Journal: PLoS ONE

Article Title: Adeno-associated virus-mediated expression of human butyrylcholinesterase to treat organophosphate poisoning

doi: 10.1371/journal.pone.0225188

Figure Lengend Snippet: (A) Schematic representation of the AAV vector construct and the variables used to optimize maximal hBChE expression. (B) In vitro expression (ng/ml) of hBChE in the supernatant of HEK293 cells transfected with plasmids expressing different codon-optimized hBChE genes under the control of the CMV promoter including un-transfected control (Ct). (C) In vivo expression (ng/ml) of hBChE in the serum of RAG KO male mice (n = 5/group) injected IM with 10 11 GC/mouse of an AAV vector expressing different codon-optimized hBChE genes expressed under the CMV promoter. (D) Expression (ng/ml) and (E) activity (units/ml) of hBChE protein in serum of RAG KO male mice (n = 5/group) injected IM with 10 11 GC/mouse of AAV8-UbC-BChE or AAV8-CB7-BChE vector. (F) In-gel activity assay and (G) native Western blot of hBChE expressed by AAV8-CB7-BChE vector in RAG KO mice serum. Data represents the average of all mice (n) ± SD. BL = Baseline.

Article Snippet: BChE expression in serially diluted mouse serum or plasma was quantified using a commercially available hBChE Quantikine ELISA kit from R&D Systems (catalog number: DBCHE0).

Techniques: Plasmid Preparation, Construct, Expressing, In Vitro, Transfection, Control, In Vivo, Injection, Activity Assay, Western Blot

Journal: PLoS ONE

Article Title: Adeno-associated virus-mediated expression of human butyrylcholinesterase to treat organophosphate poisoning

doi: 10.1371/journal.pone.0225188

Figure Lengend Snippet: (A) Butyrylcholine challenge study design in male BChE KO mice (n = 5/group) injected IM with AAV8-CB7-BChE at different doses (GC/mouse) and untreated control mice (n = 6). At study day 0, 2, and 7, (B) hBChE expression (ng/ml) in serum was analyzed. (C) Survival was determined after challenge with 500 mg/kg of butyrylcholine on day 8 post-vector injection. (D) Correlation plot between hBChE expression and percent survival. (E) VX challenge study design in ES1 KO mice (n = 10/ group) injected IM with three AAV vectors expressing hBChE individually. At study day 0, 2, 7 and weekly thereafter, hBChE expression (ng/ml) was measured in plasma of mice injected IM with (F) AAV8-UbC-BChE, (G) AAV8-CB7-BChE, or AAV9-C4/UbC-BChE at 10 12 GC/mouse. (H) Survival was observed after challenge with 2xLD 50 VX at day 70 post-vector administration. Data represents the average of all mice (n) ± SD. *Two mice were excluded from the survival analysis due to inaccurate VX dosage.

Article Snippet: BChE expression in serially diluted mouse serum or plasma was quantified using a commercially available hBChE Quantikine ELISA kit from R&D Systems (catalog number: DBCHE0).

Techniques: Injection, Control, Expressing, Plasmid Preparation, Clinical Proteomics

Journal: Journal of neuroinflammation

Article Title: Immunomodulatory effects of nicotine on interleukin 1β activated human astrocytes and the role of cyclooxygenase 2 in the underlying mechanism.

doi: 10.1186/s12974-016-0725-1

Figure Lengend Snippet: Fig. 4 Butyrylcholinesterase levels in cell culture supernatants of IL-1β activated human astrocytes following nicotine treatment. Both untreated and IL-1β-activated human astrocytes released similar BuChE protein levels into their supernatants following 20-h incubation. Nicotine dose depend- ently inhibited BuChE protein release by activated human astrocytes. Sam- ples were run as duplicates, and values are represented as mean ± SEM of four independent experiments. Data is presented in terms of fold change compared to BuChE levels following IL-1β treatment

Article Snippet: Measurement of AChE and BuChE in cell supernatants Protein concentration of AChE and BuChE in the astrocyte culture supernatants was measured using quantitative sandwich ELISA technique employed in human acetylcholinesterase (DACHE0) and human butyrylcholinesterase (DBHE0) quantikine ELISA kits (R&D Systems).

Techniques: Cell Culture, Incubation

Journal: bioRxiv

Article Title: Gut microbiota gate host exposure to metabolites from dietary Solanums

doi: 10.1101/2024.03.20.584512

Figure Lengend Snippet: A) Total steroidal alkaloid concentration in the serum of C57BL/6J mice fed potato-supplemented diet and treated with antibiotics. Aglycone concentrations were estimated using a solanidine standard curve. Data shown are the mean±SD (n=5 mice). P values were determined using a two-sided Welch’s T test. B) In vitro inhibition of recombinant human AChE activity by potato steroidal alkaloids. Data shown are normalized to AChE treated with vehicle only (DMSO for solanine, chaconine and ethanol for solanidine) with a total vehicle concentration of 1% for all treatments. Values shown are the mean±SD (n=3 assays per concentration). 5 ng of AChE was used in each reaction. C) In vitro inhibition of cholinesterase activity by 50 μM potato SAs in human protein homogenates from normal colonic tissue (n=3 donors) and Caco-2 colonic epithelial cells. Activity was normalized to the activity of the vehicle-treated sample. Values shown are the mean±SD of three replicate assays per homogenate. 5 ng each of recombinant human AChE and BChE were used for pure protein reactions, while 35 μg total protein was used for each homogenate. Multiplicity-adjusted P values were calculated using Tukey’s multiple comparison tests. D) Inhibition of recombinant human AChE activity by spent media from human stool (n=11 donors) incubated with 20 μM solanine in SAAC minimal medium. Activity was normalized to the activity of AChE treated with spent media from the same donor sample incubated with vehicle only. Values shown are the mean±SD of three AChE activity assays for each donor and compound combination. The relative abundances of select metabolites following incubation with solanine are shown below for each stool sample. Values shown are EIC peak areas normalized to the maximum peak area across all samples. P values were determined using a two-sided Welch’s T test. Ns: not significant with P>0.05.

Article Snippet: BChE activity was analogously measured using 5 ng of recombinant human BChE (R&D Systems) and butyrylthiocholine iodide.

Techniques: Concentration Assay, In Vitro, Inhibition, Recombinant, Activity Assay, Comparison, Incubation

Journal: Caspian Journal of Internal Medicine

Article Title: Occurrence of butyrylcholinesterase polymorphisms in patients undergoing surgery in Slovakia

doi: 10.22088/cjim.14.3.490

Figure Lengend Snippet: Allele and genotype frequencies of the rs1799807, rs1803274 polymorphisms of the BCHE gene in patients undergoing surgery

Article Snippet: Genotyping was realized using the TaqMan SNP Assay (C___2411904_20; C__27479669_20) (Applied Biosystems, CA, USA) according to standard protocol followed by Real-Time PCR fluorescence detection using a 7500 Fast Real-Time PCR System (Applied Biosystem).

Techniques: