Dataset Information

Human testicular matrix - Derivation and characterization of a cytocompatible scaffold from human testis

ABSTRACT: ITo derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine, we developed a method to produce a cytocompatible decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components. The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a wide variety of animal and human tissue sources. However, so far, testis scaffolds have never been considered for constructive remodelling purposes. We have therefore developed a protocol for the preparation of cell-free extracellular matrix and characterized the material extensively using a combination of proteomics, immunohistochemsitry, and cell population assays. To prepare cell-free matrix, human cadaveric testicular tissue was exposed for 24 h or 48 h to 1% Triton X-100 and/or 1% sodium dodecyl sulfate (SDS). The extent of decellularization was evaluated by histology. Confirmation of cell removal in DTM was done by a DNA quantification technique. The protein composition was analysed by LC-MS/MS. The retention of testicular tissue-specific characteristics was evaluated by immunohistochemistry, Alcian blue staining and scanning electron microscopy. Soluble toxicity and testicular cell attachment was assessed to check the cytocompatibility of DTM scaffolds. Histological analysis showed that DTM could be obtained by mechanical agitation in 1% SDS for 24 h. The resulting DTM was found to be clear of cells while retaining the typical three-dimensional structure. Proteomics analysis revelaed the presence of the major components of the native tissue scaffold, including collagen type I and IV, fibronectin, laminin and glycosaminoglycans. In addition, numerous additional ECM proteins in DTM were detected, indicating its complex nature. Importantly, we demonstrated that DTM scaffolds are not cytotoxic, as evidenced by MTT assay showing a normal fibroblast proliferation activity after indirect exposure, and support testicular cell attachment and infiltration.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Testis

SUBMITTER: Michael Landreh

LAB HEAD: Hans Jornvall

PROVIDER: PXD001524 | Pride | 2015-01-30

REPOSITORIES: Pride

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Publications

Derivation and characterization of a cytocompatible scaffold from human testis.

Baert Y Y Stukenborg J-B JB Landreh M M De Kock J J Jörnvall H H Söder O O Goossens E E

Human reproduction (Oxford, England) 20141211 2

<h4>Study question</h4>Is it possible to derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine?<h4>Summary answer</h4>We developed a method to produce a cytocompatible decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components.<h4>What is known already</h4>The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a ...[more]

PMID: 25505010

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Project description:ITo derive a scaffold from human testis for the purpose of tissue engineering and regenerative medicine, we developed a method to produce a cytocompatible decellularized testicular matrix (DTM) while maintaining the native tissue-specific characteristics and components. The potential benefits of tissue-specific scaffolds consisting of naturally-derived extracellular matrix (ECM) have been demonstrated using a wide variety of animal and human tissue sources. However, so far, testis scaffolds have never been considered for constructive remodelling purposes. We have therefore developed a protocol for the preparation of cell-free extracellular matrix and characterized the material extensively using a combination of proteomics, immunohistochemsitry, and cell population assays. To prepare cell-free matrix, human cadaveric testicular tissue was exposed for 24 h or 48 h to 1% Triton X-100 and/or 1% sodium dodecyl sulfate (SDS). The extent of decellularization was evaluated by histology. Confirmation of cell removal in DTM was done by a DNA quantification technique. The protein composition was analysed by LC-MS/MS. The retention of testicular tissue-specific characteristics was evaluated by immunohistochemistry, Alcian blue staining and scanning electron microscopy. Soluble toxicity and testicular cell attachment was assessed to check the cytocompatibility of DTM scaffolds. Histological analysis showed that DTM could be obtained by mechanical agitation in 1% SDS for 24 h. The resulting DTM was found to be clear of cells while retaining the typical three-dimensional structure. Proteomics analysis revelaed the presence of the major components of the native tissue scaffold, including collagen type I and IV, fibronectin, laminin and glycosaminoglycans. In addition, numerous additional ECM proteins in DTM were detected, indicating its complex nature. Importantly, we demonstrated that DTM scaffolds are not cytotoxic, as evidenced by MTT assay showing a normal fibroblast proliferation activity after indirect exposure, and support testicular cell attachment and infiltration.

Project description:Cholangiocarcinoma (CCA) is a hepatobiliary malignancy with dismal prognosis. Currently available models fail to recapitulate the full complexity of CCA, particularly the desmoplastic environment and the interplay between cancer cells and the extracellular matrix (ECM). We aimed to study the role of epithelial tumor cells in ECM deposition and desmoplasia by designing a model encompassing primary epithelial CCA organoids (CCAOs) and native liver and tumor scaffolds obtained by decellularization Background & Aims: Cholangiocarcinoma (CCA) is a hepatobiliary malignancy with dismal prognosis. Currently available models fail to recapitulate the full complexity of CCA, particularly the desmoplastic environment and the interplay between cancer cells and the extracellular matrix (ECM). We aimed to study the role of epithelial tumor cells in ECM deposition and desmoplasia by designing a model encompassing primary epithelial CCA organoids (CCAOs) and native liver and tumor scaffolds obtained by decellularization. Results: Decellularization resulted in effective removal of cells while preserving ECM structure and retaining important characteristics of the tissue origin. When culturing CCAOs in CCA-M, compared to TFL-M and BME, the genome-wide gene expression profile much more resembled the transcriptome of primary CCA tumor tissue in vivo, with an accompanying increase in chemoresistance. CCAOs in decellularized matrix, both CCA-M and TFL-M, exhibited the formation of complex morphological structures, and revealed environment-dependent proliferation and migration dynamics, driven by the occurrence of epithelial-mesenchymal transition (EMT). CCA-M induced specific extracellular matrix protein production in CCAOs, such as fibronectin 1 (FN1), which is related to desmoplasia and patient survival. In TFL-M, lacking the desmoplastic environment, CCAOs were able to initiate a desmoplastic reaction directly through increased production of multiple collagen types. Conclusions: This improved model of cholangiocarcinoma, combining organoids and native extracellular matrix, recapitulates key components of CCA tumor biology, including transcriptome profiles, migration patterns, EMT, and ECM protein production. The increased production of extracellular matrix proteins suggests that epithelial tumor cells can contribute to their own desmoplastic environment.

Dataset Information

Human testicular matrix - Derivation and characterization of a cytocompatible scaffold from human testis

Publications

Derivation and characterization of a cytocompatible scaffold from human testis.

Similar Datasets

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