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Scientists from the CNBP have reported an advanced imaging technique that allows the condition of joint cartilage to be examined—right down to the molecular level. The technique has potential for diagnostics and treatment-planning of cartilage disease and impairment, including for osteoarthritis.
Read more from the news item posted on the Medical Xpress web site.
CNBP scientists have reported an advanced new imaging technique that allows the condition of joint cartilage to be examined—right down to a molecular level. The technique has potential for diagnostics and treatment-planning of cartilage disease and impairment, including for osteoarthritis.
“Damage and degradation of cartilage around joints leads to severe pain and loss of mobility,” says Dr Saabah Mahbub, Research Fellow at the ARC Centre of Excellence for Nanoscale BioPhotonics (CNBP) and lead author of the published study.
“We need a tool to help us to determine objectively, the degree of problem that the joint cartilage is exhibiting. We then need a way to be able to monitor the effectiveness of any cartilage regeneration therapies that are able to be undertaken,” he says.
“Ideally we need to be able to do this monitoring at a molecular level and in a minimally invasive way.”
A cutting-edge technique termed hyperspectral imaging was used by Dr Mahbub to achieve this. This combined the power of an advanced optical microscope together with high powered data analysis, to measure and image the electromagnetic light-waves being given off by the cartilage tissue and cartilage cells known as chondrocytes.
“In this study, we applied our advanced hyperspectral microscopy to osteoarthritic human cartilage—to investigate its capacity to generate molecular data and to help us characterise the cartilage disease-state, as well as to examine potential treatment effects,” he says.
Read the full media release here.
Journal: Scientific Reports.
Publication title: Non-Invasive Monitoring of Functional state of Articular Cartilage tissue with Label-Free Unsupervised Hyperspectral Imaging.
Authors: Saabah B. Mahbub, Anna Guller, Jared M. Campbell, Ayad G. Anwer, Martin E. Gosnell, Graham Vesey & Ewa M. Goldys.
Abstract: Damage and degradation of articular cartilage leads to severe pain and loss of mobility. the development of new therapies for cartilage regeneration for monitoring their effect requires further study of cartilage, ideally at a molecular level and in a minimally invasive way. Hyperspectral microscopy is a novel technology which utilises endogenous fluorophores to non-invasively assess the molecular composition of cells and tissue. In this study, we applied hyperspectral microscopy to healthy bovine articular cartilage and osteoarthritic human articular cartilage to investigate its capacity to generate informative molecular data and characterise disease state and treatment effects. We successfully demonstrated label-free fluorescence identification of collagen type I and II – isolated in cartilage here for the first time and the co-enzymes free NADH and FAD which together give the optical redox ratio that is an important measure of metabolic activity. the intracellular composition of chondrocytes was also examined. Differences were observed in the molecular ratios within the superficial and transitional zones of the articular cartilage which appeared to be influenced by disease state and treatment. These findings show that hyperspectral microscopy could be useful for investigating the molecular underpinnings of articular cartilage degradation and repair. As it is non-invasive and non-destructive, samples can be repeatedly assessed over time, enabling true time-course experiments with in-depth molecular data. Additionally, there is potential for the hyperspectral approach to be adapted for patient examination to allow the investigation of cartilage state. this could be of advantage for assessment and diagnosis as well as treatment monitoring.
A new CNBP paper “Statistically strong label-free quantitative identification of native fluorophores in a biological sample,” by Saabah B. Mahbub (first author pictured), Martin Plöschner, Martin E. Gosnell, Ayad G. Anwer and Ewa M. Goldys has just been published in Scientific Reports and is available online.
This work addresses a genuine shortage of methods for real-time continuous monitoring of biochemistry of cells and tissues, especially live cells. Saabah Mahbub and team developed an automated and unbiased unmixing methodology to non-invasively detect the presence and spatial distributions of endogenous fluorophores in retina cells. The method was validated on artificial images, where the addition of a varying known level of noise has allowed to quantify the accuracy of spectral unmixing.
With its capability for high throughput, automation and embedded compatibility with statistical analysis this work will contribute to improved quantification and objectivity in biomedical research.
A new publication from CNBP researchers Aziz Ul Rehman (pictured), Ayad Anwer, Martin Gosnell, Saabah Mahbub, Guozhen Liu and Ewa Goldys demonstrates the validation of an innovative hyperspectral unmixing methodology, that can derive chemical information from cell colour.
The work has just been reported in the journal ‘Biomedical Optics Express’ and is accessible online.
Journal: Biomedical Optics Express.
Title: Fluorescence quenching of free and bound NADH in HeLa cells determined by hyperspectral imaging and unmixing of cell autofluorescence.
Authors: Aziz Ul Rehman, Ayad G. Anwer, Martin E. Gosnell, Saabah B. Mahbub, Guozhen Liu, and Ewa M. Goldys.
Abstract: Carbonyl cyanide-p-trifluoro methoxyphenylhydrazone (FCCP) is a well-known mitochondrial uncoupling agent. We examined FCCP-induced fluorescence quenching of reduced nicotinamide adenine dinucleotide / nicotinamide adenine dinucleotide phosphate (NAD(P)H) in solution and in cultured HeLa cells in a wide range of FCCP concentrations from 50 to 1000µM. A non-invasive label-free method of hyperspectral imaging of cell autofluorescence combined with unsupervised unmixing was used to separately isolate the emissions of free and bound NAD(P)H from cell autofluorescence. Hyperspectral image analysis of FCCP-treated HeLa cells confirms that this agent selectively quenches fluorescence of free and bound NAD(P)H in a broad range of concentrations. This is confirmed by the measurements of average NAD/NADH and NADP/NADPH content in cells. FCCP quenching of free NAD(P)H in cells and in solution is found to be similar, but quenching of bound NAD(P)H in cells is attenuated compared to solution quenching possibly due to a contribution from the metabolic and/or antioxidant response in cells. Chemical quenching of NAD(P)H fluorescence by FCCP validates the results of unsupervised unmixing of cell autofluorescence.
CNBP PhD student Saabah Mahbub has won best poster prize at the ‘Research by Degrees Showcase’, an event held by the Department of Physics and Astronomy at Macquarie University.
Saabah’s poster was titled ‘Label-free functional characterization of stem cell cartilage system by hyperspectral imaging (with unsupervised unmixing) for applications in regenerative medicine.’
Posters were assessed by external judges that included Mr Hugh Ong (Commonwealth Bank of Australia), Prof Fred Watson (Astronomer-in-Charge, AAO), Mr Steve Frisken (Finisar Corporation and Founder and CEO, Cylite Pty Ltd) and Dr Stephen Thompson (Lecia Microsystems).
Well done Saabah. Great work at showcasing your science!