Involvement of P2X7R-mediated microglia polarization and neuroinflammation in the response to electroacupuncture on post-stroke memory impairment.

PURPOSE: Post-stroke cognitive impairment (PSCI) is a common complication of ischemic stroke episodes. Memory impairment is an important component of the poststroke cognitive syndrome. Microglial activation plays a critical role in stroke-induced neuroinflammation. Previous studies have reported that electroacupuncture (EA) provides neuroprotective effects by reducing the expression levels of the Purinergic receptor P2X ligand-gated ion channel 7 (P2X7) and inhibiting neuroinflammation in rat model of ischemic stroke. Further understanding of the role and connections between P2X7R and microglial activation in EA-induced anti-inflammatory can reveal novel targets for post-stroke memory impairment treatment. METHODS: A Middle cerebral artery occlusion and reperfusion (MCAO/R) model was established. We used 2'(3')-O-(4-benzoyl) benzoyl ATP (BzATP) as a P2X7R agonist. Following MCAO/R injury, the rats underwent EA therapy at the Baihui (DU20) and Shenting (DU24) acupoints for seven consecutive days. The Barnes maze test was used to evaluate memory function. Following intervention, a T2 weighted images (T2WI) scan was performed to identify changes in cerebral infarction volume in MCAO/R rats. The levels of Interleukin-1ß (IL-1ß), Interleukin-6 (IL-6) and Interleukin-4 (IL-4), Interleukin-10 (IL-10) in the peri-infarct hippocampal were examined by ELISA. Immunofluorescence was employed to evaluate Iba-1+ / P2X7R+, Iba-1+/ iNOS+ and Iba-1+/ Arg-1+ cell populations in the peri-infarct hippocampal DG area. The protein expression of P2X7R, Nuclear factor E2-related factor 2 (Nrf2), Recombinant nlr family, pyrin domain containing protein 3 (NLRP3), Inducible nitric oxide synthase (iNOS) and Arginase-1 (Arg-1) in the peri-infarct hippocampal were investigated using western blot assays. Besides, we also measured the levels of reactive oxygen species (ROS), superoxide dismutase (SOD) and malondialdehyde (MDA). RESULTS: We found EA treatment reduced inflammation and oxidative stress, which is consistent with a decrease in P2X7R expression and improved learning and memory functions. In contrast, we found BzATP enhanced inflammation and oxidative stress. Moreover, our results showed EA treatment up-regulated Nrf2, down-regulated NLRP3, and promoted microglia M2 polarization. Finally, EA-mediated positive effects were reversed by intracerebroventricular injection of BzATP, which is consistent with an increase in P2X7R expression. CONCLUSION: EA ameliorates memory impairment in a rat model of ischemic stroke by reducing inflammation and ROS through the inhibition of P2X7R expression. In turn, this mechanism regulates Nrf2 and NLRP3 expression, suggesting EA is beneficial for ischemic stroke treatment using P2X7R as target.

CD38 x ICAM-1 Bispecific Antibody Is a Novel Approach for Treating Multiple Myeloma and Lymphoma.

The cluster of differentiation 38 (CD38) is a well-validated target for treating multiple myeloma. Although anti-CD38 mAbs have demonstrated outstanding initial responses in patients with multiple myeloma, nearly all patients eventually develop resistance and relapse. In addition, currently approved CD38 targeting therapies have failed to show monotherapy efficacy in lymphomas, where CD38 expression is present but at lower levels. To effectively target CD38 on tumor cells, we generated an antibody-dependent cellular cytotoxicity (ADCC) enhanced bispecific CD38 x intercellular cell adhesion molecule 1 (ICAM-1) antibody, VP301. This bispecific antibody targets unique epitopes on CD38 and ICAM-1 on tumor cells with reduced red blood cell binding compared with the benchmark CD38 antibody daratumumab. VP301 demonstrated potent ADCC and antibody-dependent cellular phagocytosis activities on a selected set of myeloma and lymphoma cell lines even those with low CD38 expression. In an ex vivo drug sensitivity assay, we observed responses to VP301 in multiple myeloma primary samples from relapsed/refractory patients. Moreover, VP301 demonstrated potent tumor inhibition activities in in vivo myeloma and lymphoma models. Interestingly, combination of VP301 with the immunomodulatory drug, lenalidomide, led to synergistic antitumor growth activity in an in vivo efficacy study. In conclusion, the CD38 x ICAM-1 bispecific antibody VP301 demonstrated promising efficacy and specificity toward CD38+ and ICAM-1+ tumor cells and represents a novel approach for treating multiple myeloma and lymphoma.

Modulation of entorhinal cortex-hippocampus connectivity and recognition memory following electroacupuncture on 3×Tg-AD model: Evidence from multimodal MRI and electrophysiological recordings.

Memory loss and aberrant neuronal network activity are part of the earliest hallmarks of Alzheimer's disease (AD). Electroacupuncture (EA) has been recognized as a cognitive stimulation for its effects on memory disorder, but whether different brain regions or neural circuits contribute to memory recovery in AD remains unknown. Here, we found that memory deficit was ameliorated in 3×Tg-AD mice with EA-treatment, as shown by the increased number of exploring and time spent in the novel object. In addition, reduced locomotor activity was observed in 3×Tg-AD mice, but no significant alteration was seen in the EA-treated mice. Based on the functional magnetic resonance imaging, the regional spontaneous activity alterations of 3×Tg-AD were mainly concentrated in the accumbens nucleus, auditory cortex, caudate putamen, entorhinal cortex (EC), hippocampus, insular cortex, subiculum, temporal cortex, visual cortex, and so on. While EA-treatment prevented the chaos of brain activity in parts of the above regions, such as the auditory cortex, EC, hippocampus, subiculum, and temporal cortex. And then we used the whole-cell voltage-clamp recording to reveal the neurotransmission in the hippocampus, and found that EA-treatment reversed the synaptic spontaneous release. Since the hippocampus receives most of the projections of the EC, the hippocampus-EC circuit is one of the neural circuits related to memory impairment. We further applied diffusion tensor imaging (DTI) tracking and functional connectivity, and found that hypo-connected between the hippocampus and EC with EA-treatment. These data indicate that the hippocampus-EC connectivity is responsible for the recognition memory deficit in the AD mice with EA-treatment, and provide novel insight into potential therapies for memory loss in AD.

Flexibly Controlling the Polycrystallinity and Improving the Foaming Behavior of Polylactic Acid via Three Strategies.

Controlling foamability plays the central role in preparing PLA foams with high performances. To achieve this, chain extension was often used to improve the rheological property of PLA resins; however, despite the availability of this approach, it often deteriorates the biodegradability of PLA and greatly increases the processing cost and complexity. Hence, we reported a special crystallization induction method to design PLA foams with a tunable cellular structure and a high expansion ratio. A novel crystallization-promoting agent combination (D-sorbitol, CO2, and phenylphosphonic acid zinc salt) was used to induce PLA to enhance the chain interaction force and chain mobility and to provide crystallization templets. A series of PLAs with tunable stereocomplex (Sc)/α crystallinity and rapid non-isothermal crystallization ability were obtained. The effect of various crystallization properties on the foaming behavior of PLA was studied. The results demonstrated that proper crystallization conditions (a small spherulite size, a crystallinity of 6%, and rapid crystallization ability) could virtually contribute to the optimized cellular structure with the highest cell density of 4.36 × 106 cell/cm3. When the Sc crystallinity was above 10%, PLA had a superior foamability, which thereby resulted in a high foaming expansion ratio of 16.2. A variety of cellular morphologies of PLA foams could be obtained by changing the foaming temperature and the crystallization property. The proposed crystallization-induced approach provided a useful method for controlling the cellular structure and the performances of the PLA foams.

Bivalent antibody pliers inhibit ß-tryptase by an allosteric mechanism dependent on the IgG hinge.

Human ß-tryptase, a tetrameric trypsin-like serine protease, is an important mediator of allergic inflammatory responses in asthma. Antibodies generally inhibit proteases by blocking substrate access by binding to active sites or exosites or by allosteric modulation. The bivalency of IgG antibodies can increase potency via avidity, but has never been described as essential for activity. Here we report an inhibitory anti-tryptase IgG antibody with a bivalency-driven mechanism of action. Using biochemical and structural data, we determine that four Fabs simultaneously occupy four exosites on the ß-tryptase tetramer, inducing allosteric changes at the small interface. In the presence of heparin, the monovalent Fab shows essentially no inhibition, whereas the bivalent IgG fully inhibits ß-tryptase activity in a hinge-dependent manner. Our results suggest a model where the bivalent IgG acts akin to molecular pliers, pulling the tetramer apart into inactive ß-tryptase monomers, and may provide an alternative strategy for antibody engineering.

Brain delivery of therapeutic proteins using an Fc fragment blood-brain barrier transport vehicle in mice and monkeys.

Effective delivery of protein therapeutics to the central nervous system (CNS) has been greatly restricted by the blood-brain barrier (BBB). We describe the development of a BBB transport vehicle (TV) comprising an engineered Fc fragment that exploits receptor-mediated transcytosis for CNS delivery of biotherapeutics by binding a highly expressed brain endothelial cell target. TVs were engineered using directed evolution to bind the apical domain of the human transferrin receptor (hTfR) without the use of amino acid insertions, deletions, or unnatural appendages. A crystal structure of the TV-TfR complex revealed the TV binding site to be away from transferrin and FcRn binding sites, which was further confirmed experimentally in vitro and in vivo. Recombinant expression of TVs fused to anti-ß-secretase (BACE1) Fabs yielded antibody transport vehicle (ATV) molecules with native immunoglobulin G (IgG) structure and stability. Peripheral administration of anti-BACE1 ATVs to hTfR-engineered mice and cynomolgus monkeys resulted in substantially improved CNS uptake and sustained pharmacodynamic responses. The TV platform readily accommodates numerous additional configurations, including bispecific antibodies and protein fusions, yielding a highly modular CNS delivery platform.

Target arm affinities determine preclinical efficacy and safety of anti-HER2/CD3 bispecific antibody.

Systemic cytokine release and on-target/off-tumor toxicity to normal tissues are the main adverse effects limiting the clinical utility of T cell-redirecting therapies. This study was designed to determine how binding affinity for CD3 and tumor target HER2 impact the efficacy and nonclinical safety of anti-HER2/CD3 T cell-dependent antibodies (TDBs). Affinity was found to be a major determinant for the overall tolerability. Higher affinity for CD3 associated with rapidly elevated peripheral cytokine concentrations, weight loss in mice, and poor tolerability in cynomolgus monkeys. A TDB with lower CD3 affinity was better tolerated in cynomolgus monkeys compared with a higher CD3-affinity TDB. In contrast to tolerability, T cell binding affinity had only limited impact on in vitro and in vivo antitumor activity. High affinity for HER2 was critical for the tumor-killing activity of anti-HER2/CD3 TDBs, but higher HER2 affinity also associated with a more severe toxicity profile, including cytokine release and damage to HER2-expressing tissues. The tolerability of the anti-HER2/CD3 was improved by implementing a dose-fractionation strategy. Fine-tuning the affinities for both the tumor target and CD3 is likely a valuable strategy for achieving maximal therapeutic index of CD3 bispecific antibodies.

A Novel Zirconium Modified Arylacetylene Resin: Preparation, Thermal Properties and Ceramifiable Mechanism.

With the rapid development of thermal protection systems for the aerospace industry and power electronics, polyarylacetylene (PAA) resin plays an important role because of its good mechanical properties, high glass transition temperature (Tg), low water absorption, high char yield (Yc), and the fact that there is no byproduct released in the curing process. In order to further improve the thermal property of PAA based FRP for the thermal protection field, the introduction of a zirconium element into arylacetylene is promising. In this paper, zirconium modified arylacetylene (ZAA) resin was prepared by two-step synthesis. The FTIR analysis characterized its molecular structure and confirmed the products. The viscosity of ZAA was about 6.5 Pa·s when the temperature was above 120 °C. The DSC analysis showed that the ZAA had a low curing temperature, and its apparent activation energy was 103.86 kJ/mol in the Kissinger method and 106.46 kJ/mol in the Ozawa method. The dielectric constant at 1 MHz of poly(zirconium modified arylacetylene) (PZAA) was 3.4. The TG analysis showed that the temperatures of a weight loss of 5% (Td5) and char yield (Yc) at 800 °C of PZAA were 407.5 °C and 61.4%, respectively. The XRD results showed the presence of SiO2 and ZrO2 in the PZAA residue after ablation. The XRF results showed that the contents of SiO2 and ZrO2 in PZAA residual after ablation were, respectively, 15.3% and 12.4%. The SEM showed that the surface of PZAA after ablation had been covered with a dense and rigid ceramic phase composed of ZrO2 and SiO2. Therefore, the introduction of Zr into arylacetylene greatly improved the densification of the surface after ablation, and improved the heat resistant property.

An Allosteric Anti-tryptase Antibody for the Treatment of Mast Cell-Mediated Severe Asthma.

Severe asthma patients with low type 2 inflammation derive less clinical benefit from therapies targeting type 2 cytokines and represent an unmet need. We show that mast cell tryptase is elevated in severe asthma patients independent of type 2 biomarker status. Active ß-tryptase allele count correlates with blood tryptase levels, and asthma patients carrying more active alleles benefit less from anti-IgE treatment. We generated a noncompetitive inhibitory antibody against human ß-tryptase, which dissociates active tetramers into inactive monomers. A 2.15 Å crystal structure of a ß-tryptase/antibody complex coupled with biochemical studies reveal the molecular basis for allosteric destabilization of small and large interfaces required for tetramerization. This anti-tryptase antibody potently blocks tryptase enzymatic activity in a humanized mouse model, reducing IgE-mediated systemic anaphylaxis, and inhibits airway tryptase in Ascaris-sensitized cynomolgus monkeys with favorable pharmacokinetics. These data provide a foundation for developing anti-tryptase as a clinical therapy for severe asthma.

Total Glycosides of Peony Protects Against Inflammatory Bowel Disease by Regulating IL-23/IL-17 Axis and Th17/Treg Balance.

Inflammatory bowel disease (IBD) is a group of autoimmune diseases, including ulcerative colitis and Crohn's disease, characterized by nonspecific inflammation in the gut. Total glycoside of peony (TGP) has been widely used for treatment of autoimmune diseases because of its pharmacological effects. However, it is lack of depth in whether TGP regulate T helper 17 cell (Th17) / T regulatory cell (Treg) immune balance or interleukin 23 (IL-23) / IL-17 axis to achieve the goal of treating IBD. Hence, the aim of this study was to investigate the effects of TGP on experimental colitis mice and the related mechanisms. In the present study, we demonstrated that administration of TGP effectively attenuates colonic inflammation of TNBS-induced colitis mice, mainly reflected in significantly improved clinical parameters, reduced inflammatory response and myeloperoxidase (MPO) activity, even stronger systemic immune ability and effective improvement of Th17/Treg immune disorders. In addition, there was a stronger immunosuppressive ability in a positive cluster of differentiation 4 (CD4 + ) T-lymphocytes from the TGP treated mouse colon, characterized by the inhibition of high levels of inflammatory factors and increased regulatory T cells. Importantly, high-dose TGP has similar therapeutic effects as salicylazosulfapyridine (SASP) on IBD treatment. The potential mechanisms might be, at least in part, related to the adjustment of imbalance of Th17/Treg cells and the inhibition of IL-23/IL17 inflammatory signal axis.

Characterization of the chemical composition of white chrysanthemum flowers of Hangzhou by using high-performance ion trap mass spectrometry.

In this study, high-performance liquid chromatography coupled with amaZon SL high-performance ion trap mass spectrometry was used to analyze the target components in white chrysanthemum flowers of Hangzhou. Twenty-one components were detected and identified in both white chrysanthemum flowers of Hangzhou samples by using target compound analysis. Furthermore, seven new compounds in white chrysanthemum flowers of Hangzhou were found and identified by analyzing the fragment ion behavior in the mass spectra. The established method can be expedient for the global quality investigation of complex components in herbal medicines and food.

Discovery of Novel Blood-Brain Barrier Targets to Enhance Brain Uptake of Therapeutic Antibodies.

The blood-brain barrier (BBB) poses a major challenge for developing effective antibody therapies for neurological diseases. Using transcriptomic and proteomic profiling, we searched for proteins in mouse brain endothelial cells (BECs) that could potentially be exploited to transport antibodies across the BBB. Due to their limited protein abundance, neither antibodies against literature-identified targets nor BBB-enriched proteins identified by microarray facilitated significant antibody brain uptake. Using proteomic analysis of isolated mouse BECs, we identified multiple highly expressed proteins, including basigin, Glut1, and CD98hc. Antibodies to each of these targets were significantly enriched in the brain after administration in vivo. In particular, antibodies against CD98hc showed robust accumulation in brain after systemic dosing, and a significant pharmacodynamic response as measured by brain Aß reduction. The discovery of CD98hc as a robust receptor-mediated transcytosis pathway for antibody delivery to the brain expands the current approaches available for enhancing brain uptake of therapeutic antibodies.

Adsorption of Cd(II), Cu(II) and Ni(II) ions by cross-linking chitosan/rectorite nano-hybrid composite microspheres.

Chitosan/rectorie (CTS/REC) nano-hybrid composite microsphere was prepared by changing the proportion of CTS/REC with 2:1, 3:1 and 4:1. Compared with the pure cross-linking chitosan microsphere, the nano-hybrid composite microsphere was proved to have better sorption capacity of Cd(II), Cu(II) and Ni(II), especially 2:1(CTS/REC-1). The adsorption behavior of the microsphere of Cd(II), Cu(II) and Ni(II) was investigated in single and binary metal systems. In single system, the equilibrium studies showed that the adsorption of Cd(II), Cu(II) and Ni(II) followed the Langmuir model and the pseudo-second-order kinetic model. The negative values of (ΔG) suggested that the adsorption process was spontaneous. In binary system, the combined action of the metals was found to be antagonistic and the metal sorption followed the order of Cu(II)>Cd(II)>Ni(II). The regeneration studies indicated that EDTA desorbed Cd(II), Cu(II) and Ni(II) from cross-linking microspheres better than HCl. The FT-IR and XPS spectra showed that coordination bonds were formed between Cd(II), Cu(II) and Ni(II) and the nitrogen atoms of cross-linking CTS/REC nano-hybrid composite microspheres.

Anti-CD20/CD3 T cell-dependent bispecific antibody for the treatment of B cell malignancies.

Bispecific antibodies and antibody fragments in various formats have been explored as a means to recruit cytolytic T cells to kill tumor cells. Encouraging clinical data have been reported with molecules such as the anti-CD19/CD3 bispecific T cell engager (BiTE) blinatumomab. However, the clinical use of many reported T cell-recruiting bispecific modalities is limited by liabilities including unfavorable pharmacokinetics, potential immunogenicity, and manufacturing challenges. We describe a B cell-targeting anti-CD20/CD3 T cell-dependent bispecific antibody (CD20-TDB), which is a full-length, humanized immunoglobulin G1 molecule with near-native antibody architecture constructed using "knobs-into-holes" technology. CD20-TDB is highly active in killing CD20-expressing B cells, including primary patient leukemia and lymphoma cells both in vitro and in vivo. In cynomolgus monkeys, CD20-TDB potently depletes B cells in peripheral blood and lymphoid tissues at a single dose of 1 mg/kg while demonstrating pharmacokinetic properties similar to those of conventional monoclonal antibodies. CD20-TDB also exhibits activity in vitro and in vivo in the presence of competing CD20-targeting antibodies. These data provide rationale for the clinical testing of CD20-TDB for the treatment of CD20-expressing B cell malignancies.

[Sedum sarmentosun bunge extraction ameliorated severe acute pancreatitis-induced lung injury: an experimental research]].

OBJECTIVE: To explore the effect of Sedum sarmentosum Bunge Extract (SSBE) on severe acute pancreatitis (SAP) induced acute lung injury (ALI) model rats and their excessive inflammatory reactions. METHODS: Forty-two healthy adult male Sprague-Dawley (SD) rats were randomly divided into 3 groups, the sham-operated control group (C), the SAP group (SAP), and the SSBE treated group (SSBE), 14 in each group. SAP induced ALl rat model was induced by retrograde injection of 5% sodium taurocholate (1 mL/kg) into the pancreatic duct. SSBE (100 m/kg) was administrated subcutaneously after the establishment of the SAP model. Equal dose of SSBE was injected again 12 h later. Equal volume of normal saline was administrated in the same way for rats in the C group and the SAP group. Rats were sacrificed after successful modeling and samples taken at 12 and 24 h. Pathological changes in the pancreas and the lung tissue were observed under light microscope. The ascites, serum amylase (AMS), wet/dry proportion (W/D) of the lung tissue, activities of myeloperoxidase (MPO), interleukin-1 (IL-1), IL-6, and tumor necrosis factor-alpha (TNF-alpha) were also measured. RESULTS: Ascites and serum AMS activities significantly increased; MPO, IL-1, IL-6, TNF-alpha contents, and W/D ratio also significantly increased in the SAP group, when compared with the C group (P<0.05). Compared with the SAP group, those parameters were all attenuated in the SSBE group at 12 and 24 h (P<0.05, P<0.01). Pathological changes in the pancreas and the lung tissue were alleviated in the SSBE group under light microscope. The injury degree ranged between that of the C group and the SAP group. CONCLUSION: SSBE could relieve the ALl in SAP model rats, which could be achieved through alleviating inflammation responses of SAP rats.

A new species of the genus Triplophysa (Nemacheilinae), Triplophysa qilianensis sp. nov, from Qinghai, China.

A new nemacheilid loach species, Triplophysa qilianensis, is described from Heihe River in Haibei Tibetan Autonomous Prefecture, Qinghai, China. It can be distinguished from all other species of Triplophysa by the following combination of characters: body long and compressed; skin smooth, scaleless; head short (20.1-23.1% of stand length); head convex from the position of posterior nostrils; posterior chamber of air bladder completely degenerated; intestine short, bending in zig-zag patter posterior to stomach; unbranched rays of pelvic fin ii; caudal fin forked, upper lobe and lower lobe equal in length; and pointed fin tips. A key to the known species of Triplophysa from the Heihe River is provided. 

Ultra-performance liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry analysis of the impact of bran-processing on the chemical profile of Radix Paeoniae Alba (Baishao).

In this study, a rapid and versatile ultra-performance liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry-based chemical profiling approach was applied to evaluate chemical constitution of crude and processed Radix Paeoniae Alba (RPA) samples. A total of 44 compounds were identified, among which the contents of 9 compounds in processed samples were obviously decreased and 8 compounds were increased. Furthermore, compound 28 was not found in RPA sample after stir-frying with wheat bran. The proposed method provided a chemical basis for exploring the processed mechanism of herbal medicine.

Rapid identif ication and comparative analysis of chemical constituents in herbal medicine Fufang decoction by ultra-high-pressure liquid chromatography coupled with a hybrid linear ion trap-high-resolution mass spectrometry.

This study was conducted to reveal the relation between herbal medicine Fufang decoction and a single drug in terms of material base. Da-Cheng-Qi decoction (DCQD) was used as a model. Ultrahigh-pressure liquid chromatography coupled with a hybrid linear ion trap-high-resolution mass spectrometry (UHPLC-LTQ-Orbitrap) was applied to detect and identify the main chemical compounds. This technique was also employed to determine the different chemical components. Under optimized liquid chromatography and mass spectrometry conditions, 64 components, including iridoids, flavonoids, anthraquinones and coumarins, were separated and tentatively characterized in Da-Cheng-Qi decoction. After decoction, the contents of 18 compounds were markedly changed, and two components were no longer detected in Fufang decoction compared with single-medicine decoction. The established method provided a good example for the rapid identification of complicated polar constituents in herbal medicine prescriptions.

Analysis of the influence of processing of stir-baking with glycyrrhizae on the main components of Euodiae Fructus by high-performance liquid chromatography with diode array detector.

Euodiae Fructus is one of the most commonly used Chinese herbs in China. Specifically, the crude Euodiae Fructus and its processed products of Gancao Zhi Pin are used clinically for the treatment of different diseases. In order to improve the quality control standard and evaluate the crude and processed Euodiae Fructus, in this study, a simple and sensitive high-performance liquid chromatography-diode array detector method was developed for the simultaneous determination of five major compounds in Euodiae Fructus. The results indicated that the five components had significant linear relation (r(2) ≥ 0.9997) between the peak area and the injected concentration. The average recoveries of the five components were in the range from 97.38% to 102.56%. Overall intra- and inter-day variations were less than 1.36%. The developed method can be applied to the intrinsic quality control of crude and processed Euodiae Fructus.