Cryopreservation of hepatic stellate cells

Neyzen, Svenja; Leur, Eddy Van de; Borkham-Kamphorst, Erawan; Herrmann, Jens; Hollweg, Günter; Gressner, A. M.; Weiskirchen, Ralf

doi:10.1016/j.jhep.2005.07.008

Cited by 13 publications

(7 citation statements)

References 37 publications

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“…The cryopreservation of human hepatocytes shows greatest success with a progressive freezing, with its slow freezing rate, following a 30-min preincubation at 37°C and could potentially be used in cell banking (2). Work has been done also involving the cryopreservation of hepatic stellate cells (26), which provide critical paracrine signaling for the hHBs. Here, both cooling rates and cryoprotectants were investigated and found to have a clear effect on the resulting viability and cell survival.…”

Section: Discussionmentioning

confidence: 99%

Hyaluronan-Supplemented Buffers Preserve Adhesion Mechanisms Facilitating Cryopreservation of Human Hepatic Stem/Progenitor Cells

et al. 2012

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The supply of human hepatic stem cells (hHpSCs) and other hepatic progenitors has been constrained by the limited availability of liver tissues from surgical resections, the rejected organs from organ donation programs, and the need to use cells immediately. To facilitate accessibility to these precious tissue resources, we have established an effective method for serum-free cryopreservation of the cells, allowing them to be stockpiled and stored for use as an off-the-shelf product for experimental or clinical programs. The method involves use of buffers, some serum-free, designed for cryopreservation and further supplemented with hyaluronans (HA) that preserve adhesion mechanisms facilitating postthaw culturing of the cells and preservation of functions. Multiple cryopreservation buffers were found to yield high viabilities (80–90%) of cells on thawing of the progenitor cells. Serum-free CS10 supplemented with 0.05% hyaluronan proved the most effective, both in terms of viabilities of cells on thawing and in yielding cell attachment and formation of expanding colonies of cells that stably maintain the stem/progenitor cell phenotype. Buffers to which 0.05 or 0.1% HAs were added showed cells postthaw to be phenotypically stable as stem/progenitors, as well as having a high efficiency of attachment and expansion in culture. Success correlated with improved expression of adhesion molecules, particularly CD44, the hyaluronan receptor, E-cadherin, β4 integrin in hHpSCs, and β1 integrins in hepatoblasts. The improved methods in cryopreservation offer more efficient strategies for stem cell banking in both research and potential therapy applications.

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Section: Discussionmentioning

confidence: 99%

Hyaluronan-Supplemented Buffers Preserve Adhesion Mechanisms Facilitating Cryopreservation of Human Hepatic Stem/Progenitor Cells

et al. 2012

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“…A recent report describes a cryopreservation technique for freezing primary stellate cells using dimethyl sulfoxide (442). If reproducible in other laboratories, the technique will also accelerate progress by enabling the sharing of cells between investigators, especially those without the means to isolate their own primary cells.…”

Section: B Cell Linesmentioning

confidence: 99%

Hepatic Stellate Cells: Protean, Multifunctional, and Enigmatic Cells of the Liver

Friedman

2008

Physiological Reviews

2,403

2,549

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The hepatic stellate cell has surprised and engaged physiologists, pathologists, and hepatologists for over 130 years, yet clear evidence of its role in hepatic injury and fibrosis only emerged following the refinement of methods for its isolation and characterization. The paradigm in liver injury of activation of quiescent vitamin A-rich stellate cells into proliferative, contractile, and fibrogenic myofibroblasts has launched an era of astonishing progress in understanding the mechanistic basis of hepatic fibrosis progression and regression. But this simple paradigm has now yielded to a remarkably broad appreciation of the cell's functions not only in liver injury, but also in hepatic development, regeneration, xenobiotic responses, intermediary metabolism, and immunoregulation. Among the most exciting prospects is that stellate cells are essential for hepatic progenitor cell amplification and differentiation. Equally intriguing is the remarkable plasticity of stellate cells, not only in their variable intermediate filament phenotype, but also in their functions. Stellate cells can be viewed as the nexus in a complex sinusoidal milieu that requires tightly regulated autocrine and paracrine cross-talk, rapid responses to evolving extracellular matrix content, and exquisite responsiveness to the metabolic needs imposed by liver growth and repair. Moreover, roles vital to systemic homeostasis include their storage and mobilization of retinoids, their emerging capacity for antigen presentation and induction of tolerance, as well as their emerging relationship to bone marrow-derived cells. As interest in this cell type intensifies, more surprises and mysteries are sure to unfold that will ultimately benefit our understanding of liver physiology and the diagnosis and treatment of liver disease.

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“…CRP2 antibodies were raised against the epitope GIKPESAQPHRPTTN, corresponding to amino acid 89-103 of murine CRP2, linked to KLH. 18 MMP-9 IgGs were purchased from R&D Systems (Wiesbaden, Germany), CD45 IgGs from BD (Heidelberg, Germany), F4/80 IgGs from Dianova (Hamburg, Germany), and myeloperoxidase IgGs from Abcam (Cambridge, UK). Alexa 468-and Alexa 488-conjugated secondary antibodies were purchased from Molecular Probes (Eugene, OR, USA).…”

Section: Immunofluorescencementioning

confidence: 99%

Activation of hepatic stellate cells is associated with cytokine expression in thioacetamide-induced hepatic fibrosis in mice

Palacios

Roderfeld

Hemmann

et al. 2008

Laboratory Investigation

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143

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The pathophysiological mechanisms of thioacetamide (TAA)-induced hepatic fibrogenesis are not yet fully understood. In particular, the role of hepatic stellate cells (HSCs) remains unclear. We therefore examined proliferation and transdifferentiation of HSC as well as the underlying molecular mechanisms in TAA-induced fibrosis. Hepatic fibrogenesis was induced in mice by addition of TAA to drinking water. Liver damage was determined by assessment of alanine aminotransferase and aspartate aminotransferase levels, and measurement of collagen deposition. Additionally, expression patterns of a-smooth muscle actin, glial fibrillary acidic protein (GFAP, specific hepatic biomarker for HSC), cysteine-and glycine-rich protein 2 (CRP2, specific marker of HSC transdifferentiation), tissue inhibitor of metalloproteinases-1, matrix metalloproteinase-9 (MMP-9), interleukins (IL-1b, IL-6), platelet-derived growth factors (PDGF-B, PDGF-D) , tumor necrosis factor (TNF)-a, and (transforming growth factor (TGF)-b1 were assessed by real-time PCR. Transcription of GFAP and CRP2 were transiently upregulated during TAA-induced fibrogenesis (punctum maxima (p.m.) week 10 for GFAP and week 14 for CRP2). Similar transient expression patterns were demonstrated for IL-1b, IL-6, TGF-b1, and PDGF-B (p.m. week 12) whereas TNF-a and PDGF-D continuously increased with ongoing liver injury. In particular, not only neutrophil granulocytes, but also macrophages and leukocytes served as a major source for MMP-9 expression. GFAP and CRP2 expression patterns demonstrated transiently increased HSC-activation during TAA-induced hepatic fibrogenesis. The rate of increase of transcription of GFAP correlated best with PDGF-B, whereas CRP2 levels correlated with PDGF-B, PDGF-D, and IL-1b expression. This study demonstrates for the first time that transiently increased activation patterns of HSC are observed in toxically induced hepatic fibrosis. Thus, TAA in drinking water is an effective and elegant model to induce reproducible states of liver fibrosis without parenchymal damage in mice.

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Cryopreservation of hepatic stellate cells

Cited by 13 publications

References 37 publications

Hyaluronan-Supplemented Buffers Preserve Adhesion Mechanisms Facilitating Cryopreservation of Human Hepatic Stem/Progenitor Cells

Hyaluronan-Supplemented Buffers Preserve Adhesion Mechanisms Facilitating Cryopreservation of Human Hepatic Stem/Progenitor Cells

Hepatic Stellate Cells: Protean, Multifunctional, and Enigmatic Cells of the Liver

Activation of hepatic stellate cells is associated with cytokine expression in thioacetamide-induced hepatic fibrosis in mice

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