Toxicity Impacts on Human Adipose Mesenchymal Stem/Stromal Cells Acutely Exposed to Aroclor and Non-Aroclor Mixtures of Polychlorinated Biphenyl.

Polychlorinated biphenyl (PCB) accumulates in adipose where it may impact the growth and function of cells within the tissue. This is particularly concerning during adolescence when adipocytes expand rapidly. Herein, we sought to understand how exposure to PCB mixtures found in U.S. schools affects human adipose mesenchymal stem/stromal cell (MSC) health and function. We investigated how exposure to Aroclor 1016 and Aroclor 1254, as well as a newly characterized non-Aroclor mixture that resembles the PCB profile found in cabinets, Cabinet Mixture, affects adipose MSC growth, viability, and function in vitro. We found that exposure to all three mixtures resulted in two distinct types of toxicity. At PCB concentrations >20 µM, the majority of MSCs die, while at 1-10 µM, MSCs remained viable but display numerous alterations to their phenotype. At these sublethal concentrations, the MSC rate of expansion slowed and morphology changed. Further assessment revealed that PCB-exposed MSCs had impaired adipogenesis and a modest decrease in immunosuppressive capabilities. Thus, exposure to PCB mixtures found in schools negatively impacts the health and function of adipose MSCs. This work has implications for human health due to MSCs' role in supporting the growth and maintenance of adipose tissue.

IFN-γ and TNF-α Pre-licensing Protects Mesenchymal Stromal Cells from the Pro-inflammatory Effects of Palmitate.

The use of mesenchymal stromal cell (MSC) therapy for the treatment of type 2 diabetes (T2D) and T2D complications is promising; however, the investigation of MSC function in the setting of T2D has not been thoroughly explored. In our current study, we investigated the phenotype and function of MSCs in a simulated in vitro T2D environment. We show that palmitate, but not glucose, exposure impairs MSC metabolic activity with moderate increases in apoptosis, while drastically affecting proliferation and morphology. In co-culture with peripheral blood mononuclear cells (PBMCs), we found that MSCs not only lose their normal suppressive ability in high levels of palmitate, but actively support and enhance inflammation, resulting in elevated PBMC proliferation and pro-inflammatory cytokine release. The pro-inflammatory effect of MSCs in palmitate was partially reversed via palmitate removal and fully reversed through pre-licensing MSCs with interferon-gamma and tumor necrosis factor alpha. Thus, palmitate, a specific metabolic factor enriched within the T2D environment, is a potent modulator of MSC immunosuppressive function, which may in part explain the depressed potency observed in MSCs isolated from T2D patients. Importantly, we have also identified a robust and durable pre-licensing regimen that protects MSC immunosuppressive function in the setting of T2D.

Function of Cryopreserved Mesenchymal Stromal Cells With and Without Interferon-γ Prelicensing is Context Dependent.

Tailoring MSCs to fit the disease. Fresh, cryopreserved and, prelicensed cryopreserved MSC are all being explored to treat numerous diseases, but all are not suitable to treat all conditions. injury. "*" denotes preferred therapeutic strategy when both fresh MSC and cryo-MSC have shown utility in treating the disease but one is more efficacious or logistically suitable. ABBREVIATIONS: CLI, critical limb ischemia; GvHD. graft versus host disease; I/R, ischemia reperfusion (I/R); OI, osteogenesis imperfecta.

Microstructured barbs on the North American porcupine quill enable easy tissue penetration and difficult removal.

North American porcupines are well known for their specialized hairs, or quills that feature microscopic backward-facing deployable barbs that are used in self-defense. Herein we show that the natural quill's geometry enables easy penetration and high tissue adhesion where the barbs specifically contribute to adhesion and unexpectedly, dramatically reduce the force required to penetrate tissue. Reduced penetration force is achieved by topography that appears to create stress concentrations along regions of the quill where the cross sectional diameter grows rapidly, facilitating cutting of the tissue. Barbs located near the first geometrical transition zone exhibit the most substantial impact on minimizing the force required for penetration. Barbs at the tip of the quill independently exhibit the greatest impact on tissue adhesion force and the cooperation between barbs in the 0-2 mm and 2-4 mm regions appears critical to enhance tissue adhesion force. The dual functions of barbs were reproduced with replica molded synthetic polyurethane quills. These findings should serve as the basis for the development of bio-inspired devices such as tissue adhesives or needles, trocars, and vascular tunnelers where minimizing the penetration force is important to prevent collateral damage.

Rolling circle amplification: a versatile tool for chemical biology, materials science and medicine.

Rolling circle amplification (RCA) is an isothermal enzymatic process where a short DNA or RNA primer is amplified to form a long single stranded DNA or RNA using a circular DNA template and special DNA or RNA polymerases. The RCA product is a concatemer containing tens to hundreds of tandem repeats that are complementary to the circular template. The power, simplicity, and versatility of the DNA amplification technique have made it an attractive tool for biomedical research and nanobiotechnology. Traditionally, RCA has been used to develop sensitive diagnostic methods for a variety of targets including nucleic acids (DNA, RNA), small molecules, proteins, and cells. RCA has also attracted significant attention in the field of nanotechnology and nanobiotechnology. The RCA-produced long, single-stranded DNA with repeating units has been used as template for the periodic assembly of nanospecies. Moreover, since RCA products can be tailor-designed by manipulating the circular template, RCA has been employed to generate complex DNA nanostructures such as DNA origami, nanotubes, nanoribbons and DNA based metamaterials. These functional RCA based nanotechnologies have been utilized for biodetection, drug delivery, designing bioelectronic circuits and bioseparation. In this review, we introduce the fundamental engineering principles used to design RCA nanotechnologies, discuss recently developed RCA-based diagnostics and bioanalytical tools, and summarize the use of RCA to construct multivalent molecular scaffolds and nanostructures for applications in biology, diagnostics and therapeutics.

The Impact of High Adiposity on Endometrial Progesterone Response and Metallothionein Regulation.

CONTEXT: Obesity is a disease with deleterious effects on the female reproductive tract, including the endometrium. OBJECTIVE: We sought to understand the effects of excess adipose on the benign endometrium. DESIGN: A physiologic in vitro coculture system was developed, consisting of multicellular human endometrial organoids, adipose spheroids, and menstrual cycle hormones. Native human endometrial tissue samples women with and without obesity were also analyzed. SETTING: Academic institution. PATIENTS: Benign endometrial tissues from premenopausal women were obtained following written consent. MAIN OUTCOME MEASURES: Gene expression, protein expression, chromatin binding, and expression of DNA damage and oxidative damage markers were measured. RESULTS: Under high-adiposity conditions, endometrial organoids downregulated endometrial secretory phase genes, suggestive of an altered progesterone response. Progesterone specifically upregulated the metallothionein (MT) gene family in the epithelial cells of endometrial organoids, while high adiposity significantly downregulated the MT genes. Silencing MT genes in endometrial epithelial cells resulted in increased DNA damage, illustrating the protective role of MTs. Native endometrium from women with obesity displayed increased MT expression and oxidative damage in the stroma and not in the epithelium, indicating the cell-specific impact of obesity on MT genes. CONCLUSIONS: Taken together, the in vitro and in vivo systems used here revealed that high adiposity or obesity can alter MT expression by decreasing progesterone response in the epithelial cells and increasing oxidative stress in the stroma.

Mesenchymal stem cells transmigrate between and directly through tumor necrosis factor-α-activated endothelial cells via both leukocyte-like and novel mechanisms.

Systemically administered adult mesenchymal stem cells (MSCs), which are being explored in clinical trials to treat inflammatory disease, exhibit the critical ability to extravasate at sites of inflammation. We aimed to characterize the basic cellular processes mediating this extravasation and compare them to those involved in leukocyte transmigration. Using high-resolution confocal and dynamic microscopy, we show that, like leukocytes, human bone marrow-derived MSC preferentially adhere to and migrate across tumor necrosis factor-α-activated endothelium in a vascular cell adhesion molecule-1 (VCAM-1) and G-protein-coupled receptor signaling-dependent manner. As several studies have suggested, we observed that a fraction of MSC was integrated into endothelium. In addition, we observed two modes of transmigration not previously observed for MSC: Paracellular (between endothelial cells) and transcellular (directly through individual endothelial cells) diapedesis through discrete gaps and pores in the endothelial monolayer, in association with VCAM-1-enriched "transmigratory cups". Contrasting leukocytes, MSC transmigration was not preceded by significant lateral migration and occurred on the time scale of hours rather than minutes. Interestingly, rather than lamellipodia and invadosomes, MSC exhibited nonapoptotic membrane blebbing activity that was similar to activities previously described for metastatic tumor and embryonic germ cells. Our studies suggest that low avidity binding between endothelium and MSC may grant a permissive environment for MSC blebbing. MSC blebbing was associated with early stages of transmigration, in which blebs could exert forces on underlying endothelial cells indicating potential functioning in breaching the endothelium. Collectively, our data suggest that MSC transmigrate actively into inflamed tissues via both leukocyte-like and novel mechanisms.

Tracking mesenchymal stem cells with iron oxide nanoparticle loaded poly(lactide-co-glycolide) microparticles.

Monitoring the location, distribution and long-term engraftment of administered cells is critical for demonstrating the success of a cell therapy. Among available imaging-based cell tracking tools, magnetic resonance imaging (MRI) is advantageous due to its noninvasiveness, deep penetration, and high spatial resolution. While tracking cells in preclinical models via internalized MRI contrast agents (iron oxide nanoparticles, IO-NPs) is a widely used method, IO-NPs suffer from low iron content per particle, low uptake in nonphagocytotic cell types (e.g., mesenchymal stem cells, MSCs), weak negative contrast, and decreased MRI signal due to cell proliferation and cellular exocytosis. Herein, we demonstrate that internalization of IO-NP (10 nm) loaded biodegradable poly(lactide-co-glycolide) microparticles (IO/PLGA-MPs, 0.4-3 µm) in MSCs enhances MR parameters such as the r(2) relaxivity (5-fold), residence time inside the cells (3-fold) and R(2) signal (2-fold) compared to IO-NPs alone. Intriguingly, in vitro and in vivo experiments demonstrate that internalization of IO/PLGA-MPs in MSCs does not compromise inherent cell properties such as viability, proliferation, migration and their ability to home to sites of inflammation.

Hydroxylation markedly alters how the polychlorinated biphenyl (PCB) congener, PCB52, affects gene expression in human preadipocytes.

Polychlorinated biphenyls (PCBs) accumulate in adipose tissue and are linked to obesity and diabetes. The congener, PCB52 (2,2',5,5'-tetrachorobiphenyl), is found at high levels in school air. Hydroxylation of PCB52 to 4-OH-PCB52 (4-hydroxy-2,2',5,5'-tetrachorobiphenyl) may increase its toxicity. To understand PCB52's role in causing adipose dysfunction, we exposed human preadipocytes to PCB52 or 4-OH-PCB52 across a time course and assessed transcript changes using RNAseq. 4-OH-PCB52 caused considerably more changes in the number of differentially expressed genes as compared to PCB52. Both PCB52 and 4-OH-PCB52 upregulated transcript levels of the sulfotransferase SULT1E1 at early time points, but cytochrome P450 genes were generally not affected. A set of genes known to be transcriptionally regulated by PPARα were consistently downregulated by PCB52 at all time points. In contrast, 4-OH-PCB52 affected a variety of pathways, including those involving cytokine responses, hormone responses, focal adhesion, Hippo, and Wnt signaling. Sets of genes known to be transcriptionally regulated by IL17A or parathyroid hormone (PTH) were found to be consistently downregulated by 4-OH-PCB52. Most of the genes affected by PCB52 and 4-OH-PCB52 were different and, of those that were the same, many were changed in an opposite direction. These studies provide insight into how PCB52 or its metabolites may cause adipose dysfunction to cause disease.

Updating "Dataset of transcriptomic changes that occur in human preadipocytes over a 3-day course of exposure to 3,3',4,4',5-Pentachlorobiphenyl (PCB126)" with additional data on exposure to 2,2',5,5'-tetrachlorobiphenyl (PCB52) or its 4-hydroxy metabolite (4-OH-PCB52).

Polychlorinated biphenyls (PCBs) were used extensively in building materials, including those used in schools. PCBs accumulate in fat, and exposure to PCBs is associated with the development of cancer, neurodevelopmental disorders, cardiovascular disease, obesity, and diabetes. The non-dioxin-like PCB congener, PCB52 (2,2',5,5'-tetrachlorobiphenyl), is found at one of the highest levels of any congener in school air. PCB52 is oxidized in the liver to hydroxylated forms, mainly 4-OH-PCB52 (2,2',5,5'-tetrachlorobiphenyl-4-ol). In a previous study, we reported on RNAseq data generated from exposure of human preadipocytes to the dioxin-like PCB congener, PCB126. In this new dataset, we used identical techniques to examine alterations in gene transcript levels in human preadipocytes exposed to PCB52 or 4-OH-PCB52 over a time course. This updated set of data provides a comprehensive transcriptional profile of changes that occur in preadipocytes exposed to PCB52 or 4-OH-PCB52 over time and allows for comparison of these changes between the parent compound and its hydroxy metabolite. The datasets will allow others to explore how PCB52 and 4-OH-PCB52 impact biological pathways in preadipocytes. Further studies can be performed to determine how these changes might lead to disease.

Native adiponectin plays a role in the adipocyte-mediated conversion of fibroblasts to myofibroblasts.

Adipocytes regulate tissues through production of adipokines that can act both locally and systemically. Adipocytes also have been found to play a critical role in regulating the healing process. To better understand this role, we developed a three-dimensional human adipocyte spheroid system that has an adipokine profile similar to in vivo adipose tissues. Previously, we found that conditioned medium from these spheroids induces human dermal fibroblast conversion into highly contractile, collagen-producing myofibroblasts through a transforming growth factor beta-1 (TGF-ß1) independent pathway. Here, we sought to identify how mature adipocytes signal to dermal fibroblasts through adipokines to induce myofibroblast conversion. By using molecular weight fractionation, heat inactivation and lipid depletion, we determined mature adipocytes secrete a factor that is 30-100 kDa, heat labile and lipid associated that induces myofibroblast conversion. We also show that the depletion of the adipokine adiponectin, which fits those physico-chemical parameters, eliminates the ability of adipocyte-conditioned media to induce fibroblast to myofibroblast conversion. Interestingly, native adiponectin secreted by cultured adipocytes consistently elicited a stronger level of α-smooth muscle actin expression than exogenously added adiponectin. Thus, adiponectin secreted by mature adipocytes induces fibroblast to myofibroblast conversion and may lead to a phenotype of myofibroblasts distinct from TGF-ß1-induced myofibroblasts.

Use of a polymeric implant system to assess the neurotoxicity of subacute exposure to 2,2',5,5'-tetrachlorobiphenyl-4-ol, a human metabolite of PCB 52, in male adolescent rats.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants (POPs) that ubiquitously exist in the environment. PCB exposure has been linked to cancer and multi-system toxicity, including endocrine disruption, immune inhibition, and reproductive and neurotoxicity. 2,2',5,5'-Tetrachlorobiphenyl (PCB 52) is one of the most frequently detected congeners in the environment and human blood. The hydroxylated metabolites of PCB 52 may also be neurotoxic, especially for children whose brains are still developing. However, it is challenging to discern the contribution of these metabolites to PCB neurotoxicity because the metabolism of PCB is species-dependent. In this study, we evaluated the subacute neurotoxicity of a human-relevant metabolite, 2,2',5,5'-tetrachlorobiphenyl-4-ol (4-52), on male adolescent Sprague Dawley rats, via a novel polymeric implant drug delivery system grafted subcutaneously, at total loading concentrations ranging from 0%, 1%, 5%, and 10% of the implant (w/w) for 28 days. Y-maze, hole board test, open field test, and elevated plus maze were performed on exposure days 24-28 to assess their locomotor activity, and exploratory and anxiety-like behavior. 4-52 and other possible hydroxylated metabolites in serum and vital tissues were quantified using gas chromatography with tandem mass spectrometry (GC-MS/MS). Our results demonstrate the sustained release of 4-52 from the polymeric implants into the systemic circulation in serum and tissues. Dihydroxylated and dechlorinated metabolites were detected in serum and tissues, depending on the dose and tissue type. No statistically significant changes were observed in the neurobehavioral tasks across all exposure groups. The results demonstrate that subcutaneous polymeric implants provide a straightforward method to expose rats to phenolic PCB metabolites to study neurotoxic outcomes, e.g., in memory, anxiety, and exploratory behaviors.

Efferocytosis of viable versus heat-inactivated MSC induces human monocytes to distinct immunosuppressive phenotypes.

BACKGROUND: Immunomodulation by mesenchymal stromal cells (MSCs) can occur through trophic factor mechanisms, however, intravenously infused MSCs are rapidly cleared from the body yet a potent immunotherapeutic response is still observed. Recent work suggests that monocytes contribute to the clearance of MSCs via efferocytosis, the body's natural mechanism for clearing dead and dying cells in a non-inflammatory manner. This begs the questions of how variations in MSC quality affect monocyte phenotype and if viable MSCs are even needed to elicit an immunosuppressive response. METHODS: Herein, we sought to dissect MSC's trophic mechanism from their efferocytic mechanisms and determine if the viability of MSCs prior to efferocytosis influences the resultant phenotype of monocytes. We cultured viable or heat-inactivated human umbilical cord MSCs with human peripheral blood mononuclear cells for 24 h and observed changes in monocyte surface marker expression and secretion profile. To isolate the effect of efferocytosis from MSC trophic factors, we used cell separation techniques to remove non-efferocytosed MSCs before challenging monocytes to suppress T-cells or respond to inflammatory stimuli. For all experiments, viable and heat-inactivated efferocytic-licensing of monocytes were compared to non-efferocytic-licensing control. RESULTS: We found that monocytes efferocytose viable and heat-inactivated MSCs equally, but only viable MSC-licensed monocytes suppress activated T-cells and suppression occurred even after depletion of residual MSCs. This provides direct evidence that monocytes that efferocytose viable MSCs are immunosuppressive. Further characterization of monocytes after efferocytosis showed that uptake of viable-but not heat inactivated-MSC resulted in monocytes secreting IL-10 and producing kynurenine. When monocytes were challenged with LPS, IL-2, and IFN-γ to simulate sepsis, monocytes that had efferocytosed viable MSC had higher levels of IDO while monocytes that efferocytosed heat inactivated-MSCs produced the lowest levels of TNF-α. CONCLUSION: Collectively, these studies show that the quality of MSCs efferocytosed by monocytes polarize monocytes toward distinctive immunosuppressive phenotypes and highlights the need to tailor MSC therapies for specific indications.

Utilization of commercial collagens for preparing well-differentiated human beta cells for confocal microscopy.

Introduction: With technical advances, confocal and super-resolution microscopy have become powerful tools to dissect cellular pathophysiology. Cell attachment to glass surfaces compatible with advanced imaging is critical prerequisite but remains a considerable challenge for human beta cells. Recently, Phelps et al. reported that human beta cells plated on type IV collagen (Col IV) and cultured in neuronal medium preserve beta cell characteristics. Methods: We examined human islet cells plated on two commercial sources of Col IV (C6745 and C5533) and type V collagen (Col V) for differences in cell morphology by confocal microscopy and secretory function by glucose-stimulated insulin secretion (GSIS). Collagens were authenticated by mass spectrometry and fluorescent collagen-binding adhesion protein CNA35. Results: All three preparations allowed attachment of beta cells with high nuclear localization of NKX6.1, indicating a well-differentiated status. All collagen preparations supported robust GSIS. However, the morphology of islet cells differed between the 3 preparations. C5533 showed preferable features as an imaging platform with the greatest cell spread and limited stacking of cells followed by Col V and C6745. A significant difference in attachment behavior of C6745 was attributed to the low collagen contents of this preparation indicating importance of authentication of coating material. Human islet cells plated on C5533 showed dynamic changes in mitochondria and lipid droplets (LDs) in response to an uncoupling agent 2-[2-[4-(trifluoromethoxy)phenyl]hydrazinylidene]-propanedinitrile (FCCP) or high glucose + oleic acid. Discussion: An authenticated preparation of Col IV provides a simple platform to apply advanced imaging for studies of human islet cell function and morphology.

Improved MSC Minimal Criteria to Maximize Patient Safety: A Call to Embrace Tissue Factor and Hemocompatibility Assessment of MSC Products.

The number of mesenchymal stromal/stem cell (MSC) therapeutics and types of clinical applications have greatly diversified during the past decade, including rapid growth of poorly regulated "Stem Cell Clinics" offering diverse "Unproven Stem Cell Interventions." This product diversification necessitates a critical evaluation of the reliance on the 2006 MSC minimal criteria to not only define MSC identity but characterize MSC suitability for intravascular administration. While high-quality MSC therapeutics have been safely administered intravascularly in well-controlled clinical trials, repeated case reports of mild-to-more-severe adverse events have been reported. These are most commonly related to thromboembolic complications upon infusion of highly procoagulant tissue factor (TF/CD142)-expressing MSC products. As TF/CD142 expression varies widely depending on the source and manufacturing process of the MSC product, additional clinical cell product characterization and guidelines are needed to ensure the safe use of MSC products. To minimize risk to patients receiving MSC therapy, we here propose to supplement the minimal criteria used for characterization of MSCs, to include criteria that assess the suitability of MSC products for intravascular use. If cell products are intended for intravascular delivery, which is true for half of all clinical applications involving MSCs, the effects of MSC on coagulation and hemocompatibility should be assessed and expression of TF/CD142 should be included as a phenotypic safety marker. This adjunct criterion will ensure both the identity of the MSCs as well as the safety of the MSCs has been vetted prior to intravascular delivery of MSC products.

Translating MSC Therapy in the Age of Obesity.

Mesenchymal stromal cell (MSC) therapy has seen increased attention as a possible option to treat a number of inflammatory conditions including COVID-19 acute respiratory distress syndrome (ARDS). As rates of obesity and metabolic disease continue to rise worldwide, increasing proportions of patients treated with MSC therapy will be living with obesity. The obese environment poses critical challenges for immunomodulatory therapies that should be accounted for during development and testing of MSCs. In this review, we look to cancer immunotherapy as a model for the challenges MSCs may face in obese environments. We then outline current evidence that obesity alters MSC immunomodulatory function, drastically modifies the host immune system, and therefore reshapes interactions between MSCs and immune cells. Finally, we argue that obese environments may alter essential features of allogeneic MSCs and offer potential strategies for licensing of MSCs to enhance their efficacy in the obese microenvironment. Our aim is to combine insights from basic research in MSC biology and clinical trials to inform new strategies to ensure MSC therapy is effective for a broad range of patients.

Transcriptome sequencing of 3,3',4,4',5-Pentachlorobiphenyl (PCB126)-treated human preadipocytes demonstrates progressive changes in pathways associated with inflammation and diabetes.

Polychlorinated biphenyls (PCBs) are persistent organic pollutants that accumulate in adipose tissue and have been associated with cardiometabolic disease. We have previously demonstrated that exposure of human preadipocytes to the dioxin-like PCB126 disrupts adipogenesis via the aryl hydrocarbon receptor (AhR). To further understand how PCB126 disrupts adipose tissue cells, we performed RNAseq analysis of PCB126-treated human preadipocytes over a 3-day time course. The most significant predicted upstream regulator affected by PCB126 exposure at the early time point of 9 h was the AhR. Progressive changes occurred in the number and magnitude of transcript levels of genes associated with inflammation, most closely fitting the pathways of cytokine-cytokine-receptor signaling and the AGE-RAGE diabetic complications pathway. Transcript levels of genes involved in the IL-17A, IL-1ß, MAP kinase, and NF-κB signaling pathways were increasingly dysregulated by PCB126 over time. Our results illustrate the progressive time-dependent nature of transcriptional changes caused by toxicants such as PCB126, point to important pathways affected by PCB126 exposure, and provide a rich dataset for further studies to address how PCB126 and other AhR agonists disrupt preadipocyte function. These findings have implications for understanding how dioxin-like PCBs and other dioxin-like compounds are involved in the development of obesity and diabetes.

Dataset of transcriptomic changes that occur in human preadipocytes over a 3-day course of exposure to 3,3',4,4',5-Pentachlorobiphenyl (PCB126).

Exposure to polychlorinated biphenyls (PCBs) has been associated with the development of metabolic syndrome, a cluster of diseases that includes obesity, diabetes, liver steatosis, and cardiovascular problems. PCBs accumulate and fat and are known to act on adipocytes and their precursors, termed preadipocytes. The PCB congener, PCB126, has been shown to activate the aryl hydrocarbon receptor (AhR) as well as proinflammatory genes. Here, we used RNAseq to assess gene transcript changes that occur in PCB126-exposed human preadipocytes over a time course. RNA was collected from 4 replicates of PCB126-exposed and control-treated preadipocytes at 9 h, 24 h, and 72 h post-exposure. RNA was processed for RNAseq analysis using a NovaSeq 6000 with an obtained minimum of 25 million paired-end 50 bp reads per sample. Reads were aligned using the salmon aligner and transcript expression values were summarized to the gene level using tximport. Gene transcript level counts comparing treated- versus control-treated cells were used for differential expression analysis using DESeq2. Differential expression Excel tables (one for each time point) were generated displaying average differential expression (log2 fold change) of the 4 replicates of treated versus control samples with cutoffs of 0.3 log2 fold change (increase or decrease) and p-values of less than 0.05. FastQ, raw, and differential expression tables were uploaded to GEO. A heat map of genes that were changed in common across all time points was generated using GraphPrism. The data generated from this analysis provides a full transcriptional profile of changes that occur over time in preadipocytes that have been exposed to PCB126. The rich datasets can be mined by other researchers to understand how PCB126 and other dioxin-like compounds, including other PCB congeners such as PCB77 and PCB118, affect biological pathways in preadipocytes and other cell types to cause disease.

Nanoparticle-based monitoring of cell therapy.

Exogenous cell therapy aims to replace/repair diseased or dysfunctional cells and promises to revolutionize medicine by restoring tissue and organ function. To develop effective cell therapy, the location, distribution and long-term persistence of transplanted cells must be evaluated. Nanoparticle (NP) based imaging technologies have the potential to track transplanted cells non-invasively. Here we summarize the most recent advances in NP-based cell tracking with emphasis on (1) the design criteria for cell tracking NPs, (2) protocols for cell labeling, (3) a comparison of available imaging modalities and their corresponding contrast agents, (4) a summary of preclinical studies on NP-based cell tracking and finally (5) perspectives and future directions.

Systemic Mesenchymal Stem Cell Treatment Mitigates Structural and Functional Retinal Ganglion Cell Degeneration in a Mouse Model of Multiple Sclerosis.

Purpose: The purpose of this study was to determine mesenchymal stem cell (MSC) therapy efficacy on rescuing the visual system in the experimental autoimmune encephalomyelitis (EAE) model of multiple sclerosis (MS) and to provide new mechanistic insights. Methods: EAE was induced in female C57BL6 mice by immunization with myelin oligodendrocyte glycoprotein (MOG)35-55, complete Freund's adjuvant, and pertussis toxin. The findings were compared to sham-immunized mice. Half of the EAE mice received intraperitoneally delivered stem cells (EAE + MSC). Clinical progression was monitored according to a five-point EAE scoring scheme. Pattern electroretinogram (PERG) and retinal nerve fiber layer (RNFL) thickness were measured 32 days after induction. Retinas were harvested to determine retinal ganglion cell (RGC) density and prepared for RNA-sequencing. Results: EAE animals that received MSC treatment seven days after EAE induction showed significantly lower motor-sensory impairment, improvement in the PERG amplitude, and preserved RNFL. Analysis of RNA-sequencing data demonstrated statistically significant differences in gene expression in the retina of MSC-treated EAE mice. Differentially expressed genes were enriched for pathways involved in endoplasmic reticulum stress, endothelial cell differentiation, HIF-1 signaling, and cholesterol transport in the MSC-treated EAE group. Conclusions: Systemic MSC treatment positively affects RGC function and survival in EAE mice. Better cholesterol handling by increased expression of Abca1, the cholesterol efflux regulatory protein, paired with the resolution of HIF-1 signaling activation might explain the improvements seen in PERG of EAE animals after MSC treatment. Translational Relevance: Using MSC therapy in a mouse model of MS, we discovered previously unappreciated biochemical pathways associated with RGC neuroprotection, which have the potential to be pharmacologically targeted as a new treatment regimen.