Metal-Organic Framework Based on Ratiometric dual-Fluorescent Sensor Using for Accurate Quantification and on-Site Visual Detection of Ascorbic Acid.

Ascorbic acid is very important to the metabolic process of the body, but excessive intake can lead to diarrhea, kidney calculi and stomach cramps. However, complicated production procedures and harsh experimental settings limit many detection methods, and a simpler and more accurate measurement method is needed. In this study, a smartphone-assisted ratiometric fluorescence sensor was developed for the portable analysis of ascorbic acid. Leveraging the catalytic properties of MIL-53(Fe) to expedite the conversion of H2O2 into hydroxyl radicals, thereby facilitating the oxidation of o-phenylenediamine and terephthalic acid bridging ligand. The sensor showcased exceptional sensitivity in detecting ascorbic acid within a linear range of 0.3-100 µM, boasting an impressive limit of detection at 0.15 µM. Furthermore, through the utilization of color extraction RGB values captured by smartphones, accurate detection of ascorbic acid was achieved with a detection limit of 0.4 µM. Real fruit samples exhibited robust spiked recovery rates ranging from 91 to 119%, accompanied by relative standard deviations ≤ 4.7%. The MIL-53(Fe) nanozyme-based smartphone-assisted ratiometric fluorescence sensor offers an ascorbic acid fluorescence detection device that is visible, accurate, sensitive, and reasonably priced.

Draxin inhibits chick trunk neural crest delamination and migration by increasing cell adhesion.

The neural crest is a multipotent cell population that migrates extensively to play important roles during embryonic development. After acquiring motility, trunk neural crest cells delaminate from the spinal cord and migrate to various regions of the body. Several cellular adhesion molecules, such as vinculin, are involved in the regulation of neural crest delamination and migration. In the present study, we found that draxin could inhibit delamination and migration of neural crest cells from the chick spinal cord and abnormal aggregation of the migrating neural crest cells. In the presence of draxin, the resuspended neural crest regained its adhesive ability such that it was significantly increased. Overexpression of draxin caused increased vinculin expression in vivo. Our data indicate that draxin might control delamination and migration of chick trunk neural crest by increasing cell adhesion.

Use of 1-octyl-3-methylimidazole hexafluorophosphate modified magnetic hyperbranched polyamideamine as sorbent for the extraction of pyrethroid insecticides from tea infusion.

Magnetic hyperbranched polyamideamine was carboxylated using succinic anhydride and modified further with 1-octyl-3-methylimidazole hexafluorophosphate successively. The morphology and chemical composition of the prepared material was characterized by transmission electron microscopy, scanning electron microscopy, Fourier transform infrared spectroscopy, Brunauer-Emmett-Teller measurement, X-ray photoelectron spectroscopy, etc. 1-Octyl-3-methylimidazole hexafluorophosphate modified magnetic hyperbranched polyamideamine was used as sorbent in the magnetic solid-phase extraction for the separation and enrichment of five pyrethroid insecticides from tea infusion. The magnetic solid-phase extraction method proposed in this article has low method detection limits (0.53-0.71 ng/mL), acceptable coefficient of determination (0.9992-0.9998), wide linear ranges (2.5-500.0 ng/mL), and good repeatability (intraday: 1.2-6.3%; interday: 1.6-5.4%). In the detection of five pyrethroid insecticides in tea infusion, relative recoveries were in the range from 87.7 to 114.7% with satisfactory relative standard deviations (0.2-7.4%). With the aid of quantum chemistry calculations, the interaction energy between the sorbent and five pyrethroid insecticides was calculated, which proved the necessity of the modification of 1-octyl-3-methylimidazole hexafluorophosphate.

Hyperbranched aromatic polyamide modified magnetic nanoparticles for the extraction of benzoylurea insecticides.

Herein, a novel hyperbranched aromatic polyamide-coated magnetic sorbent was prepared by in situ polymerization on the surface of amino-functionalized Fe3 O4 nanoparticles. The magnetic sorbent was characterized by transmission electron microscopy, Fourier transform infrared spectroscopy, elemental analysis, Brunauer-Emmett-Teller measurement, and X-ray photoelectron spectroscopy, etc. The new magnetic sorbent was used in the magnetic solid-phase extraction for the detection of seven benzoylurea insecticides. Under optimum conditions, low method detection limits (0.56-1.20 ng/mL), acceptable coefficient of determination (0.9967-0.9996), wide linear ranges (2.5-500.0 ng/mL), and good repeatability (intraday: 2.0-7.3%; interday: 1.9-9.2%) were achieved. The magnetic solid-phase extraction method based on the new magnetic sorbent showed good reliability in the analysis of seven benzoylurea insecticides in real water samples, as the relative recoveries were in the range from 80.1 to 116.3% with satisfactory RSDs (0.1-9.8%). By means of density functional theory and semiempirical quantum mechanical, the binding configuration and interaction energy of hyperbranched aromatic polyamide and benzoylurea insecticides were calculated. The result of theoretical calculation revealed that the adsorption of benzoylurea insecticides by hyperbranched aromatic polyamide was derived from hydrogen bonding and π-π stacking. The contribution of π-π stacking was greater than that of hydrogen bond, which was confirmed by energy decomposition analysis.

Preparation of magnetic attapulgite/polypyrrole nanocomposites for magnetic effervescence-assisted dispersive solid-phase extraction of pyrethroids from honey samples.

In this work, a novel extraction technique based on the effervescence-assisted dispersion and magnetic recovery of attapulgite/polypyrrole sorbents was developed for determining the concentrations of five pyrethroids in honey samples. The magnetic nanoparticles were synthesized by a one-pot method. Several experimental parameters that affected the extraction efficiency, including the dispersion conditions, pH, ionic strength, and desorption conditions, were investigated. Under optimal conditions, the calibration curves for the five pyrethroids in honey samples exhibited good linearity, with r2 values ranging from 0.9979 to 0.9990. The limits of detection varied between 0.21 and 0.34 µg/L. Satisfactory recoveries of 81.42-106.73% with intra- and interday relative standard deviations of less than 6.94 and 10.89%, respectively, were obtained. Moreover, the sorbents exhibited acceptable batch-to-batch repeatability in the range of 5.06-15.01%, and each sorbent could be reused for up to four extraction cycles without a significant loss in the extraction recovery.

Humic acid functionalized hyperbranched polytriazine based dispersive solid-phase extraction for acaricides determination in tea matrix.

Hyperbranched polytriazine functionalized with humic acid was prepared and developed as new sorbents for dispersive solid-phase extraction of three acaricides (clofentezine, fenpyroximate, and pyridaben) in tea samples combined with high-performance liquid chromatography detection. The sorbents were characterized by scanning electron microscopy, energy dispersive spectroscopy, Zeta-potential, and Fourier transform infrared spectroscopy. The extraction parameters (extraction time, ionic strength, desorption conditions) were optimized. The adsorption mechanism was evaluated utilizing Fourier transform infrared spectra. Under optimum conditions, satisfactory analytical performances were achieved, which included high precision (1.33-9.62%), low limits of detection (0.19-3.54 µg/L), and wide linear range (2.5-500 µg/L) for the analysis of the acaricides. Moreover, the proposed method proved highly effective for the determination of acaricides in tea samples, with the relative recoveries in the range of 65.20-108.13% and relative standard deviations < 9.87%. The method has great application potential for the detection of acaricides in tea samples.

Magnetic nanoparticles modified with hyperbranched polyamidoamine for the extraction of benzoylurea insecticides prior to their quantitation by HPLC.

A magnetic sorbent was fabricated by covalently grafting hyperbranched polyamidoamine onto the surface of Fe3O4 nanoparticles. The sorbent was used in the magnetic solid-phase extraction (MSPE) of benzoylurea insecticides (BUs) from environmental water samples. It is perceived that hydrogen bonding interactions and hydrophobic interactions are the main mechanisms for the adsorption of BUs. The sorbent was characterized by scanning electron microscopy, transmission electron microscopy, Fourier transform infrared spectroscopy, elemental analysis, Brunauer-Emmett-Teller measurement, X-ray photoelectron spectroscopy, vibrating sample magnetometry and X-ray diffraction. Various experimental parameters affecting the MSPE were optimized. Following elution with acetonitrile, the BUs were quantified by HPLC with diode array detection. The method based has a wide linear response range (2.5-500.0 ng mL-1), satisfactory coefficient of determination (0.9922-0.9976), high enrichment factors (62.8-74.4), acceptable limits of quantitation (2.5 ng mL-1) and low limits of detection (0.39-0.72 ng mL-1). The intra-day precision at concentration levels of 5.0, 50.0 and 250.0 ng mL-1 ranged from 2.3-6.4% and the inter-day precision was between 1.0 and 5.5%. The sorbent can be re-used at least 15 times. It was applied to the extraction of BUs from genuine water samples where it showed satisfactory relative recoveries (75.1-111.4%) and precision (1.0-9.1%). Graphical abstract Schematic presentation of magnetic nanoparticles modified with hyperbranched polyamidoamine and their application for extraction of benzoylurea insecticides.

In situ solvent formation microextraction combined with magnetic dispersive micro-solid-phase extraction for the determination of benzoylurea insecticides in water samples.

A simple, fast, effective, and environmentally friendly method, in situ solvent formation microextraction combined with magnetic dispersive micro-solid-phase extraction for the determination of four benzoylurea insecticides is presented herein for the first time. In the proposed method, 1-hexyl-3-methylimidazolium bis[(trifluoromethane)sulfonyl]imide was formed by the reaction between 1-hexyl-3-methylimidazolium chloride and lithium bis[(trifluoromethane)sulfonyl]imide and was used to extract benzoylurea insecticides. Then magnetic nanoparticles were added as carrier to retrieve and separate the ionic liquid from the sample solution. After the supernatant was removed, the ionic liquid was desorbed using acetonitrile and subsequently injected directly into a high-performance liquid chromatograph equipped with a variable wavelength detector for analysis. The main factors affecting the extraction efficiency were investigated by a one factor at a time approach. Under optimized conditions, the proposed method showed good repeatability (RSD = 2.2-4.5%) and linearity (2-300 µg/L), with correlation coefficients greater than 0.9994 and low limits of detection (0.67-1.46 µg/L). Finally, the method was successfully applied to the analysis of four benzoylurea insecticides in environmental water samples with good recoveries (73.2-85.8%).

Ultrasound-assisted, hybrid ionic liquid, dispersive liquid-liquid microextraction for the determination of insecticides in fruit juices based on partition coefficients.

An ultrasound-assisted, hybrid ionic liquid, dispersive liquid-liquid microextraction method coupled to high-performance liquid chromatography with a variable-wavelength detector was developed to detect ten insecticides, including diflubenzuron, triflumuron, hexaflumuron, flufenoxuron, lufenuron, diafenthiuron, transfluthrin, fenpropathrin, γ-cyhalothrin and deltamethrin, in fruit juices. In this method, an appropriate extraction solvent was chosen based on the partition coefficient of the target compounds. A mixture of 1-octyl-2,3-dimethylimidazolium bis(trifluoromethylsulfonyl)imide and 1-hexyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide was used as the extractant. The extraction efficiency was screened using Plackett-Burman design and optimized using central composite design. Under the optimal conditions, good linearity was obtained for all the analytes in the pure water model and the fruit juice samples. In pure water, the recoveries of the ten insecticides ranged from 85.7 to 108.9%, with relative standard deviations for one day ranging from 1.24 to 2.64%. The limits of detection were in the range of 0.19-0.69 µg/L, and the enrichment factors were in the range of 123-160. The logarithm of the n-octanol/water partition coefficient in this experiment is a useful reference to select a suitable extraction solvent, and the proposed technique was applied for the analysis of ten insecticides in fruit juice with satisfactory results.

Dispersive liquid-liquid microextraction based on the solidification of deep eutectic solvent for the determination of benzoylureas in environmental water samples.

We present a novel dispersive liquid-liquid microextraction method based on the solidification of deep eutectic solvent coupled with high-performance liquid chromatography with a variable-wavelength detection for the detection of five benzoylureas in real water samples. In this work, a green solvent consisting of 1-octyl-3-methylimidazolium chloride and 1-dodecanol was used as an extraction solvent, yielding the advantages of material stability, low density, and a suitable freezing point near room temperature. Parameters that significantly affect extraction efficiency were optimized by the one-factor-at-a-time approach. Under optimal conditions, the recoveries of five target compounds were obtained ranging from 87.39 to 98.05% with correlation coefficients ranging from 0.9994 to 0.9997 for pure water. The limits of detection were in the range of 0.09-0.16 µg/L. The enrichment factors were in the range of 171-188. Linearities were achieved in the range of 0.5-500 µg/L. The proposed method was successfully applied to determine benzoylureas in environmental water samples with a satisfactory recovery of approximately 81.38-97.67%.

Pipette vial dispersive liquid-liquid microextraction combined with high-performance liquid chromatography for the determination of benzoylurea insecticide in fruit juice.

A simple, sensitive, and efficient method of using a pipette vial to perform dispersive liquid-liquid microextraction based on the solidification of floating organic droplets was coupled with high-performance liquid chromatography (HPLC) and a diode array detector for the preconcentration and analysis of four benzoylurea insecticides in fruit juice. In this method, 1-dodecanol was used as an extractant, and a snipped pipette was used as an experimental vial to simplify the procedure of collecting and separating solidified extractant. The experimental parameters were optimized using a Plackett-Burman design and one-factor-at-a-time method. Under the optimal conditions in the water model, the limits of detection for analytes varied from 0.03 to 0.28 µg/L, and the enrichment factors ranged from 147 to 206. Linearity was achieved for diflubenzuron and flufenoxuron in a range of 0.5-500 µg/L, for hexaflumuron in a range of 1-500 µg/L, and for triflumuron in a range of 5-500 µg/L. The correlation coefficients for the analytes ranged from 0.9986 to 0.9994 with recoveries of 91.4-110.9%. Finally, the developed technique was successfully applied to fruit juice samples with acceptable results. The relative standard deviations of the analytes at two spiking levels (50 and 200 µg/L) varied between 0.2 and 4.5%.

Dispersive micro-solid-phase extraction of benzoylurea insecticides in honey samples with a ß-cyclodextrin-modified attapulgite composite as sorbent.

A ß-cyclodextrin-modified attapulgite composite was prepared and used as a dispersive micro-solid-phase extraction sorbent for the determination of benzoylurea insecticides in honey samples. Parameters that may influence the extraction efficiency, such as the type and volume of the eluent, the amount of the sorbent, the extraction time and the ionic strength were investigated and optimized using batch and column procedures. Under optimized conditions, good linearity was obtained for all of the tested compounds, with R(2) values of at least 0.9834. The limits of detection were determined in the range of 0.2-1.0 µg/L. The recoveries of the four benzoylurea insecticides in vitex honey and acacia honey increased from 15.2 to 81.4% and from 14.2 to 82.0%, respectively. Although the ß-cyclodextrin-modified attapulgite composite did not show a brilliant adsorption capacity for the selected benzoylurea insecticides, it exhibited a higher adsorption capacity toward relatively hydrophobic compounds, such as chlorfluazuron and hexaflumuron (recoveries in vitex honey samples ranged from 70.0 to 81.4% with a precision of 1.0-3.7%). It seemed that the logPow of the benzoylurea insecticides is related to their recoveries. The results confirmed the possibility of using cyclodextrin-modified palygorskite in the determination of relatively hydrophobic trace pharmaceutical residues.

Effervescence-assisted dispersive solid-phase extraction using ionic-liquid-modified magnetic ß-cyclodextrin/attapulgite coupled with high-performance liquid chromatography for fungicide detection in honey and juice.

In this study, a simple effervescence-assisted dispersive solid-phase extraction method was developed to detect fungicides in honey and juice. Most significantly, an innovative ionic-liquid-modified magnetic ß-cyclodextrin/attapulgite sorbent was used because its large specific surface area enhanced the extraction capacity and also led to facile separation. A one-factor-at-a-time approach and orthogonal design were employed to optimize the experimental parameters. Under the optimized conditions, the entire extraction procedure was completed within 3 min. In addition, the calibration curves exhibited good linearity, and high enrichment factors were achieved for pure water and honey samples. For the honey samples, the extraction efficiencies for the target fungicides ranged from 77.0 to 94.3% with relative standard deviations of 2.3-5.44%. The detection and quantitation limits were in the ranges of 0.07-0.38 and 0.23-1.27 µg/L, respectively. Finally, the developed technique was successfully applied to real samples, and satisfactory results were achieved. This analytical technique is cost-effective, environmentally friendly, and time-saving.

The PTEN/PI3K/Akt signaling pathway mediates HMGB1-induced cell proliferation by regulating the NF-κB/cyclin D1 pathway in mouse mesangial cells.

Our previous experiment confirmed that high-mobility group box chromosomal protein 1 (HMGB1) was involved in the pathogenesis of Lupus nephritis (LN) by upregulating the proliferation of the mouse mesangial cell line (MMC) through the cyclin D1/CDK4/p16 system, but the precise mechanism is still unknown. Therefore, in the present study, we demonstrated that HMGB1 induced the proliferation of MMC cells in a time- and concentration-dependent manner, downregulated phosphatase and tensin homolog deleted on chromosome ten (PTEN) expression, increased the level of Akt serine 473 phosphorylation, and induced p65 subunit nuclear translocation. The overexpression of PTEN prevented the upregulation of HMGB1-induced proliferation by blocking the activation of Akt. The knockdown of Akt by siRNA technology and blocking the nuclear factor-κB (NF-κB) pathway using pyrrolidine dithiocarbamate (PDTC) and SN50, inhibitors of NF-κB, both attenuated the HMGB1-induced proliferation by counteracting the activation of the cyclin D1. In addition, while sh-Akt partly blocked the nuclear translocation of the p65 subunit, PDTC did not affect the activation of the Akt induced by HMGB1 in MMC cells. These findings indicate that HMGB1 induced the proliferation of MMC cells by activating the PTEN/phosphoinositide-3-kinase (PI3K)/Akt/NF-κB signaling pathway.

Optimization of dispersive liquid-liquid microextraction based on the solidification of floating organic droplets using an orthogonal array design and its application for the determination of fungicide concentrations in environmental water samples.

A dispersive liquid-liquid microextraction method based on the solidification of floating organic droplets was developed as a simple and sensitive method for the simultaneous determination of the concentrations of multiple fungicides (triazolone, chlorothalonil, cyprodinil, and trifloxystrobin) in water by high-performance liquid chromatography with variable-wavelength detection. After an approach varying one factor at a time was used, an orthogonal array design [L25 (5(5))] was employed to optimize the method and to determine the interactions between the parameters. The significance of the effects of the different factors was determined using analysis of variance. The results indicated that the extraction solvent volume significantly affects the efficiency of the extraction. Under optimal conditions, the relative standard deviation (n = 5) varied from 2.3 to 5.5% at 0.1 µg/mL for each analyte. Low limits of detection were obtained and ranged from 0.02 to 0.2 ng/mL. In addition, the proposed method was applied to the analysis of fungicides in real water samples. The results show that the dispersive liquid-liquid microextraction based on the solidification of floating organic droplets is a potential method for detecting fungicides in environmental water samples, with recoveries of the target analytes ranging from 70.1 to 102.5%.

Single-cell sequencing of the substantia nigra reveals microglial activation in a model of MPTP.

Background: N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is a neurotoxin widely used to induce PD models, but the effect of MPTP on the cells and genes of PD has not been fully elucidated. Methods: Single-nucleus RNA sequencing was performed in the Substantia Nigra (SN) of MPTP mice. UMAP analysis was used for the dimensionality reduction visualization of the SN in the MPTP mice. Known marker genes highly expressed genes in each cluster were used to annotate most clusters. Specific Differentially Expressed Genes (DEGs) and PD risk genes analysis were used to find MPTP-associated cells. GO, KEGG, PPI network, GSEA and CellChat analysis were used to reveal cell type-specific functional alterations and disruption of cell-cell communication networks. Subset reconstruction and pseudotime analysis were used to reveal the activation status of the cells, and to find the transcription factors with trajectory characterized. Results: Initially, we observed specific DEGs and PD risk genes enrichment in microglia. Next, We obtained the functional phenotype changes in microglia and found that IGF, AGRN and PTN pathways were reduced in MPTP mice. Finally, we analyzed the activation state of microglia and revealed a pro-inflammatory trajectory characterized by transcription factors Nfe2l2 and Runx1. Conclusion: Our work revealed alterations in microglia function, signaling pathways and key genes in the SN of MPTP mice.

Dispersive micro-solid phase extraction based on self-assembling, ionic liquid-coated magnetic particles for the determination of clofentezine and chlorfenapyr in environmental water samples.

Two ionic liquid-coated-Fe3O4 magnetic particles (IL-Fe3O4 MPs) were developed for use in two types of dispersive micro-solid phase extraction (D-µ-SPE) for the high-performance liquid chromatographic analysis of clofentezine and chlorfenapyr in environmental water samples. Self-assembling IL-Fe3O4 MPs were used in D-µ-SPE as adsorbents. Two D-µ-SPE extraction methods, namely, direct dispersive micro-solid phase extraction (d-D-µ-SPE) and in situ solvent formation-based dispersive micro-solid phase extraction (ISF-D-µ-SPE), were proposed, using [C8MIM][PF6] to extract analytes through two pathways. Lower IL doses were required in the extraction process compared with those in other IL-based methods. Fe3O4 MPs can also be recycled and reused after extraction and are thus environmentally friendly. These newly developed methods were demonstrated to be feasible for use in the quantitation of clofentezine and chlorfenapyr at trace levels, with lower limit of detection values ranging from 0.4 to 0.5 ng mL(-1) for d-D-µ-SPE and 0.4 ng mL(-1) for ISF-D-µ-SPE. Finally, relative standard deviations of less than 6.0% were obtained.

Zoledronic Acid Accelerates Bone Healing in Carpal Navicular Fracture via Silencing Long Non-coding RNA Growth Arrest Specificity 5 to Modulate MicroRNA-29a-3p Expression.

Carpal navicular fractures are the most common carpal fractures. This study intends to explore the specific mechanism of Zoledronic Acid (ZA) in carpal navicular fracture healing via long non-coding RNA (lncRNA) growth arrest specificity 5 (GAS5) to mediate microRNA (miR)-29a-3p. A fractured rat model was constructed. Two weeks later, a subcutaneous injection of systemic ZA was implemented, and an injection of plasmid vectors interfered with GAS5 or miR-29a-3p expression was performed on the fracture site. Osteocalcin (OCN) and bone morphogenetic protein-2 (BMP-2) were determined, as well as serum levels of alkaline phosphatase (ALP), osteopontin (OPN) and osteoprotegerin (OPG) and bone mineral density. MC3T3-E1 cells were transfected with plasmid vectors interfering with GAS5 or miR-29a-3p, and cell proliferation and apoptosis were analyzed. GAS5 and miR-29a-3p expression in fractured rats was tested, together with their binding relationship. ZA promoted OCN and BMP-2 expression, increased bone mineral density and serum levels of ALP, OPN and OPG in fractured rats. GAS5 was upregulated and miR-29a-3p was down-regulated in fractured rats. Downregulation of GAS5 or upregulation of miR-29a-3p further promoted bone healing in fractured rats. GAS5 targets miR-29a-3p, and down-regulation of miR-29a-3p can reverse the effect of down-regulation of GAS5 on bone healing in fractured rats. ZA promoted the proliferation of MC3T3-E1 cells and inhibited apoptosis by regulating the GAS5/miR-29a-3p axis. ZA regulates miR-29a-3p expression by down-regulating GAS5 to promote carpal navicular fracture healing, promote MC3T3-E1 cell proliferation, and inhibit cell apoptosis.

Bimetallic metal-organic frameworks-based ratiometric fluorescence sensor for the quantitative detection of thiram in fruits samples.

The identification of residual thiram (Tr) in foods is vital in view of its harmful effects on human health. Herein, a ratiometric fluorescence sensor (I435/I590) based on rhodamine B/NH2-MIL-53(Al0.75Fe0.25) was constructed for the detection of Tr. Interestingly, the probe RhB/NH2-MIL-53(Bim) assisted by Cu2+ could rapidly and sensitively recognize Tr with a low detection limit of 0.11 µg/mL in 10 min. The fluorescence sensing mechanism was investigated using fluorescence spectra, UV-Vis absorption spectra, the fluorescence lifetime and quantum yield. The results showed that the excellent sensing performance was attributed to fluorescence resonance energy transfer, electrostatic interaction, and photoinduced electron transfer. In addition, the practical application of this platform showed acceptable relative recoveries for Tr (84.03-107.81 %), and precisions were also achieved (relative standard deviation ≤ 8.69 %, n = 3). These results show that the presented herein can be applied to monitor the Tr content in real fruit samples.

Draxin inhibits axonal outgrowth through the netrin receptor DCC.

Draxin, a recently identified axon guidance protein, is essential for the formation of forebrain commissures, and can mediate repulsion of netrin-stimulated spinal commissural axons. Here, we report that draxin binds multiple netrin receptors: DCC (deleted in colorectal cancer), Neogenin, UNC5s (H1, H2, H3), and DSCAM (Down's syndrome cell adhesion molecule). Since draxin and Dcc knockouts showed similar phenotype in forebrain commissures formation, we show here the functional importance of draxin/DCC interaction. Draxin interacts with subnanomolar affinity to the netrin receptor DCC, in a region of DCC distinct from its netrin-binding domain. In vitro, neurite outgrowth from cortical and olfactory bulb explants of Dcc knock-out mice is significantly less inhibited by draxin, when compared with neurites from explants of wild-type mice. Furthermore, in comparison with wild-type mice, the growth cone collapse in response to draxin is largely abolished in Dcc-deficient cortical neurons. In vivo, double heteros of draxin/Dcc mice show markedly higher frequency of complete agenesis of corpus callosum than either of the single hetero. These results identify DCC as a convergent receptor for netrin and draxin in axon growth and guidance.