Virtual surgical planning in craniomaxillofacial surgery: a structured review.

Craniomaxillofacial (CMF) surgery is a challenging and very demanding field that involves the treatment of congenital and acquired conditions of the face and head. Due to the complexity of the head and facial region, various tools and techniques were developed and utilized to aid surgical procedures and optimize results. Virtual Surgical Planning (VSP) has revolutionized the way craniomaxillofacial surgeries are planned and executed. It uses 3D imaging computer software to visualize and simulate a surgical procedure. Numerous studies were published on the usage of VSP in craniomaxillofacial surgery. However, the researchers found inconsistency in the previous literature which prompted the development of this review. This paper aims to provide a comprehensive review of the findings of the studies by conducting an integrated approach to synthesize the literature related to the use of VSP in craniomaxillofacial surgery. Twenty-nine related articles were selected as a sample and synthesized thoroughly. These papers were grouped assigning to the four subdisciplines of craniomaxillofacial surgery: orthognathic surgery, reconstructive surgery, trauma surgery and implant surgery. The following variables - treatment time, the accuracy of VSP, clinical outcome, cost, and cost-effectiveness - were also examined. Results revealed that VSP offers advantages in craniomaxillofacial surgery over the traditional method in terms of duration, predictability and clinical outcomes. However, the cost aspect was not discussed in most papers. This structured literature review will thus provide current findings and trends and recommendations for future research on the usage of VSP in craniomaxillofacial surgery.

Biomechanical and Biophysical Properties of Breast Cancer Cells Under Varying Glycemic Regimens.

Diabetes accelerates cancer cell proliferation and metastasis, particularly for cancers of the pancreas, liver, breast, colon, and skin. While pathways linking the 2 disease conditions have been explored extensively, there is a lack of information on whether there could be cytoarchitectural changes induced by glucose which predispose cancer cells to aggressive phenotypes. It was thus hypothesized that exposure to diabetes/high glucose alters the biomechanical and biophysical properties of cancer cells more than the normal cells, which aids in advancing the cancer. For this study, atomic force microscopy indentation was used through microscale probing of multiple human breast cancer cells (MCF-7, MDA-MB-231), and human normal mammary epithelial cells (MCF-10A), under different levels of glycemic stress. These were used to study both benign and malignant breast tissue behaviors. Benign cells (MCF-10A) recorded higher Young's modulus values than malignant cells (MCF-7 and MDA-231) under normoglycemic conditions, which agrees with the current literature. Moreover, exposure to high glucose (for 48 hours) decreased Young's modulus in both benign and malignant cells, to the effect that the cancer cells showed a complete loss in elasticity with high glucose. This provides a possible insight into a link between glycemic stress and cytoskeletal strength. This work suggests that reducing glycemic stress in cancer patients and those at risk can prove beneficial in restoring normal cytoskeletal structure.

3D bioprinted mammary organoids and tumoroids in human mammary derived ECM hydrogels.

The extracellular matrix (ECM) of tissues is an important mediator of cell function. Moreover, understanding cellular dynamics within their specific tissue context is also important for developmental biology, cancer research, and regenerative medicine. However, robust in vitro models that incorporate tissue-specific microenvironments are lacking. Here we describe a novel mammary-specific culture protocol that combines a self-gelling hydrogel comprised solely of ECM from decellularized rat or human breast tissue with the use of our previously described 3D bioprinting platform. We initially demonstrate that undigested and decellularized mammary tissue can support mammary epithelial and tumor cell growth. We then describe a methodology for generating mammary ECM extracts that can spontaneously gel to form hydrogels. These ECM hydrogels retain unique structural and signaling profiles that elicit differential responses when normal mammary and breast cancer cells are cultured within them. Using our bioprinter, we establish that we can generate large organoids/tumoroids in the all mammary-derived hydrogel. These findings demonstrate that our system allows for growth of organoids/tumoroids in a tissue-specific matrix with unique properties, thus providing a suitable platform for ECM and epithelial/cancer cell studies. STATEMENT OF SIGNIFICANCE: Factors within extracellular matrices (ECMs) are specific to their tissue of origin. It has been shown that tissue specific factors within the mammary gland's ECM have pronounced effects on cellular differentiation and cancer behavior. Understanding the role of the ECM in controlling cell fate has major implications for developmental biology, tissue engineering, and cancer therapy. However, in vitro models to study cellular interactions with tissue specific ECM are lacking. Here we describe the generation of 3D hydrogels consisting solely of human or mouse mammary ECM. We demonstrate that these novel 3D culture substrates can sustain large 3D bioprinted organoid and tumoroid formation. This is the first demonstration of an all mammary ECM culture system capable of sustaining large structural growths.

A 3D bioprinter platform for mechanistic analysis of tumoroids and chimeric mammary organoids.

The normal mammary microenvironment can suppress tumorigenesis and redirect cancer cells to adopt a normal mammary epithelial cell fate in vivo. Understanding of this phenomenon offers great promise for novel treatment and detection strategies in cancer, but current model systems make mechanistic insights into the process difficult. We have recently described a low-cost bioprinting platform designed to be accessible for basic cell biology laboratories. Here we report the use of this system for the study of tumorigenesis and microenvironmental redirection of breast cancer cells. We show our bioprinter significantly increases tumoroid formation in 3D collagen gels and allows for precise generation of tumoroid arrays. We also demonstrate that we can mimic published in vivo findings by co-printing cancer cells along with normal mammary epithelial cells to generate chimeric organoids. These chimeric organoids contain cancer cells that take part in normal luminal formation. Furthermore, we show for the first time that cancer cells within chimeric structures have a significant increase in 5-hydroxymethylcytosine levels as compared to bioprinted tumoroids. These results demonstrate the capacity of our 3D bioprinting platform to study tumorigenesis and microenvironmental control of breast cancer and highlight a novel mechanistic insight into the process of microenvironmental control of cancer.

Effects of protein-coated nanofibers on conformation of gingival fibroblast spheroids: potential utility for connective tissue regeneration.

Deep wounds in the gingiva caused by trauma or surgery require a rapid and robust healing of connective tissues. We propose utilizing gas-brushed nanofibers coated with collagen and fibrin for that purpose. Our hypotheses are that protein-coated nanofibers will: (i) attract and mobilize cells in various spatial orientations, and (ii) regulate the expression levels of specific extracellular matrix (ECM)-associated proteins, determining the initial conformational nature of dense and soft connective tissues. Gingival fibroblast monolayers and 3D spheroids were cultured on ECM substrate and covered with gas-blown poly-(DL-lactide-co-glycolide) (PLGA) nanofibers (uncoated/coated with collagen and fibrin). Cell attraction and rearrangement was followed by F-actin staining and confocal microscopy. Thicknesses of the cell layers, developed within the nanofibers, were quantified by ImageJ software. The expression of collagen1α1 chain (Col1α1), fibronectin, and metalloproteinase 2 (MMP2) encoding genes was determined by quantitative reverse transcription analysis. Collagen- and fibrin- coated nanofibers induced cell migration toward fibers and supported cellular growth within the scaffolds. Both proteins affected the spatial rearrangement of fibroblasts by favoring packed cell clusters or intermittent cell spreading. These cell arrangements resembled the structural characteristic of dense and soft connective tissues, respectively. Within three days of incubation, fibroblast spheroids interacted with the fibers, and grew robustly by increasing their thickness compared to monolayers. While the ECM key components, such as fibronectin and MMP2 encoding genes, were expressed in both protein groups, Col1α1 was predominantly expressed in bundled fibroblasts grown on collagen fibers. This enhanced expression of collagen1 is typical for dense connective tissue. Based on results of this study, our gas-blown, collagen- and fibrin-coated PLGA nanofibers are viable candidates for engineering soft and dense connective tissues with the required structural characteristics and functions needed for wound healing applications. Rapid regeneration of these layers should enhance healing of open wounds in a harsh oral environment.

Energy dissipation mapping of cancer cells.

The purpose of this study is to map the energy dissipation of Jurkat cells using a single 60 nanosecond pulse electric field (NsPEF), primarily through atomic force microscopy (AFM). The phase shift is generated by the sample elements that do not have a heterogeneous surface. Monitoring and manipulating the phase shift is a powerful way for determining the dissipated energy and plotting the topography. The dissipated energy is a relative value, so the silica wafer and cover slip are given a set reference while the transmission of energy between the tip of the cantilever and cell surfaces is measured. The most important finding is that the magnitude and the number of variations in the dissipated energy change with the strength of NsPEF applied. Utilizing a single low field strength NsPEF (15kV/cm), minor changes in dissipated energy were found. The application of a single high field strength NsPEF (60kV/cm) to Jurkat cells resulted in a higher dissipated energy change versus that of in the low field strength condition. Thus, the dissipated energy from the Jurkat cells changes with the strength of NsPEF. By analyzing the forces via investigation in the tapping mode of the AFM, the stabilization of the cytoskeleton and membrane of the cell are related to the strength of NsPEF applied. Furthermore, the strength of NsPEF indicates a meaningful relationship to the survival of the Jurkat cells.

Nanosecond pulsed electric field induced changes in cell surface charge density.

This study reports that the surface charge density changes in Jurkat cells with the application of single 60 nanosecond pulse electric fields, using atomic force microscopy. Using an atomic force microscope tip and Jurkat cells on silica in a 0.01M KCl ionic concentration, we were able to measure the interfacial forces, while also predicting surface charge densities of both Jurkat cell and silica surfaces. The most important finding is that the pulsing conditions varyingly reduced the cells' surface charge density. This offers a novel way in which to examine cellular effects of pulsed electric fields that may lead to the identification of unique mechanical responses. Compared to a single low field strength NsPEF (15kV/cm) application, exposure of Jurkat cells to a single high field strength NsPEF (60kV/cm) resulted in a further reduction in charge density and major morphological changes. The structural, physical, and chemical properties of biological cells immensely influence their electrostatic force; we were able to investigate this through the use of atomic force microscopy by measuring the surface forces between the AFM's tip and the Jurkat cells under different pulsing conditions as well as the interfacial forces in ionic concentrations.

Training to Improve Precision and Accuracy in the Measurement of Fiber Morphology.

An estimated $7.1 billion dollars a year is spent due to irreproducibility in pre-clinical data from errors in data analysis and reporting. Therefore, developing tools to improve measurement comparability is paramount. Recently, an open source tool, DiameterJ, has been deployed for the automated analysis of scanning electron micrographs of fibrous scaffolds designed for tissue engineering applications. DiameterJ performs hundreds to thousands of scaffold fiber diameter measurements from a single micrograph within a few seconds, along with a variety of other scaffold morphological features, which enables a more rigorous and thorough assessment of scaffold properties. Herein, an online, publicly available training module is introduced for educating DiameterJ users on how to effectively analyze scanning electron micrographs of fibers and the large volume of data that a DiameterJ analysis yields. The end goal of this training was to improve user data analysis and reporting to enhance reproducibility of analysis of nanofiber scaffolds. User performance was assessed before and after training to evaluate the effectiveness of the training modules. Users were asked to use DiameterJ to analyze reference micrographs of fibers that had known diameters. The results showed that training improved the accuracy and precision of measurements of fiber diameter in scanning electron micrographs. Training also improved the precision of measurements of pore area, porosity, intersection density, and characteristic fiber length between fiber intersections. These results demonstrate that the DiameterJ training module improves precision and accuracy in fiber morphology measurements, which will lead to enhanced data comparability.