RA-2OMe, RA-4OMe, RA-2CF3, and RA-4OCF3 showed strong antioxidant effects, while RA-2OMe exhibited potent NO and NLRP3 inhibition (~20%). RA-NAP, RA-PYD, and RA-2Q showed moderate anti-inflammatory activity. BACE-1 inhibition was significant in RA-3CF3, RA-NAP, and RA-PYD, with IC Show less

Alzheimer's disease (AD) is a multifactorial neurodegenerative disorder characterized by cognitive decline, anxiety-like behavior, β-amyloid (Aβ) accumulation, and tau hyperphosphorylation. BACE1, the enzyme critical for Aβ production, has been a major therapeutic target; however, direct BACE1 inhibition has been associated with adverse side effects. This study investigates the therapeutic potential of RA-PR058, a novel ramalin derivative, as a multi-targeted modulator of AD-related pathologies. The effects of RA-PR058 were evaluated Show less

Alzheimer's disease (AD) is the most common type of dementia. Its incidence is rising rapidly as the global population ages, leading to a significant social and economic burden. AD involves complex pathologies, including amyloid plaque accumulation, synaptic dysfunction, and neuroinflammation. This study explores the therapeutic potential of N Show less

β-secretase (BACE1) is instrumental in amyloid-β (Aβ) production, with overexpression noted in Alzheimer's disease (AD) neuropathology. The interaction of Aβ with the receptor for advanced glycation endproducts (RAGE) facilitates cerebral uptake of Aβ and exacerbates its neurotoxicity and neuroinflammation, further augmenting BACE1 expression. Given the limitations of previous BACE1 inhibition efforts, the study explores reducing BACE1 expression to mitigate AD pathology. The research reveals that the anticancer agent 6-thioguanosine (6-TG) markedly diminishes BACE1 expression without eliciting cytotoxicity while enhancing microglial phagocytic activity, and ameliorate cognitive impairments with reducing Aβ accumulation in AD mice. Leveraging advanced deep learning-based tool for target identification, and corroborating with surface plasmon resonance assays, it is elucidated that 6-TG directly interacts with RAGE, modulating BACE1 expression through the JAK2-STAT1 pathway and elevating soluble RAGE (sRAGE) levels in the brain. The findings illuminate the therapeutic potential of 6-TG in ameliorating AD manifestations and advocate for small molecule strategies to increase brain sRAGE levels, offering a strategic alternative to the challenges posed by the complexity of AD. Show less

The role of FGF is the least understood of the morphogens driving mammalian gastrulation. Here we investigated the function of FGF in a stem cell model for human gastrulation known as a 2D gastruloid. We found a ring of FGF-dependent ERK activity that closely follows the emergence of primitive streak (PS)-like cells but expands further inward. We showed that this ERK activity pattern is required for PS-like differentiation and that loss of PS-like cells upon FGF receptor inhibition can be rescued by directly activating ERK. We further demonstrated that the ERK-ring depends on localized activation of basolaterally positioned FGF receptors (FGFR) by endogenous FGF gradients. We confirmed and extended previous studies in analyzing expression of FGF pathway components, showing FGFR1 is the main receptor, FGF2 is highly expressed across several cell types, and FGF4/17 are the main FGF ligands expressed in the PS-like cells, similar to the human and monkey embryo but different from the mouse. We found that knockdown of FGF4 greatly reduced PS-like differentiation while FGF17 knockdown primarily affected subsequent mesoderm differentiation. FGF8 expression was spatially displaced from PS-markers and FGF4 expression and peaked earlier, while knockdown led to an expansion in PS-like cells, suggesting FGF8 may counteract FGF4 to limit PS-like differentiation. Thus, we have identified a previously unknown role for FGF-dependent ERK signaling in 2D gastruloids and possibly the human embryo, driven by a mechanism where FGF4 and FGF17 signal through basally localized FGFR1 to induce PS-like cells and their derivatives, potentially restricted by FGF8. Show less

Pediatric hypertrophic cardiomyopathy (HCM) is a rare condition, particularly in neonates, and is characterized by rapid and extensive myocardial hypertrophy, often leading to severe clinical outcomes. HCM can arise from variants in sarcomeric genes, which are essential for myocardial contractions, as well as non-sarcomeric gene variants. Although genetic modifiers and oligogenic inheritance have been implicated in congenital heart disease and cardiomyopathy, their complexity in HCM has not been fully elucidated, especially in familial cases with variable phenotypes. Hence, this study aims to investigate the genetic architecture in a family with a history of cardiac disease and neonatal HCM, focusing on oligogenic inheritance of non-sarcomeric variants. Clinical data and blood samples were collected for genetic analysis. Whole genome sequencing (WGS) and bioinformatic analyses identified compound heterozygous variants in the MYO19 gene. Maternally inherited variants were analyzed because the proband's mother was also diagnosed with HCM. WGS was performed on the patient's maternal grandfather and aunt, who have cardiac disease, revealing candidate genetic variants that may contribute to the cardiac phenotype. Compound heterozygous MYO19 variants were identified in the neonatal patient. Missense c.203C > G (p.A68G) and frameshift c.275₂₇₆del (p.E92Vfs*19) variants were identified, which were located in the myosin motor domain, a functionally crucial region of the MYO19 protein. Maternally inherited missense variants were identified in SURF1 and ETFDH. All three genes are associated with mitochondrial function, and in silico prediction tools suggest that these variants are likely damaging. Other candidate genetic variants possibly contributing to the cardiac phenotype were also detected in the extended maternal family. To the best of our knowledge, this study represents the first report proposing MYO19 as a candidate gene for HCM and highlights the potential role of oligogenic inheritance in the etiology of the disease. Furthermore, plausible candidate variants of other mitochondria-related genes, such as MYO19, SURF1, and ETFDH, were identified, and other family members were investigated to support the pathogenesis of HCM further. Given the limited understanding of the genetics of pediatric HCM, these findings contribute valuable insights into its genetic basis in pediatric patients. Show less

Previous studies suggest associations between the risk of developing chronic obstructive pulmonary disease (COPD) and adiponectin/leptin (ALR) and apolipoprotein B/A1 (APOR) ratios. This longitudinal observational study, using data from the Korean Genome and Epidemiology Study (KoGES), examined the rate of lung function decline, risk factors for the airflow obstruction (AFO), and the time to first AFO based on ALR and APOR groups. Among 5578 participants, high ALR and low APOR were associated with rapid decline in lung function and a shorter time to the first AFO. The high ALR group and the combined high ALR and low APOR group showed higher risk of experiencing AFO both at least once (RR 1.46, 95% CI 1.12-1.90; RR 1.74, 95% CI 1.23-2.46, respectively) and at the final follow up (RR 1.44, 95% CI 1.05-1.96; RR 1.72, 95% CI 1.14-2.60, respectively). High ALR and the combined high ALR and low APOR were identified as risk factors for earlier time to first AFO. This study highlights the potential of ALR and APOR as makers for predicting the risk of future airflow obstruction. Show less

Alzheimer's disease (AD) remains a significant public health challenge with limited effective treatment options. Ramalin, a compound derived from Antarctic lichens, has shown potential in the treatment of AD because of its strong antioxidant and anti-inflammatory properties. However, its instability and toxicity have hindered the development of Ramalin as a viable therapeutic agent. The primary objective of this study was to synthesize and evaluate novel Ramalin derivatives with enhanced stabilities and reduced toxic profiles, with the aim of retaining or improving their therapeutic potential against AD. The antioxidant, anti-inflammatory, anti-BACE-1, and anti-tau activities of four synthesized Ramalin derivatives (i.e., RA-Hyd-Me, RA-Hyd-Me-Tol, RA-Sali, and RA-Benzo) were evaluated. These derivatives demonstrated significantly improved stabilities compared to the parent compound, with RA-Sali giving the most promising results. More specifically, RA-Sali exhibited a potent BACE-1 inhibitory activity and effectively reduced tau phosphorylation, a critical factor in AD pathology. Despite exhibiting reduced antioxidant activities compared to the parent compound, these derivatives represent a potential multi-targeted approach for AD treatment, marking a significant step forward in the development of stable and effective AD therapeutics. Show less

Alzheimer's disease (AD) is a progressive neurodegenerative disorder marked by the accumulation of amyloid-beta plaques and hyperphosphorylated tau proteins, leading to cognitive decline and neuronal death. However, despite extensive research, there are still no effective treatments for this condition. In this study, a series of chloride-substituted Ramalin derivatives is synthesized to optimize their antioxidant, anti-inflammatory, and their potential to target key pathological features of Alzheimer's disease. The effect of the chloride position on these properties is investigated, specifically examining the potential of these derivatives to inhibit tau aggregation and beta-site amyloid precursor protein cleaving enzyme 1 (BACE-1) activity. Our findings demonstrate that several derivatives, particularly RA-3Cl, RA-4Cl, RA-26Cl, RA-34Cl, and RA-35Cl, significantly inhibit tau aggregation with inhibition rates of approximately 50%. For BACE-1 inhibition, Ramalin and RA-4Cl also significantly decrease BACE-1 expression in N2a cells by 40% and 38%, respectively, while RA-23Cl and RA-24Cl showed inhibition rates of 30% and 35% in SH-SY5Y cells. These results suggest that chloride-substituted Ramalin derivatives possess promising multifunctional properties for AD treatment, warranting further investigation and optimization for clinical applications. Show less

Effective intracellular delivery of therapeutic proteins can potentially treat a wide array of diseases. However, efficient delivery of functional proteins across the cell membrane remains challenging. Exosomes are nanosized vesicles naturally secreted by various types of cells and may serve as promising nanocarriers for therapeutic biomolecules. Here, we engineered exosomes equipped with a photoinducible cargo protein release system, termed mMaple3-mediated protein loading into and release from exosome (MAPLEX), in which cargo proteins can be loaded into the exosomes by fusing them with photocleavable protein (mMaple3)-conjugated exosomal membrane markers and subsequently released from the exosomal membrane by inducing photocleavage with blue light illumination. Using this system, we first induced transcriptional regulation by delivering octamer-binding transcription factor 4 and SRY-box transcription factor 2 to fibroblasts in vitro. Second, we induced in vivo gene recombination in Cre reporter mice by delivering Cre recombinase. Last, we achieved targeted epigenome editing in the brains of 5xFAD and 3xTg-AD mice, two models of Alzheimer's disease. Administration of MAPLEXs loaded with β-site amyloid precursor protein cleaving enzyme 1 ( Show less

Physical activity has been considered an important non-medication intervention in preserving mnemonic processes during aging. However, how aerobic exercise promotes such benefits for human health remains unclear. In this study, we aimed to explore the neuroprotective and anti-inflammatory effects of aerobic exercise against lipopolysaccharide (LPS)-induced amnesic C57BL/6J mice and BV-2 microglial cell models. In the in vivo experiment, the aerobic exercise training groups were allowed to run on a motorized treadmill 5 days/week for 4 weeks at a speed of 10 rpm/min, with LPS (0.1 mg/kg) intraperitoneally injected once a week for 4 weeks. We found that aerobic exercise ameliorated memory impairment and cognitive deficits among the amnesic mice. Correspondingly, aerobic exercise significantly increased the protein expressions of FNDC5, which activates target neuroprotective markers BDNF and CREB, and antioxidant markers Nrf2/HO-1, leading to inhibiting microglial-mediated neuroinflammation and reduced the expression of BACE-1 in the hippocampus and cerebral cortex of amnesic mice. We estimated that aerobic exercise inhibited neuroinflammation in part through the action of FNDC5/irisin on microglial cells. Therefore, we explored the anti-inflammatory effects of irisin on LPS-stimulated BV-2 microglial cells. In the in vitro experiment, irisin treatment blocked NF-κB/MAPK/IRF3 signaling activation concomitantly with the significantly lowered levels of the LPS-induced iNOS and COX-2 elevations and promotes the Nrf2/HO-1 expression in the LPS-stimulated BV-2 microglial cells. Together, our findings suggest that aerobic exercise can improve the spatial learning ability and cognitive functions of LPS-treated mice by inhibiting microglia-mediated neuroinflammation through its effect on the expression of BDNF/FNDC5/irisin. Show less

Differences in metabolic homeostasis, diabetes, and obesity between males and females are evident in rodents and humans. Vagal sensory neurons in the vagus nerve ganglia innervate a variety of visceral organs and use specialized nerve endings to sense interoceptive signals. This visceral organ-brain axis plays a role in relaying interoceptive signals to higher brain centers, as well as in regulating the vago-vagal reflex. I hypothesized that molecularly distinct populations of vagal sensory neurons would play a role in causing differences in metabolic homeostasis between the sexes. SnRNA-Seq was conducted on dissociated cells from the vagus nerve ganglia using the 10X Genomics Chromium platform. Single-nucleus RNA sequencing analysis of vagal sensory neurons from female and male mice revealed differences in the transcriptional profiles of cells in the vagus nerve ganglia. These differences are linked to the expression of sex-specific genes such as Understanding the physiological implications of distinct transcriptomic profiles in vagal sensory neurons on energy balance and metabolic homeostasis would help develop sex-specific treatments for obesity and metabolic dysregulation. Show less

Innate lymphoid cells (ILCs) play an important role in maintaining tissue homeostasis and various inflammatory responses. ILCs are typically classified into three subsets, as is the case for T-cells. Recent studies have reported that IL-10-producing type 2 ILCs (ILC2 Show less

Cancer cell heterogeneity is a serious problem in the control of tumor progression because it can cause chemoresistance and metastasis. Heterogeneity can be generated by various mechanisms, including genetic evolution of cancer cells, cancer stem cells (CSCs), and niche heterogeneity. Because the genetic heterogeneity of CSCs has been poorly characterized, the genetic mutation status of CSCs was examined using Exome-Seq and RNA-Seq data of liver cancer. Here we show that different surface markers for liver cancer stem cells (LCSCs) showed a unique propensity for genetic mutations. Cluster of differentiation 133 (CD133)-positive cells showed frequent mutations in the Show less

Brain aging is a complex biological process that is affected by both genetic background and environment. The transcriptomic analysis of aged human and rodent brains has been applied to identify age-associated molecular and cellular processes for which intervention could possibly restore declining brain functions induced by aging. However, whether these age-associated genetic alterations are indeed involved in the healthy aging of the brain remains unclear. We herein characterized a naturally occurring, extremely long-lived (34 months of age) but healthy mouse group retaining well-preserved motor functions. Strikingly, these long-lived mice maintained tyrosine hydroxylase expression and dopaminergic fiber densities, even in the presence of persistent neuroinflammation and expression of aging markers. Combined with Endeavor gene prioritization, we identified the following midbrain-specific longevity-associated genes in the midbrain of these mice: Show less

Increasing evidence suggests incomplete recovery of COVID-19 patients, who continue to suffer from cardiovascular diseases, including cerebral vascular disorders (CVD) and neurological symptoms. Recent findings indicate that some of the damaging effects of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, especially in the brain, may be induced by the spike protein, leading to the disruption of the initial blood-brain barrier (BBB). SARS-CoV-2-infected cells and animals exhibit age-dependent pathogenesis. In this study, we identified endothelial BACE1 as a critical mediator of BBB disruption and cellular senescence induced by the SARS-CoV-2 spike S1 subunit protein. Increased BACE1 in human brain microvascular endothelial cells (HBMVEC) decreases the levels of tight junction proteins, including ZO-1, occludin, and claudins. Moreover, BACE1 overexpression leads to the accumulation of p16 and p21, typical hallmarks of cellular senescence. Our findings show that the SARS-CoV-2 spike S1 subunit protein upregulated BACE1 expression in HBMVECs, causing endothelial leakage. In addition, the SARS-CoV-2 spike S1 subunit protein induced p16 and p21 expression, indicating BACE1-mediated cellular senescence, confirmed by β-Gal staining in HBMVECs. In conclusion, this study demonstrated that BACE1-mediated endothelial cell damage and senescence may be linked to CVD after COVID-19 infection. Show less

Obesity has emerged as a global public health challenge associated with increased risk of hyperlipidemia and hypertension. It contributes to high sympathetic activity and increased catecholamine levels. The hypothalamic melanocortin system is known to regulate the energy homeostasis. The role of melanocortin 4 receptor (MC4R) has been demonstrated pharmacologically and in animal studies, which showed that severe obesity in MC4R knockout mice was caused by increased food intake and decreased energy consumption. Over 70 multiple different mis- -sense and nonsense mutations in hMC4R have been found at a high frequency of 2-8% in severe early onset or hereditary obesity. The single amino acid variation (D90N) located in the second transmembrane domain (TM2) of MC4R results in accelerated growth and childhood onset obesity. Interestingly, the functional characterization of D90N hMC4R mutant TM2 (m-hMC4R-TM2) revealed normal cell surface expression and binding with agonist similar to the hMC4R wild-type TM2 (wt-hMC4R-TM2) but loss of signal transduction mediated via Gs/adenylyl cyclase activation. It is essential to delineate the three-dimensional structure of MC4Rs in order to elucidate their functional aspects. In this study, we demonstrate the optimized expression and isolation of wt/m-hMC4R-TM2 proteins under different chemical cleavage reaction times and purification procedures via SDS precipitation. The solid-state NMR spectroscopy was carried out to study the structure of wt/m-hMC4R- TM2 protein in the anisotropic phospholipid bicelles. The KSI-wt/m-hMC4R-TM2 fusion proteins developed in cell culture with LB medium. In order to isolate the expressed fusion protein from the cell, ultrasonication, Ni-NTA affinity chromatography, dialysis, and lyophilization techniques were used. Then, to obtain a protein with higher purity and higher yield, the CNBr chemical cleavage time was subdivided into 30 minutes, 1 h, 2 h, 3 h, and 4 h. Purification process was performed using FPLC, and 100 mM KCl and dialysis were used to remove the SDS. CD spectrometer, MALDI-TOF, solution-state NMR, and solid-state NMR were used to confirmed purity and structure of the wt/m-hMC4R-TM2. The precipitation method was used to remove the SDS bound to proteins as KCl-SDS. We optimized the 2 h cleavage reaction times for both wt-hMC4R-TM2 and m-hMC4R-TM2 depending on the purity based on mass spectra and We expressed wt/m-hMC4R-TM2 in E.coli and optimized the isolation and purification process, especially CNBr chemical cleavage time. The efficiency of KCl-SDS precipitation was confirmed via MALDI-TOF MS and the pure proteins obtained using this method were characterized by CD spectroscopy and solution-state NMR. The results of Show less

Dyslipidemia is one of the most important risk factors for cardiovascular (CV) disease. Statin therapy has dramatically improved CV outcomes and is the backbone of current lipid-lowering therapy, but despite well-controlled low-density lipoprotein cholesterol (LDL-C) levels through statin administration, up to 40% patients still experience CV disease. New therapeutic agents to tackle such residual cholesterol risk by lowering not only LDL-C but triglycerides (TG), TG-rich lipoproteins (TRL), or lipoprotein(a) (Lp(a)) are being introduced. Ezetimibe, proprotein convertase subtilisin/kexin type 9 (PCSK9) monoclonal antibodies, PCSK9 small interference RNA (siRNA), and bempedoic acid added to statin therapy have shown additional improvement to CV outcomes. Recent trials administering eicosapentaenoic acid to patients with high TG despite statin therapy have also demonstrated significant CV benefit. Antisense oligonucleotide (ASO) therapies with hepatocyte-specific targeting modifications are now being newly introduced with promising lipid-lowering effects. ASOs targeting TG/TRL, such as angiopoietin-like 3 or 4 (ANGPTL3 or ANGPTL4), apolipoprotein C-III (APOC3), or Lp(a) have effectively lowered the corresponding lipid profiles without requiring high or frequent doses. Clinical outcomes from these novel therapeutics are yet to be proven. Here, we review current and emerging therapeutics targeting LDL-C, TG, TRL, and Lp(a) to reduce the residual CV risk. Show less

The central melanocortin system plays a fundamental role in the control of feeding and body weight. Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC) also regulate overall glucose homeostasis via insulin-dependent and -independent pathways. Here, we report that a subset of ARC POMC neurons innervate the liver via preganglionic parasympathetic acetylcholine (ACh) neurons in the dorsal motor nucleus of the vagus (DMV). Optogenetic stimulation of this liver-projecting melanocortinergic pathway elevates blood glucose levels that is associated with increased expression of hepatic gluconeogenic enzymes in female and male mice. Pharmacological blockade and knockdown of the melanocortin-4 receptor gene in the DMV abolish this stimulation-induced effect. Activation of melanocortin-4 receptors inhibits DMV cholinergic neurons and optogenetic inhibition of liver-projecting parasympathetic cholinergic fibers increases blood glucose levels. This elevated blood glucose is not due to altered pancreatic hormone release. Interestingly, insulin-induced hypoglycemia increases ARC POMC neuron activity. Hence, this liver-projecting melanocortinergic circuit that we identified may play a critical role in the counterregulatory response to hypoglycemia. Show less

Proopiomelanocortin (POMC) neurons in the arcuate nucleus of the hypothalamus (ARC) plays an essential role in the control of food intake and energy expenditure. Melanocortin-4 receptors (MC4Rs) are expressed in key areas that are implicated in regulating energy homeostasis. Although the importance of MC4Rs in the paraventricular hypothalamus (PVH) has been well documented, the role of MC4Rs in the medial amygdala (MeA) on feeding remains controversial. In this study, we specifically examine the role of a novel ARC Show less

Assessment of differentiation potential is a basic requirement to obtain qualified human pluripotent stem cells (hPSCs). Here, we report a simple differentiation method using fetal bovine serum (FBS) to estimate differentiation potential and propensity of hPSCs. PluriTest using RNA-sequencing showed that cells differentiated after treatment with 5% FBS. Expression patterns of three germ layer markers revealed that cells cultured in Knockout Serum Replacement-containing medium (KSR) with mouse feeder cells had higher differentiation potential than cells cultured in a chemically defined medium (E8) with recombinant matrix proteins, especially into the mesoderm and endoderm lineages. Analysis of differentially expressed genes between KSR and E8 identified DUSP6 as a marker for where cells had been cultured. Expression of DUSP6 correlated with FGF-ERK signaling activity. Fine-tuning of FGF-ERK signaling activity to a range that can shut down DUSP6 transcription but sustain NANOG transcription partially increased the differentiation potential. Our data suggest that differentiation with 5% FBS is good to estimate differentiation potential and propensity at the early stage, and that DUSP6 is an excellent marker to monitor ERK signaling activity. Show less

The most prominent hallmarks of many neurodegenerative diseases are the accumulation of misfolded protein aggregates and the death of certain neuronal populations. Autophagy is the major intracellular mechanism that degrades protein aggregates and damaged cellular components. Many studies have reported that the dysfunction of autophagy is associated with several neurodegenerative diseases, such as Alzheimer's disease, amyotrophic lateral sclerosis (ALS), and Parkinson's disease. Here, we identified a novel mechanism of autophagy regulation. Inhibition of MEK5 reduced the level of p62 and increased the ratio of LC3-II to LC3-I, which is a marker for the activation of the autophagy-lysosome pathway (ALP). One of the most well-known regulators of the ALP is mTOR, and previous studies have reported that the major substrate of MEK5 is ERK5. However, we found that MEK5 modulates the autophagy-lysosome pathway in an mTOR- and ERK5-independent manner. Moreover, MEK5 inhibition alleviated the mislocalization of TDP-43 (an ALS-associated protein) and cell death in TDP-43-GFP-expressing neuronal cells. Taken together, these findings suggest that MEK5 is a novel autophagy modulator and that this kinase could be a therapeutic target for neurodegenerative diseases such as amyotrophic lateral sclerosis. Show less