Ficus hirta Vahl. fruit was studied herein with the aim to find the potential mechanism for significant inhibition of growth of Penicillium digitatum, a causative pathogen of citrus green mold disease. PiCB substantially inhibited mycelial growth of P. digitatum, with the observed half maximal effective concentration (EC50), minimum inhibitory concentration (MIC), and minimum fungicidal concentration (MFC) of 120.3, 200, and 400 mg/L, respectively. Moreover, PiCB altered hyphal morphology and cellular morphology by breaking and shrinking of mycelia, decomposing cell walls, cytoplasmic inclusions. In addition to, a non-targeted metabolomics analysis by UHPLC-Q-TOF/MS was also performed, which revealed that PiCB treatment notably disrupted the metabolisms of amino acids, lipids, fatty acids, TCA, and ribonucleic acids, thereby contributing to membrane peroxidation. Current findings provide a new perception into the antifungal mechanism of PiCB treatment in inhibiting P. digitatum growth through membrane peroxidation.Pinocembroside (PiCB) isolated from  Penicillium spp., Geotrichum citri-aurantii, Alternaria alternata, and Diaporthe citri being the prevalent postharvest pathogenic fungi in citrus fruit [P. digitatum, the causal agent of green mold, is considered one of the most threatening postharvest diseases in citrus fruit. The P. digitatum infection on citrus fruit has posed a series of adverse effects including toxins accumulation, postharvest deterioration, human health, environmental pollution, and 60–80% of postharvest crop loss in China [Citrus fruit is a major economic crop with a diverse range of species and varieties, including oranges, mandarins, tangerines, pummelos, lemons, citrons, kumquats, limes, and different hybrids ,2. Posthus fruit ,4,5,6. Ain China ,8. To rein China ,10. Howein China ,12. KeepF. hirta Vahl., Penthorum chinense Pursh, and Viscum articulatum [F. hirta Vahl. fruits, which is a widely used traditional medicine and a popular ingredient in soups in southern China [P. italicum in PDA media, which depicted that the drug successfully inhibits the mycelial growth of P. italicum [During the last decades, the use of natural antifungal components has been a novel alternative for the traditional synthetic fungicides used to reduce postharvest fungal rotting of citrus and other fresh fruits. In keeping with this global trend, our lab has been exploring novel antifungal phytochemicals that help reduce disease incidence and prolong the storage life of citrus fruit . Among viculatum ,15,16. Prn China ,17,18. Rrn China . PiCB shitalicum .P. digitatum in citrus fruit. In order to depict the changes in hyphal morphology and metabolism level in P. digitatum treated with PiCB, we used microscopy and UHPLC-Q-TOF/MS-based non-targeted metabolomics. The current study aimed to find the differential metabolites of P. digitatum response to PiCB, extending to explore the potential mechanism of PiCB on antifungal effects of P. digitatum.However, there is hardly any authentic and detailed study regarding the antifungal activity of PiCB against P. digitatum showed interesting results  of the P. digitatum mycelial growth. Furthermore, by calculating the MGI under different PiCB concentrations, a linear regression of the MGI of P. digitatum (Y) on the log-transformed PiCB concentrations (X) was determined  (50) of PiCB against P. digitatum was 120.3 mg/L, which indicated the mycelial growth of P. digitatum was inhibited effectively at low PiCB concentration. In the current study, the MGI of P. digitatum was obviously improved by increasing the PiCB concentrations and incubation days  . Accordiion days . Similaruarters 7.03% MGI igitatum ,22.P. digitatum mycelial growth on PDA medium incubated at 25 °C having various PiCB concentrations , 200 and 400 mg/L treatments completely inhibited P. digitatum mycelial growth on second and sixth day of incubation, respectively  in the mycelial weights (wet weight as well dry weight) of P. digitatum followed treatment with a lower concentration (12.5 mg/L) of PiCB.The effects of PiCB on mycelial weights and water-retention rate of abulated . Higher abulated . MoreoveP. digitatum (p < 0.05). Moreover, the water-retention rate decreased with increasing PiCB concentration from 25 to 400 mg/L, but 200 mg/L concentration keep 83.4% water-retention rate and is close enough to the 400 mg/L PiCB-treated group. This data is in conformity with our previous reports where carvacrol showed positive effects in the MGI of P. digitatum [P. digitatum caused by PiCB, as evidenced by a decrease in water-retention rate, was accompanied by a marked breakage and leakage of the cell membranes  and transmission electron microscope (TEM), respectively. The SEM images revealed that the hyphal morphology of the control mycelia was normal with a uniform thickness and smooth surface  exhibited notable shrinking, thinned cell walls, an insufficient cytoplasmic matrix, and reduced cell permeability  and induce aggravation of cell permeabilization [TEM images clearly differentiate between the PiCB-treated and untreated um cells C, D. Thelization ,31,32.P. digitatum mycelia exposed to PiCB at 1/2 MIC (100 mg/L), A volcano plot of all metabolite ions is shown in P. digitatum mycelia. These metabolites were significantly changed in the relative abundance in the PiCB-treated mycelia compared with the control group. Among these metabolites, 23 amino acids, 13 lipids, 10 fatty acids, 3 TCA metabolites, 4 glycolysis/gluconeogenesis metabolites, 3 galactose metabolites, 6 carbohydrate metabolites, 4 nucleic acids, 3 amino sugars and nucleotide sugars, 6 purines, 3 vitamins, and 7 miscellaneous metabolites were observed. Among these differential metabolites, 56 metabolites were up-regulated while 29 metabolites were down-regulated. Interestingly, 18 differential metabolites  were observed in both positive and negative modes  . In the igitatum A. Similaigitatum B. On theluster 2 B.P. digitatum, and their metabolic pathways may be targeted to check the fungal growth [P. digitatum mycelia, 18 amino acids were increased, while five amino acids were significantly decreased in PiCB-treated samples compared with the respective controls  cycle, purines, starch and sucrose were observed in both PiCB-treated and control mycelia . Amino al growth ,34. Amoncontrols . The conP. digitatum mycelia was associated with the increased levels of various acids  and the decreases in contents of linolenic, linoleic, and 16-hydroxypalmitic acids  was purchased from China General Microbiological Culture Collection Center , and embedded in 30% glycerol at –80 °C for its conservation. A pure culture of P. digitatum was activated on potato dextrose agar  medium at 25 ± 1 °C. After 7 d of culture, P. digitatum spore suspensions was collected, filtered, and adjusted to the desired concentration of 1.0 × 106 CFU/mL with the aid of a Countess II FL automatic cell counter .P. digitatum with some modification. Briefly, the stock solution of PiCB was diluted with moderate amount of sterile 0.5% Tween 80 and mixed with PDA for obtaining the final concentrations of 0, 12.5, 25, 50, 100, 200, and 400 mg/L. A total of 15 μL of P. digitatum spore suspensions was injected into the Oxford cup (8 mm in diameter) placed in the center of each petri dish (90 mm in diameter). Finally, all plates were incubated at 25 °C for 7 d and measured the colony diameters of PiCB-treated and control groups using the cross method. Four replicates were used per treatment and the experiment was carried out at two separate times. The following formula was used for calculating the mycelial growth inhibition (MGI): The method of Chen et al.  was emplY = 0.3396X + 9.1243, R2 = 0.9682 (50) value of PiCB against P. digitatum was 120.3 mg/L calculated based on the above linear regression.The linear regression of the MGI under different PiCB treated concentrations (12.5–200 mg/L) was determined: = 0.9682 . The halThe MIC and MFC were defined as the lowest concentration of PiCB with no visible fungal growth on a PDA plate after the second and sixth day of incubation at 25 °C, respectively .P. digitatum. The water-retention rate was calculated using the following equation: The mycelial wet and dry weights of from PiCB-treated and control samples were determined as described by Wan et al. , and theP. digitatum were observed using both scanning electron microscope (SEM) and transmission electron microscope (TEM). PiCB treatment and sample preparation were followed the method previously reported by Xin et al. [The changes of PiCB on morphology and ultra-structure of n et al. . FinallyP. digitatum spores was added to 150 mL PDB and incubated in an incubator shaker  at 160 rpm for 48 h at 25 °C. For intracellular metabolite samples, about 2.0 g of P. digitatum mycelia was collected and washed three times with sterile double-distilled water and re-suspended in a sterilized conical flask containing PiCB solution at 100 mg/L (1/2 MIC) at 160 rpm, and the mycelia treated without PiCB was used as control. The PiCB-treated and control mycelia were obtained after shaking culture of 12 h at 25 °C, placed in a vacuum freeze drier  for 24 h, and kept at −80 °C for metabolomics analysis.30 μL of v/v). After homogenizing for 60 s, the solution was ultrasonic sonic disrupted for 30 min twice and placed in −20 °C condition for 1 h. The supernate collected after centrifugation  was freeze-dried and stored at −80 °C for next detection.The freeze-dried samples (80 mg) was mixed with 20 μL of ultrapure water and homogenized for 60 s on a FastPrep-24 tissue homogenizer , and added 80 μL of methanol/acetonitrile  under the high sensitivity mode. The declustering potential (DP) was ± 60 V (positive and negative modes), and collision energy was 35 ± 15 eV.The analytes were separated using a hydrophilic interaction liquid chromatography (HILIC) column (100 mm × 3.1 mm × 1.7 μm) and detected by means of an Agilent 1290 Infinity LC Ultra-high pressure liquid chromatography  system, which was coupled to a quadrupole time-of-flight  mass spectrometer equipped with an electron spray ionization as positive (ESI+) and negative (ESI–) modes. The ESI source conditions were: Ion Source Gas1 (Gas1) as 60, Ion Source Gas2 (Gas2) as 60, curtain gas (CUR) as 30, source temperature: 600 °C, Ion Spray Voltage Floating (ISVF) ± 5500 V. The The raw data was converted to MzXML format by mean of ProteoWizard and conducted using R package XCMS. The Metabolites constructions were identified by matching to the molecular weight (<25 ppm) and secondary spectrum diagram in our standards database.Ficus hirta Vahl. fruit was demonstrated to exert a significant inhibitory effect on P. digitatum mycelial growth, and its antifungal mechanisms of membrane peroxidation might be linked with the disruption of the amino acid metabolism, lipid metabolism, fatty acid metabolism, TCA cycle, and purine metabolism. The study displays the potential of PiCB as a novel antifungal agent for controlling postharvest fungal decay in citrus fruit and emerges as a promising candidate for citrus preservatives.In summary, a flavonone PiCB isolated from


This effect of the hormone is mediated by the cellular Ca2+ signaling pathway: a sustained increase of intracellular (cytosolic) Ca2+ concentration followed by activation of Ca2+-dependent initiators and effectors of apoptosis. In recent animal studies, we demonstrated that low vitamin D status is observed in diet-induced obesity (DIO). High intake of vitamin D3 in DIO decreased the weight of white adipose tissue and improved biomarkers related to adiposity and Ca2+ regulation. The anti-obesity effect of vitamin D 2D3) in DIO was determined by the induction of Ca2+-mediated apoptosis in mature adipocytes executed by Ca2+-dependent apoptotic proteases . Thus, a high intake of vitamin D in obesity increases vitamin D nutritional status and normalizes vitamin D hormonal status that is accompanied by the reduction of adiposity. Overall, our findings imply that vitamin D may contribute to the prevention of obesity and obesity-related diseases and that the mechanism of the anti-obesity effect of 1,25(OH)2D3 includes induction of Ca2+-mediated apoptosis in adipocytes.The role of vitamin D in obesity appears to be linked to vitamin D insufficient/deficient status. However, mechanistic understanding of the role of vitamin D in obesity is lacking. We have shown earlier that the vitamin D hormonal form, 1,25-dihydroxyvitamin D The critical characteristic of the apoptotic Ca2+ signal is a sustained, prolong, globalized, and nonoscillatory increase in [Ca2+]i, reaching elevated, but not cytotoxic, levels. The mechanism of action of intracellular Ca2+ in apoptotic pathways involves interactions of the cellular Ca2+ signals with Ca2+-dependent molecular targets in cells undergoing apoptosis. The Ca2+-dependent calpains and caspases appear to be the primary Ca2+-activated apoptotic effectors i rapidly signals the cell to enter the apoptotic pathway. The activation of Ca2+-dependent protease µ-calpain followed by the activation of Ca2+/calpain-dependent caspase-12 and other effector caspases  is responsible for the execution of Ca2+-mediated apoptosis. A lack of expression or low levels of cytosolic Ca2+ binding proteins  decrease Ca2+-buffering capacity of the cell, resulting in [Ca2+]i increase and the induction of apoptosis i, and this increase led to the execution of apoptosis via activation of µ-calpain and caspase-12. The susceptibility of mature adipocytes to the Ca2+-elevating effect of 1,25(OH)2D3 appears linked to the low Ca2+-buffering capacity of these cells. These findings demonstrate that Ca2+-mediated apoptosis can be induced in mature adipocytes and that the apoptotic molecular targets activated by 1,25(OH)2D3 in these cells are Ca2+-dependent µ-calpain and caspase-12.We have shown that 1,25(OH)spase-12 ,15,16. T3 content demonstrate a decreased weight of adipose tissue and improved biomarkers of adiposity and vitamin D status i reaching elevating (200–500 nM), but not cytotoxic (>1 μM) levels—via Ca2+ release from the ER stores and Ca2+ entry through the VICC. μ-Calpain activation by the sustained cytosolic Ca2+ signal is followed by activation  of Ca2+/calpain-dependent caspase-12. Activation of these proteases appears to be sufficient for the execution of apoptosis as evaluated by the hallmarks of this process: apoptotic changes of the plasma membrane (phosphatidylserine translocation from the inner to outer surface of the plasma membrane) and nuclear condensation and DNA fragmentation .The cartoon provides a schematic representation of the possible mechanism of action of 1,25(OH)


Halide perovskite quantum dots (PQDs) are promising materials for diverse applications including displays, light‐emitting diodes, and solar cells due to their intriguing properties such as tunable bandgap, high photoluminescence quantum yield, high absorbance, and narrow emission peaks. Despite the prosperous achievements over the past several years, PQDs face severe challenges in terms of stability under different circumstances. Currently, researchers have overcome part of the stability problem, making PQDs sustainable in water, oxygen, and polar solvents for long‐term use. However, halide PQDs are easily degraded under continuous irradiation, which significantly limits their potential for conventional applications. In this study, an oleic acid/oleylamine ‐free method to fabricate perovskite quantum dot papers (PQDP) is developed by adding cellulose nanocrystals as long‐chain binding ligands that stabilize the PQD structure. As a result, the relative photoluminescence intensity of PQDP remains over ≈90% under continuous ultraviolet  irradiation for 2 months, showing negligible photodegradation. This proposed method paves the way for the fabrication of ultrastable PQDs and the future development of related applications. Solid‐state perovskite quantum dot papers are fabricated using a unique vacuum filtration growth method without a purification process. The bonding between cellulose nanocrystals and perovskite quantum dots makes the hybrid structure stable, and record high UV stability and thermal stability are achieved for perovskite quantum dot papers. However, PQDs can be susceptible to degradation under H2O and O2‐rich environment, continuous irradiation condition, or even in polar solvent due to their intrinsically unstable properties. In order to solve the problem, strategies have been provided including the adoption of alkyl ammonium with dozen‐nanometer chains and the most commonly used mixture of carboxylic acid (such as oleic acid (OA)) and alkylamines (such as oleylamine (OLA)) and alkylamines  in the perovskite precursor for stabilizing the PQDs in colloidal form. In 2016, solid‐state PQDs were stabilized via surface protection of polyhedral oligomeric silsesquioxane to exhibit water‐resistant behavior. Other methods such as using mesoporous silica/PQD core‐shell structures have also been demonstrated to prevent photooxidation and show the improved relative intensity after 96 h continuous ultraviolet (UV) irradiation from 60% to 80%. In the above methods, the protective encapsulation has usually been applied after the synthesis of PQD. Therefore, surface ligands such as OA and OLA should still be used during the synthesis stage for controlling the size of the PQD and passivating the PQD structure before the encapsulation. Although encapsulation strategies during synthesis have also been carried out to improve water and thermal resistances of PQDs, the stability under continuous irradiation has not yet evaluated.Perovskite quantum dots (PQDs) have been widely studied for developing the next‐generation display technology such as quantum dot (QD) enhancement films, QD light‐emitting diodes, and QD color filters over the past several years due to their inexpensive, color‐tunable, narrow emission peak (<20 nm), cadmium‐free, and high photoluminescence quantum yield (PLQY > 95%) properties.17 How3− and ‐O− group to synthesize the perovskite quantum dot paper (PQDP). These ‐HSO3− and ‐O− cellulose anions work as the capping ligands which limit the growth of perovskite crystals and become PQDs well‐dispersed in PQDP. In addition, we propose that the ‐O− and ‐HSO3− of the CNCs are coordinated to PbX2 empty orbital during the preparation of perovskite precursor  + MAX + PbX2 + CNCs, MA = CH3NH3+), and the PQDs are entangled between long chains of CNCs to avoid the dangling capping behavior of OA and OLA (which may be easily destroyed under high polar solvents) and become more stable.In this study, instead of mixing carboxylic acid (such as OA) and alkylamines (such as OLA) in perovskite precursor, we used the cellulose nanocrystal (CNC) with a few hundred nanometers in length which contains a large amount of ‐HSOAdditionally, we conceived a unique method of vacuum filtration growth of PQD in this study. For conventional PQD synthesis, the growth of PQD is in the colloidal solution, and the resulting PQD product is in the solution form, which requires purification procedure and spin coating to convert into solid. In the vacuum growth method, the perovskite precursor and CNC mixed solution is vacuum‐filtered to remove the DMF solvent, and the precursor/CNC residues are collected on the filter membrane. After the solvent is removed, the precursors start to crystalize into perovskite QDs with the assistance of surrounding CNC capping ligands, resulting in solid‐state PQDP. Moreover, the PQDs are confined spatially and protected in the entangled cellulose structure during the synthesis, which eliminates the need of additional encapsulation process. The differences between conventional solution growth and vacuum filtration growth are shown in Table S1, Supporting Information. The stability of PQDP was tested under different circumstances. The relative photoluminescence (PL) intensity of PQDP remains at ≈80% after exposed in an environment of 60% relative humidity (RH) and 20 °C for 8 months and ≈90% after exposed in continuous 16 W UV irradiation for 1000 h, showing unprecedentedly high stability. With superior optoelectronic properties and photostability, the PQDP reported in this work is a favorable choice for next‐generation lighting and solar‐related applications. They are composed of linear polysaccharide chains consisting of repeated β‐(1→4)‐d‐glucopyranose units. Owing to the presence of abundant hydroxyl (—OH) and sulfate (—OSO3Na) groups, CNCs are expected to exhibit a high electronegativity to complex with cations, that is, CH3NH3+, Pb2+ in metallic halide perovskites  by strong acid hydrolysis.36 The As sketched in Figure 2 and MABr dissolved in DMF solution were mixed by stirring for 30 min , and the concentration of CNC was kept constant at 0.5 wt%. The mixed solution was first vacuum‐filtered using an Anopore membrane  in a glass funnel (Wheaton) to remove unwanted DMF solvent, and the precursor/CNC mixtures were collected on the membrane. Then, the mixtures and membrane were taken from the funnel, followed by vacuum drying in a vacuum oven under the pressure of 0.36 kPa at 75 °C for 1 h to form the PQDP. Finally, the PQDP was peeled off from the membrane. Similar to commonly used papers, the resulting solid‐state PQDP is flexible and robust. Moreover, the PQDs within the nanopaper provide superior optoelectronic properties to the PQDP. Unlike colloidal PQDs, the solid‐state PQDP is no longer soluble in the solution. However, polar solvents can still degrade the PQDs within the paper due to the strong ionic property of perovskite .The PQDPs were produced using vacuum filtration followed by vacuum‐assisted heating at 75 °C.37 As Figure 3 PQD and is consistent with the previously reported lattice parameter of MAPbBr3 PQD. To further characterize the crystal structure of PQDP, X‐ray diffraction (XRD) pattern of pure CNC and PQDP were carried out . bBr3 PQD.39 To t Figure . The XRDs Figure . The TEMFigure Table  Although multiple effective strategies have been proposed to enhance humidity stability, our PQDP shows outstanding irradiation and thermal stability with <10% decay in PL intensity after 60 days of continuous UV irradiation and <40% decay after 20 days at high temperature (100 °C), respectively. To the best of our knowledge, these are the highest stabilities under continuous UV irradiation and high‐temperature conditions ever been measured for PQD materials.Because the XRD signals from PQDs are much weaker than that of CNCs for the PQDs/CNCs hybrid structure, it is difficult to characterize the stability of PQDP via XRD patterns  in PQD. Previous researches have shown that the strong bonding between capping ligands and PQD surfaces plays an important role in the stability of PQDs. Because the conventional OA and OLA ligands are weakly bound to the PQD surface, alternative ligands that can produce strong bonding were proposed to replace OA and OLA, and enhanced stability of PQDs were demonstrated. However, these achievements were mostly based on colloidal PQDs. During the transformation from colloidal to solid form, the purification process can lead to considerable ligand loss from the PQD surfaces, thus suppressing the stability. In this study, without the QD purification process, the vacuum growth method can completely avoid the ligand loss. Since the PQDs are crystalized and bound to the CNC ligands after removing the DMF solvents, the ligand bonding can be retained in solid state, leading to robust and stable PQDP. In addition, due to the long‐chain properties of CNC, the PQDP can be further protected to exhibit a higher level of stability than commonly used capping ligands such as OA and OLA.The excellent stability shown above can be attributed to the ligand integration between the high‐electronegativity long‐chain CNC and the polar cations  appear on the paper surface, thereby inhibiting the PLQY and stability. Moreover, it is expected that the synthesis procedure of PQDP can also be applied to perovskites with different halide or metal cations, which is still under investigation.To further elaborate the color tunability of PQDP, we have prepared PQDP with different ratios of perovskite precursor to CNC solution. Generally, based on the quantum confinement of QD, the emission wavelength can be slightly tuned by the QD size. It has been shown in previous studies that the size of solution‐grown PQD can be controlled by adjusting either the growth temperature49 or  the PQD‐based nanopaper exhibits a smooth surface. In addition, the thickness of PQDP can be easily tuned by the amount of CNCs and perovskite precursor during vacuum filtration (the thickness of PQDP in this study is around 40 µm). The size of PQDP is also scalable by changing the dimension of filter membrane. For example, we employed a 1.85 inch filter membrane in this study, yielding PQDP with exactly the same size  is shorter than that of solution‐grown PQDs using typical hot‐injection method (a few ns), implying that more surface defects may exist in the studied PQDs using the vacuum growth method. Therefore, more efforts are required to study the fundamental chemistry in the vacuum growth method and optimize the growth parameters such as vacuum flows, filtration time, and heating temperature to improve the quality of PQDs in the future.Surprisingly, the PQDP is semiconducting, with resistivity ≈1.3 × 10m Figure 5. OwingIn summary, by adopting the CNC for the synthesis of PQD, it is shown that inch‐scale OA and OLA‐free PQDPs with extraordinary intrinsic stability of over 60 days under continuous UV light irradiation have been successfully fabricated. The results provide a step forward to the practical application of PQD for future electronics. With consistently improved packaging techniques and polymer passivation/protection methods, it is expected that the stability can be further extended to meet the requirement of commercially used products such as QD enhancement films or QD color filters in the future.2 , CNC (Celluforce), DMF , deionized (DI) water, OA , OLA , and toluene . All the chemical reagents were used as received without further purification.MABr , PbBr2 (3.5 mmol) were loaded into a 10 mL vial with 7 mL of DMF, and the mixture was stirred until all of the powders were dissolved. Afterward, the mixture was stored in ambient under room temperature.MABr (3.5 mmol) and PbBrCNC powders were dissolved in DI water to form 5 wt% CNC in DI water, then took a suitable amount of 5 wt% CNC in DI water into DMF to prepare 0.5 wt% CNC in DMF (contains a trace amount of DI water). The mixture was stored in room temperature for later fabrication.3 precursor and 20 mL of CNC ligand solution were mixed and stirred vigorously at least 30 min at room temperature. Afterward, the mixture was vacuum filtrated for 24 h to form dried PQDP.1 mL of MAPbBrm of MAPbBr3 solution was prepared in 5 mL of DMF, then 200 µL of OA and 50 µL of OLA were added, respectively, under vigorous stirring until the solution became clear. Afterward, the precursor was pipetted into 10 mL of toluene under vigorous stirring and centrifuged at 7000 rpm for 10 min for purification.0.08 α radiation source (λ = 0.154 nm) at a scanning rate of 3° min−1 in the 2θ range of 10°–50°. TEM images were taken using a FEI Tecnai F20 G2 field emission TEM operating at 200 kV. The PL spectra, absorption spectra, and the absolute PLQY were measured with an Edinburgh FS5 spectrofluorometer. Continuous UV irradiation was performed using a Topbio UV light box with 16 W of UV light at 305 nm.The XRD patterns of the PQDP and CNC paper were measured using a Bruker D2 Phaser diffractometer with a Cu Kμ) of PQDP was obtained as ≈2.5 cm2 V−1 s−1 using equation μ = 8JL3/9ε0εrV2, where J is the current density  = 9 mm2), L is the thickness of PQDP (≈40 µm), ε0 is the free space permittivity (8.854 × 10−12 F m−1), εr is the dielectric constant of PQDP . The resistivity (ρ) of PQDP is ≈1.3 × 106 Ω cm, obtained using equation ρ = RA/L, where R is the measured electrical resistance. The SCLC measurement of pure CNC paper is shown in Figure S5, Supporting Information.SCLC and capacitance measurements were conducted using a Keithley 4200‐SCS semiconductor characterization system. The SCLC mobility (The authors declare no conflict of interest.Supporting InformationClick here for additional data file.


Hechtian adhesion between cell wall and plasma membrane transduces wall stress that opens Ca2+ channels and reorients auxin efflux “PIN” proteins; they control the auxin-activated proton pump that dissociates Ca2+ bound by periplasmic arabinogalactan proteins (AGP-Ca2+) hence the source of cytosolic Ca2+ waves that activate exocytosis of wall precursors, AGPs and PIN proteins essential for morphogenesis. This novel approach identifies the critical determinants of an algorithm that generates phyllotaxis spiral and Fibonaccian symmetry: these determinants in order of their relative contribution are: (1) size of the apical meristem and the AGP-Ca2+ capacitor; (2) proton pump activity; (3) auxin efflux proteins; (4) Ca2+ channel activity; (5) Hechtian adhesion that mediates the cell wall stress vector. Arguably, AGPs and the AGP-Ca2+ capacitor plays a decisive role in phyllotaxis periodicity and its evolutionary origins.Phyllotaxis describes the periodic arrangement of plant organs most conspicuously floral. Oscillators generally underlie periodic phenomena. A hypothetical algorithm generates phyllotaxis regulated by the Hechtian growth oscillator of the stem apical meristem (SAM) protoderm. The oscillator integrates biochemical and mechanical force that regulate morphogenetic gradients of three ionic species, auxin, protons and Ca These hydroxyproline-rich glycoproteins, especially the arabinogalactan proteins (AGPs), are involved in a hypothetical Hechtian growth oscillator. It involves transduction of the wall stress–strain to the plasma membrane where an auxin-activated proton pump dissociates AGP-Ca2+. Elevated cytosolic Ca2+-activates exocytosis thus regulating plant growth. Discussion of the Hechtian Oscillator vis-a-vis the role of the primary cell wall in plant morphogenesis [Agnes Arber  in “The 2+ release from arabinogalactan proteins (AGPs) of the cell surface; this hypothesis accounts for the origin of oscillations in molecular detail absent from previous models of tip growth [2+ waves) that regulate growth and differentiation  support that hypothesis. Recent characterization of their AGPs shows an elevated glucuronic acid content suggestive of enhanced Ca2+ binding in high salt [The Hechtian Oscillator exemplifies the pollen tube paradigm of rapid tip growth in particular ,7. The rt stress  may enhaigh salt . “cycles of auxin build-up and depletion accompany, and may direct, different stages of primordium development,” further confirmed by a more recent study of Arabidopsis embryogenesis [Heisler et al.  concludeogenesis . Auxin w+ dissociation of periplasmic AGP-Ca2+ [2+ that activates exocytosis in the AGP-rich protoderm. Indeed, ubiquitous distribution of AGPs throughout the Plant Kingdom [Thus, HAGP-Ca2+  is the i Kingdom ,35 impli Kingdom  confirm  Kingdom . Therefo+-ATPase proton pump by increasing its phosphorylation which increases the rate of proton extrusion [+-ATPase “turbo-molecular” proton pump. The molecular pathway involves successive glutamate protonation and deprotonation from the cytoplasmic side to the periplasm and cell wall  explanation invokes low pH-dependent wall loosening “enzymes” and expansins all of unknown specificity [2+ that provides an alternative reinterpretation of “acid growth” based on a visual analogy of the plasma membrane depicted as a metaphorical “molecular pin-ball machine” in . When activated by auxin, the proton pump shoots fast protons into the periplasm where they dislodge Ca2+ ions from the periplasmic AGP glycomodules; i.e., proton efflux generates Ca2+ influx. Thus, free Ca2+ ions then enter the cytosol via stretch-activated Ca2+channels. The rapid increase of cytosolic Ca2+ [2+ homeostasis is thus the major function of the proton pump rather than the regulation of wall pH.Proton extrusion and a concomitantly low wall pH associated with cell extension so-called “acid growth”  exemplifcificity  but ignohine” in  that reglic Ca2+  activatelic Ca2+  . For example in a floral phyllotaxis, N is a function of the stem apical meristem (SAM) size and the magnitude of major variables that define the symmetry and periodicity of new primordia. To sustain their growth, rapidly expanding cells demand auxin by generating the cell wall stress vector (CWsv) that redirects PIN proteins thus channeling auxin towards these incipient primordia (a) with auxin maxima and minima corresponding to future primordia and boundary tipping points, respectively. Generally, increasing the magnitude of a variable increases auxin transport towards a primordium hence rapidly depleting distal cells. Reversal of PIN protein orientation then generates a new primordium. Thus, an increased auxin depletion rate increases the number of new primordia. However, they also depend on Ca2+ availability determined by the expression of AGPs. We predict that AGPs strongly expressed in the protoderm will increase the periodicity of primordia while weaker expression will decrease it. Thus, to that extent, the algorithm is semi-quantitative and dependent on strong expression of AGPs in the protoderm of Arabidopsis meristems [Protoderm cell expansion generates new primordia ;Auxin efflux activity: A2+ channels: Cch;Ca2+ influx;Exocytosis is a complex variable regulated by Caad;Hechtian adhesion: Hsv;Cell wall stress vector: CWStem apical meristem protoderm SAMPsv vector determines auxin, proton and Ca2+ ion fluxes involving four phases of the Hechtian Oscillator proton pump that regulate cell expansion.The cell wall stress CW2+ influx and minimal activity of the oscillator.Phase I. Quiescent: Minimal cell wall stress corresponds to minimal Ca2+ channels; entry of Ca2+ initiates auxin binding to the proton pump and initial oscillator activation leading to Phase III.Phase II. Activation: Cell expansion increases wall stress, demand for auxin and opens Ca2+and supplies the Ca2+ channels thus generating cytosolic Ca2+waves that activate exocytosis, notably of wall precursors and plasticizers but also enabling dynamic redirection of PIN proteins. Addition of precursors reinforce the wall and slow its expansion, leading to Phase IV:Phase III. Maximum Activation: Occurs at the high auxin levels supplied by redirected auxin efflux PIN proteins; accelerated proton extrusion dissociates glycomodule AGP-Ca2+ channels.Phase IV. Return to quiescent phase: Stress relaxation of reinforced wall closes Ca2+ recycles via AGP precursors and Ca2+transporters replenish periplasmic AGP-Ca2+Cytosolic CaWhen attenuation of the stress vector reaches a tipping point of minimal oscillator activity and least rapid cell expansion, distant cells expand more rapidly and now exert a new stress vector in an opposing direction thus generating a new primordium that contributes to phyllotactic symmetry.a stem apical meristem protoderm generates new primordia as a function of the activity of five major variables or determinants shown in the speculative graph taking petal phyllotaxis as an example ranging from 3 to 34 petals. It assumes that the cell wall stress vector CWsv is essentially constant for a given SAMPa. SAMP2+ capacitor determines the periodicity of primordia initiation (cf. Protoderm AGP content (activity) is the primary determinant based on the simple hypothesis that size of the AGP-Cation cf.  where a The beguiling simplicity of the above plot inferred largely from biochemical observations is in strong contrast to previous complex formulations based largely on mathematical/geometrical logic.The evolutionary history of the stem apical meristem from a relatively simple arrangement of apical cells in the Bryophytes and ferns culminates in the morphogenetic protoderm of the angiosperms. Here, we conjecture that hybridization may solve the riddle of Fibonacci phyllotaxis and its evolutionary origins. 2+ signal that depends on the AGP-Ca2+ capacitor size, proton flux and Ca2+ channel status. Thus, cells of the protoderm with a large AGP-Ca2+ capacitor will increase cytosolic Ca2+ rapidly and therefore generate numerous primordia within a shorter range than in a protoderm with weaker AGP expression and therefore with more cells between primordia. For example, lower activity of the proton pump and a smaller AGP-Ca2+ capacitor increase spacing between primordia. Unlike the animal kingdom, plants have an enormous propensity for polyploidy and hybridization that we suggest provides a simple biochemical explanation for an evolutionary origin of the well-known Fibonacci series of floral organs exemplified by the 3, 5, 8, 13, 21 and 34 petals (Ranunculus ficaria (13), Erigeron canadensis (21). Can hybridization between contiguous members of the series generate a Fibonacci sequence? If so, how? Consider a hybrid expressing the sum of AGPs from both parents! If AGPs play a dominant role in defining phyllotaxis, then a hybrid of two-fold and threefold symmetry, with a corresponding increased size of the AGP-Ca2+ capacitor, would generate the most common five-fold symmetry, and so on for subsequent members of the series (“a consequence of optimal space filling in systems adding new units at a pole.” Thus, we infer that hybridization generates floral Fibonacci phyllotaxis and accounts for the evolutionary origin of a discrete series rather than a smooth arithmetic progression. This suggestion has the merit of simplicity based on Occam’s Razor in contrast to all preceding mathematical conjectures [Strong AGP expression predicts numerous closely spaced primordia  that gen4 petals , e.g., Re series . Thompsojectures  and is s“It was obviously my hope that the chemiosmotic rationale of vectorial metabolism and biological energy transfer might one day come to be generally accepted, and I have done my best to argue in favour of that state of affairs for more than twenty years…was it not the great Max Planck who remarked that a new scientific idea does not triumph by convincing its opponents, but rather because its opponents eventually die?” Although Mitchell’s unconventional ideas were initially rejected, they were finally recognized. Their universal applicability has become apparent more recently. In simple photoautotrophs, light-driven proton gradients involve bacteriorhodopsin [2+ rather than just two. Thompson’s classical “Growth and Form”  exemplifhodopsin , while i2+ channels that generate cytosolic Ca2+ gradients. Thus, the Hechtian growth oscillator is an extrapolation of Mitchell’s chemiosmosis that unifies physics and chemistry in a minimalist approach to regulating plant growth. Indeed, precursors to life surely involve proton gradients as a basis of prebiotic energy transduction and the universal proton pump of exoplanet life in the habitable zone.However, these gradients do not arise by simple diffusion but are regulated by auxin efflux “PIN” proteins whose discovery began with Rubery and Sheldrake’s  classic


Despite the success of immunotherapy in several haematological neoplasms, the effectiveness in acute myeloid leukaemia (AML) is still controversial, partially due to the lack of knowledge regarding immune‐related processes in this disease and similar neoplasias. In this study, we analysed the role and expression of histamine receptor 1 (HRH1) in haematological malignancies. Although the histamine receptor type 1 was widely expressed in healthy and malignant haematopoiesis, especially along the myeloid lineage, HRH1 lacked a relevant role in survival/proliferation and chemoresistance of AML cells, as analysed by HRH1 knockdown (KD) and pharmacological modulation. However, HRH1‐mediated signalling was critical for the activation of the differentiation process induced by several agents including all‐trans retinoic acid, establishing a role for HRH1 in myeloid differentiation. Pharmacological activation of Erk was able to partially restore differentiation capacity in HRH1 KD AML cells, suggesting that HRH1 signalling acts upstream MAPK‐Erk pathway. As an indirect consequence of our results, treatment‐related histamine release is not expected to confer a proliferative advantage in leukaemic cells. Acute myeloid leukaemia (AML) is a haematological neoplasia characterized by the accumulation in bone marrow of transformed myeloid progenitors with enhanced proliferation and a block in differentiation. Indeed, AML cells are arrested at different stages of myeloid maturation.Histamine is a pleiotropic biogenic amine acting through four different histamine receptors (HRH1 to HRH4) belonging to the G protein‐coupled receptor superfamily. Although most of the proposed effects of histamine on myeloid differentiation are mediated by HRH2,+ CD34+), T cells (CD45+ CD3+) and platelets (CD45− CD41a+), while intermediate expression was observed for B cells (CD45+ CD19+) and granulocytes (CD45+ CD15+)  showed a generalized HRH1 expression. Consistent with their molecular heterogeneity, HRH1 expression showed a wide distribution pattern in MDS and AML, ranging from 0% to 100%. Conversely, in CMML HRH1 expression was prevalent, reaching more than 50% in all 17 analysed samples  and intermediate (CD14+ CD16+) monocytes are predominant,dimCD16+ monocytes  Figure . Monocyts Figure . These rs Figure . Given ts Figure . The excs Figure .To further study the role of HRH1 expression by AML, we generated an HL60‐derived cell line stably expressing a CRISPR/Cas9‐mediated HRH1 down‐regulation system. The knockdown (KD) of HRH1 was assessed at a protein level both by flow cytometry and Western blot and was highly efficient Figure . KD of HAs one of the main features of AML is the maturation blockade and some effects of histamine have been observed on myeloid differentiation,https://hgserver1.amc.nl), performing a GSEA analysis on the top genes correlated with HRH1 in 4 different AML datasets. The KEGG retinol metabolism gene set recurrently appeared on top of the analyses  T cells have been proposed as promising strategies to eradicate AML. One of the main side effects associated with those therapies is the cytotoxicity resulting from an excessive immune activation known as cytokine release syndrome, managed, among others, with antihistamines.Taken together, the results obtained in this study suggest an involvement of HRH1 in healthy and malignant myeloid differentiation, that will need to be mechanistically elucidated in future studies.RMR is a shareholder of Leukos Biotech.Josep Maria Cornet‐Masana: Conceptualization ; Data curation (lead); Formal analysis (lead); Investigation (lead); Methodology (lead); Validation ; Writing‐original draft ; Writing‐review & editing . Antònia Banús‐Mulet: Data curation (supporting); Formal analysis (supporting); Investigation (supporting); Methodology (supporting); Validation ; Writing‐review & editing (supporting). Laia Cuesta‐Casanovas: Data curation (supporting); Formal analysis (supporting); Investigation (supporting); Methodology (supporting); Writing‐review & editing (supporting). Jose Maria Carbó: Data curation (supporting); Formal analysis (supporting); Investigation (supporting); Methodology (supporting); Validation ; Writing‐review & editing (supporting). Francesca Guijarro: Conceptualization (supporting); Methodology (supporting); Resources ; Writing‐review & editing (supporting). Miguel Ángel Torrente: Conceptualization (supporting); Formal analysis (supporting); Methodology (supporting); Resources ; Writing‐review & editing (supporting). Jordi Esteve: Conceptualization (supporting); Methodology (supporting); Resources (lead); Supervision ; Writing‐review & editing . Ruth M. Risueño: Conceptualization (lead); Data curation ; Formal analysis ; Funding acquisition (lead); Investigation (supporting); Methodology ; Project administration (lead); Resources ; Supervision (lead); Visualization (lead); Writing‐original draft ; Writing‐review & editing (lead).Supplementary MaterialClick here for additional data file.