Shedding New Light on Cancer with Non-Linear Optical Microscopy

Oesophageal cancer, one of the most aggressive cancer types is considered the seventh most common cancer in terms of incidence and the sixth most common cause of cancer deaths worldwide due to late diagnosis. In the UK, the oesophageal cancer incidence rate has increased by approximately 10% since the 1990s. At present, histopathology is the gold standard method for the diagnosis of oesophageal cancer, which rely on biopsy collection using an endoscopy procedure followed by the histological sample’s preparation. This method is invasive, time-consuming, and largely based on the pathologist's experience of diagnosis. Therefore, new diagnostic techniques are required to provide non-invasive methods for early and rapid diagnosis. Raman scattering has the potential to replace histopathology as the gold standard for diagnosis for a wide range of diseases. Raman scattering provides stain-free imaging with chemical-specificity derived from the intrinsic vibrational signatures of biomolecules. However, the low scattering cross-section severely limits the image acquisition speeds and like conventional histopathology, requires tissue sectioning to provide morphological imaging below the surface of tissue biopsies. Stimulated Raman scattering (SRS) has recently appeared as a powerful technique for (near)real-time Raman imaging in intact tissue samples. Thework in this thesis aimed to develop the stimulated Raman scattering (SRS) for rapid wavelength tuning and chemical imaging of clinical samples, such as cancer biopsies. This was achieved by making modification to a laser cavity to reduce the time of the wavelength tuning by approximately 35 times compared to the original cavity design. Furthermore, the cavity modification led to the spectra being separated efficiently and the wavelength tuning controlled by cavity length changes only. The improved design was applied to image frozen oesophageal tissues, which have four major pathology groups, normal, inflammation, columnar-lined (Barrett's) oesophagus (CLO) and low-grade dysplasia. A large area imaging was performed using the SRS technique at 2930 cm-1 for four different oesophageal tissues, which presented the morphological and structural information. However, histopathological diagnosis depends on the visualisation of the cell nucleus in the tissue. This component was not highlighted until the stimulated Raman histology approach was developed for small regions of interest in the CLO and the low-grade dysplasia sample, which required two different frequencies at 2840 cm-1 and 2930 cm-1. All SRS images were compared to haematoxlin and eosin (H&E) stained sections. Further comparisons were made between SRS and Raman imaging techniques, with SRS offering faster acquisition times and a higher spatial resolution. The spectral signature for the different pathological groups in the oesophageal tissues were explored in the high wavenumber (2800 – 2930 cm-1) region using hyperspectral SRS and compared with the spectra from the Raman. K-means clustering analysis was used to explore the morphochemical information using the CLO and low-grade dysplasia sections. Both techniques were able to demonstrate unique information such as the epithelial cells that form the oesophagus glands and surrounding connective tissue. It is concluded that SRS has the power to be one of the ideal imaging modalities to gather the molecular information in biological samples. However, it still needs more development due to the complexity of the system

Diagnosis and Treatment of Small Bowel Disorders

Over the last few decades, remarkable progress has been made in understanding the aetiology and pathophysiology of diseases and many new theories emphasize the importance of the small bowel ‘ecosystem’ in the pathogenesis of acute and chronic illness. Emerging factors such as microbiome, stem cells, innate intestinal immunity and the enteric nervous system along with mucosal and endothelial barriers have key role in the development of gastrointestinal and extra-intestinal diseases. Therefore, the small intestine is considered key player in metabolic disease development, including diabetes mellitus, and other diet-related disorders such as celiac and non-celiac enteropathies. Another major field is drug metabolism and its interaction with microbiota. Moreover, the emergence of gut-brain, gut-liver and gut-blood barriers points toward the important role of small intestine in the pathogenesis of common disorders, such as liver disease, hypertension and neurodegenerative disease. However, the small bowel remains an organ that is difficult to fully access and assess and accurate diagnosis often poses a clinical challenge. Eventually, the therapeutic potential remains untapped. Therefore, it is due time to direct our interest towards the small intestine and unravel the interplay between small-bowel and other gastrointestinal (GI) and non-GI related maladies

Antioxidant and DPPH-Scavenging Activities of Compounds and Ethanolic Extract of the Leaf and Twigs of Caesalpinia bonduc L. Roxb.

Antioxidant effects of ethanolic extract of Caesalpinia bonduc and its isolated bioactive compounds were evaluated in vitro. The compounds included two new cassanediterpenes, 1α,7α-diacetoxy-5α,6β-dihydroxyl-cass-14(15)-epoxy-16,12-olide (1)and 12α-ethoxyl-1α,14β-diacetoxy-2α,5α-dihydroxyl cass-13(15)-en-16,12-olide(2); and others, bonducellin (3), 7,4’-dihydroxy-3,11-dehydrohomoisoflavanone (4), daucosterol (5), luteolin (6), quercetin-3-methyl ether (7) and kaempferol-3-O-α-L-rhamnopyranosyl-(1Ç2)-β-D-xylopyranoside (8). The antioxidant properties of the extract and compounds were assessed by the measurement of the total phenolic content, ascorbic acid content, total antioxidant capacity and 1-1-diphenyl-2-picryl hydrazyl (DPPH) and hydrogen peroxide radicals scavenging activities.Compounds 3, 6, 7 and ethanolic extract had DPPH scavenging activities with IC50 values of 186, 75, 17 and 102 μg/ml respectively when compared to vitamin C with 15 μg/ml. On the other hand, no significant results were obtained for hydrogen peroxide radical. In addition, compound 7 has the highest phenolic content of 0.81±0.01 mg/ml of gallic acid equivalent while compound 8 showed the highest total antioxidant capacity with 254.31±3.54 and 199.82±2.78 μg/ml gallic and ascorbic acid equivalent respectively. Compound 4 and ethanolic extract showed a high ascorbic acid content of 2.26±0.01 and 6.78±0.03 mg/ml respectively.The results obtained showed the antioxidant activity of the ethanolic extract of C. bonduc and deduced that this activity was mediated by its isolated bioactive compounds

A study of Raman spectroscopy for the early detection and characterization of prostate cancer using blood plasma and prostate tissue biopsy.

Prostate cancer (PC) is the most common cancer in men after non-melanoma skin cancer in the United Kingdom (Cancer Research UK, 2019). Current diagnostic methods (PSA, DRE, MRI & prostate biopsy) have limitations as these are unable to distinguish between low-risk cancers that do not need active treatment from cancers which are more likely to progress. In addition, prostate biopsy is invasive with potential side effects. There is an urgent need to identify new biomarkers for early diagnosis and prognostication in PC. Raman spectroscopy (RS) is an optical technique that utilises molecular-specific, inelastic scattering of light photons to interrogate biological samples. When laser light is incident on a biological sample, the photons from the laser light can interact with the intramolecular bonds present within the sample. The Raman spectrum is a direct function of the molecular composition of the tissue, providing a molecular fingerprint of the phenotypic expression of the cells and tissues, which can give good objective information regarding the pathological state of the biological sample under interrogation. We applied a technique of drop coating deposition Raman (DCDR) spectroscopy using both blood plasma and sera to see if a more accurate prediction of the presence and progression of prostate cancer could be achieved than PSA which would allow for blood sample triage of patients into at risk groups. 100 participants were recruited for this study (100 blood plasma and 100 serum samples). Secondly, 79 prostate tissue samples (from the same cohort) were interrogated with the aid of Raman micro-spectroscopy to ascertain if Raman spectroscopy can provide molecular fingerprint that can be utilised for real time in vivo analysis. Multivariate analysis of support vector machine (SVM) learning and linear discriminant analysis (LDA) were utilised differently to test the performance accuracy of the discriminant model for distinguishing between benign and malignant mean plasma spectra. SVM gave a better performance accuracy than LDA with sensitivity and specificity of 96% and 97% respectively and an area under the curve (AUC) of 0.98 as opposed to sensitivity and specificity of 51% and 80% respectively with AUC of 0.74 using LDA. Slightly lower performance accuracy was also observed when blood serum mean spectra analysis was compared with blood plasma mean spectra analysis for both machine learning algorithms (SVM & LDA). Tissue spectral analysis on the other hand recorded an overall accuracy of 80.8% and AUC of 0.82 with the SVM algorithm compared to performance accuracy of 75% and AUC of 0.77 with LDA algorithm (better performance noted with the SVM algorithm). The small sample size of 79 prostate biopsy tissues was responsible for the low sensitivity and specificity. Therefore, the tissues were insufficient to describe all the variances in each group as well as the variability of the gold standard technique. Conclusion: Raman spectroscopy could be a potentially useful technique in the management of Prostate Cancer in areas such as tissue diagnosis, assessment of surgical margin after radical prostatectomy, detection of metastasis, Prostate Cancer screening as well as monitoring and prognosticating patients with Prostate Cancer. However, more needs to be done to validate the approaches outlined in this thesis using prospective collection of new samples to test the classification models independently with sufficient statistical power. At this stage only the fluid-based models are likely to be large enough for this validation process

Applications and Experiences of Quality Control

The rich palette of topics set out in this book provides a sufficiently broad overview of the developments in the field of quality control. By providing detailed information on various aspects of quality control, this book can serve as a basis for starting interdisciplinary cooperation, which has increasingly become an integral part of scientific and applied research

Perils of the Fungal Kingdom: Mycotoxins in Food and Feed

Humans and fungi have a complex relationship, especially in regards to the many fungal secondary metabolites that can be produced. When secondary metabolites are toxic to animals and humans they are defined as mycotoxins. Fungi can grow on crops pre-harvest and post-harvest, and have the potential to produce mycotoxins which occur regularly in animal feed and food for human consumption. In high doses, mycotoxins cause variety of problems that result in economic losses and extreme health issues. However, multiple mycotoxins will co-occur in nature and commonly occur in low doses. Thus chronic low daily doses of multiple mycotoxins overtime may result in wear and tear on the body, as well as disease development later in life (for both animals and humans). The goal of this thesis is to provide a broad review of current literature on mycotoxins ranging in topics from their biochemical synthesis and control to their toxicity in various organisms. To accomplish this, in this thesis we look at mycotoxin contamination from a wide perspective and from different disciplines in order to understand how our agricultural practices, trade and economics, diet and health, and regulatory policies both affect and are effected by mycotoxin contamination

Optical Diagnostics in Human Diseases

Optical technologies provide unique opportunities for the diagnosis of various pathological disorders. The range of biophotonics applications in clinical practice is considerably wide given that the optical properties of biological tissues are subject to significant changes during disease progression. Due to the small size of studied objects (from μm to mm) and despite some minimum restrictions (low-intensity light is used), these technologies have great diagnostic potential both as an additional tool and in cases of separate use, for example, to assess conditions affecting microcirculatory bed and tissue viability. This Special Issue presents topical articles by researchers engaged in the development of new methods and devices for optical non-invasive diagnostics in various fields of medicine. Several studies in this Special Issue demonstrate new information relevant to surgical procedures, especially in oncology and gynecology. Two articles are dedicated to the topical problem of breast cancer early detection, including during surgery. One of the articles is devoted to urology, namely to the problem of chronic or recurrent episodic urethral pain. Several works describe the studies in otolaryngology and dentistry. One of the studies is devoted to diagnosing liver diseases. A number of articles contribute to the studying of the alterations caused by diabetes mellitus and cardiovascular diseases. The results of all the presented articles reflect novel innovative research and emerging ideas in optical non-invasive diagnostics aimed at their wider translation into clinical practice

Near-IR plasmonic contrast agents for molecular imaging, cell tracking and clinical translation

textGold nanoparticles attain an intense focus in biomedical imaging applications due to their unique optical properties, facile conjugation with biomolecules, and biocompatibility. Although a considerable amount of work towards the development of gold nanoparticles has been completed, these promising contrast agents have not yet reached the clinic due to several challenges including efficient accumulation at the diseased site, sensitivity of detection in vivo, potential adverse effects, and clearance from the body. High signal-to-background ratio is required to enhance sensitivity of detection. Because near infrared (near-IR) light has the best tissue penetration, contrast agents designed to work in this range can significantly increase imaging sensitivity. Moreover, efficient targeting of the molecular biomarkers on diseased cells can decrease the required dosage, increase the site-specific accumulation, and enhance the imaging sensitivity. Molecular-specific contrast agents developed in this project use directional attachment of antibody molecules to the nanoparticle surface, enhancing the targeting efficacy. Additionally, cell-based delivery of diagnostic and therapeutic agents is gaining much interest due to the immune cells’ special access to the avascular, diseased regions. The contrast agents developed in this project enable detection of just a few cells per unit of imaging volume, enable multiplex imaging, and open up a possibility for tracking different cell populations with noninvasive photoacoustic and ultrasound imaging. Finally, the clearance of nanoparticles from the body dictates their clinical translation. The in vivo pharmacokinetics study along with the proposed in vitro model explored in this project will enable fast, reliable, and cost-efficient screening of promising agents and facilitate quick optimization of nanoparticles for their potential use in the clinic.Biomedical Engineerin