RCHY1 and OPTN are required for melanophagy, selective autophagy of melanosomes.

Melanosomes are specific organelles dedicated to melanin synthesis and accumulation in melanocytes. Autophagy is suggestively involved in melanosome degradation, although the potential underlying molecular mechanisms remain elusive. In selective autophagy, autophagy receptors and E3-ligases are the key factors conferring cargo selectivity. In B16F10 cells, ß-mangostin efficiently induced melanosome degradation without affecting other organelles such as mitochondria, peroxisomes, and the endoplasmic reticulum. Among various autophagy receptors, optineurin (OPTN) contributes TANK-binding kinase 1 (TBK1)-dependently to melanosome degradation and its knockdown inhibited ß-mangostin-mediated melanosome degradation. OPTN translocation to melanosomes was dependent on its ubiquitin-binding domain. Moreover, OPTN-mediated TBK1 activation and subsequent TBK1-mediated S187 OPTN phosphorylation were essential for melanosome degradation. ß-mangostin increased K63-linked melanosome ubiquitination. Finally, the E3-ligase RCHY1 knockdown inhibited the melanosome ubiquitination required for OPTN- and TBK1-phosphorylation as well as melanosome degradation. This study suggests that melanophagy, melanosome-selective autophagy, contributes to melanosome degradation, and OPTN and RCHY1 are an essential autophagy receptor and a E3-ligase, respectively, conferring cargo selectivity in melanophagy.

Optimization of cultivar, germination time and extraction for radish sprout extract with high sulforaphene content.

BACKGROUND: Cruciferous vegetable sprout has been highlighted as a promising functional material rich in bioactive compounds called isothiocyanates (ITCs) and it can be grown in very short periods in controlled indoor farms. However, because ITCs content depends on multiple factors such as cultivar, germination time and myrosinase activity, those variables need to be controlled during germination or extraction to produce functional materials enriched in ITCs. Sulforaphene (SFEN), an ITC found primarily in radishes (Raphanus sativus L.), exerts beneficial effects on obesity. However, the optimal germination and extraction conditions for radish sprout (RSP) to increase SFEN content remain unascertained, and the extract's anti-obesity effect has yet to be evaluated. RESULTS: The present study found that the SFEN content was highest in purple radish sprout (PRSP) among the six cultivars investigated. Optimal SFEN content occurred after 2 days of PRSP germination (2 days PRSP). To maximize the dry matter yield, total ITCs and SFEN contents in RSP extract, we found the optimal conditions for extracting PRSP [27.5 °C, 60 min, 1:75.52 solute/solvent (w/v), no ascorbic acid] using response surface methodology. Consistent with high SFEN content, 2 days PRSP extract significantly outperformed 3 days or 4 days PRSP extract in inhibiting lipid accumulation in 3T3-L1 cells. Moreover, 2 days PRSP extract suppressed adipogenesis and lipogenesis-related protein expression. CONCLUSION: Regarding the cultivar, germination time and extraction conditions, optimally produced PRSP extract contains high SFEN content and exerts anti-obesity effects. Thus, we suggest PRSP extract as a potent functional material for obesity prevention. © 2024 Society of Chemical Industry.

Consideration of SHP-1 as a Molecular Target for Tumor Therapy.

Abnormal activation of receptor tyrosine kinases (RTKs) contributes to tumorigenesis, while protein tyrosine phosphatases (PTPs) contribute to tumor control. One of the most representative PTPs is Src homology region 2 (SH2) domain-containing phosphatase 1 (SHP-1), which is associated with either an increased or decreased survival rate depending on the cancer type. Hypermethylation in the promoter region of PTPN6, the gene for the SHP-1 protein, is a representative epigenetic regulation mechanism that suppresses the expression of SHP-1 in tumor cells. SHP-1 comprises two SH2 domains (N-SH2 and C-SH2) and a catalytic PTP domain. Intramolecular interactions between the N-SH2 and PTP domains inhibit SHP-1 activity. Opening of the PTP domain by a conformational change in SHP-1 increases enzymatic activity and contributes to a tumor control phenotype by inhibiting the activation of the Janus kinase/signal transducer and activator of transcription (JAK/STAT3) pathway. Although various compounds that increase SHP-1 activation or expression have been proposed as tumor therapeutics, except sorafenib and its derivatives, few candidates have demonstrated clinical significance. In some cancers, SHP-1 expression and activation contribute to a tumorigenic phenotype by inducing a tumor-friendly microenvironment. Therefore, developing anticancer drugs targeting SHP-1 must consider the effect of SHP-1 on both cell biological mechanisms of SHP-1 in tumor cells and the tumor microenvironment according to the target cancer type. Furthermore, the use of combination therapies should be considered.

OptNCMiner: a deep learning approach for the discovery of natural compounds modulating disease-specific multi-targets.

BACKGROUND: Due to their diverse bioactivity, natural product (NP)s have been developed as commercial products in the pharmaceutical, food and cosmetic sectors as natural compound (NC)s and in the form of extracts. Following administration, NCs typically interact with multiple target proteins to elicit their effects. Various machine learning models have been developed to predict multi-target modulating NCs with desired physiological effects. However, due to deficiencies with existing chemical-protein interaction datasets, which are mostly single-labeled and limited, the existing models struggle to predict new chemical-protein interactions. New techniques are needed to overcome these limitations. RESULTS: We propose a novel NC discovery model called OptNCMiner that offers various advantages. The model is trained via end-to-end learning with a feature extraction step implemented, and it predicts multi-target modulating NCs through multi-label learning. In addition, it offers a few-shot learning approach to predict NC-protein interactions using a small training dataset. OptNCMiner achieved better prediction performance in terms of recall than conventional classification models. It was tested for the prediction of NC-protein interactions using small datasets and for a use case scenario to identify multi-target modulating NCs for type 2 diabetes mellitus complications. CONCLUSIONS: OptNCMiner identifies NCs that modulate multiple target proteins, which facilitates the discovery and the understanding of biological activity of novel NCs with desirable health benefits.

Amorphigenin from Amorpha fruticosa L. Root Extract Induces Autophagy-Mediated Melanosome Degradation in mTOR-Independent- and AMPK-Dependent Manner.

In this study, we investigated the depigmentation effect of Amorpha fruticosa L. root extract (RE), an herbal medicine. A. fruticosa RE significantly induced depigmentation in α-MSH-treated B16F10 cells at noncytotoxic concentrations. Further, the RE decreased the protein levels of the melanosomal proteins Tyr and Pmel without decreasing their transcript levels. We found that MG132, a proteasome complex inhibitor, was unable to rescue the protein levels, but PepA/E-64D (a lysosomal enzyme inhibitor), 3-MA (a representative autophagy inhibitor), and ATG5 knockdown effectively rescued the protein levels and inhibited the depigmentation effect following RE treatment. Among rotenoids, amorphigenin composed in the RE was identified as a functional chemical that could induce depigmentation; whereas rapamycin, an mTOR inhibitor and a nonselective autophagy inducer, could not induce depigmentation, and amorphigenin effectively induced depigmentation through the degradation of melanosomal proteins. Amorphigenin activated AMPK without affecting mTOR, and knockdown of AMPK offset the whitening effect through degradation of melanosome proteins by amorphigenin. Results from this study suggested that amorphigenin can induce degradation of the melanosome through an AMPK-dependent autophagy process, and has the potential to be used as a depigmentation agent for the treatment of hyperpigmentation.

Ca2+-permeable TRPV1 pain receptor knockout rescues memory deficits and reduces amyloid-ß and tau in a mouse model of Alzheimer's disease.

The transient receptor potential vanilloid 1 (TRPV1) protein is a pain receptor that elicits a hot sensation when an organism eats the capsaicin of red chili peppers. This calcium (Ca2+)-permeable cation channel is mostly expressed in the peripheral nervous system sensory neurons but also in the central nervous system (e.g. hippocampus and cortex). Preclinical studies found that TRPV1 mediates behaviors associated with anxiety and depression. Loss of TRPV1 functionality increases expression of genes related to synaptic plasticity and neurogenesis. Thus, we hypothesized that TRPV1 deficiency may modulate Alzheimer's disease (AD). We generated a triple-transgenic AD mouse model (3xTg-AD+/+) with wild-type (TRPV1+/+), hetero (TRPV1+/-) and knockout (TRPV1-/-) TRPV1 to investigate the role of TRPV1 in AD pathogenesis. We analyzed the animals' memory function, hippocampal Ca2+ levels and amyloid-ß (Aß) and tau pathologies when they were 12 months old. We found that compared with 3xTg-AD-/-/TRPV1+/+ mice, 3xTg-AD+/+/TRPV1+/+ mice had memory impairment and increased levels of hippocampal Ca2+, Aß and total and phosphorylated tau. However, 3xTg-AD+/+/TRPV1-/- mice had better memory function and lower levels of hippocampal Ca2+, Aß, tau and p-tau, compared with 3xTg-AD+/+/TRPV1+/+ mice. Examination of 3xTg-AD-derived primary neuronal cultures revealed that the intracellular Ca2+ chelator BAPTA/AM and the TRPV1 antagonist capsazepine decreased the production of Aß, tau and p-tau. Taken together, these results suggested that TRPV1 deficiency had anti-AD effects and promoted resilience to memory loss. These findings suggest that drugs or food components that modulate TRPV1 could be exploited as therapeutics to prevent or treat AD.

Mechanistic assessment of tolerance to iron deficiency mediated by Trichoderma harzianum in soybean roots.

AIMS: Iron (Fe) deficiency in soil is a continuing problem for soybean (Glycine max L.) production, partly as a result of continuing climate change. This study elucidates how Trichoderma harzianum strain T22 (TH) mitigates growth retardation associated with Fe-deficiency in a highly sensitive soybean cultivar. METHODS AND RESULTS: Soil TH supplementation led to mycelial colonization and the presence of UAOX1 gene in roots that caused substantial improvement in chlorophyll score, photosynthetic efficiency and morphological parameters, indicating a positive influence on soybean health. Although rhizosphere acidification was found to be a common feature of Fe-deficient soybean, the upregulation of Fe-reductase activity (GmFRO2) and total phenol secretion were two of the mechanisms that substantially increased the Fe availability by TH. Heat-killed TH applied to soil caused no improvement in photosynthetic attributes and Fe-reductase activity, confirming the active role of TH in mitigating Fe-deficiency. Consistent increases in tissue Fe content and increased Fe-transporter (GmIRT1, GmNRAMP2a, GmNRAMP2b and GmNRAMP7) mRNA levels in roots following TH supplementation were observed only under Fe-deprivation. Root cell death, electrolyte leakage, superoxide (O2 •- ) and hydrogen peroxide (H2 O2 ) substantially declined due to TH in Fe-deprived plants. Further, the elevation of citrate and malate concentration along with the expression of citrate synthase (GmCs) and malate synthase (GmMs) caused by TH suggest improved chelation of Fe in Fe-deficient plants. Results also suggest that TH has a role in triggering antioxidant defence by increasing the activity of glutathione reductase (GR) along with elevated S-metabolites (glutathione and methionine) to stabilize redox status under Fe-deficiency. CONCLUSIONS: TH increases the availability and mobilization of Fe by inducing Fe-uptake pathways, which appears to help provide resistance to oxidative stress associated with Fe-shortage in soybean. SIGNIFICANCE AND IMPACT OF THE STUDY: These findings indicate that while Fe deficiency does not affect the rate or degree of TH hyphal association in soybean roots, the beneficial effects of TH alone may be Fe deficiency-dependent.

Usefulness of three-dimensional computed tomography for patellofemoral measurement.

PURPOSE: This study aimed to establish the normal values for knee patellofemoral alignment as measured using 3-dimensional computed tomography (3D CT), to standardize the technique, and to show the inter- and intra-observer reliability of this measurement. METHODS: The present study included 62 asymptomatic volunteers (124 knees). 3D CT scanning was performed with each volunteer in the supine position with 15° of knee flexion, and consistent 3D axial images of the patellofemoral joint were obtained with alignment in the desired stereographic baseline direction in anterior-posterior, lateral, and axial rotations. Two independent observers measured patellofemoral alignment parameters, including the sulcus angle, congruence angle, lateral patellofemoral angle, condyle-patellar angle, and lateral trochlear inclination angle. RESULTS: Based on 3D CT measurement, the mean values of the parameters were 145.9° ± 9.2° for the sulcus angle, 12.6° ± 22.6° for the congruence angle, 9.2° ± 4.6° for the lateral patellofemoral angle, 14.1° ± 6.4° for the condyle-patellar (lateral facets) angle, - 8.5° ± 8.4° for condyle-patellar (patellar axis) angle, and 16.5° ± 6.3° for the lateral trochlear inclination angle. A statistically significant difference was observed between men and women in the sulcus and condyle-patellar (patellar axis) angles (p = 0.045, 0.011, respectively). All parameters showed excellent inter- and intra-observer reliability. CONCLUSION: The normal values and ranges for patellofemoral alignment parameters were evaluated using 3D CT. The results of this study provide reference information that may facilitate diagnosis and treatment planning of patellofemoral disorders in skeletally mature non-pathologic patients. LEVEL OF EVIDENCE: II.

The Function of N-Myc Downstream-Regulated Gene 2 (NDRG2) as a Negative Regulator in Tumor Cell Metastasis.

N-myc downstream-regulated gene 2 (NDRG2) is a tumor-suppressor gene that suppresses tumorigenesis and metastasis of tumors and increases sensitivity to anti-cancer drugs. In this review, we summarize information on the clinicopathological characteristics of tumor patients according to NDRG2 expression in various tumor tissues and provide information on the metastasis inhibition-related cell signaling modulation by NDRG2. Loss of NDRG2 expression is a prognostic factor that correlates with TNM grade and tumor metastasis and has an inverse relationship with patient survival in various tumor patients. NDRG2 inhibits cell signaling, such as AKT-, NF-κB-, STAT3-, and TGF-ß-mediated signaling, to induce tumor metastasis, and induces activation of GSK-3ß which has anti-tumor effects. Although NDRG2 operates as an adaptor protein to mediate the interaction between kinases and phosphatases, which is essential in regulating cell signaling related to tumor metastasis, the molecular mechanism of NDRG2 as an adapter protein does not seem to be fully elucidated. This review aims to assist the research design regarding NDRG2 function as an adaptor protein and suggests NDRG2 as a molecular target to inhibit tumor metastasis and improve the prognosis in tumor patients.

The patient-rated tennis elbow evaluation questionnaire was successfully translated to Korean.

INTRODUCTION: Lateral epicondylitis is a common tendinopathy of the elbow. An adequately validated questionnaire is essential to compare the various treatment options, because there is still controversy regarding the best treatment for lateral epicondylitis. PURPOSE OF THE STUDY: The patient-rated tennis elbow evaluation (PRTEE) was introduced to measure the degree of symptoms and the limitation of function in these patients. This English questionnaire was previously translated into several languages and validated; however, it has not been translated into Korean. The aim of this study was to translate and cross-culturally adapt the PRTEE questionnaire into Korean, and to assess the reliability and validity of the translated questionnaire. METHODS: Eighty-three patients were asked to complete the Korean version of the PRTEE and other questionnaires related to the upper extremity or to the general health condition (the Disabilities of Arm, Shoulder, and Hand questionnaire, Short Form-12, and EQ-5D). The PRTEE responses were recorded twice to evaluate test-retest reliability. Internal consistency, test-retest reliability, concurrent validity, and ceiling and flooring effects were evaluated. RESULTS: The Korean version of the PRTEE showed high internal consistency (Cronbach's alpha = 0.968). The overall test-retest reliability was good (intra-class correlation score = 0.94). High correlation efficiencies between the PRTEE and other questionnaires were observed, supporting high concurrent validity (Spearman's correlation = 0.83 for the Disabilities of Arm, Shoulder, and Hand questionnaire; -0.73 for Short Form-12; and -0.57 for EQ-5D). Ceiling and floor effects were not observed on this test. CONCLUSION: We successfully translated the PRTEE into the Korean language; this version of the PRTEE showed good reliability and construct validity in Korean-speaking patients with lateral epicondylitis.

Dog Ownership in Early Life Increased the Risk of Nonatopic Asthma in Children.

BACKGROUND: It is still debatable whether dog ownership during early childhood is a risk factor for the development of allergic diseases. OBJECTIVE: We investigated the association of dog ownership in early life with sensitization and asthma in childhood. METHODS: Data from the Cohort for Childhood Origin of Asthma and Allergic diseases were used to investigate the association between dog ownership at any time from pregnancy to 1 year of age and sensitization to aeroallergens at 3 and 7 years old, bronchial hyperresponsiveness (BHR), and asthma at 7 years old. We analyzed the cytokine levels in cord blood (CB) and indoor environmental measurement concentrations in the mother's residence obtained at 36 weeks of pregnancy. RESULTS: Sensitization to dogs at age 3 and 7 did not differ between dog ownership and nonownership, but dog ownership during early life decreased the risk of sensitization to aeroallergens at age 7 (aOR = 0.44, 95% CI 0.21-0.90). Dog ownership significantly increased the risk of nonatopic BHR (aOR = 2.86; 95% CI 1.32-6.21). In addition, dog ownership was associated with asthma, especially nonatopic asthma at 7 years old (aOR = 2.73, 95% CI 1.02-7.32; aOR = 7.05, 95% CI 1.85-26.90, respectively). There were no significant differences in the concentrations of IL-13 or interferon-γ in CB or indoor environmental measurements according to dog ownership during pregnancy. CONCLUSION: Early-life dog exposure in this birth cohort has been shown to reduce atopy but increase the risk of nonatopic BHR and nonatopic asthma at 7 years old.

Silicon induces metallochaperone-driven cadmium binding to the cell wall and restores redox status through elevated glutathione in Cd-stressed sugar beet.

Cadmium (Cd) is toxic; however, whether silicon (Si) alleviates Cd toxicity was never studied in sugar beet. The study was conducted on 2-week-old sugar beet cultivated in the presence or absence of Cd (10 µM CdSO4 ) and Si (1 mM Na2 SiO3 ) in hydroponic conditions. The morphological impairment and cellular damages observed in sugar beet upon Cd toxicity were entirely reversed due to Si. Si substantially restored the energy-providing ability, absorbed energy flux, and electron transport toward PSII, which might be correlated with the upregulation of BvIRT1 and ferric chelate reductase activity leading to the restoration of Fe status in Cd-stressed sugar beet. Although Si caused a reduction of shoot Cd, the root Cd substantially increased under Cd stress, a significant part of which was retained in the cell wall rather than in the root vacuole. While the concentration of phytochelatin and the expression of BvPCS3 (PHYTOCHELATIN SYNTHASE 3) showed no changes upon Si exposure, Si induced the expression of BvHIPP32 (HEAVY METAL-ASSOCIATED ISOPRENYLATED PLANT PROTEIN 32) in the Cd-exposed root. The BvHIPP32 and AtHIPP32 metallochaperone proteins are localized in the cell wall and they share similar sequence alignment, physiochemical properties, secondary structure, cellular localization, motif locations, domain association, and metal-binding site (cd00371) linked to the metallochaperone-like protein. It suggests that Si reduces the Cd level in shoot by retaining the excess Cd in the cell wall of roots due to the induction of BvHIPP32 gene. Also, Si stimulates glutathione-related antioxidants along with the BvGST23 expression, inferring an ascorbate-glutathione ROS detoxification pathway in Cd-exposed plants.

Metabolic stress controls mTORC1 lysosomal localization and dimerization by regulating the TTT-RUVBL1/2 complex.

The metabolism of glucose and glutamine, primary carbon sources utilized by mitochondria to generate energy and macromolecules for cell growth, is directly regulated by mTORC1. We show that glucose and glutamine, by supplying carbons to the TCA cycle to produce ATP, positively feed back to mTORC1 through an AMPK-, TSC1/2-, and Rag-independent mechanism by regulating mTORC1 assembly and its lysosomal localization. We discovered that the ATP-dependent TTT-RUVBL1/2 complex was disassembled and repressed by energy depletion, resulting in its decreased interaction with mTOR. The TTT-RUVBL complex was necessary for the interaction between mTORC1 and Rag and formation of mTORC1 obligate dimers. In cancer tissues, TTT-RUVBL complex mRNAs were elevated and positively correlated with transcripts encoding proteins of anabolic metabolism and mitochondrial function-all mTORC1-regulated processes. Thus, the TTT-RUVBL1/2 complex responds to the cell's metabolic state, directly regulating the functional assembly of mTORC1 and indirectly controlling the nutrient signal from Rags to mTORC1.

Range Dividing MIMO Waveform for Improving Tracking Performance.

A multiple-input multiple-output (MIMO) method that shares the same frequency band can efficiently increase radar performance. An essential element of a MIMO radar is the orthogonality of the waveform. Typically, orthogonality is obtained by spreading different signals into divided domains such as in time-domain multiplexing, frequency-domain multiplexing, and code domain multiplexing. This paper proposes a method of spreading the interference signals outside the range bins of interest for pulse doppler radars. This is achieved by changing the pulse repetition frequency under certain constraints, and an additional gain can be obtained by doppler processing. This method is very effective for improving the angular accuracy of the MIMO radar for a small number of air targets, although it may have limitations in use for many targets or in high clutter environments.

Topical Application of 7,3',4'-Trihydroxyisoflavone Alleviates Atopic Dermatitis-Like Symptoms in NC/Nga Mice.

Atopic dermatitis is a skin disease characterized by chronic inflammatory lesions, and new therapies are needed to address its rising prevalence. Soy isoflavone has been highlighted as a potential new cosmeceutical material that may have applications in atopic dermatitis care. We have developed a technique to attach an additional -OH group to the ortho position of -OH in the phenol ring using a special enzyme. By adding the -OH group to daidzein, 7,3',4'-trihydroxyisoflavone can be generated for possible use as a cosmeceutical and functional food material. In this study, we sought to examine the anti-atopic effects of 7,3',4'-trihydroxyisoflavone, an analog of daidzein. Topical application of 7,3',4'-trihydroxyisoflavone reduced Dermatophagoides farina extract-induced atopic dermatitis symptoms in NC/Nga mice. Histological analysis demonstrated that 7,3',4'-trihydroxyisoflavone suppressed D. farina extract-induced infiltration of eosinophils and mast cells into skin lesions. We also found that 7,3',4'-trihydroxyisoflavone significantly reduces the D. farina extract-induced increases in serum IgE and macrophage-derived chemokine (CCL22) levels. We observed that 7,3',4'-trihydroxyisoflavone suppresses atopic markers including macrophage-derived chemokine (CCL22) and thymus and activation-regulated chemokine (CCL17) in HaCaT cells. 7,3',4'-Trihydroxyisoflavone also reduced TNF-α/IFN-γ-induced phosphorylation of ERK1/2 and JNK1/2. These results highlight several desirable properties of 7,3',4'-trihydroxyisoflavone, which support its use as a cosmeceutical ingredient for the treatment of atopic dermatitis.

Arbuscular Mycorrhizal Symbiosis Mitigates Iron (Fe)-Deficiency Retardation in Alfalfa (Medicago sativa L.) Through the Enhancement of Fe Accumulation and Sulfur-Assisted Antioxidant Defense.

Iron (Fe)-deficiency is one of the major constraints affecting growth, yield and nutritional quality in plants. This study was performed to elucidate how arbuscular mycorrhizal fungi (AMF) alleviate Fe-deficiency retardation in alfalfa (Medicago sativa L.). AMF supplementation improved plant biomass, chlorophyll score, Fv/Fm (quantum efficiency of photosystem II), and Pi_ABS (photosynthesis performance index), and reduced cell death, electrolyte leakage, and hydrogen peroxide accumulation in alfalfa. Moreover, AMF enhanced ferric chelate reductase activity as well as Fe, Zn, S and P in alfalfa under Fe-deficiency. Although Fe-transporters (MsIRT1 and MsNramp1) did not induce in root but MsFRO1 significantly induced by AMF under Fe deficiency in roots, suggesting that AMF-mediated Fe enhancement is related to the bioavailability of Fe at rhizosphere/root apoplast rather than the upregulation of Fe transporters under Fe deficiency in alfalfa. Several S-transporters (MsSULTR1;1, MsSULTR1;2, MsSULTR1;3, and MsSULTR3;1) markedly increased following AMF supplementation with or without Fe-deficiency alfalfa. Our study further suggests that Fe uptake system is independently influenced by AMF regardless of the S status in alfalfa. However, the increase of S in alfalfa is correlated with the elevation of GR and S-metabolites (glutathione and cysteine) associated with antioxidant defense under Fe deficiency.

Cordycepin Resensitizes T24R2 Cisplatin-Resistant Human Bladder Cancer Cells to Cisplatin by Inactivating Ets-1 Dependent MDR1 Transcription.

Tumor cell resistance to anti-cancer drugs is a major obstacle in tumor therapy. In this study, we investigated the mechanism of cordycepin-mediated resensitization to cisplatin in T24R2 cells, a T24-derived cell line. Treatment with cordycepin or cisplatin (2 µg/mL) alone failed to induce cell death in T24R2 cells, but combination treatment with these drugs significantly induced apoptosis through mitochondrial pathways, including depolarization of mitochondrial membranes, decrease in anti-apoptotic proteins Bcl-2, Bcl-xL, and Mcl-1, and increase in pro-apoptotic proteins Bak and Bax. High expression levels of MDR1 were the cause of cisplatin resistance in T24R2 cells, and cordycepin significantly reduced MDR1 expression through inhibition of MDR1 promoter activity. MDR1 promoter activity was dependent on transcription factor Ets-1 in T24R2 cells. Although correlation exists between MDR1 and Ets-1 expression in bladder cancer patients, active Ets-1, Thr38 phosphorylated form (pThr38), was critical to induce MDR1 expression. Cordycepin decreased pThr-38 Ets-1 levels and reduced MDR1 transcription, probably through its effects on PI3K signaling, inducing the resensitization of T24R2 cells to cisplatin. The results suggest that cordycepin effectively resensitizes cisplatin-resistant bladder cancer cells to cisplatin, thus serving as a potential strategy for treatment of cancer in patients with resistance to anti-cancer drugs.

Tyrosinase Inhibition and Kinetic Details of Puerol A Having But-2-Enolide Structure from Amorpha fruticosa.

Puerol A (1) from Amorpha fruticosa showed highly potent inhibition against both monophenolase (IC50 = 2.2 µM) and diphenolase (IC50 = 3.8 µM) of tyrosinase. We tried to obtain a full story of enzyme inhibitory behavior for inhibitor 1 because the butenolide skeleton has never been reported as a tyrosinase inhibitor. Puerol A was proved as a reversible, competitive, simple slow-binding inhibitor, according to the respective parameters; k3 = 0.0279 µM-1 min-1 and k4 = 0.003 min-1. A longer lag-phase and a reduced static-state activity of the enzyme explained that puerol A had a tight formation of the complex with Emet. Dose-dependent inhibition was also confirmed by high-performance liquid chromatography (HPLC) analysis using N-acetyl-l-tyrosine as a substrate, which was completely inhibited at 20 µM. A high binding affinity of 1 to tyrosinase was confirmed by fluorescence quenching analysis. Moreover, puerol A decreased melanin content in the B16 melanoma cell dose-dependently with an IC50 of 11.4 µM.

Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

Obesity-mediated inflammation is a major cause of insulin resistance, and macrophages play an important role in this process. The 78-kDa glucose-regulated protein (GRP78) is a major endoplasmic reticulum chaperone that modulates unfolded protein response (UPR), and mice with GRP78 heterozygosity were resistant to diet-induced obesity. Here, we show that mice with macrophage-selective ablation of GRP78 (Lyz- GRP78-/-) are protected from skeletal muscle insulin resistance without changes in obesity compared with wild-type mice after 9 wk of high-fat diet. GRP78-deficient macrophages demonstrated adapted UPR with up-regulation of activating transcription factor (ATF)-4 and M2-polarization markers. Diet-induced adipose tissue inflammation was reduced, and bone marrow-derived macrophages from Lyz- GRP78-/- mice demonstrated a selective increase in IL-6 expression. Serum IL-13 levels were elevated by >4-fold in Lyz- GRP78-/- mice, and IL-6 stimulated the myocyte expression of IL-13 and IL-13 receptor. Lastly, recombinant IL-13 acutely increased glucose metabolism in Lyz- GRP78-/- mice. Taken together, our data indicate that GRP78 deficiency activates UPR by increasing ATF-4, and promotes M2-polarization of macrophages with a selective increase in IL-6 secretion. Macrophage-derived IL-6 stimulates the myocyte expression of IL-13 and regulates muscle glucose metabolism in a paracrine manner. Thus, our findings identify a novel crosstalk between macrophages and skeletal muscle in the modulation of obesity-mediated insulin resistance.-Kim, J. H., Lee, E., Friedline, R. H., Suk, S., Jung, D. Y., Dagdeviren, S., Hu, X., Inashima, K., Noh, H. L., Kwon, J. Y., Nambu, A., Huh, J. R., Han, M. S., Davis, R. J., Lee, A. S., Lee, K. W., Kim, J. K. Endoplasmic reticulum chaperone GRP78 regulates macrophage function and insulin resistance in diet-induced obesity.

Gingerenone A Attenuates Monocyte-Endothelial Adhesion via Suppression of I Kappa B Kinase Phosphorylation.

During the early stages of atherosclerosis, monocytes bind and migrate into the endothelial layer, promoting inflammation within the aorta. In order to prevent the development of atherosclerosis, it is critical to inhibit such inflammation. The therapeutic effects of ginger have been investigated in several models of cardiovascular disease. However, although a number of previous studies have focused on specific compounds, the mechanisms of action responsible remain unclear. Here, we investigated five major compounds present in ginger, and observed that gingerenone A exhibited the strongest inhibitory effects against tumor necrosis factor (TNF)-α and lipopolysaccharide (LPS) induced monocyte-endothelial adhesion. Furthermore, gingerenone A significantly suppressed the expression of TNF-α and LPS-induced vascular cell adhesion molecule-1 (VCAM-1) and chemokine (C-C motif) ligand 2 (CCL2), key mediators of the interaction between monocytes, and endothelial cells. Transactivation of nuclear factor-κB (NF-κB), which is a key transcription factor of VCAM-1 and CCL2, was induced by TNF-α and LPS, and inhibited by treatment of gingerenone A. Gingerenone A also inhibited the phosphorylation of NF-κB inhibitor (IκB) α and IκB Kinase. Taken together, these results demonstrate that gingerenone A attenuates TNF-α and LPS-induced monocyte adhesion and the expression of adhesion factors in endothelial cells via the suppression of NF-κB signaling. J. Cell. Biochem. 119: 260-268, 2018. © 2017 Wiley Periodicals, Inc.