synthesis-soft-polysiloxane-urea-elastomers-for-intraocular-lens

Synthesis of Soft Polysiloxane-urea Elastomers for Intraocular Lens Application

This study discusses a synthesis route for soft polysiloxane-based urea (PSU) elastomers for their applications as accommodating intraocular lenses ...

Fraunhofer-Institute for Interfacial Engineering and Biotechnology IGB

Noninvasive monitoring of tissue-engineered (TE) constructs during their in vitro maturation or postimplantation in vivo is highly relevant for graft evaluation. However, traditional methods for studying cell and matrix components in engineered tissues such as histology, immunohistochemistry, or biochemistry require invasive tissue processing, resulting in the need to sacrifice of TE constructs. Raman spectroscopy offers the unique possibility to analyze living cells label-free in situ and in vivo solely based on their phenotype-specific biochemical fingerprint. In this study, we aimed to determine the applicability of Raman spectroscopy for the noninvasive identification and spectral separation of primary human skin fibroblasts, keratinocytes, and melanocytes, as well as immortalized keratinocytes (HaCaT cells). Multivariate analysis of cell-type-specific Raman spectra enabled the discrimination between living primary and immortalized keratinocytes. We further noninvasively distinguished between fibroblasts, keratinocytes, and melanocytes. Our findings are especially relevant for the engineering of in vitro skin models and for the production of artificial skin, where both the biopsy and the transplant consist of several cell types. To realize a reproducible quality of TE skin, the determination of the purity of the cell populations as well as the detection of potential molecular changes are important. We conclude therefore that Raman spectroscopy is a suitable tool for the noninvasive in situ quality control of cells used in skin tissue engineering applications.

Raman spectroscopy: a noninvasive analysis tool for the discrimination of human skin cells.

Tissue-engineered skin equivalents based on primary isolated fibroblasts and keratinocytes have been shown to be useful tools for functional in vitro tests, including toxicological screenings and drug development. In this study, a commercially available squamous cell carcinoma (SCC) cell line SCC-25 was introduced into epidermal and full-thickness skin equivalents to generate human-based disease-in-a-dish model systems. Interestingly, when cultured either in the epidermis or dermis of full-thickness skin equivalents, SCC-25 cells formed hyper-keratinized tumor cell nests, a phenomenon that is frequently seen in the skin of patients afflicted with SCC. Raman spectroscopy was employed for the label-free cell phenotype characterization within the engineered skin equivalents and revealed the presence of differential protein patterns in keratinocytes and SCC-25 cells. To conclude, the here presented SSC disease-in-a-dish approaches offer the unique opportunity to model SSC in human skin in vitro, which will allow further insight into SSC disease progression, and the development of therapeutic strategies.

Design and analysis of a squamous cell carcinoma in vitro model system.

Although apoptosis and necrosis have distinct features, the identification and discrimination of apoptotic and necrotic cell death in vitro is challenging. Immunocytological and biochemical assays represent the current gold standard for monitoring cell death pathways; however, these standard assays are invasive, render large numbers of cells and impede continuous monitoring experiments. In this study, both room temperature (RT)-induced apoptosis and heat-triggered necrosis were analyzed in individual Saos-2 and SW-1353 cells by utilizing Raman microspectroscopy. A targeted analysis of defined cell death modalities, including early and late apoptosis as well as necrosis, was facilitated based on the combination of Raman spectroscopy with fluorescence microscopy. Spectral shifts were identified in the two cell lines that reflect biochemical changes specific for either RT-induced apoptosis or heat-mediated necrosis. A supervised classification model specified apoptotic and necrotic cell death based on single cell Raman spectra. To conclude, Raman spectroscopy allows a non-invasive, continuous monitoring of cell death, which may help shedding new light on complex pathophysiological or drug-induced cell death processes. 

Sibylle Thude

Fraunhofer-Institute for Interfacial Engineering and Biotechnology IGB

Synthesis of Soft Polysiloxane-urea Elastomers for Intraocular Lens Application