scholarly journals Examination of Conditions for Optimized Decellularized Liver Preparation

2019 ◽  
pp. 1-8
Author(s):  
Jaeyong Cho ◽  
Yukako Fukuda ◽  
Nana Shirakigawa ◽  
Hiroyuki Ijima
Keyword(s):  
Salt Concentration ◽  
Sodium Dodecyl ◽  
Porcine Liver ◽  
Experimental Conditions ◽  
Liver Preparation ◽  
Liver Structure ◽  
Ph Buffer ◽  
Triton X ◽  
Cellular Components ◽  
Decellularized Liver

Aims: The main aim of our study was to examine the concentration of surfactant that can cause significant disruption of the resulting decellularized liver structure. Furthermore, it is our goal to determine the suitable solvent that can boost the potential of each surfactant. Methodology: The porcine liver discs of 8-mm diameter and 2-mm thickness were prepared. These were soaked in aqueous solution of either sodium dodecyl sulfate (SDS) or Triton X-100 (TX), and placed on a rotational shaking machine (100 rpm). Results: TX was unable to completely remove the cellular components under any of our experimental conditions. The salt concentration did not affect the decellularization in TX. The pH buffer, however, was found to affect the decellularization. Also, in the solvent study, the conditions under which SDS effectively exerted power were not the salt concentration and pH, but the condition that was close to water. We also confirmed that the shrinkage of tissue occurred when decellularization with 0.1% SDS in CMF-PBS. However, 0.1% SDS in distilled water didn't cause the deformation of tissue. This is considered to be due to the low salt concentration of solvent. Conclusion: This work establishes the concentration range of the surfactant that causes the collapse of the cellular structure during decellularization. In addition, the solvent suitable for each surfactant has also been established.

Creating a tissue-specific microdispersed matrix from a decellularized porcine liver

2020 ◽  
Vol 4 ◽  
pp. 41-50
Author(s):  
A.D. Kirillova ◽  
◽  
Yu.B. Basok ◽  
A.E. Lazhko ◽  
A. M. Grigoryev ◽  
...  
Keyword(s):  
Sodium Dodecyl ◽  
Genetic Material ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Porcine Liver ◽  
Tissue Specific ◽  
Main Components ◽  
Triton X ◽  
Injectable Form

The main problem with decellularization of liver tissue as a tissue-specific matrix/scaffold in liver bioengineered structures is the need to maximize the preservation of the original three-dimensional structure of the tissue and the main components of its extracellular matrix (ECM) while removing cells and genetic material. The attempts to use the existing protocols for the decellularization of other tissues and organs have been unsuccessful. The aim of the work is to develop a method for creation of tissue-specific microdispersed matrix from decellularized porcine liver (TMM DLp). The protocol for decellularization of porcine liver (Lp) fragments has been developed based on the complex application of chemical (sodium dodecyl sulfate and Triton X-100), biochemical (DNase I), and physical (supercritical CO2) methods for treatment the initial tissue. As a result of the found optimal conditions for decellularization of Lp with subsequent cryomicronization of DLp, the injectable form of the microdispersed tissue-specific matrix was obtained, which represents DLp microparticles with the size of 100-200 microns with the residual amount of DNA no more than 10±1.5 ng/mg (less than 1.0%), with the preservation of the microstructure and basic composition of the liver ECM. According to the assessment of biocompatible properties in vitro, TMM DLp samples meet the criteria of biological safety for cytotoxicity and hemolytic activity.

scholarly journals Effect of detergent type on the performance of liver decellularized extracellular matrix-based bio-inks

2021 ◽  
Vol 12 ◽  
pp. 204173142199709
Author(s):  
Wonwoo Jeong ◽  
Min Kyeong Kim ◽  
Hyun-Wook Kang
Keyword(s):  
Extracellular Matrix ◽  
Sodium Dodecyl ◽  
Great Influence ◽  
Ammonium Hydroxide ◽  
Sodium Deoxycholate ◽  
Porcine Liver ◽  
Decellularized Extracellular Matrix ◽  
Cytochrome P450 Activity ◽  
Primary Mouse ◽  
Triton X

Decellularized extracellular matrix-based bio-inks (dECM bio-inks) for bioprinting technology have recently gained attention owing to their excellent ability to confer tissue-specific functions and 3D-printing capability. Although decellularization has led to a major advancement in bio-ink development, the effects of detergent type, the most important factor in decellularization, are still unclear. In this study, the effects of various detergent types on bio-ink performance were investigated. Porcine liver-derived dECM bio-inks prepared using widely used detergents, including sodium dodecyl sulfate (SDS), sodium deoxycholate (SDC), Triton X-100 (TX), and TX with ammonium hydroxide (TXA), were characterized in detail. SDS and SDC severely damaged glycosaminoglycan and elastin proteins, TX showed the lowest rate of decellularization, and TXA-based dECM bio-ink possessed the highest ECM content among all bio-inks. Differences in biochemical composition directly affected bio-ink performance, with TXA-dECM bio-ink showing the best performance with respect to gelation kinetics, intermolecular bonding, mechanical properties, and 2D/3D printability. More importantly, cytocompatibility tests using primary mouse hepatocytes also showed that the TXA-dECM bio-ink improved albumin secretion and cytochrome P450 activity by approximately 2.12- and 1.67-fold, respectively, compared with the observed values for other bio-inks. Our results indicate that the detergent type has a great influence on dECM damage and that the higher the dECM content, the better the performance of the bio-ink for 3D bioprinting.

scholarly journals Photocrosslinkable liver extracellular matrix hydrogels for the generation of 3D liver microenvironment models

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Akhilandeshwari Ravichandran ◽  
Berline Murekatete ◽  
Denise Moedder ◽  
Christoph Meinert ◽  
Laura J. Bray
Keyword(s):  
Mechanical Properties ◽  
Extracellular Matrix ◽  
Cancer Metastasis ◽  
Sodium Dodecyl ◽  
3D Cell Culture ◽  
Ammonium Hydroxide ◽  
Porcine Liver ◽  
Thermal Gelation ◽  
Triton X ◽  
Drug Studies

AbstractLiver extracellular matrix (ECM)-based hydrogels have gained considerable interest as biomimetic 3D cell culture environments to investigate the mechanisms of liver pathology, metabolism, and toxicity. The preparation of current liver ECM hydrogels, however, is based on time-consuming thermal gelation and limits the control of mechanical properties. In this study, we used detergent-based protocols to produce decellularized porcine liver ECM, which in turn were solubilized and functionalized with methacrylic anhydride to generate photocrosslinkable methacrylated liver ECM (LivMA) hydrogels. Firstly, we explored the efficacy of two protocols to decellularize porcine liver tissue using varying combinations of commonly used chemical agents such as Triton X-100, Sodium Dodecyl Sulphate (SDS) and Ammonium hydroxide. Then, we demonstrated successful formation of stable, reproducible LivMA hydrogels from both the protocols by photocrosslinking. The LivMA hydrogels obtained from the two decellularization protocols showed distinct mechanical properties. The compressive modulus of the hydrogels was directly dependent on the hydrogel concentration, thereby demonstrating the tuneability of mechanical properties of these hydrogels. Immortalized Human Hepatocytes cells were encapsulated in the LivMA hydrogels and cytocompatibility of the hydrogels was demonstrated after one week of culture. In summary, the LivMA hydrogel system provides a simple, photocrosslinkable platform, which can potentially be used to simulate healthy versus damaged liver for liver disease research, drug studies and cancer metastasis modelling.

Solubility and adsorption behaviors of chlorophenols in the presence of surfactant

1997 ◽  
Vol 35 (7) ◽  
pp. 123-130 ◽  
Author(s):  
J. C. Liu ◽  
P. S. Chang
Keyword(s):  
Sodium Dodecyl Sulfate ◽  
Critical Micelle Concentration ◽  
Sodium Dodecyl ◽  
Adsorption Behaviors ◽  
Brij 35 ◽  
Important Index ◽  
Dodecyl Sulfate ◽  
Triton X

The solubility of chlorophenols as affected by surfactant was investigated. Three kinds of surfactant, sodium dodecyl sulfate, Triton X-100, and Brij 35, were utilized. The solubilization of chlorophenols by surfactant follows the order of 2,4,6-trichlorophenol > 2,4-dichlorophenol > 2,6-dichlorophenol > 2-chlorophenol; and the critical micelle concentration is an important index. The adsorption reactions of 2,4-dichlorophenol and 2,4,6- trichlorophenol onto hydrous montmorillonite in the presence of surfactant were examined. The presence of surfactant decreased the adsorption of chlorophenols significantly. The roles of hydrophobicity of chlorophenols in solubilization and adsorption behaviors are discussed.

Different submicellar solubilization mechanisms revealed by 1H NMR and 2D diffusion ordered spectroscopy (DOSY)

2020 ◽  
Vol 22 (19) ◽  
pp. 11075-11085
Author(s):  
Mengjian Wu ◽  
Zhaoxia Wu ◽  
Shangwu Ding ◽  
Zhong Chen ◽  
Xiaohong Cui
Keyword(s):  
Nmr Spectroscopy ◽  
Sodium Dodecyl Sulfate ◽  
1H Nmr ◽  
Sodium Dodecyl ◽  
Butyl Methacrylate ◽  
H Nmr ◽  
Molecular Scale ◽  
Two Systems ◽  
Dodecyl Sulfate ◽  
Triton X

Different submicellar solubilization mechanisms of two systems, Triton X-100/tetradecane and sodium dodecyl sulfate (SDS)/butyl methacrylate, are revealed on the molecular scale by 1H NMR spectroscopy and 2D diffusion ordered spectroscopy (DOSY).

scholarly journals Time-dependent association between platelet-bound fibrinogen and the Triton X-100 insoluble cytoskeleton

Blood ◽  
1991 ◽  
Vol 77 (3) ◽  
pp. 508-514 ◽  
Author(s):  
EI Peerschke
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Adenosine Diphosphate ◽  
Time Dependent ◽  
Binding Studies ◽  
Fibrinogen Binding ◽  
Human Thrombin ◽  
Membrane Bound ◽  
Independent Association ◽  
Triton X

Abstract Previous studies indicated a correlation between the formation of EDTA- resistant (irreversible) platelet-fibrinogen interactions and platelet cytoskeleton formation. The present study explored the direct association of membrane-bound fibrinogen with the Triton X-100 (Sigma Chemical Co, St Louis, MO) insoluble cytoskeleton of aspirin-treated, gel-filtered platelets, activated but not aggregated with 20 mumol/L adenosine diphosphate (ADP) or 150 mU/mL human thrombin (THR) when bound fibrinogen had become resistant to dissociation by EDTA. Conversion of exogenous 125I-fibrinogen to fibrin was prevented by adding Gly-Pro-Arg and neutralizing THR with hirudin before initiating binding studies. After 60 minutes at 22 degrees C, the cytoskeleton of ADP-treated platelets contained 20% +/- 12% (mean +/- SD, n = 14) of membrane-bound 125I-fibrinogen, representing 10% to 50% of EDTA- resistant fibrinogen binding. The THR-activated cytoskeleton contained 45% +/- 15% of platelet bound fibrinogen, comprising 80% to 100% of EDTA-resistant fibrinogen binding. 125I-fibrinogen was not recovered with platelet cytoskeletons if binding was inhibited by the RGDS peptide, excess unlabeled fibrinogen, or disruption of the glycoprotein (GP) IIb-IIIa complex by EDTA-treatment. Both development of EDTA- resistant fibrinogen binding and fibrinogen association with the cytoskeleton were time dependent and reached maxima 45 to 60 minutes after fibrinogen binding to stimulated platelets. Although a larger cytoskeleton formed after platelet stimulation with thrombin as compared with ADP, no change in cytoskeleton composition was noted with development of EDTA-resistant fibrinogen binding. Examination of platelet cytoskeletons using monoclonal antibodies, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and Western blotting showed the presence of only traces of GP IIb-IIIa in the cytoskeletons of resting platelets, with no detectable increases after platelet activation or development of EDTA-resistant fibrinogen binding. These data suggest that GP IIb-IIIa-mediated fibrinogen binding to activated platelets is accompanied by time-dependent alterations in platelet- fibrinogen interactions leading to the GP IIb-IIIa independent association between bound fibrinogen and the platelet cytoskeleton.

Preparative isolation and characterization of the B875 complex from Rhodobacter sphaeroides 2.4.1

1988 ◽  
Vol 66 (5) ◽  
pp. 442-448 ◽  
Author(s):  
Rafael Picorel ◽  
Gabriel Gingras
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Phosphatidyl Ethanolamine ◽  
Sodium Dodecyl ◽  
Pigment Analysis ◽  
Preparative Isolation ◽  
Ethanolamine Phosphatidyl ◽  
Isolation And Characterization ◽  
Detergent System ◽  
Triton X

We have developed a simple and efficient method, using a mixed detergent system of sodium dodecyl sulfate and Triton X-100, for the preparative isolation of theB875 complex from Rhodobacter sphaeroides 2.4.1. As a bonus, the method allows the preparation of both the B875 and B800-850 complexes from the same batch of chromatophores. The preparations are spectrally pure, as indicated by absorption and circular dichroism spectroscopy. The latter method suggests that the Qy band of the B875 complex is due to weakly interacting bacteriochlorophyll molecules. Protein and pigment analysis shows that the B875 complex contains 2 mol of bacteriochlorophyll and 2 mol of sphaeroidene per mol of apoprotein (12 266 g), whereas the B800-850 complex contains 3 mol of bacteriochlorophyll and 1 mol of sphaeroidene per mol of apoprotein (11 497 g). While these stoichiometries are in accord with currently accepted models, they disagree with their published experimental basis. Phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl glycerol, and diphosphatidyl glycerol were found to be present in the B875 complex.

scholarly journals Evidence that coated vesicles isolated from brain are calcium-sequestering organelles resembling sarcoplasmic reticulum.

1977 ◽  
Vol 75 (1) ◽  
pp. 135-147 ◽  
Author(s):  
A L Blitz ◽  
R E Fine ◽  
P A Toselli
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Calcium Ions ◽  
Adenosine Triphosphatase ◽  
Sodium Dodecyl ◽  
Maximal Activity ◽  
Coated Vesicles ◽  
Potassium Oxalate ◽  
Triton X ◽  
Trapping Agent ◽  
The Brain

Coated vesicles from the brain have been purified to near morphological homogeneity by a modification of the method of Pearse. These vesicles resemble sarcoplasmic reticulum fragments isolated from skeletal muscle. They contain proteins with 100,000- and 55,000-dalton mol wt which co-migrate on polyacrylamide gels, in the presence of sodium dodecyl sulfate, with the two major proteins of the sarcoplasmic reticulum fragment. These vesicles contain adenosine triphosphatase (ATPase) activity which is stimulated by calcium ions in the presence of Triton X-100 (Rohm & Haas Co., Philadelphia, Pa.), displaying maximal activity at 8 x 10(-7) M Ca ++. They take up calcium ions from the medium, and this uptake is stimulated by ATP and by potassium oxalate, a calcium-trapping agent. The 100,000-dalton protein of the coated vesicles displays immunological reactivity with an antiserum directed against the 100,000-dalton, calcium-stimulated ATPase of the sarcoplasmic reticulum. As with the sarcoplasmic reticulum fragment, this protein becomes radiolabeled when coated vesicles are briefly incubated with gamma-labeled [32P]ATP. The possible functions of coated vesicles as calcium-sequestering organelles are discussed.

Solubilization and immunoprecipitation of 125I-labelled antigens from Plasmodium knowlesi schizont-infected erythrocytes using non-ionic, anionic and zwitterionic detergents

Parasitology ◽  
1984 ◽  
Vol 88 (1) ◽  
pp. 27-36 ◽  
Author(s):  
R. J. Howard ◽  
J. W. Barnwell
Keyword(s):  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Plasmodium Knowlesi ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Ionic Detergents ◽  
Class A ◽  
Anionic Detergents ◽  
Triton X

SUMMARYPlasmodium knowlesi malaria-infected erythrocytes were radio-iodinated and several non-ionic, anionic and zwitterionic detergents were compared in their capacity to extract the labelled membrane proteins. The use of these detergents for antigen identification was tested by immunoprecipitation, after addition of Triton X-100 to some detergent extracts, using hyperimmune monkey antiserum and protein A-Sepharose. 125I-labelled antigens were specifically immunoprecipitated with all detergents tested, including the anionic detergents sodium dodecyl sulphate (SDS), deoxycholate and cholate; the zwitterions Zwittergent-312 and -314, CHAPS and Empigen BB, as well as several non-ionic detergents. The SDS-polyacrylamide gel electrophoresis patterns of 125I-labelled antigens varied after extraction with different detergents, there being no consistent pattern for detergents of a particular class. A total of 14 125I-labelled antigens were identified, 11 of them using Triton X-100. Some minor antigens identified with Triton X-100 were immunoprecipitated in greater amount after extraction in other detergents. Most importantly, two antigens Mr 200000 and 180000 were detected only after extraction with deoxycholate or SDS.

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