Mechanism of Action of Oral Salmonella-Based Vaccine to Prevent and Reverse Type 1 Diabetes in NOD Mice.

A combination therapy of preproinsulin (PPI) and immunomodulators (TGFß+IL10) orally delivered via genetically modified Salmonella and anti-CD3 promoted glucose balance in in NOD mice with recent onset diabetes. The Salmonella bacteria were modified to express the diabetes-associated antigen PPI controlled by a bacterial promoter in conjunction with over-expressed immunomodulating molecules. The possible mechanisms of action of this vaccine to limit autoimmune diabetes remained undefined. In mice, the vaccine prevented and reversed ongoing diabetes. The vaccine-mediated beneficial effects were associated with increased numbers of antigen-specific CD4+CD25+Foxp3+ Tregs, CD4+CD49b+LAG3+ Tr1-cells, and tolerogenic dendritic-cells (tol-DCs) in the spleens and lymphatic organs of treated mice. Despite this, the immune response to Salmonella infection was not altered. Furthermore, the vaccine effects were associated with a reduction in islet-infiltrating lymphocytes and an increase in the islet beta-cell mass. This was associated with increased serum levels of the tolerogenic cytokines (IL10, IL2, and IL13) and chemokine ligand 2 (CCL2) and decreased levels of inflammatory cytokines (IFNγ, GM-CSF, IL6, IL12, and TNFα) and chemokines (CXCL1, CXCL2, and CXCL5). Overall, the data suggest that the Salmonella-based vaccine modulates the immune response, reduces inflammation, and promotes tolerance specifically to an antigen involved in autoimmune diabetes.

A novel approach to determine the critical survival threshold of cellular oxygen within spheroids via integrating live/dead cell imaging with oxygen modeling.

Defining the oxygen level that induces cell death within 3-D tissues is vital for understanding tissue hypoxia; however, obtaining accurate measurements has been technically challenging. In this study, we introduce a noninvasive, high-throughput methodology to quantify critical survival partial oxygen pressure (pO2) with high spatial resolution within spheroids by using a combination of controlled hypoxic conditions, semiautomated live/dead cell imaging, and computational oxygen modeling. The oxygen-permeable, micropyramid patterned culture plates created a precisely controlled oxygen condition around the individual spheroid. Live/dead cell imaging provided the geometric information of the live/dead boundary within spheroids. Finally, computational oxygen modeling calculated the pO2 at the live/dead boundary within spheroids. As proof of concept, we determined the critical survival pO2 in two types of spheroids: isolated primary pancreatic islets and tumor-derived pseudoislets (2.43 ± 0.08 vs. 0.84 ± 0.04 mmHg), indicating higher hypoxia tolerance in pseudoislets due to their tumorigenic origin. We also applied this method for evaluating graft survival in cell transplantations for diabetes therapy, where hypoxia is a critical barrier to successful transplantation outcomes; thus, designing oxygenation strategies is required. Based on the elucidated critical survival pO2, 100% viability could be maintained in a typically sized primary islet under the tissue pO2 above 14.5 mmHg. This work presents a valuable tool that is potentially instrumental for fundamental hypoxia research. It offers insights into physiological responses to hypoxia among different cell types and may refine translational research in cell therapies.NEW & NOTEWORTHY Our study introduces an innovative combinatory approach for noninvasively determining the critical survival oxygen level of cells within small cell spheroids, which replicates a 3-D tissue environment, by seamlessly integrating three pivotal techniques: cell death induction under controlled oxygen conditions, semiautomated imaging that precisely identifies live/dead cells, and computational modeling of oxygen distribution. Notably, our method ensures high-throughput analysis applicable to various cell types, offering a versatile solution for researchers in diverse fields.

Activation of ductal progenitor-like cells from adult human pancreas requires extracellular matrix protein signaling.

Ductal progenitor-like cells are a sub-population of ductal cells in the adult human pancreas that have the potential to contribute to regenerative medicine. However, the microenvironmental cues that regulate their activation are poorly understood. Here, we establish a 3-dimensional suspension culture system containing six defined soluble factors in which primary human ductal progenitor-like and ductal non-progenitor cells survive but do not proliferate. Expansion and polarization occur when suspension cells are provided with a low concentration (5% v/v) of Matrigel, a sarcoma cell product enriched in many extracellular matrix (ECM) proteins. Screening of ECM proteins identified that collagen IV can partially recapitulate the effects of Matrigel. Inhibition of integrin α1ß1, a major collagen IV receptor, negates collagen IV- and Matrigel-stimulated effects. These results demonstrate that collagen IV is a key ECM protein that stimulates the expansion and polarization of human ductal progenitor-like and ductal non-progenitor cells via integrin α1ß1 receptor signaling.

A scalable human islet 3D-culture platform maintains cell mass and function long-term for transplantation.

Present-day islet culture methods provide short-term maintenance of cell viability and function, limiting access to islet transplantation. Attempts to lengthen culture intervals remain unsuccessful. A new method was developed to permit the long-term culture of islets. Human islets were embedded in polysaccharide 3D-hydrogel in cell culture inserts or gas-permeable chambers with serum-free CMRL 1066 supplemented media for up to 8 weeks. The long-term cultured islets maintained better morphology, cell mass, and viability at 4 weeks than islets in conventional suspension culture. In fact, islets cultured in the 3D-hydrogel retained ß cell mass and function on par with freshly isolated islets in vitro and, when transplanted into diabetic mice, restored glucose balance similar to fresh islets. Using gas-permeable chambers, the 3D-hydrogel culture method was scaled up over 10-fold and maintained islet viability and function, although the cell mass recovery rate was 50%. Additional optimization of scale-up methods continues. If successful, this technology could afford flexibility and expand access to islet transplantation, especially single-donor islet-after-kidney transplantation.

One-Step Automatic Radiosynthesis and Evaluation of [18F]TM-30089 as GPR44 Radiotracer.

Recently, a G-protein coupled receptor 44 (GPR44) was discovered to play a significant role in the process of inflammation-related diseases, including cancer and diabetes. However, the precise role of GPR44 has yet to be fully elucidated. Currently, there is a strong and urgent need for the development of GPR44 radiotracers as a non-invasive methodology to explore the exact mechanism of GPR44 on inflammation-related diseases and monitor the progress of therapy. TM-30089 is a potent GPR44 antagonist that exhibits a high specificity and selectivity for GPR44. Its structure contains a fluorine nuclide, which could potentially be replaced with 18F. In the present study, we successfully took a highly effective synthesis strategy that pretreated the unprotected carboxylic acid group of the precursor and developed a feasible one-step automatic radiosynthesis strategy for [18F]TM-30089 with a high radiochemical purity and a good radiochemical yield. We further evaluated this radiotracer using mice models implanted with 1.1 B4 cell lines (GPR44-enriched cell lines) and human islets (high GPR44 expression), respectively. The results revealed the persistent and specific uptake of [18F]TM-30089 in GPR44 region, indicating that [18F]TM-30089 is a promising candidate for targeting GPR44. Further evaluation is ongoing.

Indole-Based and Cyclopentenylindole-Based Analogues Containing Fluorine Group as Potential 18F-Labeled Positron Emission Tomography (PET) G-Protein Coupled Receptor 44 (GPR44) Tracers.

Recently, growing evidence of the relationship between G-protein coupled receptor 44 (GPR44) and the inflammation-cancer system has garnered tremendous interest, while the exact role of GPR44 has not been fully elucidated. Currently, there is a strong and urgent need for the development of non-invasive in vivo GPR44 positron emission tomography (PET) radiotracers that can be used to aid the exploration of the relationship between inflammation and tumor biologic behavior. Accordingly, the choosing and radiolabeling of existing GPR44 antagonists containing a fluorine group could serve as a viable method to accelerate PET tracers development for in vivo imaging to this purpose. The present study aims to evaluate published (2000-present) indole-based and cyclopentenyl-indole-based analogues of the GPR44 antagonist to guide the development of fluorine-18 labeled PET tracers that can accurately detect inflammatory processes. The selected analogues contained a crucial fluorine nuclide and were characterized for various properties including binding affinity, selectivity, and pharmacokinetic and metabolic profile. Overall, 26 compounds with favorable to strong binding properties were identified. This review highlights the potential of GPR44 analogues for the development of PET tracers to study inflammation and cancer development and ultimately guide the development of targeted clinical therapies.

Design, Synthesis, Computational and Biological Evaluation of Novel Structure Fragments Based on Lithocholic Acid (LCA).

The regulation of bile acid pathways has become a particularly promising therapeutic strategy for a variety of metabolic disorders, cancers, and diseases. However, the hydrophobicity of bile acids has been an obstacle to clinical efficacy due to off-target effects from rapid drug absorption. In this report, we explored a novel strategy to design new structure fragments based on lithocholic acid (LCA) with improved hydrophilicity by introducing a polar "oxygen atom" into the side chain of LCA, then (i) either retaining the carboxylic acid group or replacing the carboxylic acid group with (ii) a diol group or (iii) a vinyl group. These novel fragments were evaluated using luciferase-based reporter assays and the MTS assay. Compared to LCA, the result revealed that the two lead compounds 1a-1b were well tolerated in vitro, maintaining similar potency and efficacy to LCA. The MTS assay results indicated that cell viability was not affected by dose dependence (under 25 µM). Additionally, computational model analysis demonstrated that compounds 1a-1b formed more extensive hydrogen bond networks with Takeda G protein-coupled receptor 5 (TGR5) than LCA. This strategy displayed a potential approach to explore the development of novel endogenous bile acids fragments. Further evaluation on the biological activities of the two lead compounds is ongoing.

Ubiquitous Luciferase Expression in "Firefly Rats" Does Not Alter the Pancreatic Islet Morphology, Metabolism, and Function.

"Firefly rats" ubiquitously express the luciferase reporter gene under the control of constitutively active ROSA26 promoter in inbred Lewis rats. Due to the minimal immunogenicity of luciferase, wide applications of Firefly rats have been reported in solid organ/cell transplantation studies for in vivo imaging, permitting quantitative and non-invasive tracking of the transplanted graft. ROSA26 is a non-coding gene and generally does not affect the expression of other endogenous genes. However, the effect of ubiquitous luciferase expression on islet morphology and function has not been thoroughly investigated, which is critical for the use of Firefly rats as islet donors in islet transplantation studies. Accordingly, in vivo glucose homeostasis (i.e., islet function in the native pancreas) was compared between age-matched luciferase-expressing Firefly rats and non-luciferase-expressing rats. In vivo assessments demonstrated no statistical difference between these rats in non-fasting blood glucose levels, intraperitoneal glucose tolerance tests, and glucose-stimulated serum C-peptide levels. Furthermore, islets were isolated from both rats to compare the morphology, function, and metabolism in vitro. Isolated islets from both rats exhibited similar in vitro characteristics in post-isolation islet yield, islet size, beta cell populations, insulin content per islet, oxygen consumption rate, and glucose-stimulated insulin secretion. In conclusion, ubiquitous luciferase expression in Firefly rats does not affect their islet morphology, metabolism, and function; this finding is critical and enables the use of isolated islets from Firefly rats for the dual assessment of islet graft function and bioluminescence imaging of islet grafts.

Changes in the gut microbiota of NOD mice in response to an oral Salmonella-based vaccine against type 1 diabetes.

We developed an oral Salmonella-based vaccine that prevents and reverses diabetes in non-obese diabetic (NOD) mice. Related to this, the gastrointestinal tract harbors a complex dynamic population of microorganisms, the gut microbiome, that influences host homeostasis and metabolism. Changes in the gut microbiome are associated with insulin dysfunction and type 1 diabetes (T1D). Oral administration of diabetic autoantigens as a vaccine can restore immune balance. However, it was not known if a Salmonella-based vaccine would impact the gut microbiome. We administered a Salmonella-based vaccine to prediabetic NOD mice. Changes in the gut microbiota and associated metabolome were assessed using next-generation sequencing and gas chromatography-mass spectrometry (GC-MS). The Salmonella-based vaccine did not cause significant changes in the gut microbiota composition immediately after vaccination although at 30 days post-vaccination changes were seen. Additionally, no changes were noted in the fecal mycobiome between vaccine- and control/vehicle-treated mice. Significant changes in metabolic pathways related to inflammation and proliferation were found after vaccine administration. The results from this study suggest that an oral Salmonella-based vaccine alters the gut microbiome and metabolome towards a more tolerant composition. These results support the use of orally administered Salmonella-based vaccines that induced tolerance after administration.

Integrating genetics with single-cell multiomic measurements across disease states identifies mechanisms of beta cell dysfunction in type 2 diabetes.

Dysfunctional pancreatic islet beta cells are a hallmark of type 2 diabetes (T2D), but a comprehensive understanding of the underlying mechanisms, including gene dysregulation, is lacking. Here we integrate information from measurements of chromatin accessibility, gene expression and function in single beta cells with genetic association data to nominate disease-causal gene regulatory changes in T2D. Using machine learning on chromatin accessibility data from 34 nondiabetic, pre-T2D and T2D donors, we identify two transcriptionally and functionally distinct beta cell subtypes that undergo an abundance shift during T2D progression. Subtype-defining accessible chromatin is enriched for T2D risk variants, suggesting a causal contribution of subtype identity to T2D. Both beta cell subtypes exhibit activation of a stress-response transcriptional program and functional impairment in T2D, which is probably induced by the T2D-associated metabolic environment. Our findings demonstrate the power of multimodal single-cell measurements combined with machine learning for characterizing mechanisms of complex diseases.

Methylcellulose colony assay and single-cell micro-manipulation reveal progenitor-like cells in adult human pancreatic ducts.

Progenitor cells capable of self-renewal and differentiation in the adult human pancreas are an under-explored resource for regenerative medicine. Using micro-manipulation and three-dimensional colony assays we identify cells within the adult human exocrine pancreas that resemble progenitor cells. Exocrine tissues were dissociated into single cells and plated into a colony assay containing methylcellulose and 5% Matrigel. A subpopulation of ductal cells formed colonies containing differentiated ductal, acinar, and endocrine lineage cells, and expanded up to 300-fold with a ROCK inhibitor. When transplanted into diabetic mice, colonies pre-treated with a NOTCH inhibitor gave rise to insulin-expressing cells. Both colonies and primary human ducts contained cells that simultaneously express progenitor transcription factors SOX9, NKX6.1, and PDX1. In addition, in silico analysis identified progenitor-like cells within ductal clusters in a single-cell RNA sequencing dataset. Therefore, progenitor-like cells capable of self-renewal and tri-lineage differentiation either pre-exist in the adult human exocrine pancreas, or readily adapt in culture.

Thrombospondin-1, CD47, and SIRPα display cell-specific molecular signatures in human islets and pancreata.

Thrombospondin-1 (TSP1) is a secreted protein minimally expressed in health but increased in disease and age. TSP1 binds to the cell membrane receptor CD47, which itself engages signal regulatory protein α (SIRPα), and the latter creates a checkpoint for immune activation. Individuals with cancer administered checkpoint-blocking molecules developed insulin-dependent diabetes. Relevant to this, CD47 blocking antibodies and SIRPα fusion proteins are in clinical trials. We characterized the molecular signature of TSP1, CD47, and SIRPα in human islets and pancreata. Fresh islets and pancreatic tissue from nondiabetic individuals were obtained. The expression of THBS1, CD47, and SIRPA was determined using single-cell mRNA sequencing, immunofluorescence microscopy, Western blot, and flow cytometry. Islets were exposed to diabetes-affiliated inflammatory cytokines and changes in protein expression were determined. CD47 mRNA was expressed in all islet cell types. THBS1 mRNA was restricted primarily to endothelial and mesenchymal cells, whereas SIRPA mRNA was found mostly in macrophages. Immunofluorescence staining showed CD47 protein expressed by ß cells and present in the exocrine pancreas. TSP1 and SIRPα proteins were not seen in islets or the exocrine pancreas. Western blot and flow cytometry confirmed immunofluorescent expression patterns. Importantly, human islets produced substantial quantities of secreted TSP1. Human pancreatic exocrine and endocrine tissue expressed CD47, whereas fresh islets displayed cell surface CD47 and secreted TSP1 at baseline and in inflammation. These findings suggest unexpected effects on islets from agents that intersect TSP1-CD47-SIRPα.NEW & NOTEWORTHY CD47 is a cell surface receptor with two primary ligands, soluble thrombospondin-1 (TSP1) and cell surface signal regulatory protein alpha (SIRPα). Both interactions provide checkpoints for immune cell activity. We determined that fresh human islets display CD47 and secrete TSP1. However, human islet endocrine cells lack SIRPα. These gene signatures are likely important given the increasing use of CD47 and SIRPα blocking molecules in individuals with cancer.

Integration of single-cell multiomic measurements across disease states with genetics identifies mechanisms of beta cell dysfunction in type 2 diabetes.

Altered function and gene regulation of pancreatic islet beta cells is a hallmark of type 2 diabetes (T2D), but a comprehensive understanding of mechanisms driving T2D is still missing. Here we integrate information from measurements of chromatin activity, gene expression and function in single beta cells with genetic association data to identify disease-causal gene regulatory changes in T2D. Using machine learning on chromatin accessibility data from 34 non-diabetic, pre-T2D and T2D donors, we robustly identify two transcriptionally and functionally distinct beta cell subtypes that undergo an abundance shift in T2D. Subtype-defining active chromatin is enriched for T2D risk variants, suggesting a causal contribution of subtype identity to T2D. Both subtypes exhibit activation of a stress-response transcriptional program and functional impairment in T2D, which is likely induced by the T2D-associated metabolic environment. Our findings demonstrate the power of multimodal single-cell measurements combined with machine learning for identifying mechanisms of complex diseases.

Factors associated with favourable 5 year outcomes in islet transplant alone recipients with type 1 diabetes complicated by severe hypoglycaemia in the Collaborative Islet Transplant Registry.

AIMS/HYPOTHESIS: Islet transplantation has been studied in small cohorts of recipients with type 1 diabetes complicated by severe hypoglycaemic events (SHEs). We determined factors associated with favourable outcomes in a large cohort of recipients reported to the Collaborative Islet Transplant Registry (CITR). METHODS: In 398 non-uraemic islet transplant alone (ITA) recipients with type 1 diabetes and SHEs, transplanted between 1999 and 2015 and with at least 1 year follow-up, we analysed specified favourable outcomes against each of all available characteristics of pancreas donors, islet grafts, recipients and immunosuppressive regimens, as well as immunosuppression and procedure-related serious adverse events (SAEs). RESULTS: Four factors were associated with the highest rates of favourable outcomes: recipient age ≥35 years; total infused islets ≥325,000 islet equivalents; induction immunosuppression with T cell depletion and/or TNF-α inhibition; and maintenance with both mechanistic target of rapamycin (mTOR) and calcineurin inhibitors. At 5 years after the last islet infusion, of the recipients meeting these four common favourable factors (4CFF; N=126), 95% were free of SHEs, 76% had HbA1c <53 mmol/mol (7.0%), 73% had HbA1c <53 mmol/mol (7.0%) and absence of SHEs, and 53% were insulin independent, significantly higher rates than in the remaining recipients (<4CFF; N=272). The incidence of procedural and immunosuppression-related SAEs per recipient that resulted in sequelae, disability or death was low in both the 4CFF (0.056 per person) and <4CFF (0.074 per person) groups. CONCLUSIONS/INTERPRETATION: In recipients with type 1 diabetes complicated by SHEs, islet transplantation meeting 4CFF protected 95% from SHEs at 5 years after the last islet infusion and exerted a large and significant benefit on glycaemic control, with an acceptable safety profile for this subgroup of type 1 diabetes.

Heart Uptake of [18F]Fluoro-4-Thia-Oleate in a Non-Alcoholic Fatty Liver Disease Mouse Model.

The world-wide high incidence of non-alcoholic fatty liver disease (NAFLD) is of concern for its progression to insulin resistance, steatohepatitis and cardiovascular disease (CVD). The increased uptake of fatty acids in critical organs plays a major role in NAFLD progression. Male Ceacam1−/− mice that develop NAFLD, insulin resistance and CVD on normal chow are a potential model for studying the dysregulation of fatty acid uptake. [18F]fluoro-4-thia-oleate ([18F]FTO) was chosen as a fatty acid reporter because of its higher uptake and retention in the heart in an animal model of CVD. Male wild-type (WT) or Ceacam1−/− mice fasted 4−6 h were administered [18F]FTO i.v., and dynamic PET scans were conducted in an MR/PET small animal imaging system along with terminal tissue biodistributions. Quantitative heart image analysis revealed significantly higher uptake at 35 min in Ceacam1−/− (6.0 ± 1.0% ID/cc) vs. WT (3.9 ± 0.6% ID/cc) mice (p = 0.006). Ex vivo heart uptake/retention (% ID/organ) was 2.82 ± 0.45 for Ceacam1−/− mice vs. 1.66 ± 0.45 for WT mice (p < 0.01). Higher kidney and pancreas uptake/retention in Ceacam1−/− was also evident, and the excretion of [18F]FTO into the duodenum was observed for both WT and Ceacam1−/− mice starting at 10 min. This study suggests that the administration of [18F]FTO as a marker of fatty acid uptake and retention may be an important tool in analyzing the effect of NAFLD on lipid dysregulation in the heart.

Microwell culture platform maintains viability and mass of human pancreatic islets.

Background: Transplantation of the human pancreatic islets is a promising approach for specific types of diabetes to improve glycemic control. Although effective, there are several issues that limit the clinical expansion of this treatment, including difficulty in maintaining the quality and quantity of isolated human islets prior to transplantation. During the culture, we frequently observe the multiple islets fusing together into large constructs, in which hypoxia-induced cell damage significantly reduces their viability and mass. In this study, we introduce the microwell platform optimized for the human islets to prevent unsolicited fusion, thus maintaining their viability and mass in long-term cultures. Method: Human islets are heterogeneous in size; therefore, two different-sized microwells were prepared in a 35 mm-dish format: 140 µm × 300 µm-microwells for <160 µm-islets and 200 µm × 370 µm-microwells for >160 µm-islets. Human islets (2,000 islet equivalent) were filtered through a 160 µm-mesh to prepare two size categories for subsequent two week-cultures in each microwell dish. Conventional flat-bottomed 35 mm-dishes were used for non-filtered islets (2,000 islet equivalent/2 dishes). Post-cultured islets are collected to combine in each condition (microwells and flat) for the comparisons in viability, islet mass, morphology, function and metabolism. Islets from three donors were independently tested. Results: The microwell platform prevented islet fusion during culture compared to conventional flat bottom dishes, which improved human islet viability and mass. Islet viability and mass on the microwells were well-maintained and comparable to those in pre-culture, while flat bottom dishes significantly reduced islet viability and mass in two weeks. Morphology assessed by histology, insulin-secreting function and metabolism by oxygen consumption did not exhibit the statistical significance among the three different conditions. Conclusion: Microwell-bottomed dishes maintained viability and mass of human islets for two weeks, which is significantly improved when compared to the conventional flat-bottomed dishes.

Micropyramid-patterned, oxygen-permeable bottomed dish for high density culture of pancreatic islets.

The need for maintaining cell-spheroid viability and function within high-density cultures is unmet for various clinical and experimental applications, including cell therapies. One immediate application is for transplantation of pancreatic islets, a clinically recognized treatment option to cure type 1 diabetes; islets are isolated from a donor for subsequent culture prior to transplantation. However, high seeding conditions cause unsolicited fusion of multiple spheroids, thereby limiting oxygen diffusion to induce hypoxic cell death. Here we introduce a culture dish incorporating a micropyramid-patterned surface to prevent the unsolicited fusion and oxygen-permeable bottom for optimal oxygen environment. A 400µm-thick, oxygen-permeable polydimethylsiloxane sheet topped with micropyramid pattern of 400µm-base and 200µm-height was fabricated to apply to the 24-well plate format. The micropyramid pattern separated the individual pancreatic islets to prevent the fusion of multiple islets. This platform supported the high oxygen demand of islets at high seeding density at 260 islet equivalents cm-2, a 2-3-fold higher seeding density compared to the conventional islet culture used in a preparation for the clinical islet transplantations, demonstrating improved islet morphology, metabolism and function in a 4 d-culture. Transplantation of these islets into immunodeficient diabetic mice exhibited significantly improved engraftment to achieve euglycemia compared to islets cultured in the conventional culture wells. Collectively, this simple design modification allows for high-density cultures of three-dimensional cell spheroids to improve the viability and function for an array of investigational and clinical replacement tissues.

Thyroid Carcinoma, Version 2.2022, NCCN Clinical Practice Guidelines in Oncology.

Differentiated thyroid carcinomas is associated with an excellent prognosis. The treatment of choice for differentiated thyroid carcinoma is surgery, followed by radioactive iodine ablation (iodine-131) in select patients and thyroxine therapy in most patients. Surgery is also the main treatment for medullary thyroid carcinoma, and kinase inhibitors may be appropriate for select patients with recurrent or persistent disease that is not resectable. Anaplastic thyroid carcinoma is almost uniformly lethal, and iodine-131 imaging and radioactive iodine cannot be used. When systemic therapy is indicated, targeted therapy options are preferred. This article describes NCCN recommendations regarding management of medullary thyroid carcinoma and anaplastic thyroid carcinoma, and surgical management of differentiated thyroid carcinoma (papillary, follicular, Hürthle cell carcinoma).

Biodistribution of Intra-Arterial and Intravenous Delivery of Human Umbilical Cord Mesenchymal Stem Cell-Derived Extracellular Vesicles in a Rat Model to Guide Delivery Strategies for Diabetes Therapies.

Umbilical cord mesenchymal stem cell-derived extracellular vesicles (UC-MSC-EVs) have become an emerging strategy for treating various autoimmune and metabolic disorders, particularly diabetes. Delivery of UC-MSC-EVs is essential to ensure optimal efficacy of UC-MSC-EVs. To develop safe and superior EVs-based delivery strategies, we explored nuclear techniques including positron emission tomography (PET) to evaluate the delivery of UC-MSC-EVs in vivo. In this study, human UC-MSC-EVs were first successfully tagged with I-124 to permit PET determination. Intravenous (I.V.) and intra-arterial (I.A.) administration routes of [124I]I-UC-MSC-EVs were compared and evaluated by in vivo PET-CT imaging and ex vivo biodistribution in a non-diabetic Lewis (LEW) rat model. For I.A. administration, [124I]I-UC-MSC-EVs were directly infused into the pancreatic parenchyma via the celiac artery. PET imaging revealed that the predominant uptake occurred in the liver for both injection routes, and further imaging characterized clearance patterns of [124I]I-UC-MSC-EVs. For biodistribution, the uptake (%ID/gram) in the spleen was significantly higher for I.V. administration compared to I.A. administration (1.95 ± 0.03 and 0.43 ± 0.07, respectively). Importantly, the pancreas displayed similar uptake levels between the two modalities (0.20 ± 0.06 for I.V. and 0.24 ± 0.03 for I.A.). Therefore, our initial data revealed that both routes had similar delivery efficiency for [124I]I-UC-MSC-EVs except in the spleen and liver, considering that higher spleen uptake could enhance immunomodulatory application of UC-MSC-EVs. These findings could guide the development of safe and efficacious delivery strategies for UC-MSC-EVs in diabetes therapies, in which a minimally invasive I.V. approach would serve as a better delivery strategy. Further confirmation studies are ongoing.

Critical Considerations in Bioluminescence Imaging of Transplanted Islets: Dynamic Signal Change in Early Posttransplant Phase and Signal Absorption by Tissues.

OBJECTIVES: In pancreatic islet transplantation studies, bioluminescence imaging enables quantitative and noninvasive tracking of graft survival. Amid the recent heightened interest in extrahepatic sites for islet and stem cell-derived beta-like cell transplantations, proper understanding the nature of bioluminescence imaging in these sites is important. METHODS: Islets isolated from Firefly rats ubiquitously expressing luciferase reporter gene in Lewis rats were transplanted into subcutaneous or kidney capsule sites of wild-type Lewis rats or immunodeficient mice. Posttransplant changes of bioluminescence signal curves and absorption of bioluminescence signal in transplantation sites were examined. RESULTS: The bioluminescence signal curve dynamically changed in the early posttransplantation phase; the signal was low within the first 5 days after transplantation. A substantial amount of bioluminescence signal was absorbed by tissues surrounding islet grafts, correlating to the depth of the transplanted site from the skin surface. Grafts in kidney capsules were harder to image than those in the subcutaneous site. Within the kidney capsule, locations that minimized depth from the skin surface improved the graft detectability. CONCLUSIONS: Posttransplant phase and graft location/depth critically impact the bioluminescence images captured in islet transplantation studies. Understanding these parameters is critical for reducing experimental biases and proper interpretation of data.