Auto fluorescence of intervertebral disc tissue: a new diagnostic tool

Hoell, Thomas; Huschak, Gerald; Beier, Alexander; Hüttmann, Gereon; Minkus, Yvonne; Holzhausen, Hans-Jürgen; Meisel, Hans-Joerg

doi:10.1007/s00586-006-0157-1

Cited by 15 publications

(11 citation statements)

References 27 publications

(26 reference statements)

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“…Intervertebral disc tissue typically exhibits collagen auto‐fluorescence particularly when irradiated with ultra‐violet light sources . Here, by tuning the two‐photon excitation to 730 nm, the collagen auto‐fluorescence signal was much weaker than the emission due to cellular sources, that is, NADH and FPs.…”

Section: Discussionmentioning

confidence: 98%

“…Recording auto‐fluorescent light in tissues in‐situ may be considerably more difficult due to increased light scattering caused by a dense extracellular matrix . Furthermore, mature collagenous tissues such as IVD tissue are known to exhibit strong auto‐fluorescence originating from collagen . As collagen auto‐fluorescence partially overlaps NADH emission, label‐free two‐photon imaging of live and dead cells may be negatively affected.…”

mentioning

confidence: 99%

“…6 Furthermore, mature collagenous tissues such as IVD tissue are known to exhibit strong auto-fluorescence originating from collagen. 7 As collagen auto-fluorescence partially overlaps NADH emission, label-free two-photon imaging of live and dead cells may be negatively affected. Moreover, cells in the IVD, specifically in NP, are known to be notoriously low in metabolic activity.…”

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confidence: 99%

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In situ label‐free cell viability assessment of nucleus pulposus tissue

Dittmar

Dijk

Zandvoort

et al. 2014

Journal Orthopaedic Research

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Regenerative medicine approaches aiming at treating degenerating intervertebral discs, a major cause of back pain, are increasingly tested in ex-vivo disc explant models mimicking in-vivo conditions. For assessing the efficacy of regenerative therapies, cell viability is commonly measured requiring specific labels to stain cells. Here, we demonstrate and evaluate how cellular autofluorescence can be utilized to non-invasively assess viability in disc tissue in-situ using label-free two-photon microscopy. Live and dead bovine disc cells (0% and 100% cell viability) from the nucleus pulposus were seeded into collagen gels and auto-fluorescence was characterized. Subsequently, nucleus pulposus explants were cultured for 6 days in media with different glucose supplementation (0, 0.25, 0.5, and 1 g/L) to induce different degrees of cell death. Then, samples were split and viability was assessed using label-free twophoton microscopy and conventional staining. Results show that live and dead nucleus pulposus cells systematically emit autofluorescent light with distinct characteristics. Cell viability values obtained with label-free microscopy did not significantly differ from those acquired with staining. In summary, monitoring auto-fluorescence facilitates accurate cell viability assessment in nucleus tissue requiring no additional dyes. Thus, this technique may be suitable for pre-clinical testing of regenerative therapies in nucleus pulposus cultures. ß

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Section: Discussionmentioning

confidence: 98%

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confidence: 99%

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confidence: 99%

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In situ label‐free cell viability assessment of nucleus pulposus tissue

Dittmar

Dijk

Zandvoort

et al. 2014

Journal Orthopaedic Research

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“…To quantify collagen formation, cellular auto-fluorescence was quantified using an epifluorescent microscope. Collagen, a main component of the PCM, shows substantial autofluorescence upon excitation from light ranging from UV to blue [31][32][33]. Thus collagen-based PCM formation can be quantified over time by repeated imaging of auto-fluorescence in this label-free approach.…”

Section: Pericellular Matrix Formation In Alginate Microgelsmentioning

confidence: 99%

Metabolomic Profiling and Mechanotransduction of Single Chondrocytes Encapsulated in Alginate Microgels

Fredrikson

Brahmachary

Erdoğan

et al. 2020

Preprint

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Human articular cartilage is comprised of two main components, the extracellular matrix (ECM) and the pericellular matrix (PCM). The PCM helps to protect chondrocytes in the cartilage from mechanical loads, but in patients with osteoarthritis, the PCM is weakened resulting in increased chondrocyte stress. As chondrocytes are responsible for cartilage synthesis and maintenance, it is important to understand how mechanical loads affect cellular responses of chondrocytes. Many studies have examined the chondrocyte response to in vitro mechanical loading by embedding in stiff agarose. However, these experiments are mostly performed in the absence of PCM which may obscure important responses to mechanotransduction. Here, we demonstrate that drop-based microfluidics allows culture of single chondrocytes in alginate microgels for cell-directed PCM synthesis that closely mimics the in vivo microenvironment. Chondrocytes form PCM over 10 days in these single cell microenvironments. Single cell microgels and monolayer controls were encapsulated in high stiffness agarose to mimic the cartilage PCM. After physiological dynamic compression in a custom-built bioreactor, microgels exhibited distinct metabolomic profiles from both uncompressed and monolayer controls. These results demonstrate the potential of single cell encapsulation in alginate microgels to advance cartilage tissue engineering and basic chondrocyte mechanobiology.

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“…Several different spectroscopic methods of tissue analysis are under development. The autofluorescence phenomenon allows differentiating between traumatized and degenerated disc tissue intraoperatively, and in some cases, it also allows differentiating between the bone and collagen endplate in cervical disc surgery [13], the fluorescence spectroscopy demonstrated a possibility to analyse the LOD [14]. It is well known that Raman scattering and IR absorption spectra allow tracking biochemical changes in the tissue.…”

Section: Introductionmentioning

confidence: 99%

Optical spectroscopic methods for diagnostics of intervertebral disc degeneration

Varanius¹,

Čiplys²,

Gėgžna³

et al. 2018

physics

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The level of degeneration (LOD) in the human intervertebral disc (IVD) could determine the choice of patient treatment strategy, thus there is a need for methods applicable at a point of care that enable quick medical decisions. In this paper, infrared light absorption, FT-Raman scattering and fluorescence spectroscopy were used to analyse the same specimens from different groups of IVD samples. Samples from the lumbar part of spine of 37 patients (mean age 48.2 years old) were included in the study. The gender distribution was 27 women and 10 men. The distribution by LOD in the Pfirrmann scale was as follows: grade 3 (40.5%), grade 4 (48.6%) and grade 5 (10.8%) were evaluated in our samples. Spectroscopic signals were measured and analysed. Features such as medical condition (LOD), patient’s age and gender were taken into account. All three methods of spectroscopy revealed age-dependent spectroscopic features related to LOD. For the first time a promissing possibility was found to recognize the LOD by fluorescence spectra in human IVD material taking gender and age into account. The findings create a background for the design of equipment and the design of experiment of more advanced clinical trials. FT-IR absorption and FT-Raman scattering spectra demonstrated the age dependences of a few components. The Raman peak at 2707 cm–1 seems promising for recognition of lumbar IVD LOD, but a higher number of specimens is necessary.

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Auto fluorescence of intervertebral disc tissue: a new diagnostic tool

Cited by 15 publications

References 27 publications

In situ label‐free cell viability assessment of nucleus pulposus tissue

In situ label‐free cell viability assessment of nucleus pulposus tissue

Metabolomic Profiling and Mechanotransduction of Single Chondrocytes Encapsulated in Alginate Microgels

Optical spectroscopic methods for diagnostics of intervertebral disc degeneration

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