In vitro and in vivo Biological Evaluation of Newly Tacrine-Selegiline Hybrids as Multi-Target Inhibitors of Cholinesterases and Monoamine Oxidases for Alzheimer's Disease.

Purpose: Alzheimer's disease (AD) is the most common neurodegenerative disease, and its multifactorial nature increases the difficulty of medical research. To explore an effective treatment for AD, a series of novel tacrine-selegiline hybrids with ChEs and MAOs inhibitory activities were designed and synthesized as multifunctional drugs. Methods: All designed compounds were evaluated in vitro for their inhibition of cholinesterases (AChE/BuChE) and monoamine oxidases (MAO-A/B) along with their blood-brain barrier permeability. Then, further biological activities of the optimizing compound 7d were determined, including molecular model analysis, in vitro cytotoxicity, acute toxicity studies in vivo, and pharmacokinetic and pharmacodynamic property studies in vivo. Results: Most synthesized compounds demonstrated potent inhibitory activity against ChEs/MAOs. Particularly, compound 7d exhibited good and well-balanced activity against ChEs (hAChE: IC50 = 1.57 µM, hBuChE: IC50 = 0.43 µM) and MAOs (hMAO-A: IC50 = 2.30 µM, hMAO-B: IC50 = 4.75 µM). Molecular modeling analysis demonstrated that 7d could interact simultaneously with both the catalytic active site (CAS) and peripheral anionic site (PAS) of AChE in a mixed-type manner and also exhibits binding affinity towards BuChE and MAO-B. Additionally, 7d displayed excellent permeability of the blood-brain barrier, and under the experimental conditions, it elicited low or no toxicity toward PC12 and BV-2 cells. Furthermore, 7d was not acutely toxic in mice at doses up to 2500 mg/kg and could improve the cognitive function of mice with scopolamine-induced memory impairment. Lastly, 7d possessed well pharmacokinetic characteristics. Conclusion: In light of these results, it is clear that 7d could potentially serve as a promising multi-functional drug for the treatment of AD.

Cytotoxic phenyl polyketides from Hypericum curvisepalum N. Robson.

(±)-Hypecurvone A (1) and B (2), two new undescribed phenyl polyketides, along with seven known analogues (3-9) were isolated from the whole plant of Hypericum curvisepalum. Chiral separation of 1 and 2 yielded two pairs of enantiomers 1a/1b and 2a/2b, respectively. The structures of these compounds were elucidated by extensive spectroscopic analyses and ECD spectra simulations. All isolates exhibited moderate cytotoxicity against human hepatocellular carcinoma SMMC-7721 cells, and compound 3 also showed weak cytotoxicity toward MGC-803 cells. The cytotoxicity of these compounds was found to be related to enhanced mitochondria-mediated apoptosis and inhibition of the G2/M phase of the cell cycle.

Three New Xanthones from Hypericum scabrum and Their Quorum Sensing Inhibitory Activities against Chromobacterium violaceum.

Quorum sensing (QS) plays an important role in the production of virulence factors and pathogenicity in pathogenic bacteria and is, therefore, a hopeful target to fight against bacterial infections. During our search for natural QS inhibitors, two new xanthonolignoids (1 and 2), each existing as a racemic mixture, one new simple oxygenated xanthone (7), and eight known analogs (3-6, 8-11) were isolated from Hypericum scabrum Linn. Chiral separation of 1 yielded a pair of enantiomers 1a and 1b. The structures of these compounds were elucidated by spectroscopic analysis and ECD (electrostatic circular dichroism) calculations. All isolates were evaluated for their QS inhibitory activity against Chromobacterium violaceum. Both 9 and 10 exhibited the most potent QS inhibitory effects with minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) values of 31.25 and 62.5 µM, respectively. Crystal violet staining was used to further evaluate the biofilm inhibition potential of compounds 7, 9 and 10, and the formation of biofilms increased with decreasing drug concentration in a classic dose-dependent manner. The results of a cytotoxicity assay revealed that compounds 7, 9 and 10 exhibited no cytotoxic activity on PC-12 cells at the tested concentration.

Streptomyces: Derived Active Extract Inhibits Candida albicans Biofilm Formation.

Candida albicans is an opportunistic pathogen that causes biofilm-associated infections. C. albicans biofilms are known to display reduced susceptibility to antimicrobials and high rates of acquired antibiotic resistance, and biofilm forming in C. albicans further hampers treatment options and highlights the need for new antibiofilm strategies. Identifying active components from desert actinomycetes strains to inhibit the formation of C. albicans biofilms represents an effective treatment strategy. In this study, actinomycetes that can inhibit C. albicans biofilm formation were isolated from the Taklimakan Desert, and the underlying mechanisms were explored. After screening the anti-C.albicans biofilm activities of culture supernatants from 170 Actinomycete strains, six strains showed significant inhibition of C. albicans biofilm formation. Microscopic examination showed a reduction in biofilm formation of C. albicans treated with supernatants from actinomycetes. Scanning electron microscopy showed that the morphological changes in biofilm cells were caused by cell membrane rupture and cell material leakage. Then, C.albicans biofilms were destroyed by changing the content of extracellular polysaccharides or degrading extracellular DNA. Finally, a preliminary study on active substances extracted from a new species (TRM43335) showed that the substances that inhibited the formation of biofilms might be peptides. This study provides preliminary evidence that desert actinomyces strains have inhibitory effects on the biofilm development of C. albicans.

Etiological Analysis of Viral Encephalitis in Children in Zhejiang Province from 2018 to 2019.

OBJECTIVE: To investigate the common pathogens of viral encephalitis (VE) in children, and to provide guidance for the empirical diagnosis and treatment of patients with VE. METHODS: A total of 227 cerebrospinal fluid (CSF) samples were collected from pediatric patients with VE in Zhejiang province from January 2018 to December 2019. The samples were tested using multiplex and singleplex Reverse Transcription-Polymerase Chain Reaction (RT-PCR) with primers specific to enterovirus (EV), varicella-zoster virus (VZV), mumps virus (MuV), cytomegalovirus (CMV), herpes simplex virus type 1 (HSV-1)/type 2 (HSV-2), Epstein-Barr virus (EBV), and human herpesvirus 6 (HHV-6). The data of the two analyses were compared and then verified using Sanger sequencing. RESULTS: Of the 227 CSF samples, 90 were shown to be positive for multiplex RT-PCR with a positivity rate of 39.65% and a 95% confidence interval (33.2%, 46.1%). EV was the most common cause of VE, followed by EBV, HHV-6, MuV, CMV, VZV, and HSV-1. Most included cases occurred in summer, accounting for 49.78% of all cases. For EV, EBV, and HSV-2, multiplex RT-PCR showed a positivity rate of 34.36%, which was not statistically different from that of 30.4% shown by singleplex RT-PCR. The sequences of EV, EBV, VZV, MuV, CMV, HSV-1, HHV-6, and HSV-2 were confirmed by sequencing the PCR products obtained from multiplex and singleplex PCR. CONCLUSIONS: In children, VE is more prevalent in the summer than in other seasons in Zhejiang province, and EV may be the most common causative pathogen.

Niga-ichigoside F1 ameliorates high-fat diet-induced hepatic steatosis in male mice by Nrf2 activation.

Hepatic lipid accumulation and oxidative stress (OS) lead to non-alcoholic fatty liver disease (NAFLD). Thus, we hypothesized that antihyperlipidemic and antioxidant activities of niga-ichigoside F1 (NI) would ameliorate events leading to NAFLD. Lanbuzheng (Geum japonicum Thunb. var. chinense), a type of wild vegetable found in Southwest China, was used to extract NI. Male C57BL/6J mice were fed a standard diet (Con) or a high-fat diet (HFD) (denoted as diet) with or without 40 mg kg-1 NI (defined as treatment) for 12 weeks. Diet-treatment interactions were observed in the final body weight, fat pad mass, respiratory exchange ratio (RER) in the daytime, and energy expenditure during the whole day. Moreover, NI alleviated hepatic steatosis, possibly by significantly interacting with HFD to regulate lipid metabolism genes (including Srebp1c, Acc1, Fasn, Scd1, Cpt1a and Fabp5). We also found significant diet-treatment interactions on superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT) activities, and thiobarbituric acid reactive substance (TBARS) levels, as well as the nuclear and cellular Nrf2 protein levels. Significant free fatty acid (FFA)-treatment interactions on Nrf2 nuclear translocation, antioxidant enzymes activities, genes in lipogenesis (Srebp1c, Acc1, Fasn, and Scd1), and fatty acid oxidation (Pparα) and transport (Fabp5 and Cd36) were also detected in 1 mM FFA-treated HepG2 cells with or without 20 µM NI. These beneficial effects of NI on oxidative stress and lipid accumulation were abolished by Nrf2 siRNA. Our data revealed that dietary NI could prevent HFD-induced hepatic steatosis, possibly via interacting with HFD to activate Nrf2 nuclear translocation to maintain a redox status, thus regulating lipid metabolism genes expressions.

[Clinical and laboratory features of macrophage activation syndrome].

OBJECTIVE: To study the clinical and laboratory features of macrophage activation syndrome (MAS) at the early stage of diagnosis, and to explore a method for early identification of MAS. METHODS: A retrospective analysis was performed for the demographic data, clinical and laboratory features, and treatment outcomes of 21 MAS patients. RESULTS: Of the 21 MAS patients, 14 had systemic juvenile idiopathic arthritis, 5 had Kawasaki disease (KD), and 2 had connective tissue disease (CTD) as primary diseases. The median time of MAS onset was 19 days. The KD patients had the shortest time of MAS onset, while the CTD patients had the longest onset time (P=0.009). The top 10 clinical symptoms were fever (95%), rash (86%), lymph node enlargement (67%), hemophagocytic phenomenon in bone marrow (63%), pulmonary disease (62%), serous effusion (62%), hepatomegaly (52%), cerebrospinal fluid abnormalities (50%), central nervous system damage (43%), and splenomegaly (38%). The median of hemoglobin level was lower than the normal value. The medians of C-reactive protein level and erythrocyte sedimentation rate were higher than the normal values. There were significant increases in serum ferritin, glutamic-pyruvic transaminase, aspartate aminotransferase, lactate dehydrogenase, and triglyceride. The median of fibrinogen level was lower than the normal value. There were significant increases in D-dimer, interleukin-6 (IL-6), interleukin-10 (IL-10), and interferon-γ (IFN-γ). Of the 21 patients, 20 were improved and discharged. CONCLUSIONS: If patients with rheumatic disease have persistent fever, hepatic dysfunction, coagulation disorders, multiple organ impairment, significantly increased IL-10 and IFN-γ, and a persistent increase in serum ferritin, the development of MAS should be considered.

One-pot construction of boronate ester based pH-responsive micelle for combined cancer therapy.

In this study, one-pot strategy for the construction of micelles loaded with two types of anticancer drugs (i.e., doxorubicin and methotrexate) together is reported. On the basis of the reaction between boronic acid and 1,2-diol to form boronate ester, the formation of amphiphiles, their self-assembly into micelles and drug encapsulation occurs simultaneously under simple dialysis at the appropriate pH condition. In the one-pot strategy, the micelle yield is high (78.2%) and the drug encapsulation efficiency of the two drugs is improved compared with that of the traditional method. The micelles can selectively increase the drug release ratio at acidic pH, showing the pH-responsive behavior inherited from the property of boronate ester. By combining doxorubicin and methotrexate, the half-maximum inhibition concentrations of the two drugs are obviously reduced, showing synergistic efficacy against cancer cells. This strategy is promising and may be expanded to various applications.

[Association of vitamin D concentrations with juvenile idiopathic arthritis].

OBJECTIVE: 25-Hydroxyvitamin D3 [25(OH)D3] is the main product of vitamin D and can reflect the absolute concentration of active vitamin D in the body. This study examined serum 25(OH)D3 levels in children with juvenile idiopathic arthritis (JIA) in order to explore the association of vitamin D concentrations with the pathogenesis and disease activity of JIA. METHODS: Serum samples were collected from 53 children confirmed as having JIA between January 2013 and March 2014, as well as 106 healthy children (control group) who underwent physical examination in the same period. Serum concentrations of 25(OH)D3 were measured using ELISA and compared between the cases and healthy controls. The association of serum 25(OH)D3 levels with JIA subtypes, ACR Pediatric 30 Score, peripheral blood C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR) were analyzed in children with JIA. RESULTS: Compared with the control group, the JIA group had significantly reduced serum 25(OH)D3 levels (median: 42.6 nmol/L vs 49.9 nmol/L; P<0.01). The percentage of subjects with severe deficiency of vitamin D in the JIA group was significantly higher than that in the control group (17.0% vs 6.6%; P<0.05). Serum 25(OH)D3 showed no significant correlations with JIA subtypes, ACR Pediatric 30 Score, CRP, and ESR in children with JIA. CONCLUSIONS: Vitamin D concentrations are significantly decreased in children with JIA. Decreased vitamin D concentrations may be associated with the pathogenesis of JIA. However, vitamin D concentrations may have no correlations with JIA subtypes, disease severity, and disease activity.

[Serum cytokine levels in children with newly diagnosed active systemic juvenile idiopathic arthritis].

OBJECTIVE: To study the changes in serum cytokines levels in children with newly diagnosed active systemic juvenile idiopathic arthritis (SJIA) and to explore the role of cytokines in the development and progression of SJIA. METHODS: Seventy-four pediatric patients with active SJIA between January 2010 and December 2013 were included in the study. Serum levels of interleukin-2 (IL-2), interleukin-4 (IL-4), interleukin-6 (IL-6), interleukine-10 (IL-10), tumor necrosis factor (TNF), and interferon gamma (IFN-γ) were measured by flow cytometry in these patients. The levels of cytokines were also determined in 202 healthy children as the control group. Routine laboratory parameters including white blood cell (WBC) count, percentage of neutrophils, hemoglobin level, platelet count, hypersensitive C-reactive protein (hs-CRP), and erythrocyte sedimentation rate (ESR) were monitored in the patient group. RESULTS: The WBC count, percentage of neutrophils, hs-CRP, and ESR in 74 cases of SJIA were significantly above the normal range, their platelet counts were within the normal range, whereas hemoglobin levels were below the normal range. Compared with the control group, the patient group showed a significantly increased level of IL-6 (P<0.01) and significantly reduced levels of IL-4, IL-10, and TNF (P<0.01). However, there were no significant changes in serum levels of IL-2 and IFN-γ in the patient group (P>0.05). In SJIA children, IL-6 level, which was significantly elevated, was negatively correlated with hemoglobin level, which was significantly reduced (r=-0.244, P<0.05). CONCLUSIONS: Serum level of IL-6 is significantly increased in children with SJIA, and it has a negative correlation with anemia.

[Risk factors for pleural lung disease in children with juvenile idiopathic arthritis].

OBJECTIVE: To investigate the risk factors for pleural lung disease (PLD) in children with juvenile idiopathic arthritis (JIA) and to provide a basis for the early diagnosis and timely treatment of this disease. METHODS: A total of 360 children with a confirmed diagnosis of JIA were enrolled, and their clinical data were retrospectively analyzed. All patients underwent a chest X-ray. The patients with PLD were assigned to PLD group, while those without PLD were assigned to non-PLD group. The clinical, imaging, and laboratory results of JIA patients with PLD were analyzed. RESULTS: Among the 360 JIA patients, 43 (11.9%) had PLD, and 9 (21%) of them had respiratory symptoms. Chest X-ray findings mainly included interstitial pneumonitis (53.5%) and pleurisy and/or pleural effusion (38.1%). In the 43 cases of JIA-PLD, 4 (9.3%) had normal chest X-ray findings but abnormal chest CT findings. The incidence of PLD was relatively high in patients aged under 3 years and those aged 12 years or above. Children with systemic JIA had a relatively high incidence of PLD. Compared with the non-PLD group, the PLD group had a significantly higher incidence of anemia, elevated white blood cell (WBC) count and IgG levels in peripheral blood, and positive rheumatoid factors or antinuclear antibodies (P<0.05). CONCLUSIONS: Among children with JIA, PLD is mostly seen in patients with systemic JIA or aged <3 years or ≥ 12 years, especially those with anemia, elevated WBC count and IgG levels, and positive rheumatoid factors or antinuclear antibodies. For JIA patients with PLD, interstitial pneumonitis is usually seen on chest X-ray or CT, but respiratory symptoms are rarely observed. Routine use of high-resolution chest CT is recommended for early diagnosis and timely treatment of PLD in children with JIA.

Expression and localization of mesothelin in developing rat pancreas.

To define a genetic network that regulates development of the pancreas, we used high-density microarray (Affymetrix) to generate transcriptional profiles of rat pancreas from five biologically significant stages of development: embryonic day 12.5 (E12.5), E15.5, E18.5, postnatal day 0 (P0) and adult. Many genes were notably highly expressed in the later gestation when islet architecture and function are gradually forming. The expression and localization of mesothelin, one of these genes, was further examined. Reverse transcription-polymerase chain reaction and Western blot analysis revealed that mRNA and protein levels of mesothelin were high from later gestation to 2-3 weeks after birth, and with relatively low but detectable expression levels in adult rat pancreas. Immunolocalization indicated that mesothelin localized not only in islet beta-cells but also in the mesenchyme of developing rat pancreas. Transient mesothelin expression was concomitant with the development of islets architecture formation, remodeling and maturation. These findings indicate that mesothelin is dynamically expressed in the developing rat pancreas and that mesothelin might be involved in some developmental events during development of rat pancreas.