Intrinsic Phase Stability and Inherent Bandgap of Formamidinium Lead Triiodide Perovskite Single Crystals.

Understanding the intrinsic phase stability and inherent band gap of formamidinium lead triiodide (FAPbI3 ) perovskites is crucial to further improve the performance of perovskite solar cells (PSCs). Herein, we explored the α- to δ-phase transition and band gap of FAPbI3 single crystals grown by an inverse temperature solubility method. We found that the residual γ-butyrolactone solvents in the inner empty space of the FAPbI3 single crystal accelerate the phase transition at kinetics. By adopting 2-methoxyethanol as the solvent, over 2000 h of stable α-FAPbI3 crystals could be acquired. This proves that although FAPbI3 is regarded as unstable at thermodynamics, it could own excellent kinetic stability without any doping or additives because of the slow solid to solid phase transition instead of the fast phase transition assisted by the solvents. Furthermore, we revealed that the bulk FAPbI3 single crystal with a size above 100 µm can have an inherent band gap of 1.41 eV. Thus, our work provides key scientific guidance for high-performance FAPbI3 -based PSCs.

Histopathological characteristics of interstitial cystitis/bladder pain syndrome without Hunner lesion.

AIMS: To assess the distinct histopathological characteristics and their clinical significance between non-Hunner-type and Hunner-type interstitial cystitis (IC)/bladder pain syndrome (BPS). METHODS AND RESULTS: We prospectively enrolled and classified IC/BPS patients, on the basis of cystoscopic findings, as having non-Hunner-type IC and Hunner-type IC. Specimens obtained from the posterior wall in non-Hunner-type IC cases during hydrodistension or from Hunner/non-Hunner lesions in Hunner-type IC cases during transurethral resection were evaluated. Stress urinary incontinence patients with microscopic haematuria were selected as controls. Biopsy specimens were obtained from 15 non-Hunner-type IC, 15 Hunner-type IC and 5 non-IC patients. Severe and moderate fibrosis was more frequently observed in non-Hunner-type IC than in Hunner-type IC and non-IC cases. However, severe and moderate inflammation was more frequently observed in Hunner-type IC than in non-Hunner-type IC cases. The remnant urothelium was significantly decreased in Hunner-type IC cases as compared with non-Hunner-type IC and non-IC cases (P < 0.05), and non-Hunner-type IC cases showed a higher number of mast cells than Hunner-type IC and non-IC cases (P = 0.035). Accordingly, several fibrosis-promoting genes were highly expressed in bladder tissues of non-Hunner-type IC, as compared with Hunner-type IC. Patients with severe fibrosis showed significantly higher urinary frequency and smaller bladder capacity than those with moderate and mild fibrosis (all P < 0.05). CONCLUSIONS: Non-Hunner-type IC is characterized by severe fibrosis and increased mast cell infiltration, whereas Hunner-type IC is characterized by severe inflammation and urothelial denudation in the entire bladder. Fibrosis in the bladder of IC/BPS patients was correlated with increased urinary frequency and decreased bladder capacity.

Cystamine induces AIF-mediated apoptosis through glutathione depletion.

Cystamine and its reduced form cysteamine showed protective effects in various models of neurodegenerative disease, including Huntington's disease and Parkinson's disease. Other lines of evidence demonstrated the cytotoxic effect of cysteamine on duodenal mucosa leading to ulcer development. However, the mechanism for cystamine cytotoxicity remains poorly understood. Here, we report a new pathway in which cystamine induces apoptosis by targeting apoptosis-inducing factor (AIF). By screening of various cell lines, we observed that cystamine and cysteamine induce cell death in a cell type-specific manner. Comparison between cystamine-sensitive and cystamine-resistant cell lines revealed that cystamine cytotoxicity is not associated with unfolded protein response, reactive oxygen species generation and transglutaminase or caspase activity; rather, it is associated with the ability of cystamine to trigger AIF nuclear translocation. In cystamine-sensitive cells, cystamine suppresses the levels of intracellular glutathione by inhibiting γ-glutamylcysteine synthetase expression that triggers AIF translocation. Conversely, glutathione supplementation completely prevents cystamine-induced AIF translocation and apoptosis. In rats, cysteamine administration induces glutathione depletion and AIF translocation leading to apoptosis of duodenal epithelium. These results indicate that AIF translocation through glutathione depletion is the molecular mechanism of cystamine toxicity, and provide important implications for cystamine in the neurodegenerative disease therapeutics as well as in the regulation of AIF-mediated cell death.

Priming with ceramide-1 phosphate promotes the therapeutic effect of mesenchymal stem/stromal cells on pulmonary artery hypertension.

Some molecules enriched in damaged organs can contribute to tissue repair by stimulating the mobilization of stem cells. These so-called "priming" factors include bioactive lipids, complement components, and cationic peptides. However, their therapeutic significance remains to be determined. Here, we show that priming of mesenchymal stromal/stem cells (MSCs) with ceramide-1 phosphate (C1P), a bioactive lipid, enhances their therapeutic efficacy in pulmonary artery hypertension (PAH). Human bone marrow (BM)-derived MSCs treated with 100 or 200 µM C1P showed improved migration activity in Transwell assays compared with non-primed MSCs and concomitantly activated MAPK(p42/44) and AKT signaling cascades. Although C1P priming had little effect on cell surface marker phenotypes and the multipotency of MSCs, it potentiated their proliferative, colony-forming unit-fibroblast, and anti-inflammatory activities. In a monocrotaline-induced PAH animal model, a single administration of human MSCs primed with C1P significantly attenuated the PAH-related increase in right ventricular systolic pressure, right ventricular hypertrophy, and thickness of α-smooth muscle actin-positive cells around the vessel wall. Thus, this study shows that C1P priming increases the effects of MSC therapy by enhancing the migratory, self-renewal, and anti-inflammatory activity of MSCs and that MSC therapy optimized with priming protocols might be a promising option for the treatment of PAH patients.

Activating transcription factor-2 supports the antioxidant capacity and ability of human mesenchymal stem cells to prevent asthmatic airway inflammation.

Glutathione (GSH), an abundant nonprotein thiol antioxidant, participates in several biological processes and determines the functionality of stem cells. A detailed understanding of the molecular network mediating GSH dynamics is still lacking. Here, we show that activating transcription factor-2 (ATF2), a cAMP-response element binding protein (CREB), plays a crucial role in maintaining the level and activity of GSH in human mesenchymal stem cells (MSCs) by crosstalking with nuclear factor erythroid-2 like-2 (NRF2), a well-known master regulator of cellular redox homeostasis. Priming with ascorbic acid 2-glucoside (AA2G), a stable vitamin C derivative, increased the expression and activity of ATF2 in MSCs derived from human embryonic stem cells and umbilical cord. Subsequently, activated ATF2 crosstalked with the CREB1-NRF2 pathway to preserve the GSH dynamics of MSCs through the induction of genes involved in GSH synthesis (GCLC and GCLM) and redox cycling (GSR and PRDX1). Accordingly, shRNA-mediated silencing of ATF2 significantly impaired the self-renewal, migratory, proangiogenic, and anti-inflammatory capacities of MSCs, and these defects were rescued by supplementation of the cells with GSH. In addition, silencing ATF2 attenuated the ability of MSCs to alleviate airway inflammatory responses in an ovalbumin-induced mouse model of allergic asthma. Consistently, activation of ATF2 by overexpression or the AA2G-based priming procedure enhanced the core functions of MSCs, improving the in vivo therapeutic efficacy of MSCs for treating asthma. Collectively, our findings suggest that ATF2 is a novel modulator of GSH dynamics that determines the core functionality and therapeutic potency of MSCs used to treat allergic asthma.

Glutathione dynamics is a potential predictive and therapeutic trait for neoadjuvant chemotherapy response in bladder cancer.

Radical cystectomy with preoperative cisplatin-based neoadjuvant chemotherapy (NAC) is the standard care for muscle-invasive bladder cancers (MIBCs). However, the complete response rate to this modality remains relatively low, and current clinicopathologic and molecular classifications are inadequate to predict NAC response in patients with MIBC. Here, we demonstrate that dysregulation of the glutathione (GSH) pathway is fundamental for MIBC NAC resistance. Comprehensive analysis of the multicohort transcriptomes reveals that GSH metabolism and immune-response genes are enriched in NAC-resistant and NAC-sensitive MIBCs, respectively. A machine-learning-based tumor/stroma classifier is applied for high-throughput digitalized immunohistochemistry analysis, finding that GSH dynamics proteins, including glutaminase-1, are associated with NAC resistance. GSH dynamics is activated in cisplatin-resistant MIBC cells, and combination treatment with a GSH dynamics modulator and cisplatin significantly suppresses tumor growth in an orthotopic xenograft animal model. Collectively, these findings demonstrate the predictive and therapeutic values of GSH dynamics in determining the NAC response in MIBCs.

The CDK1/TFCP2L1/ID2 cascade offers a novel combination therapy strategy in a preclinical model of bladder cancer.

Aberrant activation of embryogenesis-related molecular programs in urothelial bladder cancer (BC) is associated with stemness features related to oncogenic dedifferentiation and tumor metastasis. Recently, we reported that overexpression of transcription factor CP2-like protein-1 (TFCP2L1) and its phosphorylation at Thr177 by cyclin-dependent kinase-1 (CDK1) play key roles in regulating bladder carcinogenesis. However, the clinical relevance and therapeutic potential of this novel CDK1-TFCP2L1 molecular network remain elusive. Here, we demonstrated that inhibitor of DNA binding-2 (ID2) functions as a crucial mediator by acting as a direct repressive target of TFCP2L1 to modulate the stemness features and survival of BC cells. Low ID2 and high CDK1 expression were significantly associated with unfavorable clinical characteristics. TFCP2L1 downregulated ID2 by directly binding to its promoter region. Consistent with these findings, ectopic expression of ID2 or treatment with apigenin, a chemical activator of ID2, triggered apoptosis and impaired the proliferation, suppressed the stemness features, and reduced the invasive capacity of BC cells. Combination treatment with the specific CDK1 inhibitor RO-3306 and apigenin significantly suppressed tumor growth in an orthotopic BC xenograft animal model. This study demonstrates the biological role and clinical utility of ID2 as a direct target of the CDK1-TFCP2L1 pathway for modulating the stemness features of BC cells.

Intravital imaging and single cell transcriptomic analysis for engraftment of mesenchymal stem cells in an animal model of interstitial cystitis/bladder pain syndrome.

Mesenchymal stem cell (MSC) therapy is a promising treatment for various intractable disorders including interstitial cystitis/bladder pain syndrome (IC/BPS). However, an analysis of fundamental characteristics driving in vivo behaviors of transplanted cells has not been performed, causing debates about rational use and efficacy of MSC therapy. Here, we implemented two-photon intravital imaging and single cell transcriptome analysis to evaluate the in vivo behaviors of engrafted multipotent MSCs (M-MSCs) derived from human embryonic stem cells (hESCs) in an acute IC/BPS animal model. Two-photon imaging analysis was performed to visualize the dynamic association between engrafted M-MSCs and bladder vasculature within live animals until 28 days after transplantation, demonstrating the progressive integration of transplanted M-MSCs into a perivascular-like structure. Single cell transcriptome analysis was performed in highly purified engrafted cells after a dual MACS-FACS sorting procedure and revealed expression changes in various pathways relating to pericyte cell adhesion and cellular stress. Particularly, FOS and cyclin dependent kinase-1 (CDK1) played a key role in modulating the migration, engraftment, and anti-inflammatory functions of M-MSCs, which determined their in vivo therapeutic potency. Collectively, this approach provides an overview of engrafted M-MSC behavior in vivo, which will advance our understanding of MSC therapeutic applications, efficacy, and safety.

A Preclinical Study of Human Embryonic Stem Cell-Derived Mesenchymal Stem Cells for Treating Detrusor Underactivity by Chronic Bladder Ischemia.

BACKGROUND: The therapeutic effects of human embryonic stem cell-derived multipotent mesenchymal stem cells (M-MSCs) were evaluated for detrusor underactivity (DUA) in a rat model with atherosclerosis-induced chronic bladder ischemia (CBI) and associated mechanisms. METHODS: Sixteen-week-old male Sprague-Dawley rats were divided into five groups (n = 10). The DUA groups underwent 30 bilateral repetitions of endothelial injury to the iliac arteries to induce CBI, while the sham control group underwent a sham operation. All rats used in this study received a 1.25% cholesterol diet for 8 weeks. M-MSCs at a density of 2.5, 5.0, or 10.0 × 105 cells (250 K, 500 K, or 1000 K; K = a thousand) were injected directly into the bladder 7 weeks post-injury, while the sham and DUA group were treated only with vehicle (phosphate buffer solution). One week after M-MSC injection, awake cystometry was performed on the rats. Then, the bladders were harvested, studied in an organ bath, and prepared for histological and gene expression analyses. RESULTS: CBI by iliac artery injury reproduced voiding defects characteristic of DUA with decreased micturition pressure, increased micturition interval, and a larger residual volume. The pathological DUA properties were improved by M-MSC treatment in a dose-dependent manner, with the 1000 K group producing the best efficacy. Histological analysis revealed that M-MSC therapy reduced CBI-induced injuries including bladder fibrosis, muscular loss, and apoptosis. Transplanted M-MSCs mainly engrafted as vimentin and NG2 positive pericytes rather than myocytes, leading to increased angiogenesis in the CBI bladder. Transcriptomes of the CBI-injured bladders were characterized by the complement system, inflammatory, and ion transport-related pathways, which were restored by M-MSC therapy. CONCLUSIONS: Single injection of M-MSCs directly into the bladder of a CBI-induced DUA rat model improved voiding profiles and repaired the bladder muscle atrophy in a dose-dependent manner.

Estimation of spatially continuous daytime particulate matter concentrations under all sky conditions through the synergistic use of satellite-based AOD and numerical models.

Satellite-derived aerosol optical depth (AOD) products are one of main predictors to estimate ground-level particulate matter (PM10 and PM2.5) concentrations. Since AOD products, however, are only provided under high-quality conditions, missing values usually exist in areas such as clouds, cloud shadows, and bright surfaces. In this study, spatially continuous AOD and subsequent PM10 and PM2.5 concentrations were estimated over East Asia using satellite- and model-based data and auxiliary data in a Random Forest (RF) approach. Data collected from the Geostationary Ocean Color Imager (GOCI; 8 times per day) in 2016 were used to develop AOD and PM models. Three schemes (i.e. G1, A1, and A2) were proposed for AOD modeling according to target AOD data (GOCI AOD and AERONET AOD) and the existence of satellite-derived AOD. The A2 scheme showed the best performance (validation R2 of 0.74 and prediction R2 of 0.73 when GOCI AOD did not exist) and the resultant AOD was used to estimate spatially continuous PM concentrations. The PM models with location information produced successful estimation results with R2 of 0.88 and 0.90, and rRMSE of 26.9 and 27.2% for PM10 and PM2.5, respectively. The spatial distribution maps of PM well captured the seasonal and spatial characteristics of PM reported in the literature, which implies the proposed approaches can be adopted for an operational estimation of spatially continuous AOD and PMs under all sky conditions.

Transglutaminase 2 mediates hypoxia-induced selective mRNA translation via polyamination of 4EBPs.

Hypoxia selectively enhances mRNA translation despite suppressed mammalian target of rapamycin complex 1 activity, contributing to gene expression reprogramming that promotes metastasis and survival of cancer cells. Little is known about how this paradoxical control of translation occurs. Here, we report a new pathway that links hypoxia to selective mRNA translation. Transglutaminase 2 (TG2) is a hypoxia-inducible factor 1-inducible enzyme that alters the activity of substrate proteins by polyamination or crosslinking. Under hypoxic conditions, TG2 polyaminated eukaryotic translation initiation factor 4E (eIF4E)-bound eukaryotic translation initiation factor 4E-binding proteins (4EBPs) at conserved glutamine residues. 4EBP1 polyamination enhances binding affinity for Raptor, thereby increasing phosphorylation of 4EBP1 and cap-dependent translation. Proteomic analyses of newly synthesized proteins in hypoxic cells revealed that TG2 activity preferentially enhanced the translation of a subset of mRNA containing G/C-rich 5'UTRs but not upstream ORF or terminal oligopyrimidine motifs. These results indicate that TG2 is a critical regulator in hypoxia-induced selective mRNA translation and provide a promising molecular target for the treatment of cancers.

Glutathione dynamics determine the therapeutic efficacy of mesenchymal stem cells for graft-versus-host disease via CREB1-NRF2 pathway.

Glutathione (GSH), the most abundant nonprotein thiol functioning as an antioxidant, plays critical roles in maintaining the core functions of mesenchymal stem cells (MSCs), which are used as a cellular immunotherapy for graft-versus-host disease (GVHD). However, the role of GSH dynamics in MSCs remains elusive. Genome-wide gene expression profiling and high-throughput live-cell imaging assays revealed that CREB1 enforced the GSH-recovering capacity (GRC) of MSCs through NRF2 by directly up-regulating NRF2 target genes responsible for GSH synthesis and redox cycling. MSCs with enhanced GSH levels and GRC mediated by CREB1-NRF2 have improved self-renewal, migratory, anti-inflammatory, and T cell suppression capacities. Administration of MSCs overexpressing CREB1-NRF2 target genes alleviated GVHD in a humanized mouse model, resulting in improved survival, decreased weight loss, and reduced histopathologic damages in GVHD target organs. Collectively, these findings demonstrate the molecular and functional importance of the CREB1-NRF2 pathway in maintaining MSC GSH dynamics, determining therapeutic outcomes for GVHD treatment.

Phosphorylation of TFCP2L1 by CDK1 is required for stem cell pluripotency and bladder carcinogenesis.

Molecular programs involved in embryogenesis are frequently upregulated in oncogenic dedifferentiation and metastasis. However, their precise roles and regulatory mechanisms remain elusive. Here, we showed that CDK1 phosphorylation of TFCP2L1, a pluripotency-associated transcription factor, orchestrated pluripotency and cell-cycling in embryonic stem cells (ESCs) and was aberrantly activated in aggressive bladder cancers (BCs). In murine ESCs, the protein interactome and transcription targets of Tfcp2l1 indicated its involvement in cell cycle regulation. Tfcp2l1 was phosphorylated at Thr177 by Cdk1, which affected ESC cell cycle progression, pluripotency, and differentiation. The CDK1-TFCP2L1 pathway was activated in human BC cells, stimulating their proliferation, self-renewal, and invasion. Lack of TFCP2L1 phosphorylation impaired the tumorigenic potency of BC cells in a xenograft model. In patients with BC, high co-expression of TFCP2L1 and CDK1 was associated with unfavorable clinical characteristics including tumor grade, lymphovascular and muscularis propria invasion, and distant metastasis and was an independent prognostic factor for cancer-specific survival. These findings demonstrate the molecular and clinical significance of CDK1-mediated TFCP2L1 phosphorylation in stem cell pluripotency and in the tumorigenic stemness features associated with BC progression.

Ascorbic Acid 2-Glucoside Stably Promotes the Primitiveness of Embryonic and Mesenchymal Stem Cells Through Ten-Eleven Translocation- and cAMP-Responsive Element-Binding Protein-1-Dependent Mechanisms.

Aims: The naive or primitive states of stem cells (SCs) residing in specific niches are unstable and difficult to preserve in vitro. Vitamin C (VitC), in addition to suppressing oxygen radicals, exerts pleiotropic effects to preserve the core functions of SCs. However, this compound is labile and readily oxidized, resulting in cellular toxicity and preventing its reliable application in this context. We found that a VitC derivative, ascorbic acid 2-glucoside (AA2G), stably maintains the naive pluripotency of murine embryonic SCs (mESCs) and the primitiveness of human mesenchymal SCs (hMSCs) without cellular toxicity. Results: The beneficial effects of AA2G and related molecular mechanisms were evaluated in mESCs, induced pluripotent-SCs (iPSCs), and hMSCs. AA2G was stable in aqueous solution and barely induced cellular toxicity in cultured SCs, unlike VitC. AA2G supplementation recapitulated the well-known effects of VitC, including induction of ten-eleven translocation-dependent DNA demethylation in mESCs and suppression of p53 during generation of murine iPSCs. Furthermore, supplementation of hMSCs with AA2G improved therapeutic outcomes in an asthma mouse model by promoting their self-renewal, engraftment, and anti-inflammatory properties. Particularly, activation of the cAMP-responsive element-binding protein-1 (CREB1) pathway contributed to the ability of AA2G to maintain naive pluripotency of mESCs and functionality of hMSCs. Innovation and Conclusion: Given its long-lasting effects and low cellular toxicity, AA2G supplementation is useful to support the naive pluripotency of mESCs and the primitiveness of hMSCs, affecting their developmental potency and therapeutic efficacy. Furthermore, we demonstrate the significance of the CREB1 pathway in the mechanism of action of AA2G.

Efficient, stable solar cells by using inherent bandgap of α-phase formamidinium lead iodide.

In general, mixed cations and anions containing formamidinium (FA), methylammonium (MA), caesium, iodine, and bromine ions are used to stabilize the black α-phase of the FA-based lead triiodide (FAPbI3) in perovskite solar cells. However, additives such as MA, caesium, and bromine widen its bandgap and reduce the thermal stability. We stabilized the α-FAPbI3 phase by doping with methylenediammonium dichloride (MDACl2) and achieved a certified short-circuit current density of between 26.1 and 26.7 milliamperes per square centimeter. With certified power conversion efficiencies (PCEs) of 23.7%, more than 90% of the initial efficiency was maintained after 600 hours of operation with maximum power point tracking under full sunlight illumination in ambient conditions including ultraviolet light. Unencapsulated devices retained more than 90% of their initial PCE even after annealing for 20 hours at 150°C in air and exhibited superior thermal and humidity stability over a control device in which FAPbI3 was stabilized by MAPbBr3.

Induction of detrusor underactivity by extensive vascular endothelial damages of iliac arteries in a rat model and its pathophysiology in the genetic levels.

We tried to establish a reliable detrusor underactivity (DUA) rat model and to investigate pathophysiology of chronic bladder ischemia (CBI) on voiding behavior and bladder function. Adult male rats were divided into five groups. The arterial injury (AI) groups (AI-10, AI-20, AI-30) underwent vascular endothelial damage (VED) of bilateral iliac arteries (with 10, 20, and 30 bilateral repetitions of injury, respectively) and received a 1.25% cholesterol diet. The sham group underwent sham operation and received the same diet. Controls received a regular diet. After 8 weeks, all rats underwent unanesthetized cystometrogram. Bladder tissues were processed for organ bath investigation, immunohistochemistry staining, and genome-wide gene expression analysis. Awake cystometry analysis showed that frequency of voiding contractions and micturition pressure were lower in the AI-30 group than in sham group (p < 0.01). Contractile responses to various stimuli were lower in AI-20 and AI-30 groups (both p < 0.001). In the AI-20 and AI-30 groups, atherosclerotic occlusion in the iliac arteries, tissue inflammation, fibrosis, denervation, and apoptosis of bladder muscle were prominent compared to the sham. Mechanistically, the expression of purinergic receptor P2X-1 was reduced in the AI-30 group, and the genome-wide gene expression analysis revealed that genes related to IL-17 and HIF-1 signaling pathways including INF-γ receptor-1 and C-X-C motif chemokine ligand-2 were upregulated in the CBI-induced DUA rat model. A rat model of progressive VED successfully induced DUA. Abnormal tissue inflammation, fibrosis, denervation, and bladder muscle tissue apoptosis may be involved in CBI-induced DUA pathophysiology.

N-acetylcysteine prevents bladder tissue fibrosis in a lipopolysaccharide-induced cystitis rat model.

Therapeutic options for non-Hunner type interstitial cystitis (IC), which is histologically characterized by fibrosis and mast cell infiltration, are limited. We developed a rat model that replicates chronic inflammation and fibrosis and evaluated the therapeutic effect of N-acetylcysteine (NAC), a well-known anti-fibrotic agent, on the model. Intravesical instillation of lipopolysaccharide (LPS, 750 µg) after protamine sulfate (10 mg) was conducted twice per week for five consecutive weeks. One week after final instillation, 200 mg/kg NAC (n = 10, IC + NAC group) or phosphate-buffered saline (n = 10, IC group) was daily injected intraperitoneally once daily for 5 days. LPS instillation induced bladder fibrosis, mast cell infiltration, and apoptotic tissue damage. Functionally, LPS insult led to irregular micturition, decreased inter-contraction intervals, and decreased micturition volume. NAC significantly improved most of the voiding parameters and reversed histological damages including fibrosis. NAC inhibited the induction and nuclear localization of phospho-Smad2 protein in bladder tissues and the upregulation of genes related to fibrosis, such as Tgfb2, Tgfb3, Smad2, Smad3, Cxcl10, and Card10. This is the first study to demonstrate the beneficial effects on NAC in restoring voiding function, relieving tissue fibrosis and related bladder injuries, in the LPS-induced cystitis rat model.

Analysis by HPLC and LC/MS of pungent piperamides in commercial black, white, green, and red whole and ground peppercorns.

Pepper plants accumulate pungent bioactive alkaloids called piperamides. To facilitate studies in this area, high-performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometry methods were developed and used to measure the following piperamides in 10 commercial whole (peppercorns) and in 10 ground, black, white, green, and red peppers: piperanine, piperdardine, piperine, piperlonguminine, and piperettine. Structural identification of individual compounds in extracts was performed by associating the HPLC peak of each compound with the corresponding mass spectrum. The piperanine content of the peppers (in mg/g piperine equivalents) ranged from 0.3 for the ground white pepper to 1.4 in black peppercorns. The corresponding range for piperdardine was from 0.0 for seven samples to 1.8 in black peppercorns; for four isomeric piperines, from 0.7 for red to 129 in green peppercorns; for piperlonguminine, from 0.0 in red peppercorns to 1.0 in black peppercorns; and for piperyline, from 0.9 in ground black pepper to 5.9 for red peppercorn. Four well-separated stereoisomeric forms of piperettine with the same molecular weight were present in 19 peppers. The sums of the piperamides ranged from 6.6 for red to 153 for green peppercorns. In contrast to large differences in absolute concentrations among the peppers, the ratios of piperines to total piperamide were quite narrow, ranging from 0.76 for black to 0.90 for white peppercorns, with an average value of 0.84 +/- 0.04 ( n = 19). Thus, on average, the total piperamide content of the peppers consists of 84% piperines and 16% other piperamides. These results demonstrate the utility of the described extraction and analytical methods used to determine the wide-ranging individual and total piperamide contents of widely consumed peppers.

Analysis of phenolic compounds by high-performance liquid chromatography and liquid chromatography/mass spectrometry in potato plant flowers, leaves, stems, and tubers and in home-processed potatoes.

Potato plants synthesize phenolic compounds as protection against bruising and injury from bacteria, fungi, viruses, and insects. Because antioxidative phenolic compounds are also reported to participate in enzymatic browning reactions and to exhibit health-promoting effects in humans, a need exists for accurate methods to measure their content in fresh and processed potatoes. To contribute to our knowledge about the levels of phenolic compounds in potatoes, we validated and used high-performance liquid chromatography and liquid chromatography/mass spectrometry to measure levels of chlorogenic acid, a chlorogenic isomer, and caffeic acid in flowers, leaves, stems, and tubers of the potato plant and in home-processed potatoes. The total phenolic acid content of flowers (626 mg/100 g fresh wt) was 21 and 59 times greater than that of leaves and stems, respectively. For all samples, chlorogenic acid and its isomer contributed 96-98% to the total. Total phenolic acid levels (in g/100 g fresh wt) of peels of five potato varieties grown in Korea ranged from 6.5 to 42.1 and of the flesh (pulp) from 0.5 to 16.5, with peel/pulp ratios ranging from 2.6 to 21.1. The total phenolic acid content for 25 American potatoes ranged from 1.0 to 172. The highest amounts were present in red and purple potatoes. Home processing of pulp with various forms of heat induced reductions in the phenolic content. The described methodology should facilitate future studies on the role of potato phenolic compounds in the plant and the diet.

Downregulation of WNT11 is associated with bladder tissue fibrosis in patients with interstitial cystitis/bladder pain syndrome without Hunner lesion.

This study assessed the functional role of WNT genes and the association between WNT signalling cascades and fibrosis in interstitial cystitis/bladder pain syndrome (IC/BPS) patients. Twenty-five patients (3 males, 22 females; mean age 59.7 ± 10.9 years), included 7 non-Hunner-type IC (NHIC), 18 Hunner-type IC (HIC), and 5 non-IC (control) groups. The expression of sonic hedgehog, WNT gene family, and genes previously reported as biomarkers for IC/BPS were examined using RT-PCR in biopsy specimens from the mucosa and submucosa layer of the bladder. WNT2B, WNT5A, WNT10A, and WNT11 functions in the urothelium were evaluated by silencing in an HBlEpC cell line. Pelvic Pain and Urgency/Frequency Patient Symptom Scale scores, O'Leary-Sant Symptom and Problem Index scores, and Visual Analogue Scores did not differ between the NHIC and HIC groups. However, HIC patients had significantly shorter symptom duration (30.9 vs 70.8 months, p = 0.046), higher daily urinary frequency (16.1 versus 8.5 times, p = 0.006), and smaller bladder capacity (208.6 versus 361.4 ml, p = 0.006) than NHIC patients. Overall WNT gene expression was lower in NHIC than HIC patients. Bladder epithelial tissues from HIC patients were characterised by the downregulation of WNT11. Silencing of WNT11, WNT2B, WNT5A, and WNT10A in HBlEpCs resulted in fibrotic changes, indicated by fibrotic morphology, increased fibrosis-related gene expression, and nuclear localisation of phosphorylated SMAD2, and increased vimentin and fibronectin levels. Downregulation of WNT11 results in fibrotic changes of bladder epithelial cells and is associated with the pathogenesis and differential diagnosis of NHIC. Decreased expression of WNT11 is a potential biomarker for predicting NHIC.