GM-CSF impairs erythropoiesis by disrupting erythroblastic island formation via macrophages.

Anemia is a significant complication of chronic inflammation and may be related to dysregulated activities among erythroblastic island (EBI) macrophages. GM-CSF was reported to be upregulated and attracted as a therapeutic target in many inflammatory diseases. Among EBIs, we found that the GM-CSF receptor is preferentially and highly expressed among EBI macrophages but not among erythroblasts. GM-CSF treatment significantly decreases human EBI formation in vitro by decreasing the adhesion molecule expression of CD163. RNA-sequence analysis suggests that GM-CSF treatment impairs the supporting function of human EBI macrophages during erythropoiesis. GM-CSF treatment also polarizes human EBI macrophages from M2-like type to M1-like type. In addition, GM-CSF decreases mouse bone marrow (BM) erythroblasts as well as EBI macrophages, leading to a reduction in EBI numbers. In defining the molecular mechanism at work, we found that GM-CSF treatment significantly decreases the adhesion molecule expression of CD163 and Vcam1 in vivo. Importantly, GM-CSF treatment also decreases the phagocytosis rate of EBI macrophages in mouse BM as well as decreases the expression of the engulfment-related molecules Mertk, Axl, and Timd4. In addition, GM-CSF treatment polarizes mouse BM EBI macrophages from M2-like type to M1-like type. Thus, we document that GM-CSF impairs EBI formation in mice and humans. Our findings support that targeting GM-CSF or reprogramming EBI macrophages might be a novel strategy to treat anemia resulting from inflammatory diseases.

Performance evaluation of a 71Ga(n,γ)72Ga reaction-based epithermal neutron flux detector at an AB-BNCT device.

The 71Ga(n,γ)72Ga reaction-based epithermal neutron flux detectors are novel instruments developed to measure the epithermal neutron flux of boron neutron capture therapy (BNCT) treatment beams. In this study, a spherical epithermal neutron flux detector using 71Ga(n,γ)72Ga reaction was prototyped. The performance of the detector was experimentally evaluated at an accelerator-based BNCT (AB-BNCT) device developed by Lanzhou University, China. Based on the experimental results and related analysis, we demonstrated that the detector is a reliable tool for the quality assurance of BNCT treatment beams.

(2S)-5, 2', 5'-trihydroxy-7-methoxyflavanone, a natural product from Abacopteris penangiana, presents neuroprotective effects in vitro and in vivo.

The aim of this study was to investigate the neuroprotective effects of (2S)-5, 2', 5'-trihydroxy-7-methoxyflavanone (TMF), a natural product from Abacopteris penangiana (Hook.) Ching, in oxidative stress-induced neurodegeneration models in vitro and in vivo. In PC12 cells, preincubation of TMF (3-20 µM) for 24 h decreased the dopamine-induced toxicity and attenuated the redox imbalance in PC12 cells through regulating the ratio of reduced glutathione/oxidized glutathione (GSH/GSSG), which is a sensitive marker of oxidative stress. Additionally, long-term intraperitoneal (i.p.) injection of TMF (4 or 8 mg/kg/day) for 2 weeks significantly improved the behavioral performance of D-galactose (D-gal) treated mice in a Morris water maze test. Biochemical analysis revealed that TMF inhibited the activation of AP-1 (activator protein-1) and upregulated the level of BDNF (brain derived neurophic factor) as well as the ratio of GSH/GSSG in the hippocampus of D-gal treated mice. Furthermore, western blotting analysis indicated that TMF increased phosphorylation of cAMP-response element-binding protein (CREB). Therefore, the natural product TMF possessed a potential for the treatment of neurodegenerative diseases.

Determination of protoapigenone in rat plasma by high-performance liquid chromatography with UV detection and its application in pharmacokinetic studies.

A simple and sensitive HPLC method using UV detection was developed to determine the concentration of protoapigenone in rat plasma. Chromatographic separation was conducted on a C18 column with a mobile phase consisting of an acetonitrile-methanol-aqueous phase (containing 0.2% acetic acid, pH 3.0) system at a flow rate of 1.0 mL/min. The UV detector was set at 248 nm. The calibration curve was linear over the range of 0.031-10.0 µg/mL. The lower limit of quantification was 31 ng/mL. The recoveries for plasma samples ranged from 70.3 to 82.5%. The intra- and inter-day accuracy and precision fulfilled the international standards. This method was successfully applied to a pharmacokinetic study of protoapigenone in rats after oral administration of protoapigenone. It was shown that protoapigenone could be absorbed rapidly after oral administration and could reach the maximum concentration within 1 h.

Genetic and functional analysis of the NKX2-5 gene promoter in patients with ventricular septal defects.

The ventricular septal defect (VSD) is the most common type of congenital heart disease (CHD). The morbidity and mortality of CHD patients are significantly higher due to late cardiac complications, likely caused by genetic defects. Mutations in cardiac transcription factor genes such as GATA-4, TBX5, and NKX2-5 have been implicated in CHD cases. The NKX2-5 gene, a homeobox gene, is expressed in the developing heart and the adult heart. Because NKX2-5 is a dosage-sensitive regulator during embryonic development, the authors hypothesized that the expression levels of the NKX2-5 gene rather than the mutant protein may play important roles in CHD. In this study, the promoter regions and exon regions of the NKX2-5 gene were bidirectionally sequenced in large cohorts of VSD patients and healthy control subjects. The results showed that a novel sequence variant (g.4574c>deletion), found only in one VSD patient, and a single nucleotide polymorphism (rs118026695), the frequency of which was significantly higher in VSD patients, were identified within the promoter region. Functional analysis confirmed that these sequence variants significantly enhanced the transcriptional activities of the NKX2-5 gene promoter, altering the expression of the NKX2-5 gene and the cardiac gene regulatory network. In addition, a synonymous mutation in the second exon of the NKX2-5 gene was identified in one VSD patient, which may affect the translation process. Therefore, the authors' data provide supportive evidence that mutations in the coding region of the NKX2-5 gene and sequence variants within its promoter region may be among the contributors to the CHD etiology.

Nephroprotective activity of Macrothelypteris oligophlebia rhizomes ethanol extract.

CONTEXT: Macrothelypteris oligophlebia (Bak.) Ching (Thelypteridaceae) is a Chinese herbal medicine used traditionally for the treatment of diseases such as edema, boils, burns, and roundworms. However, research about the nephroprotective potential of this plant is not available. OBJECTIVE: Present study was designed to evaluate the protective effect of ethanol extract of M. oligophlebia rhizomes (EMO) on gentamicin (GM)-induced nephrotoxicity. MATERIALS AND METHODS: Rats were intraperitoneal (i.p.) injected with GM (100 mg/kg) to induce nephrotoxicity and simultaneously EMO (250 and 500 mg/kg) was orally given to GM-treated rats for 8 days. Blood urea nitrogen (BUN), serum creatinine (Cr), malondialdehyde (MDA), nitric oxide (NO), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) were evaluated in renal tissues. Histopathological analysis was used for evaluation of the renal damage. RESULTS: Administration with GM-induced renal dysfunction in rats. Pre-treatment with EMO (500 mg/kg) significantly decreased the levels of BUN, Cr, MDA and NO (decreased BUN from 12.71 ± 1.28 to 7.19 ± 0.23 mmol/l, Cr from 39.77 ± 5.34 to 19.17 ± 0.90 µmol/l, MDA from 5.60 ± 0.37 to 2.63 ± 0.24 nmol/ml, and NO from 868.17 ± 22.67 to 589.51 ± 8.83 µmol/ml), and also restored the activities of renal antioxidant enzymes (SOD, CAT, and GSH-Px) (restored SOD from 1.59 ± 0.17 to 2.94 ± 0.13 U/mg protein, CAT from 3.22 ± 0.34 to 10.57 ± 0.27 U/mg protein, and GSH-Px from 9.11 ± 1.29 to 20.72 ± 1.83 U/mg protein). DISCUSSION AND CONCLUSION: Our results suggest that the rhizomes of M. oligophlebia potentially have a protective role in renal tissue against oxidative stress in acute renal failure.

Effect of Fuel Physicochemical Properties on Spray and Particulate Emissions.

To investigate the effect of fuel physicochemical properties on spray and particulate emissions, fuel spray characteristics were tested on a constant volume chamber (CVC) test rig using a high-speed camera method to investigate the effect of different injection and ambient pressures on spray characteristics. In the engine bench tests, the effects of particulate emissions from five different diesel fuels with different physicochemical properties were analyzed under low-, medium-, and high-load steady-state conditions and 5 s transient loading conditions. The test results showed that the spray tip penetration of different CNs results from the combined effect of the fuel properties. The spray cone angle of the five fuels increased with the increase of injection and ambient pressure, and the impact of ambient pressure on the spray cone angle was more prominent. Spray tip penetration and spray projection area increase with increased injection pressure and decrease with increased ambient pressure; compared with spray tip penetration, the spray cone angle has more influence on spray projection area, especially near-field spray cone angle as the primary influence factor. Fuels with different ignition characteristics have other effects on particulates at different loads. At low loads, choosing CN = 55.3 fuel improved the number and mass of particulates; at medium and high loads, choosing CN = 51 fuel reduced the number of particulate emissions. Fuels with different volatilities have different effects on particulates at other loads. At low loads, CN = 54.9 fuel was chosen with moderate volatility and aromatic content. At medium and high loads, the volatility of the fuel had a lower weight on particulates, and the aromatic content had a higher weight. Under the transient loading condition of 5 s, using fuel with a higher CN, good volatility, and lower aromatic content can appropriately reduce the number of particulate emissions.

Decreased gene expression of LC3 in peripheral leucocytes of patients with coronary artery disease.

BACKGROUND: Coronary artery disease (CAD) is a common and multifactorial arterial disease that is mainly caused by atherosclerosis. Macrophages, lymphocytes and neutrophils have been implicated in atherosclerotic plaque development. Autophagy, a highly conserved cellular process for the removal of long-lived protein and organelles, plays a variety of pathophysiological roles. However, the roles of autophagy in peripheral leucocytes in atherosclerosis and CAD have not been explored. MATERIALS AND METHODS: LC3 is a marker gene for autophagy, and LC3-II, a conjugated form of LC3 protein, is a membrane marker for autophagosome and autophagolysosomes. In this study, LC3 gene expression levels and LC3-II protein levels in peripheral leucocytes were measured in patients with CAD (n = 146) and healthy controls (n = 87). RESULTS: In patients with CAD, LC3 gene expression levels in the peripheral leucocytes were significantly decreased compared with age- and sex-matched healthy controls (P < 0·01). LC3-II protein levels were also significantly decreased in patients with CAD (P < 0·01). Multivariate logistic analyses showed that decreased LC3 gene expression levels were strongly associated with CAD. There were no differences in LC3 transcripts and LC3-II protein levels between subgroups of patients with CAD. CONCLUSIONS: LC3 gene expression in the peripheral leucocytes was significantly decreased in patients with CAD, indicating that autophagosome formation is decreased. These data suggest that autophagy in circulating leucocytes may be involved in the pathogenesis of atherosclerosis and CAD.

Decreased expression of lysosomal alpha-galactosiase A gene in sporadic Parkinson's disease.

Parkinson's disease (PD) is a progressive neurodegenerative disease. To date, the causal genes and variants associated with sporadic PD are largely unknown. Accumulating evidence demonstrates that autophagy delivers alpha-syncuclein proteins to lysosome for degradation and dysfunctional autophagy is involved in the PD pathogenesis. We have previously screened a group of lysosomal hydrolases and found that alpha-galactosidase A (GLA) activity is significantly decreased in the peripheral leukocytes of sporadic PD patients. In this study, GLA transcript and protein levels were semi-quantitatively examined. The GLA transcript (P = 0.020) and protein (P = 0.027) levels in the peripheral leukocytes of sporadic PD patients were significantly decreased, compared to age- and sex-matched healthy controls. Furthermore, decreased GLA gene expression levels were strongly associated with sporadic PD (OR 3.33, 95%CI 1.17-9.52, P = 0.024). Therefore, our data suggest that insufficient GLA activity may contribute to the pathogenesis of sporadic PD. The underlying molecular mechanisms remain to be determined.

Neuropective constituents from the rhizomes of Abacopteris penangiana.

Two neuropective compounds were isolated from the rhizomes of Abacopteris penangiana, one was a new flavone and the other was a flavanone. Both compounds were firstly separated from natural plant. The isolation work was guided by the antioxidant activity. Both the compounds showed a significant antioxidant activity in vitro and a protective effect on dopamine-induced neurotoxicity in PC12 cells.

[Preliminary study on the efficacy and safety of oral sildenafil in pulmonary arterial hypertension in China].

OBJECTIVE: Sildenafil has been shown to be effective in pulmonary arterial hypertension (PAH). However, the impact of sildenafil on PAH has been under-investigated in China. The aim of the present study was to evaluate the efficacy and safety of oral sildenafil in PAH patients in China. METHODS: In this prospective, open-label and multi-center study, 90 patients were recruited from 14 centers to receive oral sildenafil (75 mg/d) for 12 weeks. They underwent a six-minute walk test (SMWT) and cardiac catheterization at the beginning and the end of 12 weeks. The primary endpoint was the changes in exercise capacity as assessed by SMWT. And the secondary endpoints included assessment of functional class, evaluation of cardiopulmonary hemodynamics and clinical deterioration (defined as death, transplantation and re-hospitalization for PAH). Drug safety and tolerability were also examined. RESULTS: There were 19 males and 71 females with an average age of 32.5 ± 12.1 years old (range: 18 - 61). Their etiologies were idiopathic (n = 15), related with congenital heart disease (n = 60), or related with connective tissue disease (n = 9) and chronic thromboembolic pulmonary hypertension (n = 6). Oral sildenafil significantly increased the SMWT distances [(342 ± 93) m vs. (403 ± 88) m, P < 0.001]. There was also remarkable improvement in Borg dyspnea score (2.9 ± 2.6 vs. 2.4 ± 2.0, P = 0.005). Furthermore, significant improvements in World Healthy Organization (WHO) functional class and cardiopulmonary hemodynamics were also found (mean pulmonary artery pressure, P < 0.001; cardiac index, P < 0.001; pulmonary vascular resistance, P < 0.001). Side effects were mild and consistent with other reports. CONCLUSION: This study confirms and extends previous studies. Oral sildenafil is both safe and effective for the treatment of adult PAH patients in China.

[Flavonoids with special B-ring from Macrothelypteris viridifrons and their anti-proliferative effects on tumor cell].

OBJECTIVE: To study the chemical constituents of Macrothelypteris viridifrons and their anti-proliferative effects on tumor cell. METHOD: The compounds were isolated by column chromatography with silica gel, C18 reverse-phase silica gel, sephadex LH-20, and their structures were elucidated on the basis of physiochemical propertities and spectral analysis. The antitumor activities of all compounds were tested with MOLT4, Hep G2, A-549, MCF-7, HT-29, PC-3 tumor cell lines. RESULT: Five compounds were isolated and identified as protoapigenone (1), protoapigenin (2), protoapigenin-4'-O-beta-D-glucopyanoside (3), 5,7-dihydroxy-2-(1,2-isopropyldioxy-4-oxo-cyclohex-5-enyl) -chromen-4-one (4), 5,7-dihydroxy-2-(1-hydroxy-2,6-dimethoxy-cyclohex-4-oxo) -chromen-4-one (5), respectively. CONCLUSION: All compounds were obtained from this plant for the first time. Compounds 1, 4 and 5 showed strong anti-proliferative effects on six tumor cells, which were in concentration-dependent manner.

Single-cell map of diverse immune phenotypes in the acute myeloid leukemia microenvironment.

BACKGROUND: Knowledge of immune cell phenotypes, function, and developmental trajectory in acute myeloid leukemia (AML) microenvironment is essential for understanding mechanisms of evading immune surveillance and immunotherapy response of targeting special microenvironment components. METHODS: Using a single-cell RNA sequencing (scRNA-seq) dataset, we analyzed the immune cell phenotypes, function, and developmental trajectory of bone marrow (BM) samples from 16 AML patients and 4 healthy donors, but not AML blasts. RESULTS: We observed a significant difference between normal and AML BM immune cells. Here, we defined the diversity of dendritic cells (DC) and macrophages in different AML patients. We also identified several unique immune cell types including T helper cell 17 (TH17)-like intermediate population, cytotoxic CD4+ T subset, T cell: erythrocyte complexes, activated regulatory T cells (Treg), and CD8+ memory-like subset. Emerging AML cells remodels the BM immune microenvironment powerfully, leads to immunosuppression by accumulating exhausted/dysfunctional immune effectors, expending immune-activated types, and promoting the formation of suppressive subsets. CONCLUSION: Our results provide a comprehensive AML BM immune cell census, which can help to select pinpoint targeted drug and predict efficacy of immunotherapy.

Parathelypteriside attenuates cognition deficits in d-galactose treated mice by increasing antioxidant capacity and improving long-term potentiation.

Parathelypteriside (PG), a stilbenoid compound, was extracted from Parathelypteris glanduligera (kze.) ching that exhibits antioxidative and anti-inflammatory effects. The aim of this study was to investigate the protective effect of PG against the d-galactose (d-gal)-induced neurotoxicity in mice. It was found that long-term intraperitoneal (i.p.) injection of PG (5 or 10 mg/(kg day)) for two weeks significantly improved the behavioral performance of d-gal-treated mice in both Morris water maze test and step-down avoidance test. Biochemical examination revealed that PG reduced the increased levels of malondialdehyde (MDA), and attenuated the decreased activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase in the hippocampus of d-gal-treated mice. Furthermore, the electrophysiological assay showed that PG significantly rescued the long-term potentiation (LTP) impairment in mice hippocampus, and western blotting analysis indicated that the effects of PG on LTP might be attributed to the activation of cAMP-response element-binding protein (CREB). Together, these results suggested that the natural product PG represented a potential source of medicine for the treatment of the neurodegenerative diseases.

Distinctive Gene Expression Profiles and Effectors Consistent With Host Specificity in Two Formae Speciales of Marssonina brunnea.

The knowledge on the host specificity of a pathogen underlying an interaction is becoming an urgent necessity for global warming. In this study, the gene expression profiles and the roles of effectors in host specificity were integrally characterized with two formae speciales, multigermtubi and monogermtubi, of a hemibiotrophic pathogen Marssonina brunnea when they were infecting respective susceptible poplar hosts. With a functional genome comparison referring to a de novo transcriptome of M. brunnea and Pathogen-Host Interaction database functional annotations, the multigermtubi strain showed abundant and significant differentially expressed unigenes (DEGs) (more than 40%) in colonizing the initial invasion stage and in the necrotrophic stage. The monogermtubi strain induced less than 10% of DEGs in the initial invasion stage but which abruptly increased to more than 80% DEGs in the necrotrophic stage. Both strains induced the least DEGs in the biotrophic stage compared to the initial invasion and necrotrophic stages. The orthologs of the effector genes Ecp6, PemG1, XEG1, ACE1, and Mg3LysM were exclusively induced by one of the two formae speciales depending on the infection stages. Some unigenes homologous to carbohydrate lytic enzyme genes, especially pectate lyases, were notably induced with multigermtubi forma specialis infection but not expressed in the monogermtubi forma specialis at an earlier infection stage. The extraordinary differences in the functional genome level between the two formae speciales of M. brunnea could be fundamental to exploring their host specificity determinant and evolution. This study also firstly provided the fungal transcriptome of the monogermtubi forma specialis for M. brunnea.

Decisions of the Dual-Channel Supply Chain under Double Policy Considering Remanufacturing.

Considering the preference of green consumers for remanufactured products, a dual-sale-channel supply chain model with government non-intervention, government remanufacturing subsidy policy, and carbon tax policy is constructed, respectively. The difference of the optimal decision between the firm and the government under the two policies is discussed in this paper. Meanwhile, we analyze the influence of green consumers on the government's optimal decision, based on social welfare maximization. It is found that without government intervention, social welfare is the lowest. The carbon tax policy is better when the proportion of green consumers and the environmental coefficient are extreme or moderate at the same time. Otherwise, the subsidy policy is better. The carbon tax policy is more effective than the subsidy policy in controlling carbon emissions. Profit-sharing contracts should be established by enterprises and governments to achieve win⁻win results.

The Optimal Concentration of Formaldehyde is Key to Stabilizing the Pre-Fusion Conformation of Respiratory Syncytial Virus Fusion Protein.

BACKGROUND: To date, there is no licensed vaccine available to prevent respiratory syncytial virus (RSV) infection. The valuable pre-fusion conformation of the fusion protein (pre-F) is prone to lose high neutralizing antigenic sites. The goals of this study were to stabilize pre-F protein by fixatives and try to find the possibility of developing an inactivated RSV vaccine. METHODS: The screen of the optimal fixative condition was performed with flow cytometry. BALB/c mice were immunized intramuscularly with different immunogens. The serum neutralizing antibody titers of immunized mice were determined by neutralization assay. The protection and safety of these immunogens were assessed. RESULTS: Fixation in an optimal concentration of formaldehyde (0.0244%-0.0977%) or paraformaldehyde (0.0625%-1%) was able to stabilize pre-F. Additionally, BALB/c mice inoculated with optimally stabilized pre-F protein (opti-fixed) induced a higher anti-RSV neutralization (9.7 log2, mean value of dilution rate) than those inoculated with unstable (unfixed, 8.91 log2, p < 0.01) or excessively fixed (exce-fixed, 7.28 log2, p < 0.01) pre-F protein. Furthermore, the opti-fixed immunogen did not induce enhanced RSV disease. CONCLUSIONS: Only the proper concentration of fixatives could stabilize pre-F and the optimal formaldehyde condition provides a potential reference for development of an inactivated RSV vaccine.

Decreased activities of lysosomal acid alpha-D-galactosidase A in the leukocytes of sporadic Parkinson's disease.

Parkinson's disease (PD) is a progressive neurodegenerative disease that affects aged people. Although a number of genes have been linked to familial PD, the genetic causes of sporadic PD that accounts for 90% of all PD cases remain unclear. Accumulating evidence has demonstrated that alpha-synuclein aggregation is essential to the pathogenesis of PD. Recent studies suggest that autophagic-lysosomal system play major roles in the process of alpha-synuclein aggregation. We hypothesized that lysosomal acid hydrolases may be involved in the alpha-synuclein degradation and aggregation. In this study, we examined the activities of 11 lysosomal acid hydrolases in peripheral blood leukocytes of 38 sporadic PD patients and 258 age- and sex-matched healthy controls. The activities of alpha-D-galactosidase A were significantly decreased in sporadic PD patients, compared to age- and sex-matched controls. In contrast, no significant differences of the activities of other 10 lysosomal acid hydrolases was observed. This initial study suggests that decreased activities of lysosomal alpha-D-galactosidase A in the central nervous system may be involved in the degradation and aggregation of alpha-synuclein protein and contribute to the pathogenesis of sporadic PD as a risk factor.

Smart fluorescent probes for in situ imaging of enzyme activity: design strategies and applications.

Enzymes play critical roles in the physiological and pathological processes of living systems. To provide detailed pictures of enzyme activity at the molecular and cellular levels, interdisciplinary studies of chemistry and biology have led to the emergence of many smart fluorescent probes, which emit fluorescence or show a shifted signal only upon interaction with their targets. With distinct advantage of a higher signal-to-noise ratio than traditional 'always on' probes, smart fluorescent probes enable sensitive detection of enzymes with clinical significance. In this review, we summarize the design strategies and selected applications of smart fluorescent probes for in situ imaging of enzyme activity. Current challenges and future developments in this field are also discussed.

Universal primers for HBV genome DNA amplification across subtypes: a case study for designing more effective viral primers.

BACKGROUND: The highly heterogenic characteristic of viruses is the major obstacle to efficient DNA amplification. Taking advantage of the large number of virus DNA sequences in public databases to select conserved sites for primer design is an optimal way to tackle the difficulties in virus genome amplification. RESULTS: Here we use hepatitis B virus as an example to introduce a simple and efficient way for virus primer design. Based on the alignment of HBV sequences in public databases and a program BxB in Perl script, our method selected several optimal sites for HBV primer design. Polymerase chain reaction showed that compared with the success rate of the most popular primers for whole genome amplification of HBV, one set of primers for full length genome amplification and four sets of walking primers showed significant improvement. These newly designed primers are suitable for most subtypes of HBV. CONCLUSION: Researchers can extend the method described here to design universal or subtype specific primers for various types of viruses. The BxB program based on multiple sequence alignment not only can be used as a separate tool but also can be integrated in any open source primer design software to select conserved regions for primer design.