Bulk Droplet Vitrification for Primary Hepatocyte Preservation

10.3791/60250 ◽  
2019 ◽  
Author(s):  
Reinier J. de Vries ◽  
Peony D. Banik ◽  
Sonal Nagpal ◽  
Lindong Weng ◽  
Sinan Ozer ◽  
...  
Keyword(s):  
Primary Hepatocyte ◽  
Droplet Vitrification

scholarly journals Marine Fish Primary Hepatocyte Isolation and Culture: New Insights to Enzymatic Dissociation Pancreatin Digestion

2021 ◽  
Vol 18 (4) ◽  
pp. 1380
Author(s):  
Neusa Figueiredo ◽  
Beatriz Matos ◽  
Mário Diniz ◽  
Vasco Branco ◽  
Marta Martins
Keyword(s):  
Cell Viability ◽  
Cell Cultures ◽  
Marine Fish ◽  
Sparus Aurata ◽  
Cost Effective ◽  
Primary Hepatocyte ◽  
Digestion Method ◽  
Trypan Blue Exclusion ◽  
And Function

Primary cell cultures from wild organisms have been gaining relevance in ecotoxicology as they are considered more sensitive than immortalized cell lines and retain the biochemical pathways found in vivo. In this study, the efficacy of two methods for primary hepatocyte cell isolation was compared using liver from two marine fish (Sparus aurata and Psetta maxima): (i) two-step collagenase perfusion and (ii) pancreatin digestion with modifications. Cell cultures were incubated in L-15 medium at 17 ± 1 °C and monitored for up to six days for cell viability and function using the trypan blue exclusion test, MTT test, lactate dehydrogenase (LDH) activity, and ethoxyresorufin O-deethylase (EROD) activity after Benzo[a]Pyrene exposure. The results showed significant differences between the number of viable cells (p < 0.05), the highest number being obtained for the pancreatin digestion method (average = 4.5 ± 1.9 × 107 cells). Moreover, the hepatocytes showed solid adherence to the culture plate and the rounded shape, changing into a triangular/polygonal shape. The cell viability and function obtained by pancreatin digestion were maintained for five days, and the EROD induction after exposure to the B[a]P showed that cells were metabolically active. This study shows that the optimized pancreatin digestion method is a valid, cost-effective, and simple alternative to the standard perfusion method for the isolation of primary hepatocytes from fish and is suitable for ecotoxicological studies involving marine pollutants, such as PAHs.

Assessing Mechanisms of Ketoconazole and Amiodarone Induced Steatohepatitis in an Organotypic Human Primary Hepatocyte Liver Model

2016 ◽  
Vol 64 (2) ◽  
pp. S226-S227
Author(s):  
R. Figler ◽  
S. Marukian ◽  
M. Collado ◽  
D. Manka ◽  
M. Lawson ◽  
...  
Keyword(s):  
Primary Hepatocyte ◽  
Liver Model

scholarly journals Substrate stiffness regulates primary hepatocyte functions

RSC Advances ◽  
2015 ◽  
Vol 5 (99) ◽  
pp. 80956-80966 ◽  
Author(s):  
Vaishaali Natarajan ◽  
Eric J. Berglund ◽  
Dorothy X. Chen ◽  
Srivatsan Kidambi
Keyword(s):  
Liver Fibrosis ◽  
Alcohol Abuse ◽  
Metabolic Disorders ◽  
Viral Infections ◽  
Substrate Stiffness ◽  
Primary Hepatocyte ◽  
Chronic Injuries

Liver fibrosis occurs as a consequence of chronic injuries from viral infections, metabolic disorders, and alcohol abuse.

scholarly journals In vitro Conservation and Cryopreservation of Nandina domestica, an Outdoor Ornamental Shrub

2013 ◽  
Vol 41 (2) ◽  
pp. 638 ◽  
Author(s):  
Aylin OZUDOGRU ◽  
Diogo Pedrosa Corrêa Da SILVA ◽  
Ergun KAYA ◽  
Giuliano DRADI ◽  
Renato PAIVA ◽  
...  
Keyword(s):  
Liquid Nitrogen ◽  
Storage Temperature ◽  
Sucrose Concentration ◽  
Aluminum Foil ◽  
Shoot Tips ◽  
In Vitro Conservation ◽  
Shoot Cultures ◽  
Storage Medium ◽  
Droplet Vitrification

The study focused on an economically-important ornamental outdoor shrub, Nandina domestica, with the aims to (i) optimize an effective in vitro conservation method, and (ii) develop a cryopreservation protocol for shoot tips by the PVS2 vitrification and droplet-vitrification techniques. For in vitro conservation of shoot cultures, the tested parameters were sucrose content in the storage medium (30, 45, 60 g/L) and storage temperature (4 °C or 8 °C). Cryopreservation was performed by applying the PVS2 vitrification solution, in 2-ml cryovials or in drops over aluminum foil strips, for 15, 30, 60 or 90 min at 0 °C, followed by the direct immersion in liquid nitrogen of shoot tips. Results show that N. domestica shoots can be conserved successfully for 6 months at both the temperatures tested, especially when 60 g/L sucrose is used in the storage medium. However, conservation at 4 °C showed to be more appropriate, as hyperhydricity was observed in post-conservation of shoots coming from storage at 8 °C. As for cryopreservation, a daily gradual increase of sucrose concentration (from 0.25 to 1.0 M) produced better protection to the samples that were stored in liquid nitrogen. Indeed, with this sucrose treatment method, a 30-min PVS2 incubation time was enough to produce, 60 days after thawing, the best recovery (47% and 50%) of shoot tips, cryopreserved with PVS2 vitrification and droplet-vitrification, respectively.

Primary hepatocyte survival on non-integrin-recognizable matrices without the activation of Akt signaling

Biomaterials ◽  
2007 ◽  
Vol 28 (6) ◽  
pp. 1093-1104 ◽  
Author(s):  
Takashi Hoshiba ◽  
Hikaru Nagahara ◽  
Chong-Su Cho ◽  
Yoh-ichi Tagawa ◽  
Toshihiro Akaike
Keyword(s):  
Akt Signaling ◽  
Primary Hepatocyte
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