3D U-Net for automated detection of multiple sclerosis lesions: utility of transfer learning from other pathologies.

Background and purpose: Deep learning algorithms for segmentation of multiple sclerosis (MS) plaques generally require training on large datasets. This manuscript evaluates the effect of transfer learning from segmentation of another pathology to facilitate use of smaller MS-specific training datasets. That is, a model trained for detection of one type of pathology was re-trained to identify MS lesions and active demyelination. Materials and methods: In this retrospective study using MRI exams from 149 patients spanning 4/18/2014 to 7/8/2021, 3D convolutional neural networks were trained with a variable number of manually-segmented MS studies. Models were trained for FLAIR lesion segmentation at a single timepoint, new FLAIR lesion segmentation comparing two timepoints, and enhancing (actively demyelinating) lesion segmentation on T1 post-contrast imaging. Models were trained either de-novo or fine-tuned with transfer learning applied to a pre-existing model initially trained on non-MS data. Performance was evaluated with lesionwise sensitivity and positive predictive value (PPV). Results: For single timepoint FLAIR lesion segmentation with 10 training studies, a fine-tuned model demonstrated improved performance [lesionwise sensitivity 0.55 ± 0.02 (mean ± standard error), PPV 0.66 ± 0.02] compared to a de-novo model (sensitivity 0.49 ± 0.02, p = 0.001; PPV 0.32 ± 0.02, p < 0.001). For new lesion segmentation with 30 training studies and their prior comparisons, a fine-tuned model demonstrated similar sensitivity (0.49 ± 0.05) and significantly improved PPV (0.60 ± 0.05) compared to a de-novo model (sensitivity 0.51 ± 0.04, p = 0.437; PPV 0.43 ± 0.04, p = 0.002). For enhancement segmentation with 20 training studies, a fine-tuned model demonstrated significantly improved overall performance (sensitivity 0.74 ± 0.06, PPV 0.69 ± 0.05) compared to a de-novo model (sensitivity 0.44 ± 0.09, p = 0.001; PPV 0.37 ± 0.05, p = 0.001). Conclusion: By fine-tuning models trained for other disease pathologies with MS-specific data, competitive models identifying existing MS plaques, new MS plaques, and active demyelination can be built with substantially smaller datasets than would otherwise be required to train new models.

Optimisation of Storage and Transportation Conditions of Cultured Corneal Endothelial Cells for Cell Replacement Therapy.

As the cornea is one of the most transplanted tissues in the body it has placed a burden on the provision of corneas from cadaveric donors. Corneal endothelial dysfunction is the leading indication for cornea transplant. Therefore, tissue engineering is emerging as an alternative approach to overcome the global shortage of transplant-grade corneas. The propagation and expansion of corneal endothelial cells has been widely reported. However, one obstacle to overcome is the transport and storage of corneal endothelial cells. In this study we investigated whether tissue engineered corneal endothelial cells can be preserved in hypothermic conditions. Human corneal endothelial cells (HCEnCs) were exposed to various temperatures (4 °C, 23 °C, and 37 °C) in both adherent and suspension storage models. Optimal storage media and storage duration was tested along with post-storage viability. Following storage and subsequent recovery at 37 °C, cell phenotype was assessed by immunofluorescence, gene and protein expression, and proliferative capacity analysis. Functionality was also assessed within a rabbit model of bullous keratopathy. Our data support our hypothesis that functional HCEnCs can be preserved in hypothermic conditions.

Characterization of Human Transition Zone Reveals a Putative Progenitor-Enriched Niche of Corneal Endothelium.

: The corneal endothelium regulates corneal hydration to maintain the transparency of cornea. Lacking regenerative capacity, corneal endothelial cell loss due to aging and diseases can lead to corneal edema and vision loss. There is limited information on the existence of corneal endothelial progenitors. We conducted ultrastructural examinations and expression analyses on the human transition zone (TZ) at the posterior limbus of corneal periphery, to elucidate if the TZ harbored progenitor-like cells, and to reveal their niche characteristics. Within the narrow TZ (~190 µm width), the inner TZ-adjacent to the peripheral endothelium (PE)-contained cells expressing stem/progenitor markers (Sox2, Lgr5, CD34, Pitx2, telomerase). They were located on the inner TZ surface and in its underlying stroma. Lgr5 positive cells projected as multicellular clusters into the PE. Under transmission electron microscopy and serial block face-scanning electron microscopy and three-dimensional (3D) reconstruction, the terminal margin of Descemet's membrane was inserted beneath the TZ surface, with the distance akin to the inner TZ breadth. Porcine TZ cells were isolated and proliferated into a confluent monolayer and differentiated to cells expressing corneal endothelial markers (ZO1, Na+K+ATPase) on cell surface. In conclusion, we have identified a novel inner TZ containing progenitor-like cells, which could serve the regenerative potential for corneal endothelium.

Quantification of the Posterior Cornea Using Swept Source Optical Coherence Tomography.

PURPOSE: We define optical coherence tomography (OCT) measurement parameters of the corneal endothelium/Descemet's membrane (DM) complex and peripheral transition zone (TZ) and describe these measurements in an ethnically Chinese population. METHODS: OCT images of the anterior segment and iridocorneal angle were obtained from 129 healthy Chinese subjects (129 eyes), aged 40 to 81 years. The scleral spur (SS) and Schwalbe's line (SL) were identified in each image. Endothelium/DM diameter, referred to as endothelial arc length (EAL), is the SL-to-SL distance. The SS-to-SL distance encompasses the TZ and trabecular meshwork (TM). Since the TZ cannot be visualized by OCT, a ratio of TZ-to-TZ+TM width was calculated from scanning electron microscopy (SEM) images obtained from 5 cadaveric corneas. The SS-to-SL distance was multiplied by this ratio to approximate in vivo TZ width. RESULTS: From SEM measurements, the relationship TZ = 0.20*(TZ+TM) was determined. From OCT measurements, mean EAL was 12.15 ± 0.58 mm and mean TZ width was 156 ± 20 µm. For eyes with horizontal and vertical images, vertical EAL was significantly greater than horizontal EAL (P = 0.03). CONCLUSIONS: Corneal endothelium/DM diameter and TZ width can be obtained from OCT images. Although only combined TZ+TM is visualized on OCT, TZ width can be reasonably approximated. TRANSLATIONAL RELEVANCE: Emerging procedures, like endothelial cell injection and DM transplantation (DMT), require accurate measurements of endothelium/DM size for preoperative planning. Size of the TZ, which may contain progenitor cells, also could contribute to endothelial regeneration in these procedures.

Functional role of peroxiredoxin 6 in the eye.

Peroxiredoxin 6 (Prdx6) is the only mammalian 1-Cys member of the Prdx family, a group of enzymes which share the ability to reduce peroxides. In addition to its peroxidase function, Prdx6 also demonstrates phospholipase A2 and lysophosphatidylcholine acyl transferase (LPCAT) activities. These enzymatic activities play an important role in regenerating oxidized membrane phospholipids and maintaining an appropriate balance of intracellular reactive oxygen species. Development of clinical pathologies, including those within the eye, have been linked to dysregulation of Prdx6 function. Interplay between external stressors like exposure to UV light, transforming growth factor ß (TGF-ß), and hyperglycemia in conjunction with diminished Prdx6 levels and loss of redox balance is associated with cellular changes in a variety of ophthalmic pathologies including cataracts, glaucoma, and retinal degeneration. Many of these cellular abnormalities can be rescued through supplementation with exogenous Prdx6. Additionally, corneal endothelial cells have been found to express high levels of Prdx6 in the plasma membrane. These findings highlight the importance of Prdx6 as an essential regulator of oxidative stress in the eye.

Targeting Stromal Glutamine Synthetase in Tumors Disrupts Tumor Microenvironment-Regulated Cancer Cell Growth.

Reactive stromal cells are an integral part of tumor microenvironment (TME) and interact with cancer cells to regulate their growth. Although targeting stromal cells could be a viable therapy to regulate the communication between TME and cancer cells, identification of stromal targets that make cancer cells vulnerable has remained challenging and elusive. Here, we identify a previously unrecognized mechanism whereby metabolism of reactive stromal cells is reprogrammed through an upregulated glutamine anabolic pathway. This dysfunctional stromal metabolism confers atypical metabolic flexibility and adaptive mechanisms in stromal cells, allowing them to harness carbon and nitrogen from noncanonical sources to synthesize glutamine in nutrient-deprived conditions existing in TME. Using an orthotopic mouse model for ovarian carcinoma, we find that co-targeting glutamine synthetase in stroma and glutaminase in cancer cells reduces tumor weight, nodules, and metastasis. We present a synthetic lethal approach to target tumor stroma and cancer cells simultaneously for desirable therapeutic outcomes.

Metabolic shifts toward glutamine regulate tumor growth, invasion and bioenergetics in ovarian cancer.

Glutamine can play a critical role in cellular growth in multiple cancers. Glutamine-addicted cancer cells are dependent on glutamine for viability, and their metabolism is reprogrammed for glutamine utilization through the tricarboxylic acid (TCA) cycle. Here, we have uncovered a missing link between cancer invasiveness and glutamine dependence. Using isotope tracer and bioenergetic analysis, we found that low-invasive ovarian cancer (OVCA) cells are glutamine independent, whereas high-invasive OVCA cells are markedly glutamine dependent. Consistent with our findings, OVCA patients' microarray data suggest that glutaminolysis correlates with poor survival. Notably, the ratio of gene expression associated with glutamine anabolism versus catabolism has emerged as a novel biomarker for patient prognosis. Significantly, we found that glutamine regulates the activation of STAT3, a mediator of signaling pathways which regulates cancer hallmarks in invasive OVCA cells. Our findings suggest that a combined approach of targeting high-invasive OVCA cells by blocking glutamine's entry into the TCA cycle, along with targeting low-invasive OVCA cells by inhibiting glutamine synthesis and STAT3 may lead to potential therapeutic approaches for treating OVCAs.