Smart Surgical Microscope based on Optical Coherence Domain Reflectometry

Department of Biomedical EngineeringOver the several decades, there have been clinical needs that requires advanced technologies in medicine. Optical coherence tomography (OCT), one of the newly emerged medical imaging devices, provides non-invasive cross-sectional images in high resolution which is mainly used in ophthalmology. However, due to the limited penetration depth of 1-2 mm in bio-samples, there is a limit to be widely used. In order to easily integrate with existing medical tools and be convenient to users, it is necessary that the sample unit of OCT should be compact and simple. In this study, we developed high-speed swept-source OCT (SS-OCT) for advanced screening of otolaryngology. Synchronized signal sampling with a high-speed digitizer using a clock signal from a swept laser source, its trigger signal is also used to synchronize with the movement of the scanning mirror. The SS-OCT system can reliably provide high-throughput images, and two-axis scanning of galvano mirrors enables real-time acquisition of 3D data. Graphic processing unit (GPU) can performs high-speed data processing through parallel programming, and can also implement perspective projection 3D OCT visualization with optimal ray casting techniques. In the Clinical Study of Otolaryngology, OCT was applied to identify the microscopic extrathyroidal extension (mETE) of papillary thyroid cancer (PTC). As a result to detect the mETE of around 60% in conventional ultrasonography, it could be improved to 84.1% accuracy in our study. The detection ratio of the mETE was calculated by the pathologist analyzing the histologic image. In chapter 3, we present a novel study using combined OCT system integrated with a conventional surgical microscope. In the current set-up of surgical microscope, only two-dimensional microscopic images through the eyepiece view are provided to the surgeon. Thus, image-guided surgery, which provides real-time image information of the tissues or the organs, has been developed as an advanced surgical technique. This study illustrate newly designed optical set-up of smart surgical microscope that combined sample arm of the OCT with an existing microscope. Specifically, we used a beam projector to overlay OCT images on existing eyepiece views, and demonstrated augmented reality images. In chapter 4, in order to develop novel microsurgical instruments, optical coherence domain reflectometry (OCDR) was applied. Introduces smart surgical forceps using OCDR as a sensor that provides high-speed, high-resolution distance information in the tissue. To attach the sensor to the forceps, the lensed fiber which is a small and high sensitivity sensor was fabricated and the results are shown to be less affected by the tilt angle. In addition, the piezo actuator compensates the hand tremor, resulting in a reduction in the human hand tremor of 5 to 15 Hz. Finally, M-mode OCT needle is proposed for microsurgery guidance in ophthalmic surgery. Stepwise transitional core (STC) fiber was applied as a sensor to measure information within the tissue and attached to a 26 gauge needle. It shows the modified OCT system and the position-guided needle design of the sample stage and shows the algorithm flowchart of M-mode OCT imaging software. The developed M-mode OCT needle has been applied to animal studies using rabbit eyes and demonstrates the big-bubble deep anterior lamellar keratoplasty (DALK) surgery for corneal transplantation. Through this study, we propose a novel microsurgical instrument for lamellar keratoplasty and evaluate its feasibility with conventional regular OCT system images. In conclusion, for fundamental study required new augmented reality guided surgery with smart surgical microscope, it is expected that OCT combined with surgical microscope can be widely used. We demonstrated a novel microsurgical instrument to share with light source and the various optical components. Acquired information throughout our integrated system would be a key method to meet a wide range of different clinical needs in the real world.ope

Characterisations of Pre-Descemet’s (Dua’s) layer for its clinical application in keratoplasty

There exists a newly discovered, well defined, acellular, strong layer, termed pre-Descemets layer or Dua’s layer (PDL), in the cornea just anterior to the Descemets membrane. This, with the Descemets membrane, separates along the last row of keratocytes in most cases of deep anterior lamellar keratoplasty with the big bubble technique. Recognition of this layer has considerable impact on lamellar corneal surgery, understanding of posterior corneal biomechanics and posterior corneal pathology, such as descemetocele, acute hydrops and pre-Descemets dystrophies. The aim of this work was to understand the dynamics of big bubble formation in the context of the known architecture of the cornea stroma, ascertain how type 1 (air between deep stroma and PDL), type 2 (air between PDL and Descemets membrane) and mixed bubbles (combination of type 1 and type 2) form and measure the pressure and volume of air required to produce big bubbles in vitro, including the intra-bubble pressure and volume for the different types of big bubbles. We also aimed to characterise the optical coherence tomography characteristics of the different layers in the wall of the big bubbles to help surgeons identify bubbles and understand the structures seen by intra-operative OCT. Finally we evaluated the endothelial cell density and viability in tissue samples obtained for Descemets membrane endothelial keratoplasty (DMEK) and pre-Descemets endothelial keratoplasty (PDEK) by the pneumodissection technique. Air was injected in 145 corneo-scleral samples, which were unsuitable for transplantation. Samples were obtained in organ culture medium from the UK eye banks and transferred to balanced salt solution ready for injection. Different types of big bubble formed were ascertained. Air pressure and volume required to create the big bubble in simulated deep anterior lamellar keratoplasty were measured. It was found that PDL could withstand a high pressure before bursting at around 700 mm of Hg. Accurate measurements of type-2 big bubble proved challenging. The volume of the type-1 BB was fairly consistent at 0.1ml. The movement of air injected in the corneal stroma was studied from the point of exit from the needle tip to complete aeration of the stroma and formation of a BB. This was video recorded and analysed. A very consistent pattern of air movement was observed. The initial movement was predominantly radial from the needle tip to the limbus, then circular in a clock-wise and counter clock-wise direction circumferentially along the limbus, then centripetally to fill the stroma. All type 1 BB started in the centre as multiple small bubbles which coalesced to form a BB. Almost all type 2 BB started at the periphery near the limbus. Ultrastructural examination of the point of commencement of type 2 BB revealed the presence of clusters of fenestrations, which most likely allow air to escape from the otherwise impervious PDL to access the plane between PDL and DM. This was a novel discovery and explained how type 2 BB formed and why they almost always start at the periphery. The consistent pattern of passage of air was in concordance with the known microarchitecture of the central and peripheral corneal stroma. Optical coherence tomography (OCT) characteristics of different types of big bubbles were studied. Samples obtained from the UK eye banks were scanned with Fourier-domain (FD-OCT), while that obtained from Canada eye bank were scanned with Time-domain (TD-OCT). A special clamp was used to affix the corneo-scleral sample on the OCT table with its posterior surface face the machine and mounted on artificial anterior chamber. It was found that FD-OCT could demonstrate type 1 BB wall as two parallel, double contour, hyper-reflective lines with hypo-reflective space in between. It also revealed that in type-2 BB, the posterior wall showed a parallel, double-contour curved hyper-reflective line with a dark space in between. This probably corresponds to the banded and non-banded zones of DM. Dua’s layer presents as a single hyper-reflective line. In TD-OCT, the posterior wall of type-1 and type-2 BB showed a single hyper-reflective curved line rather than the double-contour line. This finding will help cornea surgeons to identify and interpret different layers of big bubble intra-operatively with high resolution OCT devices. Endothelial cell density of PDEK and DMEK tissue were calculated. Endothelial cells were counted using light microscope before pneumodissection. Air was then injected to ascertain the creation of type-1 and type-2 BB. Tissue was then harvested by trephination and endothelial cell density of both types were calculated again. It was found that the corneal endothelial cell count in PEDK tissue preparation is no worse, if not slightly better than, in DMEK tissue prepared by pneumodissection. Therefore, PDEK preparation represents a viable graft preparation technique

Characterisations of Pre-Descemet’s (Dua’s) layer for its clinical application in keratoplasty

There exists a newly discovered, well defined, acellular, strong layer, termed pre-Descemets layer or Dua’s layer (PDL), in the cornea just anterior to the Descemets membrane. This, with the Descemets membrane, separates along the last row of keratocytes in most cases of deep anterior lamellar keratoplasty with the big bubble technique. Recognition of this layer has considerable impact on lamellar corneal surgery, understanding of posterior corneal biomechanics and posterior corneal pathology, such as descemetocele, acute hydrops and pre-Descemets dystrophies. The aim of this work was to understand the dynamics of big bubble formation in the context of the known architecture of the cornea stroma, ascertain how type 1 (air between deep stroma and PDL), type 2 (air between PDL and Descemets membrane) and mixed bubbles (combination of type 1 and type 2) form and measure the pressure and volume of air required to produce big bubbles in vitro, including the intra-bubble pressure and volume for the different types of big bubbles. We also aimed to characterise the optical coherence tomography characteristics of the different layers in the wall of the big bubbles to help surgeons identify bubbles and understand the structures seen by intra-operative OCT. Finally we evaluated the endothelial cell density and viability in tissue samples obtained for Descemets membrane endothelial keratoplasty (DMEK) and pre-Descemets endothelial keratoplasty (PDEK) by the pneumodissection technique. Air was injected in 145 corneo-scleral samples, which were unsuitable for transplantation. Samples were obtained in organ culture medium from the UK eye banks and transferred to balanced salt solution ready for injection. Different types of big bubble formed were ascertained. Air pressure and volume required to create the big bubble in simulated deep anterior lamellar keratoplasty were measured. It was found that PDL could withstand a high pressure before bursting at around 700 mm of Hg. Accurate measurements of type-2 big bubble proved challenging. The volume of the type-1 BB was fairly consistent at 0.1ml. The movement of air injected in the corneal stroma was studied from the point of exit from the needle tip to complete aeration of the stroma and formation of a BB. This was video recorded and analysed. A very consistent pattern of air movement was observed. The initial movement was predominantly radial from the needle tip to the limbus, then circular in a clock-wise and counter clock-wise direction circumferentially along the limbus, then centripetally to fill the stroma. All type 1 BB started in the centre as multiple small bubbles which coalesced to form a BB. Almost all type 2 BB started at the periphery near the limbus. Ultrastructural examination of the point of commencement of type 2 BB revealed the presence of clusters of fenestrations, which most likely allow air to escape from the otherwise impervious PDL to access the plane between PDL and DM. This was a novel discovery and explained how type 2 BB formed and why they almost always start at the periphery. The consistent pattern of passage of air was in concordance with the known microarchitecture of the central and peripheral corneal stroma. Optical coherence tomography (OCT) characteristics of different types of big bubbles were studied. Samples obtained from the UK eye banks were scanned with Fourier-domain (FD-OCT), while that obtained from Canada eye bank were scanned with Time-domain (TD-OCT). A special clamp was used to affix the corneo-scleral sample on the OCT table with its posterior surface face the machine and mounted on artificial anterior chamber. It was found that FD-OCT could demonstrate type 1 BB wall as two parallel, double contour, hyper-reflective lines with hypo-reflective space in between. It also revealed that in type-2 BB, the posterior wall showed a parallel, double-contour curved hyper-reflective line with a dark space in between. This probably corresponds to the banded and non-banded zones of DM. Dua’s layer presents as a single hyper-reflective line. In TD-OCT, the posterior wall of type-1 and type-2 BB showed a single hyper-reflective curved line rather than the double-contour line. This finding will help cornea surgeons to identify and interpret different layers of big bubble intra-operatively with high resolution OCT devices. Endothelial cell density of PDEK and DMEK tissue were calculated. Endothelial cells were counted using light microscope before pneumodissection. Air was then injected to ascertain the creation of type-1 and type-2 BB. Tissue was then harvested by trephination and endothelial cell density of both types were calculated again. It was found that the corneal endothelial cell count in PEDK tissue preparation is no worse, if not slightly better than, in DMEK tissue prepared by pneumodissection. Therefore, PDEK preparation represents a viable graft preparation technique

New Trends and Applications in Femtosecond Laser Micromachining

This book contains the scientific contributions to the Special Issue entitled: "New Trends and Applications in Femtosecond Laser Micromachining". It covers an array of subjects, from the basics of femtosecond laser micromachining to specific applications in a broad spectra of fields such biology, photonics and medicine

The corneal endothelium reflected: Studies on surgical damage tot the corneal endothelium and on endothelial specular microscopy

The endothelium is the innermost layer of the cornea. It is a mosaic of hexagonal cells that is only one cell thick. These endothelial cells actively maintain the corneal hydration equilibrium, and hence are very important for its transparency. However, they are vulnerable to trauma, disease, and intra-ocular surgery, because they have a very restricted capacity for cell division. In part II, studies are presented on the reliability of in-vivo examinations of the endothelium with a specific type of specular microscope. We found that after correct calibration and with adequate assessment methods, valid and reproducible measurements of the endothelial cell density (ECD) could be obtained. However, there is a systematic difference between specular microscopic ECDs and donor cornea ECDs that are measured with a different technique. This difference can only in part be explained by optical factors. In part III, in a clinical and an experimental study, no toxic effects on the endothelium of current common applications of the dye trypan blue, in cataract surgery and in eye banks on donor corneas, could be observed. However, caution is warranted, as higher concentrations or longer exposures were found to cause substantial toxicity. In part IV, endothelial cell loss patterns were investigated after selective transplantation of different parts of the cornea (deep anterior and posterior lamellar keratoplasty, DALK and PLK). After DALK, ECD-loss approached normal levels after an initial drop. When however in PLK a posterior lamella including the endothelium was transplanted, ECD-loss continued at an increased level for up to 7 years. The possible consequences of this result for graft survival were discussed

Clamp-assisted retractor advancement for lower eyelid involutional entropion

Scientific Poster 144PURPOSE: To describe a novel approach to internal repair of lower lid entropion using the Putterman clamp. METHODS: Retrospective, consecutive case series of patients with entropion who underwent retractor advancement using the clamp. RESULTS: Seven eyes of 6 patients (average age: 80; 4 women and 2 men) were analyzed. Complete resolution was achieved in 5 of the 6 patients (83.3%). The 1 patient with recurrence had 2 previous entropion surgeries on each eye over the past 4 years; there was lid laxity, and horizontal tightening was needed. No severe adverse events occurred in the patients. CONCLUSION: Clamp-assisted lower lid retractor advancement offers a safe and effective, minimally invasive approach to involutional entropion. Further study is needed to assess its role in recurrent entropion.postprin

Human corneal endothelial cell culture and corneal transplantation

Cornea is the front transparent window of the eye which is responsible for optimal and clear vision. Transparency of this tissue is highly inevitable and cannot be compromised. Human cornea is made up of multiple layers out of which the posterior layer ‘endothelium’ is responsible for the transparency of the cornea. Endothelium is a monolayer of cells that allow the ions and solutes to transport from aqueous humour to the cornea and back which in turn maintains the transparency of the cornea by preserving the homeostasis between the anterior and posterior cornea. Earlier, it was observed that the endothelium had non regenerating capability however; recent studies have shown that these cells could be proliferated in vitro. Currently, the only method of treatment is the replacement of the diseased endothelium with the healthy donor endothelium. Penetrating keratoplasty which transplants a full thickness cornea was the only solution a decade ago. However, with the new advancements in the field of corneal transplants, specific surgical techniques like DMEK and DSAEK which replace only a part of the cornea have been identified. DSAEK replaces a part of the stroma along with the Descemet’s membrane and endothelium whereas DMEK only replaces the Descemet’s membrane and the endothelium and does not involve stroma. The results in terms of visual rehabilitation and outcomes have been found to be advantageous in these specific surgical procedures. However, DMEK is more challenging then DSAEK as DMEK is not yet a widespread technique, associated with steep learning curves and difficult donor tissue preparation. Despite DMEK is a challenging procedure it is becoming more popular because of the significant advantages in term of faster visual recovery, less postoperative astigmatism and reduced risk of transplant rejection, as compared to the other EK procedures. DMEK has several advantages in terms of rehabilitation rate and post-operative visual outcomes and therefore it is necessary to further refine this technique for a higher uptake of such surgeries and also considering that this is the only possible treatment for treating the patients suffering from endothelial dysfunctions. Although the corneal transplantation is well advanced, due to a limited supply of donor corneas for the transplantation purposes, alternative approaches like culturing corneal endothelium in vitro play an important role. Culturing the endothelium is not the only problem in EK but transplanting a 20 micron thick graft inside the recipient eye is another challenge. Moreover, the donor availability for culturing the corneal endothelium is less, making this strategy further more complicated. The thesis is therefore structured to highlight two significantly important issues in current scenario of endothelial keratoplasty, 1) posterior corneal transplantation or EK which is the on-going method of treatment for EK and 2) Human corneal endothelial cell culture which is the future of EK. Chapter 1 is an introduction to the world of eye banking, its current nature and development in the modern world and as a support to the surgeons not only in terms of new techniques but also devices for selective surgeries. It also highlights the preservation of the corneal tissues which is an important element in the field of eye banking. Eye banks play a significant role in the field of corneal transplants as they collect the human corneas and process them for transplantation. The corneas that are rejected for transplantation can be used for research and therefore development of eye banking and its research can change the field of corneal transplantation. Chapter 2 introduces the field of corneal cell culture and current techniques that are followed for culturing and possible transplantation of the cultured cells. To understand the reason and requirement of tissue engineering, it is important to study the human cornea, its extracellular matrix and its behaviour in different media. The biomechanical behaviour of the thin tissue i.e. the DM in different conditions becomes a relevant part of this study for future engineering which is studied in chapter 3. It is also important to standardize the currently available treatment options to reduce the burden of endothelial compromised patients in the future and avoid damages or tissue wastage that is currently occurring in the surgical theatres by providing standardized tissues in validated preservation medium which is studied in chapter 4. DMEK promises to become a more popular technique for the replacement of unhealthy corneal endothelium as it shows advantages like early rehabilitation rate and visual outcomes. Chapter 5 highlights the importance of new technique in rolling the DMEK tissue for easy insertion and unfolding in the recipient eye compared to the currently used technique with endothelium rolled in opposite direction. Presently, the DMEK tissues are either prepared in the surgical theatre or are stripped in the eye bank and shipped to the surgeons. However, there is no standardized procedure that could help validate a graft before surgery and provide a ready-to-use graft to the surgeons. Chapter 6 describes about a new technique of pre-loading a graft in a commercially available IOL cartridge which can be used as a preservation, transportation and transplantation device. This technique will further reduce graft wastage and will provide the surgeons a pre-validated graft further reducing the overall time in the surgical theatre and related costs. Thus different approaches for standardizing the DMEK technique were studied in the first phase of the thesis. HCECs are currently being cultured using young donor corneas. There are two major issues, firstly, the availability of the young donor corneas is less compared to the old donor corneas and secondly, there is no standard method of culturing the HCECs obtained so far. Therefore, to reduce the global tissue demand, there is a strong need to culture the HCECs from the old donor corneas which are less proliferative and less robust in nature but with high availability of the donor source. Chapter 7 is a study on isolation of HCECs and further culture of these cells from old donor corneas. Once the protocol was obtained, a full length study was performed with high sample size to prove the consistency of this technique which is highlighted in chapter 8. Meanwhile it was also noted that cells from old donors can be cultured using ROCK inhibitor in combination with Hyaluronic Acid (HA). HA induces mechanical force to the cells attaching them forcefully on the base and allows a higher proliferation of old donor cells which was studied in chapter 9. The second part of the thesis therefore investigates the culturing technique of HCECs from old donor corneas. However, once the cells are cultured, another challenge is to transplant them in the anterior chamber of the eye. This can be performed using two strategies, first, to implant the cells as suspension in the anterior chamber which is already been proposed, but the clinical evidence is still not confirmed yet, and second, to develop a carrier to transport the cultured cells. In chapter 10, we identified fish scales as a great source of collagen and therefore have investigated it as a potential scaffold to be used for HCECs culture and transplant in the future. It is also important to understand the regulations that govern the scientific studies and its use for clinical applications. Therefore, we also identified rHSA as a source to replace FCS for preserving human corneas in chapter 11. This will also help to create a synthetic media that could be used for GMP purposes for HCECs culture in the future. In conclusion, it was observed that pre-loading the tissues with endothelium-flapped inwards and preserved in dextran based medium could be a potential solution for providing a validated and standardized DMEK graft for the treatment of current endothelial dysfunction. Eye banks play a major role in the development of these surgical techniques and related devices which will change the face of corneal transplantation in the future. Alternatives like HCECs culture has a potential for the treatment of endothelial disorders and carriers like FSS could be used for culturing and transplanting these cells. However, the efficacy of these cells will only be validated after the clinical study. Considering the regulatory issues, synthetic medium would help both, the eye banks for preserving the corneas and its new products like pre-loaded DMEK and for cell culture in the future