A CD25×TIGIT bispecific antibody induces anti-tumor activity through selective intratumoral Treg cell depletion.

Intratumoral regulatory T cells (Tregs) express high levels of CD25 and TIGIT, which are also recognized as markers of effector T cell (Teff) activation. Targeting these molecules each alone with monoclonal antibodies (mAbs) poses a risk of concurrently depleting both Teffs and peripheral Tregs, thereby compromising the effectiveness and selectivity of intratumoral Treg depletion. Here, leveraging the increased abundance of CD25+ TIGIT+ double-positive Tregs in the solid tumor microenvironment (but not in peripheral tissues), we explore the feasibility of using a CD25×TIGIT bispecific antibody (bsAb) to selectively deplete intratumoral Tregs. We initially constructed a bsAb co-targeting mouse CD25 and TIGIT, NSWm7210, and found that NSWm7210 conferred enhanced intratumoral Treg depletion, Teff activation, and tumor suppression as compared to the parental monotherapies in mouse models. We subsequently constructed a bsAb co-targeting human CD25 and TIGIT (NSWh7216), which preferentially eliminated CD25+ TIGIT+ double-positive cells over single-positive cells in vitro. NSWh7216 exhibited enhanced anti-tumor activity without toxicity of peripheral Tregs in CD25 humanized mice compared to the parental monotherapies. Our study illustrates the use of CD25×TIGIT bsAbs as effective agents against solid tumors based on selective depletion of intratumoral Tregs.

Propelling Multi-Modal Therapeutics of PEEK Implants through the Power of NO evolving Covalent Organic Frameworks (COFs).

The design and fabrication of NO-evolving core-shell nanoparticles (denoted as NC@Fe), comprised of BNN6-laden COF@Fe3 O4 nanoparticles, are reported. This innovation extends to the modification of 3D printed polyetheretherketone scaffolds with NC@Fe, establishing a pioneering approach to multi-modal bone therapy tailored to address complications such as device-associated infections and osteomyelitis. This work stands out prominently from previous research, particularly those relying on the use of antibiotics, by introducing a bone implant capable of simultaneous NO gas therapy and photothermal therapy (PPT). Under NIR laser irradiation, the Fe3 O4 NP core (photothermal conversion agent) within NC@Fe absorbs photoenergy and initiates electron transfer to the loaded NO donor (BNN6), resulting in controlled NO release. The additional heat generated through photothermal conversion further propels the NC@Fe nanoparticles, amplifying the therapeutic reach. The combined effect of NO release and PPT enhances the efficacy in eradicating bacteria over a more extensive area around the implant, presenting a distinctive solution to conventional challenges. Thorough in vitro and in vivo investigations validate the robust potential of the scaffold in infection control, osteogenesis, and angiogenesis, emphasizing the timeliness of this unique solution in managing complicated bone related infectious diseases.

CAV3 alleviates diabetic cardiomyopathy via inhibiting NDUFA10-mediated mitochondrial dysfunction.

BACKGROUND: The progression of diabetic cardiomyopathy (DCM) is noticeably influenced by mitochondrial dysfunction. Variants of caveolin 3 (CAV3) play important roles in cardiovascular diseases. However, the potential roles of CAV3 in mitochondrial function in DCM and the related mechanisms have not yet been elucidated. METHODS: Cardiomyocytes were cultured under high-glucose and high-fat (HGHF) conditions in vitro, and db/db mice were employed as a diabetes model in vivo. To investigate the role of CAV3 in DCM and to elucidate the molecular mechanisms underlying its involvement in mitochondrial function, we conducted Liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis and functional experiments. RESULTS: Our findings demonstrated significant downregulation of CAV3 in the cardiac tissue of db/db mice, which was found to be associated with cardiomyocyte apoptosis in DCM. Importantly, cardiac-specific overexpression of CAV3 effectively inhibited the progression of DCM, as it protected against cardiac dysfunction and cardiac remodeling associated by alleviating cardiomyocyte mitochondrial dysfunction. Furthermore, mass spectrometry analysis and immunoprecipitation assays indicated that CAV3 interacted with NDUFA10, a subunit of mitochondrial complex I. CAV3 overexpression reduced the degradation of lysosomal pathway in NDUFA10, restored the activity of mitochondrial complex I and improved mitochondrial function. Finally, our study demonstrated that CAV3 overexpression restored mitochondrial function and subsequently alleviated DCM partially through NDUFA10. CONCLUSIONS: The current study provides evidence that CAV3 expression is significantly downregulated in DCM. Upregulation of CAV3 interacts with NDUFA10, inhibits the degradation of lysosomal pathway in NDUFA10, a subunit of mitochondrial complex I, restores the activity of mitochondrial complex I, ameliorates mitochondrial dysfunction, and thereby protects against DCM. These findings indicate that targeting CAV3 may be a promising approach for the treatment of DCM.

Tuning the Emission of a Nonconventional Aggregation-Induced Emission Polymer via Silicon-Bridged Twisted Intramolecular Charge Transfer for Targeted Delivery and Visualized Drug Release.

The design of tunable luminescent biomaterials with large Stokes shifts is usually pursued by a twisted intramolecular charge transfer (TICT) effect with switchable emission colors in response to various external stimuli. However, such a strategy is usually realized in conjugated molecules containing benzene or its derivatives and consequently suffers from poor biocompatibility. In this work, a hyperbranched polysiloxane (HBPSi)-based non-conjugated fluorescent polymer with TICT and aggregation-induced emission (AIE) features is developed, and its luminescent properties, fluorescence mechanism, and potential applications are investigated. Initially, the non-conjugated HBPSi exhibits remarkable AIE characteristics due to the formation of through-space conjugation. With the introduction of the sulfur atom, a non-conjugated D-A type AIE material, HBPSi-Cys, that exhibits a dual-state emission with a large Stokes shift of 213 nm, is obtained. The correlation of the lower-energy emission band with solvent polarity suggests the existence of the TICT state. TICT and AIE characteristics direct different properties of HBPSi-Cys, with TICT regulating solvatochromic emission wavelengths and AIE manipulating the emission intensity with a compensation effect. Density functional theory calculations reveal that the non-conjugated D-A structure in HBPSi-Cys was formed across the silicon bridge, with auxochromic sulfhydryl groups and adjacent amide groups as acceptor units and amine and hydroxyl groups as donor units. Additionally, the AIE-active HBPSi could be utilized as a fluorescent probe for the analysis of metal ions. After grafting the AS1411 aptamer to HBPSi-Cys as the recognition motif, HBPSi-Apt possesses excellent targeted bioimaging, drug loading, pH/GSH dual-responsive drug release, and visualized drug delivery performance. This work provides a new way to design functional AIE polymers with tunable optical properties, and the synthesized HBPSi-Cys shows great potential as a smart fluorescent biomaterial.

Attributes underlying patient choice of treatment modality for low-grade squamous intraepithelial lesion complicated by high-risk human papillomavirus infection.

OBJECTIVE: To use logistic regression to analyze the attributes underlying patients' treatment options for low-grade squamous intraepithelial lesion (LSIL) complicated with high-risk human papillomavirus (HR-HPV) infection, and identify the best benefit group of different treatment options. METHODS: Clinical data of 197 LSIL patients with HR-HPV infection between June 2009 and February 2022 were collected. According to the treatment options chosen by the patients, they were divided into the interferon, photodynamic therapy, follow-up observation, and focused ultrasound (FUS) treatment groups. One-way analysis of variance (ANOVA) and multivariate logistic regression analysis were used to analyze the influencing factors, including age, occupation, education level, maternity history, reason for encounter, route of consultation, annual personal and household income, screening for related risk factors, and identifying the best benefit group of different treatment options. RESULTS: One-way ANOVA revealed a statistically significant difference in age, education level, maternity history, reason for encounter, and annual household income (p < 0.05). Multivariate logistic regression analysis was performed on these five factors, indicating that age ≤35 years, high school educational level or higher, and no childbirth history were independent risk factors influencing patients' choices of FUS treatment. The receiver operating characteristic curve was used to determine the age threshold of 31 years. CONCLUSION: Age, educational level, and maternity history were independent risk factors influencing patients' choice of treatment modality for LSIL complicated with HR-HPV infection. Age ≤31 years, high school, equivalent, or higher educational level, and no childbirth yielded a higher rate of choosing FUS treatment for LSIL patients with HR-HPV infection.

Comparison of the efficacy of focused ultrasound at different focal depths in treating vulvar lichen sclerosus.

OBJECTIVE: This study aimed to compare the efficacy and safety of focused ultrasound (FU) at different focal depths in treating vulvar lichen sclerosus (VLS). METHODS: A retrospective study was conducted on 84 patients with VLS. Among them, 43 cases were treated with FU at a focal depth of 2.5 mm and 41 cases at a focal depth of 4.0 mm. Therapeutic time, treatment energy, postoperative efficacy, complications and recurrence rates were compared. RESULTS: No statistically substantially differences in age, disease course, history of immune system diseases, lesion size and severity of symptoms were found between the two groups. All patients successfully received FU therapy. No significant difference in curative rate was observed between the two groups at 3, 6 and 12 months after FU therapy. At 12 months after FU therapy, the recurrence rate of the experimental group (FU treatment at 2.5 mm focal depth) was lower than the control group (FU treatment at 4.0 mm focal depth) (7.0% vs 24.4%, p = 0.027). The experimental group was treated for a shorter period of time [22.69 ± 0.64 (min) vs 24.93 ± 0.72(min), p = 0.022] and at a lower dose[5,026.05 ± 148.00(J) vs 5,484.26 ± 160.60(J) p = 0.039]. CONCLUSION: Compared with that at the routine focal depth (4.0 mm), FU therapy at a low treatment depth (2.5 mm) can achieve a similar therapeutic effect but lower recurrence rate, therapeutic time and treatment energy. This work provides insight into the optimization of clinical protocols.

DROOPY LEAF1 controls leaf architecture by orchestrating early brassinosteroid signaling.

Leaf architecture directly determines canopy structure, and thus, grain yield in crops. Leaf droopiness is an agronomic trait primarily affecting the cereal leaf architecture but the genetic basis and underlying molecular mechanism of this trait remain unclear. Here, we report that DROOPY LEAF1 (DPY1), an LRR receptor-like kinase, plays a crucial role in determining leaf droopiness by controlling the brassinosteroid (BR) signaling output in Setaria, an emerging model for Panicoideae grasses. Loss-of-function mutation in DPY1 led to malformation of vascular sclerenchyma and low lignin content in leaves, and thus, an extremely droopy leaf phenotype, consistent with its preferential expression in leaf vascular tissues. DPY1 interacts with and competes for SiBAK1 and as a result, causes a sequential reduction in SiBRI1-SiBAK1 interaction, SiBRI1 phosphorylation, and downstream BR signaling. Conversely, DPY1 accumulation and affinity of the DPY1-SiBAK1 interaction are enhanced under BR treatment, thus preventing SiBRI1 from overactivation. As such, those findings reveal a negative feedback mechanism that represses leaf droopiness by preventing an overresponse of early BR signaling to excess BRs. Notably, plants overexpressing DPY1 have more upright leaves, thicker stems, and bigger panicles, suggesting potential utilization for yield improvement. The maize ortholog of DPY1 rescues the droopy leaves in dpy1, suggesting its conserved function in Panicoideae. Together, our study provides insights into how BR signaling is scrutinized by DPY1 to ensure the upward leaf architecture.

Corralarenes: A Family of Conjugated Tubular Hosts.

Despite the advances in host-guest chemistry, macrocyclic hosts with deep cavities are far from abundant among the large number of wholly synthetic hosts described in the literature. Herein, we describe the design and synthesis of two new tubular hosts, namely, corral[4]arene and corral[5]arene. The former has been isolated and characterized as two conformational diastereoisomers, one is centrosymmetric and the other asymmetric. The latter, a fivefold symmetrical and flexible host, has also been investigated in detail. It is composed of five 4,4'-dimethoxybiphenyl units bridged by ethynylene linkers at their 2,2'-positions and adopts a pentagonal conformation with a tubular-shaped cavity in the presence of guests. This structure endows corral[5]arene not only with a conjugated backbone, capable of bright fluorescent emission (quantum yield, 56%), but also a deep π-electron-rich aromatic cavity with remarkable conformational flexibility. The adaptive cavity of corral[5]arene allows it to accommodate a wide range of neutral and positively charged electron-deficient guests with different molecular sizes and shapes. Binding constants between this host and these guests in three different nonpolar organic solvents lie in the range of 103 to 107 M-1. Moreover, corral[5]arene exhibits dynamic chirality on account of the axes of chirality associated with each of the five biphenyl units and displays first-order transformation as exhibited by circular dichroism in response to the addition of chiral guests. All these stereochemical features render corral[5]arene an attractive host for a variety of supramolecular and nanotechnological applications.

Pteris vittata plantation decrease colloidal phosphorus contents by reducing degree of phosphorus saturation in manure amended soils.

The agricultural use of manure fertilizer increases the phosphorus (P) saturation of soils and the risk of colloidal P (Pcoll) release to aquatic ecosystems. Two experiments were conducted to identify whether Pteris vittata plantation can decrease Pcoll contents in two soils (Cambisol and Anthrosol) amended with various manure P rates (0, 10, 25, and 50 mg P kg-1 of soil). The total Pcoll contents in manured soil without P. vittata were 1.14-3.37 mg kg-1 (Cambisol), and 0.01-2.83 mg kg-1 (Anthrosol) across manure-P rates. The corresponding values with P. vittata were 0.97-2.33 mg kg-1 (Cambisol) and 0.005-1.6 mg kg-1 (Anthrosol). Experimentally determined colloidal minerals (Fe, Al, Ca), colloidal total organic carbon, Mehlich-3 nutrients (Fe, Al, and Ca), and the degree of P saturation were good predictors of Pcoll concentrations in both soils with and without P. vittata plantation. In unplanted soils, P adsorption decreased and the degree of P saturation increased which released more Pcoll. However, P. vittata plantation decreased the Pcoll release and P loss risk due to the increase of P adsorption and reduced DPS in both soils. The P fractions (NaOH, NH4F, and HCl-P) contributed to increase the P pool in planted soils which enhanced the bioavailability of Pcoll and increased the P. vittata biomass. It suggested that P. vittata plantation was an effective approach to reduce Pcoll release from manure amended soils.

Hybrid Metal-Boron Diatomic Site Embedded in C2 N Monolayer Promotes C-C Coupling in CO2 Electroreduction.

Double-atom catalyst (DAC) has gained much interest for its versatile tuning and synergistic effect of dual-atom active sites. Metal (M)-metal (M) diatomic sites, either homo- or heteronuclear, are typically researched. Hybrid metal-non-metal combined sites have rarely been studied and even the viability of such active sites are unknown. Herein, CO2 electroreduction (CO2 RR) is explored on M@X-C2 N (M = Fe, Co, Ni, and Cu; X = S, P, and B) which renders naturally generated M-X diatomic site. Using spin-polarized density functional theory coupled with computational hydrogen electrode model, it is demonstrated that the functionality of hybrid M-B dual-atom center is superior over that of a single- or double-M center in driving CO2 RR especially C-C coupling. Among metal-boron DACs studies, Fe@B-C2 N (µ = 2µB ) exhibits the lowest free energy barrier of 0.17 eV in C-C coupling whereas Ni@B-C2 N (µ = 0µB ) mainly produces CH4 with the lowest barrier of 0.42 eV. Hence, the electronic spin state of M can be particularly important in modulating selectivity and C-C coupling barrier in CO2 RR. Fe@B-C2 N is predicted as the promising catalyst for CO2 RR towards C2+ products owing partially to its enhanced spin state. The findings can enrich the design strategy of electrocatalysts normally running at ambient conditions.

Titanium-interlayer mediated hydroxyapatite coating on polyetheretherketone: a prospective study in patients with single-level cervical degenerative disc disease.

BACKGROUND: Currently, there are limited reports regarding investigation of the biological properties of polyetheretherketone (PEEK) coated with titanium (Ti) and hydroxyapatite (HA) in human. The objective of this study is to evaluate the in vivo response of the PEEK cages coated with Ti and HA versus uncoated PEEK cages after anterior cervical discectomy and fusion (ACDF) in patients with single-level cervical degenerative disc disease (CDDD). METHODS: Twenty-four patients with PEEK cages coated with Ti and HA (PEEK/Ti/HA group) were matched one-to-one with patients with uncoated PEEK cages (PEEK group) based on age, gender, and operative segment. All patients had been followed up for more than 2 years. Radiological assessments included intervertebral height (IH), C2-7 angle (C2-7a), segmental alignment (SA), and fusion rate. Clinical parameters included Visual Analogue Scale (VAS) and Japanese Orthopedic Association (JOA) scores. RESULTS: There was no statistical difference in SA, IH, and C2-7a between the two groups before and after surgery and all these parameters were restored postoperatively. The fusion rate of PEEK/Ti/HA group was significantly higher than PEEK group at 3-month post-operation (87.5% vs. 62.5%). At the last follow-up, the fusion rate of the both groups achieved 100%. The VAS and JOA scores were comparable between two groups and improved postoperatively. CONCLUSIONS: In patients with single-level ACDF, PEEK cage coated with Ti and HA provided a higher fusion rate than uncoated PEEK cage at 3-month post-operation, while both two cages could achieve solid osseous fusion at the last follow up. Compared with the uncoated PEEK cage, PEEK/Ti/HA cage yielded similar favorable segmental and overall cervical lordosis, IH, and clinical outcomes after the surgery.

3D-Printing Biodegradable PU/PAAM/Gel Hydrogel Scaffold with High Flexibility and Self-Adaptibility to Irregular Defects for Nonload-Bearing Bone Regeneration.

A three-dimensional (3D) printed biodegradable hydrogel scaffold with a strong self-expanding ability to conform to the contour of irregular bone defects and be closely adjacent to host tissues is reported herein. The scaffold has a triple cross-linked network structure consisting of photo-cross-linked polyacrylamide (PAAM) and polyurethane (PU) as the primary IPN network and chemical cross-linked gelatin (Gel) as the secondary network, which confers the scaffold with good mechanical properties. The addition of PU in the polymerization process of acrylamide (AAM) can improve the ultraviolet (UV) photocuring efficiency of the hydrogel and incorporate abundant hydrogen bonds between the PAAM copolymer chain and the PU chain. The results show that the hydrogel scaffold contains regular structures with smooth morphology, excellent dimensional stability, and uniform aperture. The degradation rate of the hydrogel scaffold is controllable through adjusting cross-linking agents and can be up to about 60% after degradation for 28 days. More importantly, the rapid self-inflating characteristic of the scaffold in water, that is, the volume of hydrogel scaffold can increase to about 8 times that of their own in an hour and can generate a slight compressive stress on the surrounding host tissue, thus stimulating the reconstruction and growth of new bone tissues. The in vitro experiment indicates that the scaffold is nontoxic and biocompatible. The in vivo experiment shows that the PU/PAAM/Gel chemically cross-linked scaffold displays the desirable osteogenic capability. This UV-curable 3D printed self-adaptive and degradable hydrogel scaffold holds great potential for nonload-bearing bone repair.

Titanium interlayer-mediated hydroxyapatite-coated polyetheretherketone cage in transforaminal lumbar interbody fusion surgery.

BACKGROUND: The variance in clinical responses to polyetheretherketone (PEEK) cages with titanium (Ti) and hydroxyapatite (HA) coatings (PEEK-Ti-HA cages) is still not clear. In this study, we aimed to evaluate the radiographic and clinical outcomes of patients undergoing TLIF using PEEK-Ti-HA cages with a particular focus on fusion rate. METHODS: A prospective and nonrandomized study was conducted to compare the outcomes of PEEK-Ti-HA cages (group A, n = 32) and uncoated PEEK cages (group B, n = 32). The follow up time was at least 2 years. The radiographic assessments included the regional lordosis (RL), disc height (DH), and fusion rate. The clinical indexes included the Japanese Orthopedic Association (JOA) scores and visual analog scale (VAS) scores (back and leg). RESULTS: No significant differences were found in the pre- and postoperative RL and DH between Group A and Group B. And RL and DH, even if there were any variance initially, were restored not long after surgery in both groups. Though Group A had a significantly higher fusion rate than group B at 3 months post-surgery (93.7% vs. 75.0%), the fusion rates for the two groups reached the same level (100%) when it comes to the final follow-up. Additionally, differences of VAS and JOA scores for the two groups in general approximate. CONCLUSIONS: PEEK-Ti-HA cages, in contrast with uncoated PEEK cages, produced a better fusion rate at 3 months after single-level TLIF. The fusion rates of both groups could get 100% at the final follow-up. PEEK-Ti-HA cages could achieve similar RL, DH, JOA scores and VAS scores in comparison with uncoated PEEK cages post-surgery.

Ion momentum imaging study of the ion-molecule reaction Ar+ + O2→ Ar + O2.

Charge exchange reactions between Ar+(2P) and O2 (X3Σ) are investigated in the collision energy range of 3.40-9.24 eV within the center-of-mass coordinate, by using the ion momentum imaging technique. The internal energy of the product O2+ is enhanced gradually with the increase of collision energy, and the forward-scattered O2+ ions are distributed in the broader range of scattering angle at higher collision energies. At the low collision energy of 3.40 eV, the resonant charge transfer, similar to a photon ionization process, leads to the Franck-Condon-like vibrational state population of O2+ at the a4Πu state. At the higher collision energies, besides a4Πu and the high-lying states that are visible in the photoionization process, the O2+ products could be populated at some electronically bound states in the non-Franck-Condon region. The present observations indicate again the strong collision-energy dependences of the charge exchange reactions, but distinctly different from our previous findings for Ar+ + NO → Ar + NO+.

Collision-Energy Dependence of the Ion-Molecule Charge-Exchange Reaction Ar+ + CO → Ar + CO.

Ion-molecule charge-exchange reactions Ar+ + CO → Ar + CO+ at the center-of-mass collision energies of 4.40, 6.40, and 8.39 eV are investigated using ion velocity map imaging technique. Although multiple electronically excited states of CO+ are accessed, the population of CO+ at the A2Π state is predominant in the present collision-energy range. In contrast to our previous study for NO, but similar to the case of O2, the forward-scattered CO+ yields show a broader angular distribution at the higher collision energy. Typically, the Franck-Condon-region charge transfer, energy resonant charge transfer, and intimate collision are three different mechanisms in which the intimate collision experiences an intermediate complex, and this mechanism usually plays an essential role in the thermal-energy reactions. However, the present observations indicate that this mechanism, concerning the intermediate (Ar-CO)+, is still of utmost importance in a relatively high collision-energy range.

Cloning and characterization of a glycosyltransferase from Catharanthus roseus for glycosylation of cardiotonic steroids and phenolic compounds.

OBJECTIVES: To characterize a glycosyltransferase (UGT74AN3) from Catharanthus roseus and investigate its specificity toward cardiotonic steroids and phenolic compounds. RESULTS: UGT74AN3, a novel permissive GT from C. roseus, displayed average high conversion rate (> 90%) toward eight structurally different cardiotonic steroids. Among them, resibufogenin, digitoxigenin, and uzarigenin gave 100% yield. Based on LC-MS, 1H-NMR and 13C-NMR analysis, structure elucidation of eight glycosides was consistent with 3-O-ß-D-glucosides. We further confirmed UGT74AN3 was permissive enough to glycosylate curcumin, resveratrol, and phloretin. The cDNA sequence of UGT74AN3 contained an ORF of 1,425 nucleotides encoding 474 amino acids. UGT74AN3 performed the maximum catalytic activity at 40 °C, pH 8.0, and was divalent cation-independent. Km values of UGT74AN3 toward resibufogenin, digitoxigenin, and uzarigenin were 7.0 µM, 12.3 µM, and 17.4 µM, respectively. CONCLUSIONS: UGT74AN3, a glycosyltransferase from a noncardenolide-producing plant, displayed catalytic efficiency toward cardiotonic steroids and phenolic compounds, which would make it feasible for glycosylation of bioactive molecules.

Ion Momentum Imaging Study of the Ion-Molecule Charge Exchange Reaction of Ar+ + CO2.

Three-dimensional ion momentum imaging is developed in a combination of ion velocity map imaging technique and delay-line anode ion detection, and it is applied for the ion-molecule charge exchange reaction between Ar+ and CO2. In a center-of-mass collision energy range of 7.23-15.96 eV, CO2+ products are primarily populated at the ground state X2Πg and the single-electron excited states A2Πu, B2Σu+, and C2Σg+; the multielectron excited states of CO2+ are also found at the higher collision energies. The production efficiency profiles of CO2+ are distinctly different from the photoionization electron spectrum of CO2, implying that the charge transfer from Ar+ would be not fast as expected. The strong electron correlations in the short-lived intermediate (Ar-CO2)+ should be responsible for the CO2+ yields at the multielectron excited states.

An RNAi-mediated screen identifies novel targets for next-generation antiepileptic drugs based on increased expression of the homeostatic regulator pumilio.

Despite availability of a diverse range of anti-epileptic drugs (AEDs), only about two-thirds of epilepsy patients respond well to drug treatment. Thus, novel targets are required to catalyse the design of next-generation AEDs. Manipulation of neuron firing-rate homoeostasis, through enhancing Pumilio (Pum) activity, has been shown to be potently anticonvulsant in Drosophila. In this study, we performed a genome-wide RNAi screen in S2R + cells, using a luciferase-based dPum activity reporter and identified 1166 genes involved in dPum regulation. Of these genes, we focused on 699 genes that, on knock-down, potentiate dPum activity/expression. Of this subgroup, 101 genes are activity-dependent based on comparison with genes previously identified as activity-dependent by RNA-sequencing. Functional cluster analysis shows these genes are enriched in pathways involved in DNA damage, regulation of cell cycle and proteasomal protein catabolism. To test for anticonvulsant activity, we utilised an RNA-interference approach in vivo. RNAi-mediated knockdown showed that 57/101 genes (61%) are sufficient to significantly reduce seizure duration in the characterized seizure mutant, parabss. We further show that chemical inhibitors of protein products of some of the genes targeted are similarly anticonvulsant. Finally, to establish whether the anticonvulsant activity of identified compounds results from increased dpum transcription, we performed a luciferase-based assay to monitor dpum promoter activity. Third instar larvae exposed to sodium fluoride, gemcitabine, metformin, bestatin, WP1066 or valproic acid all showed increased dpum promoter activity. Thus, this study validates Pum as a favourable target for AED design and, moreover, identifies a number of lead compounds capable of increasing the expression of this homeostatic regulator.

Physicochemical and biological characteristics of BMP-2/IGF-1-loaded three-dimensional coaxial electrospun fibrous membranes for bone defect repair.

Coaxial electrospun fibrous membranes show favorable mechanical properties for use in guided bone regeneration (GBR). We used coaxial electrospinning technology to fabricate three-dimensional nanofiber membranes loaded with BMP-2 and IGF-1, and assessed the physicochemical and biological properties of these novel membranes in vitro. We fabricated four experimental groups of BMP-2/IGF-1/BSA-loaded membranes with different flow ratios (shell/core). Membrane characteristics were assessed by scanning and transmission electron microscopy, and laser confocal microscopy. Physicochemical and drug release properties were evaluated based on contact angle, mechanical property testing, X-ray diffraction analysis, and ELISA. The membranes were seeded with bone marrow-derived mesenchymal stem cells (BMMSCs) to estimate their biological properties based on cell viability and alkaline phosphatase (ALP) activity. The four membrane groups presented uniform diameters and core-shell structures. Acceleration of the shell solution flow rate increased the contact angle and mechanical properties of the fibrous membrane, while dual-factor addition did not impact fiber structure. Each drug-loaded membrane showed a gradually increasing release curve, with varying degrees of burst and sustained release. Compared to the other groups, the membranes with a core-shell flow ratio of 1:10 showed better drug-loading capacity and sustained release performance, higher biological properties and good barrier function. Optimal parameters were chosen based on the physical and chemical characteristics and biological properties of the membrane. Our results imply that the BMP-2/IGF-1/BSA-loaded coaxial electrospun fibrous membrane with optimum parameters is a suitable barrier membrane for GBR, and releases multiple factors promoting osteoconduction and osteoinduction.

Structure and Molecular Dynamics Simulations of Protein Tyrosine Phosphatase Non-Receptor 12 Provide Insights into the Catalytic Mechanism of the Enzyme.

Protein tyrosine phosphatase non-receptor 12 (PTPN12) is an important protein tyrosine phosphatase involved in regulating cell adhesion and migration as well as tumorigenesis. Here, we solved a crystal structure of the native PTPN12 catalytic domain with the catalytic cysteine (residue 231) in dual conformation (phosphorylated and unphosphorylated). Combined with molecular dynamics simulation data, we concluded that those two conformations represent different states of the protein which are realized during the dephosphorylation reaction. Together with docking and mutagenesis data, our results provide a molecular basis for understanding the catalytic mechanism of PTPN12 and its role in tumorigenesis.