Reduced maximal range of ocular movements and its response to acute levodopa challenge in Parkinson's disease.

Introduction: Although restriction of vertical ocular range of motion is known to be the hallmark of progressive supranuclear palsy (PSP), the maximal amplitude of ocular movement has not been quantitatively assessed despite of accumulating evidences of oculomotor dysfunction in Parkinson's disease (PD). Here, we evaluated the maximal oculomotor range and its response to levodopa in PD, and compare findings to atypical parkinsonism. Methods: We recruited 159 healthy controls (HC) as well as 154 PD, 30 PSP, and 16 multiple system atrophy (MSA) patients. Oculomotor range was assessed using a kinetic perimeter-adapted device for the vertical and horizontal axes (four positions). Parameters were reassessed after levodopa challenge and compared among PD, PSP, and MSA patients. Results: Maximum oculomotor range in PD patients was reduced as compared to HC. Levodopa improved oculomotor range in all directions; corrective effects of upward range positively correlated with improvements in Unified Parkinson's Disease Rating Scale III and bradykinesia sub-scores among PD patients. Although oculomotor range was markedly restricted among PSP and MSA patients, the beneficial effects of levodopa was less pronounced. Reduced oculomotor range of motion was more significant among PSP as compared to PD or MSA patients; MSA patients did not significantly differ from PD patients. The range of upward gaze was optimally sensitive for differentiating among PD, PSP, and MSA patients. Conclusion: Maximum oculomotor range was reduced among PD patients significantly improved by levodopa treatment. Variations in, as well as the positively effects of levodopa on, the range of upward gaze assist diagnostic differentiation among PD, PSP, and MSA patients.

Development of a therapeutic monoclonal antibody against circulating adipocyte fatty acid binding protein to treat ischaemic stroke.

BACKGROUND AND PURPOSE: Adipocyte fatty acid-binding protein (A-FABP) exacerbates cerebral ischaemia injury by disrupting the blood-brain barrier (BBB) through inducing expression of MMP-9. Circulating A-FABP levels positively correlate with infarct size in stroke patients. We hypothesized that targeting circulating A-FABP by a neutralizing antibody would alleviate ischaemic stroke outcome. EXPERIMENTAL APPROACH: Monoclonal antibodies (mAbs) against A-FABP were generated using mouse hybridoma techniques. Binding affinities of a generated mAb named 6H2 towards various FABPs were determined using Biacore. Molecular docking studies were performed to characterize the 6H2-A-FABP complex structure and epitope. The therapeutic potential and safety of 6H2 were evaluated in mice with transient middle cerebral artery occlusion (MCAO) and healthy mice, respectively. KEY RESULTS: Replenishment of recombinant A-FABP exaggerated the stroke outcome in A-FABP-deficient mice. 6H2 exhibited nanomolar to picomolar affinities to human and mouse A-FABP, respectively, with minimal cross-reactivities with heart and epidermal FABPs. 6H2 effectively neutralized JNK/c-Jun activation elicited by A-FABP and reduced MMP-9 production in macrophages. Molecular docking suggested that 6H2 interacts with the "lid" of the fatty acid binding pocket of A-FABP, thus likely hindering the binding of its substrates. In mice with transient MCAO, 6H2 significantly attenuated BBB disruption, cerebral oedema, infarction, neurological deficits, and decreased mortality associated with reduced cytokine and MMP-9 production. Chronic 6H2 treatment showed no obvious adverse effects in healthy mice. CONCLUSION AND IMPLICATIONS: These results establish circulating A-FABP as a viable therapeutic target for ischaemic stroke, and provide a highly promising antibody drug candidate with high affinity and specificity.

The novel CDK9 inhibitor, XPW1, alone and in combination with BRD4 inhibitor JQ1, for the treatment of clear cell renal cell carcinoma.

BACKGROUND: Clear cell renal cell carcinoma (ccRCC) is a highly lethal malignancy with few therapeutic options. Cyclin­dependent kinase 9 (CDK9), a potential therapeutic target of many cancers, has been recently observed to be upregulated in ccRCC patients. Therefore, we aimed to investigate the therapeutic potential of CDK9 in ccRCC and develop a novel CDK9 inhibitor with low toxicity for ccRCC treatment. METHODS: The expression of CDK9 in ccRCC was checked using the online database and tissue microarray analysis. shRNA-mediated CDK9 knockdown and CDK inhibitor were applied to evaluate the effect of CDK9 on ccRCC. Medicinal chemistry methods were used to develop a new CDK9 inhibitor with drugability. RNA-seq and ChIP-seq experiments were conducted to explore the mechanism of action. MTS, western blotting, and colony formation assays were performed to evaluate the anti-ccRCC effects of CDK9 knockdown and inhibition in vitro. The in vivo anti-tumour efficacy was evaluated in a xenograft model. RESULTS: CDK9 is overexpressed and associated with poor survival in ccRCC. Knockdown or inhibition of CDK9 significantly suppressed ccRCC cells. XPW1 was identified as a new potent and selective CDK9 inhibitor with excellent anti-ccRCC activity and low toxicity. In mechanism, XPW1 transcriptionally inhibited DNA repair programmes in ccRCC cells, resulting in an excellent anti-tumour effect. CDK9 and BRD4 were two highly correlated transcriptional regulators in ccRCC patients, and the BRD4 inhibitor JQ1 enhanced XPW1's anti-ccRCC effects in vitro and in vivo. CONCLUSIONS: This work provides valuable insights into the therapeutic potential of CDK9 in ccRCC. The CDK9 inhibitor XPW1 would be a novel therapeutic agent for targeting ccRCC, alone or in rational combinations.

Synthesis of P-Stereogenic Phosphinamides via Pd(II)-Catalyzed Enantioselective C-H Alkynylation.

P-Stereogenic phosphinamides represent important structural elements in chiral organocatalysts and bioactive compounds. Herein, we report Pd(II)-catalyzed enantioselective C-H alkynylation using cheap commercially available l-pyroglutamic acid as a chiral ligand. A range of structurally diverse P-stereogenic phosphinamides was prepared in good yields with high enantioselectivities via desymmetrization and kinetic resolution. A tailor-made congested directing group, N-ethyl-N-(3-methylpyridin-2-yl)amino, was crucial for the reactivity.

Assessing the decision quality of artificial intelligence and oncologists of different experience in different regions in breast cancer treatment.

Background: AI-based clinical decision support system (CDSS) has important prospects in overcoming the current informational challenges that cancer diseases faced, promoting the homogeneous development of standardized treatment among different geographical regions, and reforming the medical model. However, there are still a lack of relevant indicators to comprehensively assess its decision-making quality and clinical impact, which greatly limits the development of its clinical research and clinical application. This study aims to develop and application an assessment system that can comprehensively assess the decision-making quality and clinical impacts of physicians and CDSS. Methods: Enrolled adjuvant treatment decision stage early breast cancer cases were randomly assigned to different decision-making physician panels (each panel consisted of three different seniority physicians in different grades hospitals), each physician made an independent "Initial Decision" and then reviewed the CDSS report online and made a "Final Decision". In addition, the CDSS and guideline expert groups independently review all cases and generate "CDSS Recommendations" and "Guideline Recommendations" respectively. Based on the design framework, a multi-level multi-indicator system including "Decision Concordance", "Calibrated Concordance", " Decision Concordance with High-level Physician", "Consensus Rate", "Decision Stability", "Guideline Conformity", and "Calibrated Conformity" were constructed. Results: 531 cases containing 2124 decision points were enrolled; 27 different seniority physicians from 10 different grades hospitals have generated 6372 decision opinions before and after referring to the "CDSS Recommendations" report respectively. Overall, the calibrated decision concordance was significantly higher for CDSS and provincial-senior physicians (80.9%) than other physicians. At the same time, CDSS has a higher " decision concordance with high-level physician" (76.3%-91.5%) than all physicians. The CDSS had significantly higher guideline conformity than all decision-making physicians and less internal variation, with an overall guideline conformity variance of 17.5% (97.5% vs. 80.0%), a standard deviation variance of 6.6% (1.3% vs. 7.9%), and a mean difference variance of 7.8% (1.5% vs. 9.3%). In addition, provincial-middle seniority physicians had the highest decision stability (54.5%). The overall consensus rate among physicians was 64.2%. Conclusions: There are significant internal variation in the standardization treatment level of different seniority physicians in different geographical regions in the adjuvant treatment of early breast cancer. CDSS has a higher standardization treatment level than all physicians and has the potential to provide immediate decision support to physicians and have a positive impact on standardizing physicians' treatment behaviors.

Targeting early proximal-rod component substrate FlgB to FlhB for flagellar-type III secretion in Salmonella.

The Salmonella flagellar secretion apparatus is a member of the type III secretion (T3S) family of export systems in bacteria. After completion of the flagellar motor structure, the hook-basal body (HBB), the flagellar T3S system undergoes a switch from early to late substrate secretion, which results in the expression and assembly of the external, filament propeller-like structure. In order to characterize early substrate secretion-signals in the flagellar T3S system, the FlgB, and FlgC components of the flagellar rod, which acts as the drive-shaft within the HBB, were subject to deletion mutagenesis to identify regions of these proteins that were important for secretion. The ß-lactamase protein lacking its Sec-dependent secretion signal (Bla) was fused to the C-terminus of FlgB and FlgC and used as a reporter to select for and quantify the secretion of FlgB and FlgC into the periplasm. Secretion of Bla into the periplasm confers resistance to ampicillin. In-frame deletions of amino acids 9 through 18 and amino acids 39 through 58 of FlgB decreased FlgB secretion levels while deleting amino acid 6 through 14 diminished FlgC secretion levels. Further PCR-directed mutagenesis indicated that amino acid F45 of FlgB was critical for secretion. Single amino acid mutagenesis revealed that all amino acid substitutions at F45 of FlgB position impaired rod assembly, which was due to a defect of FlgB secretion. An equivalent F49 position in FlgC was essential for assembly but not for secretion. This study also revealed that a hydrophobic patch in the cleaved C-terminal domain of FlhB is critical for recognition of FlgB at F45.

Synthesis of Chiral Sulfoxides via Pd(II)-Catalyzed Enantioselective C-H Alkynylation/Kinetic Resolution of 2-(Arylsulfinyl)pyridines.

A Pd(II)-catalyzed enantioselective C-H alkynylation of 2-(arylsulfinyl)pyridines via kinetic resolution using cheap and commercially available l-pGlu-OH as a chiral ligand is reported. A wide range of 2-(arylsulfinyl)pyridines were compatible with this protocol, giving the alkynylation products and recovered sulfoxides in high yields with high enantioselectivities (up to 99% ee). Furthermore, the enantioenriched products can be easily transformed to several other types of chiral sulfoxide scaffolds with the retention of enantiopurity.

SNAPIN Regulates Cell Cycle Progression to Promote Pancreatic ß Cell Growth.

In diabetes mellitus, death of ß cell in the pancreas occurs throughout the development of the disease, with loss of insulin production. The maintenance of ß cell number is essential to maintaining normoglycemia. SNAPIN has been found to regulate insulin secretion, but whether it induces ß cell proliferation remains to be elucidated. This study aimed to explore the physiological roles of SNAPIN in ß cell proliferation. SNAPIN expression increases with the age of mice and SNAPIN is down-regulated in diabetes. KEGG pathway and GO analysis showed that SNAPIN- interacting proteins were enriched in cell cycle regulation. B cell cycle was arrested in the S phase, and cell proliferation was inhibited after SNAPIN knockdown. The expression of CDK2, CDK4 and CCND1 proteins in the S phase of the cell cycle were reduced after SNAPIN knockdown, whereas they were increased after overexpression of SNAPIN. In addition, insulin protein and mRNA levels also increased or decreased after SNAPIN knockdown or overexpression, respectively. Conclusions: Our data indicate that SNAPIN mediates ß cells proliferation and insulin secretion, and provide evidences that SNAPIN might be a pharmacotherapeutic target for diabetes mellitus.

Pd(II)-Catalyzed enantioselective arylation of unbiased methylene C(sp3)-H bonds enabled by a 3,3'-F2-BINOL ligand.

Palladium-catalyzed asymmetric functionalization of unbiased methylene C(sp3)-H bonds is a long-standing challenge. Here, we report a Pd(ii)-catalyzed highly enantioselective arylation of unbiased methylene C(sp3)-H bonds enabled by a strongly coordinating bidentate 2-pyridinylisopropyl (PIP) directing group and an easily accessible 3,3'-F2-BINOL chiral ligand. The use of aryl iodides with the combination of 3,3'-F2-BINOL was beneficial for high enantiocontrol. A range of aliphatic amides and aryl iodides were tolerated, providing the desired arylated products in high enantioselectivities (up to 96% ee). The PIP directing group could be removed under mild conditions without erosion of enantiopurity.

Pd(II)-Catalyzed Enantioselective Intramolecular Arylation of Unbiased C(sp3)-H Bonds to Construct Chiral Benzo-ring Compounds.

The asymmetric synthesis of chiral benzo-ring containing compounds through enantioselective intramolecular arylation of unbiased methylene C(sp3)-H bonds was reported. Judicious choice of non-C2-symmetric chiral phosphoric acid (CPA) ligand is crucial for the high reactivity and enantioselectivity. The slight decrease in enantioselectivity at the late stage of the reaction was attributed to the hydrolysis of CPA ligands to the corresponding BINOL.

Late-stage functionalization of peptides via a palladium-catalyzed C(sp3)-H activation strategy.

Peptides hold great promise in proteomics, diagnostics and drug discovery. While natural peptides continue to be of key importance, chemically modified unnatural peptides have been found to show enhanced biological activities and improved therapeutic capabilities compared to their natural counterparts. Therefore, the development of efficient and versatile strategies to enable easy access to unnatural peptides is in high demand. In recent years, palladium-catalyzed direct functionalization of inert C(sp3)-H bonds has emerged as a powerful and straightforward synthetic strategy for late-stage modification of peptides. In this review, we summarize recent progress in this emerging field. For clarity, this review is organized into three sections according to the functionalization of the peptide side-chains at ß-, γ-, and δ-positions.

After In Vitro Digestion, Jackfruit Flake Affords Protection against Acrylamide-Induced Oxidative Damage.

Some studies have demonstrated that acrylamide (AA) was correlated with oxidative stress, resulting in physical damage. The jackfruit flake was an immature pulp that contained a high level of antioxidant activity. This study aimed to assess the defensive efficacy of jackfruit flake in AA-induced oxidative stress before and after simulated gastrointestinal digestion. Our results indicate that the total polyphenol content of Jackfruit flake digest (Digestive products of jackfruit flake after gastrointestinal, JFG) was diminished; however, JFG had raised the relative antioxidant capacity compared to Jackfruit flake extract (JFE). Additionally, the results of High Performance Liquid Chromatography-Mass Spectrometry (HPLC-MS) implied that a proportion of compounds were degraded/converted into other unknown and/or undetected metabolites. Further, by high content analysis (HCA) techniques, JFG markedly reduced cytotoxicity and excessive production of reactive oxygen species (ROS) in cells, thereby alleviating mitochondrial disorders. In this study, it may be converted active compounds after digestion that had preferable protective effects against AA-induced oxidative damage.

Scalable Formal Synthesis of (-)-Quinocarcin.

A scalable and unified strategy is described for the synthesis of (-)-quinocarcin, an important tetrahydroisoquinoline antitumor alkaloid. The strategy allows the practical formal synthesis of (-)-quinocarcin in 13 steps and 4.8% overall yield using N-phthaloyl-l-alanine as a chiral pool. It features the gram-scale and stereoselective synthesis of the tetrahydroisoquinoline moiety (AB ring) via Pd-catalyzed C(sp3)-H arylation and Pictet-Spengler condensation and a Cu(I)-catalyzed exo-selective [C + NC + CC] coupling reaction to generate the chiral pyrrolidine motif (D ring).

Analysis of CRISPR/Cas system of Proteus and the factors affected the functional mechanism.

BACKGROUND: The Proteus is one of the most common human and animal pathogens. Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR/Cas) are inheritable genetic elements found in a variety of archaea and bacteria in the evolution, providing immune function against foreign invasion. OBJECTIVES: To analyze the characteristics and functions of the CRISPR/Cas system in Proteus genomes, as well as the internal and external factors affecting the system. METHODS: CRISPR loci were identified and divided into groups based on the repeat sequence in 96 Proteus strains by identification. Compared the RNA secondary structure and minimum free energy of CRISPR loci through bioinformatics, the evolution of cas genes, and the effects of related elements were also discussed. RESULTS: 85 CRISPR loci were identified and divided into six groups based on the sequence of repeats, and the more stable the secondary structure of RNA, the smaller the minimum free energy, the fewer base mutations in the repeat, the more stable the CRISPR and the more complete the evolution of the system. In addition, Cas1 gene can be a symbol to distinguish species to some extent. Of all the influencing factors, CRISPR/Cas had the greatest impact on plasmids. CONCLUSIONS: This study examined the diversity of CRISPR/Cas system in Proteus and found statistically significant positive/negative correlations between variety factors (the RNA stability, free energy, etc.) and the CRISPR locus, which played a vital role in regulating the CRISPR/Cas system.

Curcumin ameliorates gestational diabetes in mice partly through activating AMPK.

CONTEXT: In vitro and in vivo research has shown that curcumin can alleviate diabetes and the relevant complications. OBJECTIVE: To investigate the effect of curcumin on gestational diabetes (GD). MATERIALS AND METHODS: C57 BL/KsJdb/+(db/+) mice and C57 BL/KsJ+/+ mice (10-12 weeks old) were divided into four groups (n = 15): normal pregnancy (C57 BL/KsJ+/+), GD (C57 BL/KsJdb/+), GD plus low dose curcumin (50 mg/kg, orally gavage every day) and GD plus high dose curcumin (100 mg/kg, orally gavage every day). The tolerance of glucose and insulin were measured on gestation day 10. Body weight at birth and litter size of offspring were investigated, and the expression of oxidative stress factors [thiobarbituric acid reactive substance (TBARS), glutathione (GSH), superoxide dismutase (SOD) and catalase (CAT)] and AMP-activated protein kinase (AMPK), phospho-AMPK, histone deacetylases 4 (HDAC4), pHDAC4 and glucose-6-phosphatase (G6Pase) in the livers were analyzed by ELISA and Western blot on gestation day 20. RESULTS: High dose curcumin could partly ameliorate the intolerance of glucose and insulin, and completely restore the litter size and the body weight of GD mice through decreased TBARS expression (p < 0.05) and increased GSH, SOD and CAT expression (p < 0.05). Enhanced AMPK activation, accompanied with decreased HDAC4 and G6Pase expression (p < 0.05) were partly contributed to the alleviation of GD mediated by curcumin. CONCLUSIONS: Although further detailed mechanism needs to be deciphered, curcumin can be considered as an alternative treatment for gestational diabetes.

Synergistic Effects of The Enhancements to Mitochondrial ROS, p53 Activation and Apoptosis Generated by Aspartame and Potassium Sorbate in HepG2 Cells.

The safety of food additives has been widely concerned. Using single additives in the provisions of scope is safe, but the combination of additives, may induce additive, synergy, antagonism and other joint effects. This study investigated the cytotoxicity of aspartame (AT) together with potassium sorbate (PS). Thiazolyl Blue Tetrazolium Bromide (MTT) assay indicated that AT and PS had IC50 values of 0.48 g/L and 1.25 g/L at 24 h, respectively. High content analysis (HCA) showed that both AT and PS had a negative effect on mitochondrial membrane potential (MMP), reactive oxygen species (ROS) and DNA damage while the joint group behaved more obviously. The biochemical assays revealed typical cell morphological changes and the activation of cytochrome c and caspase-3 verified apoptosis induced by AT together with PS. With dissipation of MMP and increase of cell membrane permeability (CMP), it indicated AT together with PS-induced apoptosis was mediated by mitochondrial pathway. Meanwhile, p53 were involved in DNA damage, and the ratio of Bax/Bcl-2 was increased. Moreover, excessive ROS induced by AT together with PS is a key initiating factor for apoptosis. All these results proved that p53 was involved in apoptosis via mitochondria-mediated pathway and the process was regulated by ROS.