Deficiency of leucine-rich repeat kinase 2 aggravates thioacetamide-induced acute liver failure and hepatic encephalopathy in mice.

BACKGROUND: Hepatic encephalopathy (HE) is closely associated with inflammatory responses. However, as a crucial regulator of the immune and inflammatory responses, the role of leucine-rich repeat kinase 2 (LRRK2) in the pathogenesis of HE remains unraveled. Herein, we investigated this issue in thioacetamide (TAA)-induced HE following acute liver failure (ALF). METHODS: TAA-induced HE mouse models of LRRK2 wild type (WT), LRRK2 G2019S mutation (Lrrk2G2019S) and LRRK2 knockout (Lrrk2-/-) were established. A battery of neurobehavioral experiments was conducted. The biochemical indexes and pro-inflammatory cytokines were detected. The prefrontal cortex (PFC), striatum (STR), hippocampus (HIP), and liver were examined by pathology and electron microscopy. The changes of autophagy-lysosomal pathway and activity of critical Rab GTPases were analyzed. RESULTS: The Lrrk2-/--HE model reported a significantly lower survival rate than the other two models (24% vs. 48%, respectively, p < 0.05), with no difference found between the WT-HE and Lrrk2G2019S-HE groups. Compared with the other groups, after the TAA injection, the Lrrk2-/- group displayed a significant increase in ammonium and pro-inflammatory cytokines, aggravated hepatic inflammation/necrosis, decreased autophagy, and abnormal phosphorylation of lysosomal Rab10. All three models reported microglial activation, neuronal loss, disordered vesicle transmission, and damaged myelin structure. The Lrrk2-/--HE mice presented no severer neuronal injury than the other genotypes. CONCLUSIONS: LRRK2 deficiency may exacerbate TAA-induced ALF and HE in mice, in which inflammatory response is evident in the brain and aggravated in the liver. These novel findings indicate a need of sufficient clinical awareness of the adverse effects of LRRK2 inhibitors on the liver.

A Single Strain of Lactobacillus (CGMCC 21661) Exhibits Stable Glucose- and Lipid-Lowering Effects by Regulating Gut Microbiota.

Type 2 diabetes (T2D) is usually accompanied by obesity and nonalcoholic fatty-liver-related insulin resistance. The link between T2D and dysbiosis has been receiving increasing attention. Probiotics can improve insulin sensitivity by regulating imbalances in microbiota, but efficacy varies based on the probiotic used. This study screened the main strain in the feces of healthy adult mice and found it to be a new Lactobacillus (abbreviated as Lb., named as CGMCC No. 21661) after genetic testing. We designed the most common Bifidobacterium longum subsp. longum (CGMCC1.2186, abbreviated as B. longum. subsp.), fecal microbiota transplantation (FMT), and Lb. CGMCC No. 21661 protocols to explore the best way for modulating dysbiosis to improve T2D. After 6 weeks of gavage in T2D mice, it was found that all three protocols had a therapeutic alleviating effect. Among them, compared with the B. longum. subsp. and FMT, the Lb. CGMCC No. 21661 showed a 1- to 2-fold decrease in blood glucose (11.84 ± 1.29 mmol/L, p < 0.05), the lowest HOMA-IR (p < 0.05), a 1 fold increase in serum glucagon-like peptide-1 (5.84 ± 1.1 pmol/L, p < 0.05), and lowest blood lipids (total cholesterol, 2.21 ± 0.68 mmol/L, p < 0.01; triglycerides, 0.4 ± 0.15 mmol/L, p < 0.01; Low-density lipoprotein cholesterol, 0.53 ± 0.16 mmol/L, p < 0.01). In addition, tissue staining in the Lb. CGMCC No. 21661 showed a 2- to 3-fold reduction in T2D-induced fatty liver (p < 0.0001), a 1- to 2-fold decrease in pancreatic apoptotic cells (p < 0.05), and a significant increase in colonic mucus layer thickness (p < 0.05) compared with the B. longum. subsp. and FMT. The glucose and lipid lowering effects of this Lb. CGMCC No. 21661 indicate that it may provide new ideas for the treatment of diabetes.

Immunotherapy and Prognosis of Non-small Cell Lung Carcinoma by Monomethoxy polyethylene glycol-hyaluronic acid-platinum Combined with Immune CT4+ and CT8+ Detection.

To investigate the changes in CT4+ and CT8+ lymphocyte subpopulations of patients with non-small cell lung carcinoma (NSCLC) by monomethoxy polyethylene glycol-hyaluronic acid-platinum (MPEG-HA-Pt) and the correlation between efficacy evaluation and the changes in T lymphocyte subpopulation, 76 NSCLC patients treated at oncology department of Chengdu First People's Hospital were selected and randomly divided into the treatment group and the control group (38 cases in each). mPEG-HA-Pt was used for the treatment of the included patients in the research. The patients in the control group were performed with traditional chemotherapy for 2 treatment courses. The changes in the T-lymphocyte subpopulation before and after the treatment were detected and the therapeutic effects on the patients in the two groups were compared. The particle size of mPEG-HA-Pt ranged between 78nm and 100nm with an average of 84.6±7.5nm. After that, a transmission electron microscope (TEM) was used to observe the spheres with uniform size. The drug loading capacity and entrapped efficiency of mPEG-HA-Pt were 18.7% and 87.4%, respectively. After the treatment for NSCLC patients by nanomicelle, cellular immune functions were all improved. In particular, cellular immune functions of the patients with good efficacy evaluation were improved more apparently.

Gynura segetum induces hepatic sinusoidal obstruction syndrome in mice by impairing autophagy.

PURPOSE: To investigate the underlying mechanism of hepatic sinusoidal obstruction syndrome (HSOS) induced by Gynura segetum by measuring autophagy in mouse models. METHODS: The model group was administered G. segetum (30 g/kg/d) by gavage, while the normal control group was administered an equal volume of saline daily for five weeks. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), hepatic histopathological examinations, and Masson staining were performed to evaluate liver injury. Liver intercellular adhesion molecule-1 (ICAM-1) and P-selectin were evaluated by immunohistochemistry. Hepatocellular apoptosis was assessed using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Protein expression levels of autophagy markers were measured using Western blot analysis. RESULTS: Gynura segetum was found to significantly induce liver injury compared with control mice, as evidenced by the increase of serum transaminases, a decrease in triglyceride levels, and histopathological changes in mice. Gynura segetum remarkably induced hepatocellular apoptosis and upregulated the expressions of ICAM-1 and P-selectin and also downregulated the protein expression levels of LC3, Atg12 and cytoplasmic polyadenylation element binding protein. CONCLUSIONS: Our results suggested that G. segetum induced liver injury with HSOS, and it was partly due to its ability to impair the autophagy pathway.

Clinical analysis of the therapeutic effect of plasma exchange on hypertriglyceridemic acute pancreatitis: A retrospective study.

BACKGROUND: The therapeutic effect of plasma exchange (PE) on hypertriglyceridemic acute pancreatitis (HTGAP) is unclear. Therefore, we aimed to explore this therapeutic effect. STUDY DESIGN AND METHODS: This study included 204 patients with HTGAP who underwent treatment at two provincial tertiary grade A hospitals in Fujian Province from October 2012 to May 2021. Patients were divided into a conventional group and a PE group. The Student's t-test and chi-square test were used for data analysis. RESULTS: Among 204 patients, 56 and 148 were included in the PE and conventional groups, respectively. After propensity score matching (PSM), the PE and conventional groups each had 42 patients. There was no significant difference in age; sex; pregnancy; comorbidities; laboratory findings; incidences of complications, and multiple organ dysfunction syndrome (MODS); organ support treatment; surgical rate; mortality; and hospital stay between the groups (p > 0.05). The total expenses were significantly higher in the PE group than in the conventional group (p < 0.05). There was no statistically significant difference in the times of PE; total volume of PE; incidences of complications, and MODS; organ support treatment; surgical rate; mortality; and hospital stay between the early PE and delayed PE groups (p > 0.05). All patients in the PE group and conventional group with acute renal failure had significantly higher D-dimer levels than those without acute renal failure (p < 0.05). DISCUSSION: Compared with conventional treatment, PE does not have a better therapeutic effect on HTGAP. The D-dimer level can predict whether patients with HTGAP will have acute renal failure.

Nilotinib, a Discoidin domain receptor 1 (DDR1) inhibitor, induces apoptosis and inhibits migration in breast cancer.

Overexpression of discoidin domain receptor 1 (DDR1) is known to enhance the malignancy of breast cancer considerably. This study reports the identification of a potent DDR1 inhibitor, nilotinib, for the treatment of breast cancer. MTT assay was used to evaluate the inhibitory activity of nilotinib and meantime we used flow cytometry to evaluate the pro-apoptotic activity of nilotinib in MCF-7 and MDA-MB-231 cells. Expression of DDR1 was manipulated in MDA-MB-231 and MCF-7 cell lines with low-level DDR1 expression by transfecting with plasmids containing shRNA. The effect of DDR1 or treatment with nilotinib on cell migration was assayed. The expression of p-DDR1, DDR1, p-ERK1/2, ERK1/2 and E-cadherin, Vimentin, Snail1, and caspase 3 were detected by western blot and immunofluorescent staining. Nilotinib in MCF-7 (IC50=0.403 µM) and MDA-MB-231 (IC50=0.819 µM) also indicated induced apoptotic cell death. After co-culturing with nilotinib (500 nM), apoptosis rate is 29.60±2.19% and 18.75±2.30%, respectively. Moreover, nilotinib effectually blocked the cellular migration of MCF-7 cells. Interestingly, the knock-down DDR1 could significantly block the migration of breast cancer, while the sensitivity of MCF-7 and MDA-MB-231 cells to nilotinib was reduced. Targeting DDR1 therapeutically could potentially affect survival and influence metabolism in breast cancer, and nilotinib could be used as a candidate for the treatment of breast cancer.

IFI16 directly senses viral RNA and enhances RIG-I transcription and activation to restrict influenza virus infection.

The retinoic acid-inducible gene I (RIG-I) receptor senses cytoplasmic viral RNA and activates type I interferons (IFN-I) and downstream antiviral immune responses. How RIG-I binds to viral RNA and how its activation is regulated remains unclear. Here, using IFI16 knockout cells and p204-deficient mice, we demonstrate that the DNA sensor IFI16 enhances IFN-I production to inhibit influenza A virus (IAV) replication. IFI16 positively upregulates RIG-I transcription through direct binding to and recruitment of RNA polymerase II to the RIG-I promoter. IFI16 also binds to influenza viral RNA via its HINa domain and to RIG-I protein with its PYRIN domain, thus promoting IAV-induced K63-linked polyubiquitination and RIG-I activation. Our work demonstrates that IFI16 is a positive regulator of RIG-I signalling during influenza virus infection, highlighting its role in the RIG-I-like-receptor-mediated innate immune response to IAV and other RNA viruses, and suggesting its possible exploitation to modulate the antiviral response.

Gynura segetum induces hepatic sinusoidal obstruction syndrome in mice by impairing autophagy

ABSTRACT Purpose: To investigate the underlying mechanism of hepatic sinusoidal obstruction syndrome (HSOS) induced by Gynura segetum by measuring autophagy in mouse models. Methods: The model group was administered G. segetum (30 g/kg/d) by gavage, while the normal control group was administered an equal volume of saline daily for five weeks. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), hepatic histopathological examinations, and Masson staining were performed to evaluate liver injury. Liver intercellular adhesion molecule-1 (ICAM-1) and P-selectin were evaluated by immunohistochemistry. Hepatocellular apoptosis was assessed using the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Protein expression levels of autophagy markers were measured using Western blot analysis. Results: Gynura segetum was found to significantly induce liver injury compared with control mice, as evidenced by the increase of serum transaminases, a decrease in triglyceride levels, and histopathological changes in mice. Gynura segetum remarkably induced hepatocellular apoptosis and upregulated the expressions of ICAM-1 and P-selectin and also downregulated the protein expression levels of LC3, Atg12 and cytoplasmic polyadenylation element binding protein. Conclusions: Our results suggested that G. segetum induced liver injury with HSOS, and it was partly due to its ability to impair the autophagy pathway.

Sensitive Colorimetric Hg2+ Detection via Amalgamation-Mediated Shape Transition of Gold Nanostars.

Reliable and sensitive methods to monitor mercury levels in real samples are highly important for environment protection and human health. Herein, a label-free colorimetric sensor for Hg2+ quantitation using gold nanostar (GNS) has been demonstrated, based on the formation of Au-Hg amalgamate that leads to shape-evolution of the GNS and changes in its absorbance. Addition of ascorbic acid (AA) to GNS solution is important for quantitation of Hg2+, mainly because it can reduce Hg2+ to Hg to enhance amalgamation on the GNSs and stabilize GNSs. In addition to transmission electron microscopy images, the distribution of circular ratios of GNSs in the presence of 2 mM AA and various concentrations of Hg2+ are used to show the morphology changes of the GNSs. Upon increasing the concentration of Hg2+, the average circular ratio of GNSs decreases, proving GNS is approaching to sphere. The morphology change alters the longitudinal localized surface plasmonic resonance (LSPR) absorbance of the GNSs significantly. Under the optimum conditions, our sensor exhibits a dynamic response for Hg2+ in the range of 1-4,000 nM with a detection limit of 0.24 nM. Upon Increasing Hg2+ concentration, the solution color changes from greenish-blue, purple to red, which can be distinguished by the naked eye when the Hg2+ concentration is higher than 250 nM. Owing to having a high surface-to-volume ratio and affinity toward Hg0, the GNS is sensitive and selective (at least 50-fold over tested metal ions like Pb2+) toward Hg2+ in the presence of AA. Practicality of this assay has been validated by the analysis of water samples without conducting tedious sample pretreatment.

Synthesis and Evaluation of Anti-inflammatory N-Substituted 3,5-Bis(2-(trifluoromethyl)benzylidene)piperidin-4-ones.

A total of 24 N-substituted 3,5-bis(2-(trifluoromethyl)benzylidene)piperidin-4-one derivatives were synthesized via aldol condensation, and their anti-inflammatory activities were evaluated. These compounds were found to have no significant cytotoxicity against mouse bone marrow cells in vitro. However, some compounds, such as c6 (N-(3-methylbenzoyl)-3,5-bis-(2-(trifluoromethyl)benzylidene)piperidin-4-one) and c10 (N-(2-chlorobenzoyl)-3,5-bis-(2-(trifluoromethyl)benzylidene)piperidin-4-one), displayed potent anti-inflammatory activity by inhibiting lipopolysaccharide (LPS)-stimulated tumor necrosis factor (TNF)-α, interleukin-6 (IL-6), IL-1ß, prostaglandin E2 (PGE2), and nitric oxide (NO) production in RAW 264.7 cells. Treatment with c6 or c10 at 2.5 or 10 mg kg-1 significantly decreased the paw edema induced by carrageenan in rats, and the anti-inflammatory effects of these compounds were found to be better than those of celecoxib or indomethacin as well as their parent compound C66 (2,6-bis-(2-(trifluoromethyl)benzylidene)cyclohexanone). Pharmacokinetic analysis indicated that c6 has better bioavailability than curcumin. Therefore, these compounds may be valuable leads for the development of new anti-inflammatory drugs.

Treatment and Management of Osteoporotic Fractures: A Nation-wide Survey of 484 Senior Orthopaedists in China.

OBJECTIVE: To investigate the treatment given to osteoporotic fracture patients by orthopaedists at major hospitals in China. METHODS: A 25-item quantitative questionnaire survey, categorized into five domains, including primary purpose of osteoporosis treatment, anti-osteoporosis therapy, calcium and vitamin D supplement, monitoring of osteoporosis, and knowledge of osteoporosis treatment, was designed to elicit information on orthopaedists' views on the treatment of osteoporotic fracture. A pre-survey test was conducted with a sample of 40 orthopaedic specialists to confirm the reliability and validity of the questionnaire. Each interview of the survey took approximately 15 min and did not directly involve any patients. The survey was conducted through face-to-face interviews at 119 tier 3 hospitals in 28 cities across Mainland China. RESULTS: A total of 484 valid responses were received. Seven in ten respondents have ≥10 years of professional practice. While two-thirds believed that osteoporosis treatment was to prevent fractures or re-fractures, 95.0% agreed that anti-osteoporosis medication should be administered to patients with a history of fragility fractures. Three in four would prescribe anti-osteoporosis medication perioperatively. Of these, 79.0% regarded bisphosphonates as the first-line drug. Approximately 86.0% of the 21-30 years cohort chose bisphosphonates compared to 71.4% for those with ≤10 years. More of the younger (≤10 years) cohort chose calcitonin compared to their older (21-30 years) colleagues (25.7% vs 11.6%). The most commonly prescribed daily dose is 800 IU for vitamin D supplements and 600 mg/day for calcium. CONCLUSIONS: Our respondents generally adhered to guidelines for the treatment and management of osteoporosis. A significant number had recommended lower dosages of vitamin D and calcium. Some differences exist between the younger cohort and their older colleagues in the prescription of pharmacological therapies. The criteria for initiating therapy should be more holistic and include other factors besides bone mineral density (BMD). Our results demonstrated that more comprehensive guidelines for osteoporosis management and a greater awareness of these guidelines by orthopaedists are needed to enable them to better manage their patients.

Design, synthesis, and biological evaluation of 3-vinyl-quinoxalin-2(1H)-one derivatives as novel antitumor inhibitors of FGFR1.

FGFR1 is well known as a molecular target in anticancer drug design. TKI258 plays an important role in RTK inhibitors. Utilizing TKI258 as a lead compound that contains a quinazolinone nucleus, we synthesized four series of 3-vinyl-quinoxalin-2(1H)-one derivatives, a total of 27 compounds. We further evaluated these compounds for FGFR1 inhibition ability as well as cytotoxicity against four cancer cell lines (H460, B16-F10, Hela229, and Hct116) in vitro. Some compounds displayed good-to-excellent potency against the four tested cancer cell lines compared with TKI258. Structure-activity relationship analyses indicated that small substituents at the side chain of the 3-vinyl-quinoxalin-2(1H)-one were more effective than large substituents. Lastly, we used molecular docking to obtain further insight into the interactions between the compounds and FGFR1.

Synthesis and Evaluation of Biological and Antitumor Activities of Tetrahydrobenzothieno[2,3-d]Pyrimidine Derivatives as Novel Inhibitors of FGFR1.

A series of tetrahydrobenzothieno[2,3-d]pyrimidine derivatives were designed, synthesized, and evaluated as inhibitors of FGFR1. These analogs were synthesized via Gewald's reaction under mild conditions. The structures of the synthesized compounds were characterized by spectroscopic data (IR, (1) H NMR and MS). Their antitumor activities were evaluated against H460, A549 and U251 cell lines in vitro. Results revealed that the tested compounds showed moderate antitumor activities. Structure-activity relationship analyses indicated that compounds with an aromatic ring substituted in the C-2 position or with larger molecules such as 3g, 4c, and 7 were more effective than others. The compound, 3g (78.8% FGFR1 inhibition at 10 µm), was identified to have the most potent antitumor activities, with IC50 values of 7.7, 18.9, and 13.3 µm against the H460, A549, and U251 cell lines, respectively. Together, the results suggested that tetrahydrobenzothieno[2,3-d]pyrimidine derivatives may serve as a potential agent for the treatment of FGFR1-mediated cancers.

Design, synthesis and preliminary biological evaluation of C-8 substituted guanine derivatives as small molecular inhibitors of FGFRs.

Two series of C-8 substituted guanine derivatives were synthesized, one bearing 2-amino substitutions and the other bearing 2-acetamide substitutions. Biological activity tests showed that almost all of them possessed some extent of antitumor activities, and were with lower toxicity against normal human liver HL7702 cells than AZD4547 (the positive control). Among them, N-[8-(4-bromo-1H-indol-3-yl)-6-hydroxy-9H-purin-2-yl]-acetamide exhibited a relatively satisfied inhibition against FGFR1 kinase with IC50 of 1.56 µM and specifically against A549 cells with IC50 of 8.28 µM and B16-F10 cells with IC50 of 6.59 µM. Above all, the introduction of large substituents such as indolyl groups at 8-position of the guanine scaffold probably achieves higher selectivity for FGFR1 as compared with AZD4547.

Synthesis and evaluation of biological and antitumor activities of 5,7-dimethyl- oxazolo[5,4-d]pyrimidine-4,6(5H,7H)-dione derivatives as novel inhibitors of FGFR1.

A series of 5,7-dimethyl-oxazolo[5,4-d]pyrimidine-4,6(5H,7H)-dione derivatives, N5a-5l, was designed, synthesized and evaluated for their FGFR1-inhibition ability as well as cytotoxicity against three cancer cell lines (H460, B16F10 and A549) in vitro. Several compounds displayed good-to-excellent potency against these cancer cell lines compared to SU5402. Structure-activity relationship analyses indicated that compounds with a rigid structure and more heteroatoms at the side chain of the parent ring were more effective than those without these substitutions. The compound N5g (37.4% FGFR1 inhibition at 1.0 µM) was identified to have the most potent antitumor activities, with IC50 values of 5.472, 4.260 and 5.837 µM against H460, B16F10 and A549 cell lines, respectively. Together, our results suggest that 5,7-dimethyl-oxazolo[5,4-d]pyrimidine-4,6(5H,7H)-dione derivatives may serve as potential agents for the treatment of FGFR1-mediated cancers.

Magnetic and pH-sensitive nanoparticles for antitumor drug delivery.

A dually responsive nanocarrier with multilayer core-shell architecture was prepared based on Fe(3)O(4)@SiO(2) nanoparticles coated with mPEG-poly(l-Asparagine). Imidazole groups (pK(a)∼6.0) were tethered to the side chains of poly(l-Asparagine) segments by aminolysis. These nanoparticles were expected to be sensitive to both magnetic field and pH environment. The obtained materials were characterized with FTIR, dynamic light scattering, ζ-potential, TEM, TGA and hysteresis loop analysis. It was found that this Fe(3)O(4)@SiO(2)-polymer complex can form nano-scale core-shell-corona trilayer particles (∼250 nm) in aqueous solution. The Fe(3)O(4)@SiO(2), poly(L-Asparagine) and mPEG segments serve as a super-paramagnetic core, a pH-sensitive shell, and a hydrophilic corona, respectively. An antitumor agent, doxorubicin (DOX), was successfully loaded into the nanocarrier via combined actions of hydrophobic interaction and hydrogen bonding. The drug release profiles displayed a pH-dependent behavior. DOX release rate increased significantly as the ambient pH dropped from the physiological pH (7.4) to acidic (5.5). This is most likely due to protonation and a change in hydrophilicity of the imidazole groups in the poly(l-Asparagine) segments. This new approach may serve as a promising platform to formulate magnetic targeted drug delivery systems.

Nanowell-array surfaces prepared by argon plasma etching through a nanopore alumina mask.

A method for preparing a glass surface containing an ordered array of nanowells is described. These nanowell arrays are prepared via a plasma-etch method using a nanopore alumina film as the etch mask. A replica of the pore structure of the alumina mask is etched into the glass. We demonstrate that chemical information in the form of negatively charged latex nanoparticles can be selectively stored within these nanowells and not indiscriminately deposited on the surface surrounding the nanowells. To accomplish this, the chemistry of the glass surfaces within these nanowells (walls and bottoms) must be different from the chemistry of the surface surrounding the nanowells. Two different procedures were developed to make the inside vs. surrounding surface chemistries different. Atomic force microscopy (AFM) was used to image the nanowells and, via friction-force measurements, to prove that the inner nanowell surfaces can be made chemically different from the surface surrounding the nanowells.

Conical nanopore membranes. Preparation and transport properties.

We have been investigating applications of nanopore membranes in analytical chemistry-specifically in membrane-based bioseparations, in electroanalytical chemistry, and in the development of new approaches to biosensor design. Membranes that have conically shaped pores (as opposed to the more conventional cylindrical shape) may offer some advantages for these applications. We describe here a simple plasma-etch method that converts cylindrical nanopores in track-etched polymeric membranes into conically shaped pores. This method allows for control of the shape of the resulting conical nanopores. For example, the plasma-etched pores may be cylindrical through most of the membrane thickness blossoming into cones at one face of the membrane (trumpet-shaped), or they may be nearly perfect cones. The key advantage of the conical pore shape is a dramatic enhancement in the rate of transport through the membrane, relative to an analogous cylindrical pore membrane. We demonstrate this here by measuring the ionic resistances of the plasma-etched conical pore membranes.