Ex vivo normothermic machine perfusion (NMP) is a new clinical strategy to assess and resuscitate organs likely to be declined for transplantation, thereby increasing the number of viable organs available. Short periods of NMP provide a window of opportunity to deliver therapeutics directly to the organ and, in particular, to the vascular endothelial cells (ECs) that constitute the first point of contact with the recipient’s immune system. ECs are the primary targets of both ischemia-reperfusion injury and damage from preformed antidonor antibodies, and reduction of perioperative EC injury could have long-term benefits by reducing the intensity of the host’s alloimmune response. Using NMP to administer therapeutics directly to the graft avoids many of the limitations associated with systemic drug delivery. We have previously shown that polymeric nanoparticles (NPs) can serve as depots for long-term drug release, but ensuring robust NP accumulation within a target cell type (graft ECs in this case) remains a fundamental challenge of nanomedicine. We show that surface conjugation of an anti-CD31 antibody enhances targeting of NPs to graft ECs of human kidneys undergoing NMP. Using a two-color quantitative microscopy approach, we demonstrate that targeting can enhance EC accumulation by about 5-to 10-fold or higher in discrete regions of the renal vasculature. In addition, our studies reveal that NPs can also non-specifically accumulate within obstructed regions of the vasculature that are poorly perfused. These quantitative preclinical human studies demonstrate the therapeutic potential for targeted nanomedicines delivered during ex vivo NMP.

Human endothelial cells are initiators and targets of the rejection response. Pre-operative modification of endothelial cells by small interfering RNA transfection could shape the nature of the host response post-transplantation. Ablation of endothelial cell class II major histocompatibility complex molecules by small interfering RNA targeting of class II transactivator can reduce the capacity of human endothelial cells to recruit and activate alloreactive T cells. Here, we report the development of small interfering RNA-releasing poly(amine-co-ester) nanoparticles, distinguished by their high content of a hydrophobic lactone. We show that a single transfection of small interfering RNA targeting class II transactivator attenuates major histocompatibility complex class II expression on endothelial cells for at least 4 to 6 weeks after transplantation into immunodeficient mouse hosts. Furthermore, silencing of major histocompatibility complex class II reduces allogeneic T-cell responses in vitro and in vivo. These data suggest that poly(amine-co-ester) nanoparticles, potentially administered during ex vivo normothermic machine perfusion of human organs, could be used to modify endothelial cells with a sustained effect after transplantation.

To investigate the variations of T-helper 17 (Th17) and regulatory T (Treg) cells in patients with lupus nephritis (LN), a total of 60 systemic lupus erythematosus patients and 28 healthy controls (HCs) were enrolled. The frequency of Th17 cells and Treg cells in peripheral blood mononuclear cells (PBMCs) was evaluated by flow cytometric analysis. The serum concentrations of interleukin-17 (IL-17) and transforming growth factor-beta 1 (TGF-β1) were measured by enzyme-linked immunosorbent assay (ELISA). The results demonstrated in LN patients a significant decrease in the frequency of CD4+CD25(high) and CD4+CD25+FoxP3+ T cells and a significant increase in the frequency of Th17 cells in peripheral blood, and the ratio of Th17 to Treg cell frequency was significantly increased along with increased SLEDAI scores. LN patients had a lower percentage and expression of FoxP3 in CD4+CD25(high) T cells than SLE patients without nephritis. The concentration of TGF-β1 was found decreased in SLE patients compared with that from healthy controls, though no significant difference was found between LN patients and SLE patients without nephritis. The expression of IL-17 levels in LN patients exhibited a significant increase compared with patients without nephritis and healthy controls. Based on our results, the significantly elevated Th17 cells are accompanied by FoxP3+ Treg cells decrease in lupus nephritis, suggesting that Th17/Treg functional imbalance may be involved in the pathogenesis of renal damage in SLE patients.

Laryngeal squamous cell carcinoma (LSCC) is a common form of head and neck cancer with poor prognosis. However, the mechanism underlying the pathogenesis of LSCC remains unclear. Here, we demonstrated increased expression of fascin actin-bundling protein 1 ( FSCN1 ) and decreased expression of microRNA-145-5p (miR-145-5p) in a clinical cohort of LSCC. Luciferase assay revealed that miR-145-5p is a negative regulator of FSCN1 . Importantly, low miR-145-5p expression was correlated with TNM (tumor, node, metastasis) status and metastasis. Moreover, cases with low miR-145-5p/high FSCN1 expression showed poor prognosis, and these characteristics together served as independent prognostic indicators of survival. Gain- and loss-of-function studies showed that miR-145-5p overexpression or FSCN1 knockdown inhibited LSCC migration, invasion, and growth by suppressing the epithelial-mesenchymal transition along with inducing cell-cycle arrest and apoptosis. Additionally, hypermethylation of the miR-145-5p promoter suggested that repression of miR-145-5p arises through epigenetic inactivation. LSCC tumor growth in vivo could be inhibited by using miR-145-5p agomir or FSCN1 small interfering RNA (siRNA), which highlights the potential for clinical translation. Collectively, our findings indicate that miR-145-5p plays critical roles in inhibiting the progression of LSCC by suppressing FSCN1 . Both miR-145-5p and FSCN1 are important potential prognostic markers and therapeutic targets for LSCC.

Nanoparticles are of long-standing interest for the treatment of neurological diseases such as glioblastoma. Most past work focused on methods to introduce nanoparticles into the brain, suggesting that reaching the brain interstitium will be sufficient to ensure therapeutic efficacy. However, optimized nanoparticle design for drug delivery to the central nervous system is limited by our understanding of their cellular deposition in the brain. Here, we investigated the cellular fate of poly(lactic acid) nanoparticles presenting different surface chemistries, after administration by convection-enhanced delivery. We demonstrate that nanoparticles with ‘stealth' properties mostly avoid internalization by all cell types, but internalization can be enhanced by functionalization with bio-adhesive end-groups. We also show that association rates measured in cultured cells predict the extent of internalization of nanoparticles in cell populations. Finally, evaluating therapeutic efficacy in an orthotopic model of glioblastoma highlights the need to balance significant uptake without inducing adverse toxicity.

Human choice behavior exhibits many paradoxical and challenging patterns. Traditional explanations focus on how values are represented, but little is known about how values are integrated. Here we outline a psychophysical task for value integration that can be used as a window on high-level, multiattribute decisions. Participants choose between alternative rapidly presented streams of numerical values. By controlling the temporal distribution of the values, we demonstrate that this process underlies many puzzling choice paradoxes, such as temporal, risk, and framing biases, as well as preference reversals. These phenomena can be explained by a simple mechanism based on the integration of values, weighted by their salience. The salience of a sampled value depends on its temporal order and momentary rank in the decision context, whereas the direction of the weighting is determined by the task framing. We show that many known choice anomalies may arise from the microstructure of the value integration process. decision making | decoy effects | value psychophysics | expanded judgement R ecent research on the psychology and neuroscience of simple, evidence-based choices (e.g., integrating perceptual or reward information) has made impressive progress, leading to the conclusion that the brain is optimized to make the fastest decision for a specified accuracy (1-5). Accordingly, the observer is assumed to infer the most probable cause of a perceived experience by sequentially accumulating samples of noisy evidence until a response criterion is reached. The idea that simple, evidence-based decision making is optimal contrasts with findings in more complex, motivation-based decisions, focused on multiple goals with tradeoffs (e.g., choices among cars or flats). Here, a number of paradoxical and puzzling choice behaviors (6-8) have been revealed, posing a serious challenge to the development of a unified theory of choice.Can a common theoretical framework between evidence-based and motivation-based decisions be established? A natural starting point is to propose that, in the latter, the cognitive system integrates subjective values (rather than, say, pieces of perceptual evidence), that depend on how each alternative matches the decision maker's goals (9). In particular, when alternatives are characterized by different attributes (e.g., product price and quality), preference is shaped through shifting attention across these attributes (8, 10), assessing an item's subjective value on each attribute, integrating these values across time, and finally making a choice when some threshold is reached (11-13). A detailed understanding of these computations might explain the systematic anomalies observed in motivation-based decisions.This line of research has been difficult to pursue, however, because classical laboratory preference tasks provide little control of the moment-by-moment processes of value sampling and integration. This stands in contrast with psychophysical paradigms for studying evidence-based perceptual choice wher...

Glioblastoma (GBM) is the most common and deadly form of malignant brain tumor in the United States, and current therapies fail to provide significant improvement in survival. Local delivery of nanoparticles is a promising therapeutic strategy that bypasses the blood-brain barrier, minimizes systemic toxicity, and enhances intracranial drug distribution and retention. Here, we developed nanoparticles loaded with agents that inhibit miR-21, an oncogenic microRNA (miRNA) that is strongly overexpressed in GBM compared to normal brain tissue. We synthesized, engineered, and characterized two different delivery systems. One was designed around an anti-miR-21 composed of RNA and employed a cationic poly(amine-co-ester) (PACE). The other was designed around an anti-miR-21 composed of peptide nucleic acid (PNA) and employed a block copolymer of poly(lactic acid) and hyperbranched polyglycerol (PLA-HPG). We show that both nanoparticle products facilitate efficient intracellular delivery and miR-21 suppression that leads to PTEN upregulation and apoptosis of human GBM cells. Further, when administered by convectionenhanced delivery (CED) to animals with intracranial gliomas, they both induced significant miR-21 knockdown and provided chemosensitization, resulting in improved survival when combined with chemotherapy. The challenges involved in optimizing the two delivery systems differed, and despite offering distinct advantages and limitations, results showed significant therapeutic efficacy with both methods of treatment. This study demonstrates the feasibility and promise of local administration of miR-21 inhibiting nanoparticles as an adjuvant therapy for GBM.

Regulatory T (Treg) cells play an important role in the maintenance of immune tolerance to self and in the pathogenesis of autoimmune disease. Transforming growth factor-beta 1(TGF-β1) is a regulatory cytokine with pleiotropic properties in immune responses. This study was to investigate the role of Treg cells and TGF-β1 in the pathogenesis of patients with lupus nephritis (LN). A total of 42 new-onset systemic lupus erythematosus patients and 22 healthy controls were enrolled. The proportion of Treg cells in peripheral blood mononuclear cells (PBMCs) was evaluated by flow cytometric analysis. The serum and urinary TGF-β1 levels were measured by enzyme-linked immunosorbent assay (ELISA). The results demonstrated a significant decrease in the frequency of CD4(+)CD25(high) and CD4(+)CD25(+)FoxP3(+) T cells in LN patients. The concentration of serum TGF-β1 was found decreased in SLE patients, while urinary TGF-β1 levels were significantly higher in LN patients. Based on our results, decreased Treg cells were accompanied with lower serum TGF-β1 levels and higher urinary TGF-β1 levels in LN patients. TGF-ß1 levels in serum may play a key role in the pathogenesis of renal impairment while the significantly increased urinary TGF-β1 levels may be used as a biological marker in prediction of lupus nephritis.