| 1. |
- Beal, Jacob, et al.
(författare)
-
Robust estimation of bacterial cell count from optical density
- 2020
-
Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
-
Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
|
|
| 2. |
|
|
| 3. |
- Klionsky, Daniel J., et al.
(författare)
-
Guidelines for the use and interpretation of assays for monitoring autophagy
- 2012
-
Ingår i: Autophagy. - : Informa UK Limited. - 1554-8635 .- 1554-8627. ; 8:4, s. 445-544
-
Forskningsöversikt (refereegranskat)abstract
- In 2008 we published the first set of guidelines for standardizing research in autophagy. Since then, research on this topic has continued to accelerate, and many new scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Accordingly, it is important to update these guidelines for monitoring autophagy in different organisms. Various reviews have described the range of assays that have been used for this purpose. Nevertheless, there continues to be confusion regarding acceptable methods to measure autophagy, especially in multicellular eukaryotes. A key point that needs to be emphasized is that there is a difference between measurements that monitor the numbers or volume of autophagic elements (e.g., autophagosomes or autolysosomes) at any stage of the autophagic process vs. those that measure flux through the autophagy pathway (i.e., the complete process); thus, a block in macroautophagy that results in autophagosome accumulation needs to be differentiated from stimuli that result in increased autophagic activity, defined as increased autophagy induction coupled with increased delivery to, and degradation within, lysosomes (in most higher eukaryotes and some protists such as Dictyostelium) or the vacuole (in plants and fungi). In other words, it is especially important that investigators new to the field understand that the appearance of more autophagosomes does not necessarily equate with more autophagy. In fact, in many cases, autophagosomes accumulate because of a block in trafficking to lysosomes without a concomitant change in autophagosome biogenesis, whereas an increase in autolysosomes may reflect a reduction in degradative activity. Here, we present a set of guidelines for the selection and interpretation of methods for use by investigators who aim to examine macroautophagy and related processes, as well as for reviewers who need to provide realistic and reasonable critiques of papers that are focused on these processes. These guidelines are not meant to be a formulaic set of rules, because the appropriate assays depend in part on the question being asked and the system being used. In addition, we emphasize that no individual assay is guaranteed to be the most appropriate one in every situation, and we strongly recommend the use of multiple assays to monitor autophagy. In these guidelines, we consider these various methods of assessing autophagy and what information can, or cannot, be obtained from them. Finally, by discussing the merits and limits of particular autophagy assays, we hope to encourage technical innovation in the field.
|
|
| 4. |
- Lin, Yi-Ting, et al.
(författare)
-
Differences in the Microbial Composition of Hemodialysis Patients Treated with and without β-Blockers
- 2021
-
Ingår i: Journal of Personalized Medicine. - : MDPI. - 2075-4426. ; 11:3
-
Tidskriftsartikel (refereegranskat)abstract
- β-blockers are commonly prescribed to treat cardiovascular disease in hemodialysis patients. Beyond the pharmacological effects, β-blockers have potential impacts on gut microbiota, but no study has investigated the effect in hemodialysis patients. Hence, we aim to investigate the gut microbiota composition difference between β-blocker users and nonusers in hemodialysis patients. Fecal samples collected from hemodialysis patients (83 β-blocker users and 110 nonusers) were determined by 16S ribosomal RNA amplification sequencing. Propensity score (PS) matching was performed to control confounders. The microbial composition differences were analyzed by the linear discriminant analysis effect size, random forest, and zero-inflated Gaussian fit model. The α-diversity (Simpson index) was greater in β-blocker users with a distinct β-diversity (Bray–Curtis Index) compared to nonusers in both full and PS-matched cohorts. There was a significant enrichment in the genus Flavonifractor in β-blocker users compared to nonusers in full and PS-matched cohorts. A similar finding was demonstrated in random forest analysis. In conclusion, hemodialysis patients using β-blockers had a different gut microbiota composition compared to nonusers. In particular, the Flavonifractor genus was increased with β-blocker treatment. Our findings highlight the impact of β-blockers on the gut microbiota in hemodialysis patients.
|
|
| 5. |
- Liao, Shih-Fen, et al.
(författare)
-
Immunization of fucose-containing polysaccharides from Reishi mushroom induces antibodies to tumor-associated Globo H-series epitopes.
- 2013
-
Ingår i: Proceedings of the National Academy of Sciences of the United States of America. - : Proceedings of the National Academy of Sciences. - 0027-8424 .- 1091-6490. ; 110:34, s. 13809-13814
-
Tidskriftsartikel (refereegranskat)abstract
- Carbohydrate-based vaccines have shown therapeutic efficacy for infectious disease and cancer. The mushroom Ganoderma lucidum (Reishi) containing complex polysaccharides has been used as antitumor supplement, but the mechanism of immune response has rarely been studied. Here, we show that the mice immunized with a l-fucose (Fuc)-enriched Reishi polysaccharide fraction (designated as FMS) induce antibodies against murine Lewis lung carcinoma cells, with increased antibody-mediated cytotoxicity and reduced production of tumor-associated inflammatory mediators (in particular, monocyte chemoattractant protein-1). The mice showed a significant increase in the peritoneal B1 B-cell population, suggesting FMS-mediated anti-glycan IgM production. Furthermore, the glycan microarray analysis of FMS-induced antisera displayed a high specificity toward tumor-associated glycans, with the antigenic structure located in the nonreducing termini (i.e., Fucα1-2Galβ1-3GalNAc-R, where Gal, GalNAc, and R represent, respectively, D-galactose, D-N-acetyl galactosamine, and reducing end), typically found in Globo H and related tumor antigens. The composition of FMS contains mainly the backbone of 1,4-mannan and 1,6-α-galactan and through the Fucα1-2Gal, Fucα1-3/4Man, Fucα1-4Xyl, and Fucα1-2Fuc linkages (where Man and Xyl represent d-mannose and d-xylose, respectively), underlying the molecular basis of the FMS-induced IgM antibodies against tumor-specific glycans.
|
|
| 6. |
- Wu, Ping-Hsun, et al.
(författare)
-
Exploring the Benefit of 2-Methylbutyric Acid in Patients Undergoing Hemodialysis Using a Cardiovascular Proteomics Approach
- 2019
-
Ingår i: Nutrients. - : MDPI. - 2072-6643. ; 11:12
-
Tidskriftsartikel (refereegranskat)abstract
- Short-chain fatty acids (SCFAs) can reduce pro-inflammatory parameters and oxidative stress, providing potential cardiovascular (CV) benefits. Although some evidence links SCFAs with host metabolic health via several biological mechanisms, the role of SCFA on CV disease in patients with kidney disease remains unclear. Herein, we investigate the association between a SCFA, 2-methylbutyric acid, and target CV proteomics to explore the potential pathophysiology of SCFA-related CV benefit in patients with kidney disease. Circulating 2-methylbutyric acid was quantified by high-performance liquid chromatography and 181 CV proteins by a proximity extension assay in 163 patients undergoing hemodialysis (HD). The associations between 2-methylbutyric acid and CV proteins were evaluated using linear regression analysis with age and gender, and multiple testing adjustment. The selected CV protein in the discovery phase was further confirmed in multivariable-adjusted models and evaluated by continuous scale association. The mean value of circulating 2-methylbutyric acid was 0.22 +/- 0.02 mu m which was negatively associated with bone morphogenetic protein 6 (BMP-6) according to the false discovery rate (FDR) multiple testing adjustment method. The 2-methylbutyric acid level remained negatively associated with BMP-6 (beta coefficient -1.00, 95% confidence interval -1.45 to -0.55, p < 0.001) after controlling for other CV risk factors in multivariable models. The cubic spline curve demonstrated a linear relationship. In conclusion, circulating 2-methylbutyric acid level was negatively associated with BMP-6, suggesting that this pathway maybe involved in vascular health in patients undergoing HD. However, further in vitro work is still needed to validate the translation of the mechanistic pathways.
|
|
| 7. |
- Wu, Ping-Hsun, et al.
(författare)
-
β-blocker dialyzability and the risk of mortality and cardiovascular events in patients undergoing hemodialysis
- 2020
-
Ingår i: Nephrology, Dialysis and Transplantation. - : Oxford University Press (OUP). - 0931-0509 .- 1460-2385. ; 35:11, s. 1959-1965
-
Tidskriftsartikel (refereegranskat)abstract
- Backgroundβ-blocker (BB) dialyzability has been proposed to limit their efficacy among hemodialysis (HD) patients. We attempted to confirm this hypothesis by comparing health outcomes associated with the initiation of dialyzable or nondialyzable BBs in a nationwide cohort of HD patients.MethodsWe created a prospective cohort study of 15 699 HD patients who initiated dialyzable BBs (atenolol, acebutolol, metoprolol and bisoprolol) and 20 904 hemodialysis patients who initiated nondialyzable BBs (betaxolol, carvedilol and propranolol) between 2004 and 2011 in Taiwan healthcare. We compared the risk of all-cause mortality and major adverse cardiovascular events (MACEs, a composite of the acute coronary syndrome, ischemic stroke and heart failure) between users of dialyzable versus nondialyzable BBs during a 2-year follow-up.ResultsNew users of dialyzable BBs were younger, more often men, with diabetes mellitus, hypertension and hyperlipidemia compared with users of nondialyzable BBs. Compared with nondialyzable BBs, initiation of dialyzable BBs was associated with lower all-cause mortality {hazard ratio [HR] 0.82 [95% confidence interval (CI) 0.75–0.88]} and lower risk of MACEs [HR 0.89 (95% CI 0.84–0.93)]. Results were confirmed in subgroup analyses, censoring at BB discontinuation or switch, after 1:1 propensity score matching, reclassifying bisoprolol or excluding bisoprolol/carvedilol users.ConclusionsThis study does not offer support for the hypothesis that the dialyzability of BBs reduces their efficacy in HD patients.
|
|
| 8. |
- Hsu, Yu-Cheng, et al.
(författare)
-
One-Pot Tandem Photoredox and Cross-Coupling Catalysis with a Single Palladium Carbodicarbene Complex
- 2018
-
Ingår i: Angewandte Chemie International Edition. - : Wiley. - 1433-7851 .- 1521-3773. ; 57:17, s. 4622-4626
-
Tidskriftsartikel (refereegranskat)abstract
- The combination of conventional transition-metal-catalyzed coupling (2e(-) process) and photoredox catalysis (1e(-) process) has emerged as a powerful approach to catalyze difficult cross-coupling reactions under mild reaction conditions. Reported is a palladium carbodicarbene (CDC) complex that mediates both a Suzuki-Miyaura coupling and photoredox catalysis for C-N bond formation upon visible-light irradiation. These two catalytic pathways can be combined to promote both conventional transition-metal-catalyzed coupling and photoredox catalysis to mediate C-H arylation under ambient conditions with a single catalyst in an efficient one-pot process.
|
|
| 9. |
- Purnomo, Kartiko Arif, et al.
(författare)
-
Decoding Multiple Biofunctions of Maca on Its Anti-allergic, Anti-inflammatory, Anti-thrombotic, and Pro-angiogenic Activities
- 2021
-
Ingår i: Journal of Agricultural and Food Chemistry. - : American Chemical Society (ACS). - 0021-8561 .- 1520-5118. ; 69:40, s. 11856-11866
-
Tidskriftsartikel (refereegranskat)abstract
- Four active partition layers and ten isolates, including (5R)- and (5S)-macapyrrolidone A (1a, 1b), and four new alkaloids, (5R)- and (5S)-macapyrrolidone B (2a, 2b) and macapyrrolins D, E (3, 4), were isolated from maca (Lepidium meyenii Walp.), an indigenous food plant from Peru. Derived from the n-hexane layer, the macamide-rich fraction exhibited pro-angiogenic activity on EPC and HUVEC cells. Anti-thrombotic activity was displayed by the polar part of maca extracts (n-butanol and water layers). Both 75% methanol aq. (midlower polar part) and n-hexane (low polar part) layers, which showed signs of fatty acid content, markedly inhibited superoxide and elastase release in an anti-inflammatory assay. The 75% methanol aq. layer showed strong anti-allergic activity, and macapyrrolin A (5) was found active based on beta-hexosaminidase release inhibition assays and a ChemGPS-NP experiment. These valuable bioactivity results suggest that maca is a food plant with good benefits for human health.
|
|
| 10. |
- Korinek, Michal, et al.
(författare)
-
Anti-allergic Hydroxy Fatty Acids from Typhonium blumei Explored through ChemGPS-NP
- 2017
-
Ingår i: Frontiers in Pharmacology. - : FRONTIERS MEDIA SA. - 1663-9812. ; 8
-
Tidskriftsartikel (refereegranskat)abstract
- Increasing prevalence of allergic diseases with an inadequate variety of treatment drives forward search for new alternative drugs. Fatty acids, abundant in nature, are regarded as important bioactive compounds and powerful nutrients playing an important role in lipid homeostasis and inflammation. Phytochemical study on Typhonium blumei Nicolson and Sivadasan (Araceae), a folk anti-cancer and anti-inflammatory medicine, yielded four oxygenated fatty acids, 12R-hydroxyoctadec-9Z, 13E-dienoic acid methyl ester (1) and 10R-hydroxyoctadec-8E, 12Z-dienoic acid methyl ester (2), 9R-hydroxy-10E-octadecenoic acid methyl ester (3), and 12R *-hydroxy-10E-octadecenoic acid methyl ester (4). Isolated compounds were identified by spectroscopic methods along with GC-MS analysis. Isolated fatty acids together with a series of saturated, unsaturated and oxygenated fatty acids were evaluated for their anti-inflammatory and anti-allergic activities in vitro. Unsaturated (including docosahexaenoic and eicosapentaenoic acids) as well as hydroxylated unsaturated fatty acids exerted strong anti-inflammatory activity in superoxide anion generation (IC50 2.14-3.73 mu M) and elastase release (IC50 1.26-4.57 mu M) assays. On the other hand, in the anti-allergic assays, the unsaturated fatty acids were inactive, while hydroxylated fatty acids showed promising inhibitory activity in A23187-and antigen-induced degranulation assays (e.g., 9S-hydroxy-10E, 12Z-octadecadienoic acid, IC50 92.4 and 49.7 mu M, respectively). According to our results, the presence of a hydroxy group in the long chain did not influence the potent anti-inflammatory activity of free unsaturated acids. Nevertheless, hydroxylation of fatty acids (or their methyl esters) seems to be a key factor for the anti-allergic activity observed in the current study. Moreover, ChemGPS-NP was explored to predict the structure-activity relationship of fatty acids. The anti-allergic fatty acids formed different cluster distant from clinically used drugs. The bioactivity of T. blumei, which is historically utilized in folk medicine, might be related to the content of fatty acids and their metabolites.
|
|
