Early Changes in Liver Distribution Following Implementation of Share 35

In June 2013, a change to the liver waitlist priority algorithm was implemented. Under Share 35, regional candidates with MELD ≥ 35 receive higher priority than local candidates with MELD < 35. We compared liver distribution and mortality in the first 12 months of Share 35 to an equivalent time period before. Under Share 35, new listings with MELD ≥ 35 increased slightly from 752 (9.2% of listings) to 820 (9.7%, p = 0.3), but the proportion of deceased‐donor liver transplants (DDLTs) allocated to recipients with MELD ≥ 35 increased from 23.1% to 30.1% (p < 0.001). The proportion of regional shares increased from 18.9% to 30.4% (p < 0.001). Sharing of exports was less clustered among a handful of centers (Gini coefficient decreased from 0.49 to 0.34), but there was no evidence of change in CIT (p = 0.8). Total adult DDLT volume increased from 4133 to 4369, and adjusted odds of discard decreased by 14% (p = 0.03). Waitlist mortality decreased by 30% among patients with baseline MELD > 30 (SHR = 0.70, p < 0.001) with no change for patients with lower baseline MELD (p = 0.9). Posttransplant length‐of‐stay (p = 0.2) and posttransplant mortality (p = 0.9) remained unchanged. In the first 12 months, Share 35 was associated with more transplants, fewer discards, and lower waitlist mortality, but not at the expense of CIT or early posttransplant outcomes.     

ATG16L1 and NOD2 polymorphisms enhance phagocytosis in monocytes of Crohn’s disease patients

AIM:To investigate if the presence of relevant genetic polymorphisms has effect on the effectual clearance of bacteria by monocytes and granulocytes in patients with Crohn’s disease(CD).METHODS:In this study,we assessed the differential responses in phagocytosis by measuring the phagocytic activity and the percentage of active phagocytic monocytes and granulocytes in inflammatory bowel disease patients as well as healthy controls.As both autophagy related like 1(ATG16L1)and immunityrelated guanosine triphosphatase gene are autophagy genes associated with CD and more recently nucleo-tide-binding ligomerization domain-containing protein2(NOD2)has been identified as a potent inducer of autophagy we genotyped the patients for these variants and correlated this to the phagocytic reaction.The genotyping was done with restriction fragment length polymorphisms analysis and the phagocytosis was determined with the pHrodo?Escherichia coli Bioparticles Phagocytosis kit for flowcytometry.RESULTS:In this study,we demonstrate that analysis of the monocyte and granulocyte populations of patients with CD and ulcerative colitis showed a comparable phagocytic activity(ratio of mean fluorescence intensity)between the patient groups and the healthy controls.CD patients show a significantly higher phagocytic capacity(ratio mean percentage of phagocytic cells)compared to healthy controls(51.91%±2.85%vs 37.67%±7.06%,P=0.05).The extend of disease was not of influence.However,variants of ATG16L1(WT:2.03±0.19 vs homozygoot variant:4.38±0.37,P<0.009)as well as NOD2(C-ins)(heterozygous variant:42.08±2.94 vs homozygous variant:75.58±4.34(P=0.05)are associated with the phagocytic activity in patients with CD.CONCLUSION:Monocytes of CD patients show enhanced phagocytosis associated with the presence of ATG16L1 and NOD2 variants.This could be part of the pathophysiological mechanism resulting in the disease.     

Phosphoglucan-bound structure of starch phosphatase Starch Excess4 reveals the mechanism for C6 specificity

Plants use the insoluble polyglucan starch as their primary glucose storage molecule. Reversible phosphorylation, at the C6 and C3 positions of glucose moieties, is the only known natural modification of starch and is the key regulatory mechanism controlling its diurnal breakdown in plant leaves. The glucan phosphatase Starch Excess4 (SEX4) is a position-specific starch phosphatase that is essential for reversible starch phosphorylation; its absence leads to a dramatic accumulation of starch in Arabidopsis, but the basis for its function is unknown. Here we describe the crystal structure of SEX4 bound to maltoheptaose and phosphate to a resolution of 1.65 Å. SEX4 binds maltoheptaose via a continuous binding pocket and active site that spans both the carbohydrate-binding module (CBM) and the dual-specificity phosphatase (DSP) domain. This extended interface is composed of aromatic and hydrophilic residues that form a specific glucan-interacting platform. SEX4 contains a uniquely adapted DSP active site that accommodates a glucan polymer and is responsible for positioning maltoheptaose in a C6-specific orientation. We identified two DSP domain residues that are responsible for SEX4 site-specific activity and, using these insights, we engineered a SEX4 double mutant that completely reversed specificity from the C6 to the C3 position. Our data demonstrate that the two domains act in consort, with the CBM primarily responsible for engaging glucan chains, whereas the DSP integrates them in the catalytic site for position-specific dephosphorylation. These data provide important insights into the structural basis of glucan phosphatase site-specific activity and open new avenues for their biotechnological utilization.Starch is the primary carbohydrate storage molecule in plants and is an essential constituent of human and animal diets. Starch granules are composed of the glucose homopolymers amylose (10–25%) and amylopectin (75–90%) (1, 2). Amylose is a linear molecule formed from α-1,4-glycosidic–linked chains, whereas amylopectin is formed from α-1,4-glycosidic–linked chains with α-1,6-glycosidic–linked branches (3, 4). Adjacent amylopectin chains interact to form double helices that cause starch granules to be water insoluble, which is an essential feature for its function as a glucose storage molecule (1, 3, 5). However, the outer granular surface of transitory starch must be solubilized during nonphotosynthetic periods so that glycolytic enzymes can access and degrade starch glucans and meet the metabolic needs of the plant (6, 7). Plants regulate the solubility of the starch granular surface via reversible starch phosphorylation that results in a cyclic degradative process: phosphorylation by dikinases, degradation by starch hydrolyzing amylases, and dephosphorylation by phosphatases (1, 8–11). Phosphorylation of amylopectin chains causes helical unwinding and local solubilization of the outer starch granule (12–14). The local solubilization and helix unwinding permits degradation of surface, linear α-1,4 glucan chains by β-amylase, which sequentially removes maltosyl units from the nonreducing end (1, 8, 15). Although glucan phosphorylation of the starch surface is necessary for degradation, the removal of these phosphate groups is required because β-amylase is unable to degrade past the phosphate (6, 15–17). Therefore, glucan phosphatases must release phosphate from starch to reset the degradation cycle, allowing processive starch degradation (8, 16).Recent studies have established that plants use a two-enzyme system for both starch phosphorylation and dephosphorylation. α-Glucan water dikinase phosphorylates the hydroxyl group of starch glucose at the C6 position. This event triggers phosphorylation of the hydroxyl group at the C3 position by phosphoglucan water dikinase (18–21). Similarly, two glucan phosphatases release phosphate from starch. Starch Excess4 (SEX4) preferentially dephosphorylates the C6 position of starch glucose and Like Sex Four2 (LSF2) exclusively dephosphorylates the C3 position (22–26). SEX4 activity is essential for starch catabolism and its mutation in Arabidopsis leads to an excess of leaf starch, a decrease in plant growth, and an accumulation of soluble phosphoglucans produced by the activity of α-amylase 3 and the debranching enzyme isoamylase 3 (16, 25, 27). Conversely, lsf2 mutant Arabidopsis plants display normal levels of leaf starch and plant growth, but the starch contains increased levels of C3-phosphate (22). The difference in plant vitality between sex4 and lsf2 mutants is likely due to SEX4 possessing some compensatory C3-position phosphatase activity (22). Cumulatively, the process of reversible phosphorylation requires the concerted activity of dikinases and phosphatases with SEX4 activity being essential for normal patterns of starch metabolism and plant growth.Glucan phosphatases are members of the protein tyrosine phosphatase (PTP) superfamily characterized by a conserved Cx₅R catalytic motif (24, 28, 29). The glucan phosphatases belong to a heterogeneous subset of PTPs called dual-specificity phosphatases (DSPs), with some DSPs dephosphorylating p-Tyr and p-Ser/Thr residues of proteinaceous substrates and other DSPs dephosphorylating lipids, nucleic acids, or glucans (30–32). In addition to SEX4 and LSF2, a glucan phosphatase called laforin has been identified that dephosphorylates glycogen and influences its solubility in vertebrates (24, 33, 34). Loss of laforin function in humans results in a fatal, neurodegenerative epilepsy called Lafora disease (35–37). Due to their critical function in complex carbohydrate metabolism, understanding the structural basis of glucan phosphatase activity is of particular interest. Toward this goal, we previously determined the ligand-free structure of SEX4 and identified an extensive interdomain interface between its DSP domain and carbohydrate-binding module (CBM) that is maintained in part by a previously unrecognized C-terminal (CT) motif (38). However, the structural mechanism for domain coupling, glucan interaction, and specific C6 dephosphorylation in SEX4 activity is unclear.Starch granule solubilization depends on phosphorylation of starch glucose on the hydroxyl group at both the C6 and C3 positions (6, 13). These phosphorylation events are critical for normal transitory starch degradation, but also directly impact the melting temperature, viscosity, and hydration of starch in industrial settings (39, 40). Developing a means to manipulate starch phosphorylation patterns via enzymatic modification is relevant to agricultural and industrial applications that use starch as a feedstock (9, 12, 14, 41). Therefore, understanding the basis for the site specificity of glucan phosphatases is of particular interest. We recently determined the structure of LSF2 with a glucan bound in a C3-specific orientation and identified unique noncatalytic surface-binding sites (SBSs) not found in other glucan phosphatases (26). SEX4 lacks SBSs and preferentially dephosphorylates the C6 position. The present study was designed to define the fundamental basis for SEX4 substrate binding and understand preferential C6-position specificity in SEX4.Herein, we elucidate the structural mechanism of SEX4-specific activity by determining the structure of SEX4 bound to the phosphoglucan products maltoheptaose and phosphate. SEX4 engages glucan chains via an extended interface of aromatic and hydrophilic residues that spans the CBM and DSP domains. Moreover, the SEX4 CBM is primarily responsible for glucan binding whereas the SEX4 DSP active site is uniquely adapted to engage the phosphoglucan substrate, positioning it in a C6-specific orientation. Structure-guided mutagenesis of DSP active-site residues resulted in a complete reversal from C6 to preferential C3 dephosphorylation by SEX4. Cumulatively, this study establishes the molecular basis for both SEX4 substrate engagement and SEX4 specificity and provides a method for engineering glucan phosphatase activity with modified site specificity.     

An unusual case of vaccine-associated paralytic poliomyelitis

The present article reports a case involving an immunocompetent, previously well child who, despite two previous doses of inactivated poliovirus vaccine, developed severe flaccid paralysis consistent with polio after receiving oral polio vaccine.     

The impact of periodontitis on oral health‐related quality of life: a review of the evidence from observational studies

Modern population based oral health management requires a complete understanding of the impact of disease in order to provide efficient and effective oral health care and guidance. Periodontitis is an important cause of tooth loss and has been shown to be associated with a number of systemic conditions. The impact of oral conditions and disorders on quality of life has been extensively studied. However, the impact of periodontitis on quality of life has received less attention. This review summarizes the literature on the impact of periodontitis on oral health‐related quality of life (OHRQoL). Relevant publications were identified after searching the MEDLINE and EMBASE electronic databases. Screening of titles and abstracts and data extraction was conducted. Only observational studies were included in this review. Most of the reviewed studies reported a negative impact of periodontitis on OHRQoL. However, the reporting standards varied across studies. Moreover, most of the studies were conducted in developed countries.     

Addressing Geographic Disparities in Liver Transplantation Through Redistricting

Severe geographic disparities exist in liver transplantation; for patients with comparable disease severity, 90‐day transplant rates range from 18% to 86% and death rates range from 14% to 82% across donation service areas (DSAs). Broader sharing has been proposed to resolve geographic inequity; however, we hypothesized that the efficacy of broader sharing depends on the geographic partitions used. To determine the potential impact of redistricting on geographic disparity in disease severity at transplantation, we combined existing DSAs into novel regions using mathematical redistricting optimization. Optimized maps and current maps were evaluated using the Liver Simulated Allocation Model. Primary analysis was based on 6700 deceased donors, 28 063 liver transplant candidates, and 242 727 Model of End‐Stage Liver Disease (MELD) changes in 2010. Fully regional sharing within the current regional map would paradoxically worsen geographic disparity (variance in MELD at transplantation increases from 11.2 to 13.5, p = 0.021), although it would decrease waitlist deaths (from 1368 to 1329, p = 0.002). In contrast, regional sharing within an optimized map would significantly reduce geographic disparity (to 7.0, p = 0.002) while achieving a larger decrease in waitlist deaths (to 1307, p = 0.002). Redistricting optimization, but not broader sharing alone, would reduce geographic disparity in allocation of livers for transplant across the United States.     

Dynamic Challenges Inhibiting Optimal Adoption of Kidney Paired Donation: Findings of a Consensus Conference

While kidney paired donation (KPD) enables the utilization of living donor kidneys from healthy and willing donors incompatible with their intended recipients, the strategy poses complex challenges that have limited its adoption in United States and Canada. A consensus conference was convened March 29–30, 2012 to address the dynamic challenges and complexities of KPD that inhibit optimal implementation. Stakeholders considered donor evaluation and care, histocompatibility testing, allocation algorithms, financing, geographic challenges and implementation strategies with the goal to safely maximize KPD at every transplant center. Best practices, knowledge gaps and research goals were identified and summarized in this document.     

Neural progenitor cell transplantation and imaging in a large animal model

To evaluate neural stem/progenitor cell (NPC) transplantation therapy in cat models of neurodegenerative diseases, we have isolated, expanded and characterized feline NPCs (fNPCs) from normal fetal cat brain. Feline NPCs responsive to both human epidermal growth factor (hEGF) and human fibroblast growth factor 2 (hFGF2) proliferated as neurospheres, which were able to differentiate to neurons and glial cells. The analysis of growth factors indicated that both hEGF and hFGF2 were required for proliferation of fNPCs. In contrast to the effect on human NPCs, human leukemia inhibitory factor (hLIF) enhanced differentiation of fNPCs. Expanded fNPCs were injected into the brains of normal adult cats. Immunohistochemical analysis showed that the majority of transplanted cells were located adjacent to the injection site and some fNPCs differentiated into neurons. The survival of transplanted fNPCs over time was monitored using non-invasive bioluminescent imaging technology. This study provided the first evidence of allotransplantation of fNPCs into feline CNS. Cats have heterogeneous genetic backgrounds and possess neurological diseases that closely resemble analogous human diseases. The characterization of fNPCs and exploration of non-invasive bioluminescent imaging to track transplanted cells in this study will allow evaluation of NPC transplantation therapy using feline models of human neurological diseases.