Identification of novel dimers and chemical profiling of acid amide alkaloids in Piper nigrum.

Peppers (Piper nigrum L.) are distinguished by their pungent flavor and aroma. Piperine is a major acid-amide alkaloid with a piperidine ring that gives pepper its flavor and scent. In plant metabolomics research, the accessibility of the chemical standards is critical for scientific credibility. We isolated and identified 10 novel dimers of acid amide alkaloids (9-15 and 20-22), along with 12 known monomers (1-6) and dimers (7, 8, 16-19) from black pepper. Subsequently, we found the distribution of monomers and dimers of acid amide alkaloids in black and white peppers by twenty-two acid amide alkaloids which we obtained using the molecular networking technique and multivariate analysis to reveal the molecular relationships between the acid amide alkaloids in black and white peppers. Our research delved into the chemical diversity of acid amide alkaloids in black and white peppers, which could help inform future culinary and potential medicinal utilization of pepper.

Tetramic acid-motif natural products from a marine fungus Tolypocladium cylindrosporum FB06 and their anti-Parkinson activities.

Tetramic acid-containing natural products are attracting significantly increasing attention from biologists and chemists due to their intriguing structures and biological activities. In the present study, two new tetramic acid alkaloids tolypyridone I (1) and tolypyridone J (2), together with five known ones (3-7), were isolated from cultures of a marine fungus Tolypocladium cylindrosporum FB06 isolate obtained from a marine sediment in Beaufort sea of North Alaska. Their structures were elucidated using 1D, 2D NMR, and HRESIMS. Their configurations were established on the basis of 1H coupling constants, ROESY correlations and DP4 calculations. Compound 2 was isolated as mixtures of rotational isomers with C-3 to C-7 axis between 4-hydroxy-2-pyridone and 1-ethyl-3,5-dimethylcyclohexane, hindering rotation. In our unbiased screening to discover neuroprotective compounds in an in vitro Parkinson's disease (PD) model, SH-SY5Y dopaminergic cells were treated with isolated compounds followed by treatment with 1-methyl-4-phenylpyridinium (MPP+), a parkinsonian neurotoxin. Among tested compounds, F-14329 (7) significantly protected cells from MPP+-induced cytotoxicity. MPP+-mediated cell death is known to be related to the regulation of Bcl-2 family proteins, specifically the down-regulation of anti-apoptotic Bcl-2 and the up-regulation of pro-apoptotic Bax levels. Treatment with 2 mmol/L of MPP+ for 24 h significantly reduced Bcl-2 levels compared to control treated with vehicle. However, treatment with F-14329 (7) attenuated such reduction. This study demonstrates that tetramic acid-motif compounds could be potential lead compounds for treating PD. Supplementary Information: The online version contains supplementary material available at 10.1007/s42995-023-00198-7.

Effects of Poly-L-Lactic Acid on Adipogenesis and Collagen Gene Expression in Cultured Adipocytes Irradiated with Ultraviolet B Rays.

BACKGROUND: Poly-L-lactic acid (PLLA), a synthetic, biocompatible, and biodegradable polymer, has been safely used in several clinical applications. Recently, PLLA has been widely used in the field of dermatology to treat wrinkles in aging skin. Reportedly, PLLA directly acts on dermal fibroblasts causing a significant increase in the expression of type I collagen. However, little is known about the effect of PLLA on adipocytes. OBJECTIVE: This study aimed to analyze the effect of PLLA on adipocytes and examine its potential in treating deep wrinkles engendered by the loss of subcutaneous fat because of aging and photoaging. METHODS: To elucidate the effect of PLLA on skin photoaging, cultured 3T3-L1 adipocytes were irradiated with ultraviolet B (UVB) rays. Oil red O staining was used to detect lipid accumulation in the adipocytes. Real-time quantitative polymerase chain reaction and Western blotting were performed to detect types IV and VI collagen mRNA and protein levels, respectively, under different conditions. RESULTS: The differentiation of 3T3-L1 cells enhanced adipogenesis and the expression of types IV and VI collagens, both of which were inhibited by UVB irradiation. Following this irradiation, PLLA stimulated adipogenesis and the expression of types IV and VI collagens. CONCLUSION: PLLA may provide the beneficial effect on adipocytes from the aspect of adipogenesis and collagen expression in the subcutaneous adipose tissues.

On-site applicable diagnostic fluorescent probe for fire blight bacteria.

Fire blight is a representative plant infection that contaminates edible plants and causes socio-economic problems in agricultural and livestock industries globally. It is caused by the pathogen Erwinia amylovora (E. amylovora) creates lethal plant necrosis and spreads rapidly across plant organs. We newly disclose the fluorogenic probe B-1 for real-time on-site detection of fire blight bacteria for the first time. B-1 exhibited no emission signals but manifested bright emission properties in the presence of fire blight bacteria. Based on these features, fluorescence imaging of the fire blight bacteria and its real-time detection from the infected host plant tissues were conducted. The detection limit against E. amylovora was 102 CFU/mL, which had excellent sensitivity. The fluorogenic probe-based on-site diagnostic technology was supplemented by introducing a new portable UV device. This work holds enormous potential to be a new advanced tool for detecting fire blight in agricultural and livestock industries.

Real-time and label-free biosensing using moiré pattern generated by bioresponsive hydrogel.

Bioresponsive hydrogels are smart materials that respond to various external stimuli and exhibit great potential as biosensors owing to their capability of real-time and label-free detection. Here, we propose a sensing platform based on bioresponsive hydrogels, employing the concept of moiré patterns. Two sets of line patterns with different pitch sizes are prepared; a hydrogel grating whose pitch size changes according to external stimuli and a reference grating with constant pitch size. The volume changes of the hydrogel caused by external stimuli changes the pitch size of the hydrogel grating, and subsequently, the pitch sizes of the moiré patterns (moiré signal), whose values can be obtained in a real-time and label-free manner through customized moiré microscopy and signal processing. After confirming that the pH-induced swelling of hydrogel could be monitored using moiré patterns, we performed moiré pattern-based detection of specific proteins using protein-responsive hydrogel that underwent shrinking via interaction with target proteins. Brain-derived neurotrophic factor and platelet-derived growth factor were selected as the model proteins, and our proposed system successfully detected both proteins at nanomolar levels. In both cases, the pitch size change of hydrogel grating was monitored much more sensitively using moiré patterns than through direct measurements. The changes in the moiré signals caused by target proteins were detected in ex-vivo environments using a custom-made intraocular lens incorporating the hydrogel grating, demonstrating the capability of the proposed system to detect various markers in intraocular aqueous humor, when implanted in the eye.

Porous Silicon-Based Nanomedicine for Simultaneous Management of Joint Inflammation and Bone Erosion in Rheumatoid Arthritis.

The lack of drugs that target both disease progression and tissue preservation makes it difficult to effectively manage rheumatoid arthritis (RA). Here, we report a porous silicon-based nanomedicine that efficiently delivers an antirheumatic drug to inflamed synovium while degrading into bone-remodeling products. Methotrexate (MTX) is loaded into the porous silicon nanoparticles using a calcium silicate based condenser chemistry. The calcium silicate-porous silicon nanoparticle constructs (pCaSiNPs) degrade and release the drug preferentially in an inflammatory environment. The biodegradation products of the pCaSiNP drug carrier are orthosilicic acid and calcium ions, which exhibit immunomodulatory and antiresorptive effects. In a mouse model of collagen-induced arthritis, systemically administered MTX-loaded pCaSiNPs accumulate in the inflamed joints and ameliorate the progression of RA at both early and established stages of the disease. The disease state readouts show that the combination is more effective than the monotherapies.

Development of a fluorescent nanoprobe based on an amphiphilic single-benzene-based fluorophore for lipid droplet detection and its practical applications.

Lipid droplets (LDs) are crucial biological organelles connected with metabolic pathways in biological systems and diseases. To monitor the locations and accumulation of LDs in lipid-related diseases, the development of a visualization tool for LDs has gained importance. In particular, LD visualization using fluorescent probes has gained attention. Herein, a new fluorescent nanoprobe, BMeS-Ali, is developed that can sense LDs based on an amphiphilic single benzene-based fluorophore (SBBF). BMeS-Ali consists of hydrophilic (-NH2) and hydrophobic (-C12H25) moieties and exists as a micelle nanostructure in aqueous media. BMeS-Ali has a weak fluorescence, but its emission was dramatically enhanced upon exposure to the LD components such as oleic acids (OA) by reassembling its nano-formulation. BMeS-Ali showed a selective LD staining ability and great biocompatibility in cells (cancer cells and stem cells). It also showed a practical sensing ability towards biologically derived lipids and can be applied to the visualization of human fingerprints. We found that the nanoprobe BMeS-Ali has significant potential to serve as a practical dye and sensor for lipids, especially for LD imaging in the biomedical research area and broader industrial applications.

Recent advances in hybrid system of porous silicon nanoparticles and biocompatible polymers for biomedical applications.

Hybrid systems of nanoparticles and polymers have emerged as a new material in the biomedical field. To date, various kinds of hybrid systems have been introduced and applied to drug delivery, regenerative medicine, therapeutics, disease diagnosis, and medical implantation. Among them, the hybridization of nanostructured porous silicon nanoparticles (pSiNPs) and biocompatible polymers has been highlighted due to its unique biological and physicochemical properties. This review focuses on the recent advances in the hybrid systems of pSiNPs and biocompatible polymers from an engineering aspect and its biomedical applications. Representative hybrid formulations, (i) Polymer-coated pSiNPs, (ii) pSiNPs-embedded polymeric nanofibers, are outlined along with their preparation methods, biomedical applications, and future perspectives. We believe this review provides insight into a new hybrid system of pSiNPs and biocompatible polymers as a promising nano-platform for further biomedical applications. Recently developed and representative hybrid systems of porous silicon nanoparticles and biocompatible polymers and their biomedical applications are introduced.

Red-Emitting SBBF (Single-Benzene-Based Fluorophore)-Silica Hybrid Material: One-Pot Synthesis, Characterization, and Biomedical Applications.

We report, for the first time, a new red-emitting hybrid material based on a single-benzene-based fluorophore (SBBF) and silica. This robust formulation shows several features, including bright emissions at a red wavelength (>600 nm), high scalability (>gram-scale), facile synthesis (one-pot reaction; SBBF formation, hydrolytic condensation, propagation), high stability (under different humidity, pH, light), bio-imaging applicability with low cellular toxicity, and an antibacterial effect within Gram-negative/Gram-positive strains. Based on our findings, we believe that these hybrid materials can pave the way for the further development of dye-hybrid materials and applications in various fields.

A metastasis suppressor Pt-dendrimer nanozyme for the alleviation of glioblastoma.

Nanozymes are nanostructure-based materials which mimic the enzymatic characteristics of natural enzymes. Biological applications of nanozymes have been highlighted in basic research, industry, and translational medicine as a new cutting-edge tool. In this work, and for the first time, we disclose a tumor alleviation property of a nanozyme that is made up of amine-terminated sixth-generation polyamidoamine dendrimers with encapsulated tiny platinum nanoparticles. We systematically conducted the synthesis and characterization of the dendrimer-encapsulated Pt nanoparticles (denoted Pt-dendrimer) and confirmed their enzymatic function (hydrogen peroxide (H2O2) decomposition) within various cell lines (normal, cancerous), including glioblastoma (GBM) cells. By understanding the effects of the Pt-dendrimer at the gene level, especially related to cancer cell metastasis, we have thoroughly demonstrated its ability for tumor alleviation and suppressing GBM migration, invasion, and adhesion. The present findings show great promise for the application of the nanozyme for use in GBM-related basic research as well as at clinical sites.

Membrane-Targeting Triphenylphosphonium Functionalized Ciprofloxacin for Methicillin-Resistant Staphylococcus aureus (MRSA).

Multidrug-resistant (MDR) bacteria have become a severe problem for public health. Developing new antibiotics for MDR bacteria is difficult, from inception to the clinically approved stage. Here, we have used a new approach, modification of an antibiotic, ciprofloxacin (CFX), with triphenylphosphonium (TPP, PPh3) moiety via ester- (CFX-ester-PPh3) and amide-coupling (CFX-amide-PPh3) to target bacterial membranes. In this study, we have evaluated the antibacterial activities of CFX and its derivatives against 16 species of bacteria, including MDR bacteria, using minimum inhibitory concentration (MIC) assay, morphological monitoring, and expression of resistance-related genes. TPP-conjugated CFX, CFX-ester-PPh3, and CFX-amide-PPh3 showed significantly improved antibacterial activity against Gram-positive bacteria, Staphylococcus aureus, including MDR S. aureus (methicillin-resistant S. aureus (MRSA)) strains. The MRSA ST5 5016 strain showed high antibacterial activity, with MIC values of 11.12 µg/mL for CFX-ester-PPh3 and 2.78 µg/mL for CFX-amide-PPh3. The CFX derivatives inhibited biofilm formation in MRSA by more than 74.9% of CFX-amide-PPh3. In the sub-MIC, CFX derivatives induced significant morphological changes in MRSA, including irregular deformation and membrane disruption, accompanied by a decrease in the level of resistance-related gene expression. With these promising results, this method is very likely to combat MDR bacteria through a simple TPP moiety modification of known antibiotics, which can be readily prepared at clinical sites.

Penta-fluorophenol: a Smiles rearrangement-inspired cysteine-selective fluorescent probe for imaging of human glioblastoma.

Two of the most critical factors for the survival of glioblastoma (GBM) patients are precision diagnosis and the tracking of treatment progress. At the moment, various sophisticated and specific diagnostic procedures are being used, but there are relatively few simple diagnosis methods. This work introduces a sensing probe based on a turn-on type fluorescence response that can measure the cysteine (Cys) level, which is recognized as a new biomarker of GBM, in human-derived cells and within on-site human clinical biopsy samples. The Cys-initiated chemical reactions of the probe cause a significant fluorescence response with high selectivity, high sensitivity, a fast response time, and a two-photon excitable excitation pathway, which allows the imaging of GBM in both mouse models and human tissue samples. The probe can distinguish the GBM cells and disease sites in clinical samples from individual patients. Besides, the probe has no short or long-term toxicity and immune response. The present findings hold promise for application of the probe to a relatively simple and straightforward following of GBM at clinical sites.

Visualizing mitochondria and mouse intestine with a fluorescent complex of a naphthalene-based dipolar dye and serum albumin.

We have explored a new research field of fluorophores through the manipulation of fluorophore-binding proteins. The development of a new imaging agent for tracing a specific organelle or a particular site within a living organism has been of great interest in the field of basic science as well as translational medicine. In this work and for the first time, we will disclose a new naphthalene-based dipolar dye and its complex, with serum albumin (SA), and show their applicability for the selective imaging of mitochondria in cells and the intestine in a mouse. The SA-binding dipolar dye, IPNHC, was synthesized straightforwardly, and we identified its photophysical properties and binding mode with SA. IPNHC-SA complex showed a bright emission in the blue wavelength range with a high quantum yield and stability. In the fluorescence imaging study, bright fluorescence images of mouse intestines were observed under a UV light, as well as two-photon (TP) deep tissue imaging after intravenous injection of IPNHC and IPNHC-SA complex. The present findings hold great promise for the application of the fluorescent complex for use in the tracing and tracking of intestine-related diseases at clinical sites.

Thermoresponsive poly(N-isopropylacrylamide) hydrogel substrates micropatterned with poly(ethylene glycol) hydrogel for adipose mesenchymal stem cell spheroid formation and retrieval.

Cell spheroid formation is necessary to develop three-dimensional (3D) cellular environments that provide appropriate cell-cell and cell-matrix interactions similar to in vivo environments without additional substrates. Although some methods including stirring culture, low adhesion plate culture, hanging drop, and microfluidics are used to construct cell spheroids, there is no method to fulfill all of the mass production of uniform spheroids, simple media change, and easy retrievability. Here, bulk poly(N-isopropylacrylamide) (PNIPAAm) hydrogel substrate (PHS) was used to fabricate, culture, and retrieve cell spheroids. Adipose-derived stem cells (ASCs) were cultured on bulk PHS to form spheroids. ASCs formed cell spheroids directly on substrates without additional manipulation. These spheroids adhered to the semi-adhesive substrate, while the spheroids fabricated using the nonadhesive surface method floated without getting fixed to the surface. Bulk PHS stiffness was evaluated using the compressive test (compressive modulus: 153 ± 11 kPa). A poly(ethylene glycol) (PEG) hydrogel microwell pattern was created on PHS to control the spheroid size, forming uniform ASC spheroids between 100 and 150 µm in diameter on 200 and 300 µm well-patterned substrates. Cell-cell interactions in the resulting ASC spheroids were evaluated based on fibronectin and laminin expression; fluorescence intensities of fibronectin- and laminin-immunostained images of ASC spheroids were 10.9 and 7.3 times higher than those of ASCs cultured on the tissue culture plate, respectively. ASC spheroids were detached following incubation at 4 °C for 10 min (retrieval efficiency: 74 ± 19%). Retrieved spheroid cell viability was over 97.5%. The PEG hydrogel microwell-patterned PHS is a convenient spheroid fabrication and retrieval platform that can increase cell spheroid usage.

Argonaute system of Kordia jejudonensis is a heterodimeric nucleic acid-guided nuclease.

We purified and characterized a prokaryotic argonaute (pAgo) (KjMP) and its associated protein (KjAA) from a bacterium Kordia jejudonensis. The two proteins present as a complex were revealed by the copurification of KjAA with His-tagged KjMP by Ni-NTA affinity column. The KjAA/KjMP complex was a heterodimer evaluated from the molecular weight estimated using size exclusion chromatography. The pAgo complex presented a guide-dependent target DNA cleavage. RNA was the preferred guide; however, DNA also functioned, albeit weakly. Additionally, 5'-phosphorylate or non-phosphorylated guide was equally effective. The purified complex exhibited nonspecific nuclease activity on dsDNA and ssDNA. This is the first study to report that short pAgo and its associated protein form a complex, which has a nucleic acid-guided target recognition and cleavage.

Ultraviolet B Downregulated Aquaporin 1 Expression via the MEK/ERK pathway in the Dermal Fibroblasts.

BACKGROUND: Aquaporin 1 (AQP1) is a transmembrane channel protein that allows rapid transposition of water and gases, in recent discoveries of AQP1 function involve cell proliferation, differentiation, wound healing, inflammation and infection in different cell types, suggesting that AQP1 plays key roles in diverse biologic process. Until now, less is known about the function of AQP1 on ultraviolet radiation induced photoaged skin. OBJECTIVE: In this study we set out to examine whether AQP1 expression may be influenced by repeated irradiation of ultraviolet B (UVB) in cultured dermal fibroblasts. METHODS: To elucidate the function of AQP1 in skin photoaging, human dermal fibroblasts (HS68) were irradiated by a series of 4 sub-cytotoxic doses of UVB which are known as UV-induced cell premature senescence model. Reverse transcription polymerase chain reaction and Western blotting were conducted to detect AQP1 expression from different groups. Then, cells were transfected with AQP1-targeting small interfering RNA. The activities of signaling proteins upon UVB irradiation were investigated to determine which pathways are involved in AQP1 expression. RESULTS: AQP1 expression was increased by 100 mJ/cm2 of UVB irradiation, but decreased by 200 mJ/cm2. Depletion of the AQP1 increased the apoptotic sensitivity of cells to UVB, as judged by upregulation of the p53, p21, poly (adenosine diphosphate [ADP]-ribose) polymerase and Bax together with the increased Bax/Bcl2 ratio. UVB induced downregulation of AQP1 was significantly attenuated by pretreatment with the MEK/ERK inhibitor (PD98059). CONCLUSION: We concluded that AQP1 expression was down-regulated by repeated exposure of UVB via MEK/ERK activation pathways. The AQP1 reduction by UVB lead to changes of physiological functions in dermal fibroblasts, which might be associated with the occurrence and development of UVB induced photoaging.

Hybrid Composite of Silver Nanoparticle-Porous Silicon Microparticles as an Image-Guided Localization Agent for Computed Tomography Scan of the Lungs.

A hybrid composite of silver nanoparticles (AgNPs) and porous silicon microparticles (pSiMPs) was developed and applied for the computed tomography (CT) scanning of the lungs as an image-guided localization agent. We confirmed the grafting of AgNPs on oxidized pSiMPs template using various analytical equipment, including a scanning electron microscope (SEM), Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDS). The hybrid composite showed a high CT contrast intensity (>1000 HU) that enabled us to produce and view images of the lungs. In addition, it showed the ability to maintain a strong CT signal at the injected area of the rabbit's lungs, up to an hour, without spreading. The lack of toxicity and immune response indicated that the composite could be fully utilized as a new image-guided localization agent of CT scans for lung cancer surgery.

Latent turn-on fluorescent probe for the detection of toxic malononitrile in water and its practical applications.

A latent turn-on fluorescent probe for the detection of malononitrile (NCCH2CN), a precursor of hydrogen cyanide (HCN) in the mammalian tissue metabolism, is developed based on reaction-based fluorophore generation for the first time. Malononitrile is utilized within a wide spectrum of academic and industrial applications, and it is a key reagent to make o-chlorobenzylidene malononitrile (CS gas; tear gas), which is used for riot control. Due to its extensive use as well as potential health risks and the environmental pollution, malononitrile monitoring method has been required. In this paper, we discovered that our key sensing platform, 6-(dimethylamino)-3-hydroxy-2-naphthaldehyde (named Mal-P1), responds sensitively and selectively towards malononitrile. The Knoevenagel condensation induced benzo [g]coumarin formation of Mal-P1 with malononitrile showed significant fluorescence turn-on response. In addition, Mal-P1 showed the malononitrile sensing ability in environmental samples (real water, CS gas) and imaging ability in biological sample (HeLa cell line) using fluorescence microscopy with low cytotoxicity. The successful demonstrations will facilitate further applications in a variety of fields.

Hydrazine-Selective Fluorescent Turn-On Probe Based on Ortho-Methoxy-Methyl-Ether (o-MOM) Assisted Retro-aza-Henry Type Reaction.

Hydrazine (N2H4) is one of the most widely used industrial chemicals that can be utilized as a precursor of pesticides, pharmaceutics, and rocket propellant. Due to its biological and environmental toxicity with potential health risks, various sensing tools have been developed. Among them, fluorescence-based molecular sensing systems have been highlighted due to its simple-operation, high selectivity and sensitivity, and biocompatibility. In our recent report, we disclosed a ratiometric type fluorescent probe, called HyP-1, for the detection of hydrazine, which is based on ortho-methoxy-methyl-ether (o-MOM) moiety assisted hydrazone-formation of the donor (D)-acceptor (A) type naphthaldehyde backbone. As our follow-up research, we disclose a turn-on type fluorescent probe, named HyP-2, as the next-generation hydrazine probe. The sensing rational of HyP-2 is based on the o-MOM assisted retro-aza-Henry type reaction. The dicyanovinyl moiety, commonly known as a molecular rotor, causes significant emission quenching of a fluorescent platform in aqueous media, and its cleavage with hydrazone-formation, which induces a significant fluorescence enhancement. The high selectivity and sensitivity of HyP-2 shows practical explicabilities, including real-time paper strip assay, vapor test, soil analysis, and real water assay. We believe its successful demonstrations suggest further applications into a wide variety of fields.

Activity of hippocampal adult-born neurons regulates alcohol withdrawal seizures.

Alcohol withdrawal (AW) after chronic alcohol exposure produces a series of symptoms, with AW-associated seizures being among the most serious and dangerous. However, the mechanism underlying AW seizures has yet to be established. In our mouse model, a sudden AW produced 2 waves of seizures: the first wave includes a surge of multiple seizures that occurs within hours to days of AW, and the second wave consists of sustained expression of epileptiform spikes and wave discharges (SWDs) during a protracted period of abstinence. We revealed that the structural and functional adaptations in newborn dentate granule cells (DGCs) in the hippocampus underlie the second wave of seizures but not the first wave. While the general morphology of newborn DGCs remained unchanged, AW increased the dendritic spine density of newborn DGCs, suggesting that AW induced synaptic connectivity of newborn DGCs with excitatory afferent neurons and enhanced excitability of newborn DGCs. Indeed, specific activation and suppression of newborn DGCs by the chemogenetic DREADD method increased and decreased the expression of epileptiform SWDs, respectively, during abstinence. Thus, our study unveiled that the pathological plasticity of hippocampal newborn DGCs underlies AW seizures during a protracted period of abstinence, providing critical insight into hippocampal neural circuits as a foundation to understand and treat AW seizures.