The Metabolomic-Gut-Clinical Axis of Mankai Plant-Derived Dietary Polyphenols.

BACKGROUND: Polyphenols are secondary metabolites produced by plants to defend themselves from environmental stressors. We explored the effect of Wolffia globosa 'Mankai', a novel cultivated strain of a polyphenol-rich aquatic plant, on the metabolomic-gut clinical axis in vitro, in-vivo and in a clinical trial. METHODS: We used mass-spectrometry-based metabolomics methods from three laboratories to detect Mankai phenolic metabolites and examined predicted functional pathways in a Mankai artificial-gut bioreactor. Plasma and urine polyphenols were assessed among the 294 DIRECT-PLUS 18-month trial participants, comparing the effect of a polyphenol-rich green-Mediterranean diet (+1240 mg/polyphenols/day, provided by Mankai, green tea and walnuts) to a walnuts-enriched (+440 mg/polyphenols/day) Mediterranean diet and a healthy controlled diet. RESULTS: Approximately 200 different phenolic compounds were specifically detected in the Mankai plant. The Mankai-supplemented bioreactor artificial gut displayed a significantly higher relative-abundance of 16S-rRNA bacterial gene sequences encoding for enzymes involved in phenolic compound degradation. In humans, several Mankai-related plasma and urine polyphenols were differentially elevated in the green Mediterranean group compared with the other groups (p < 0.05) after six and 18 months of intervention (e.g., urine hydroxy-phenyl-acetic-acid and urolithin-A; plasma Naringenin and 2,5-diOH-benzoic-acid). Specific polyphenols, such as urolithin-A and 4-ethylphenol, were directly involved with clinical weight-related changes. CONCLUSIONS: The Mankai new plant is rich in various unique potent polyphenols, potentially affecting the metabolomic-gut-clinical axis.

Wolffia globosa-Mankai Plant-Based Protein Contains Bioactive Vitamin B12 and Is Well Absorbed in Humans.

BACKGROUND: Rare plants that contain corrinoid compounds mostly comprise cobalamin analogues, which may compete with cobalamin (vitamin B12 (B12)) metabolism. We examined the presence of B12 in a cultivated strain of an aquatic plant: Wolffia globosa (Mankai), and predicted functional pathways using gut-bioreactor, and the effects of long-term Mankai consumption as a partial meat substitute, on serum B12 concentrations. METHODS: We used microbiological assay, liquid-chromatography/electrospray-ionization-tandem-mass-spectrometry (LC-MS/MS), and anoxic bioreactors for the B12 experiments. We explored the effect of a green Mediterranean/low-meat diet, containing 100 g of frozen Mankai shake/day, on serum B12 levels during the 18-month DIRECT-PLUS (ID:NCT03020186) weight-loss trial, compared with control and Mediterranean diet groups. RESULTS: The B12 content of Mankai was consistent at different seasons (p = 0.76). Several cobalamin congeners (Hydroxocobalamin(OH-B12); 5-deoxyadenosylcobalamin(Ado-B12); methylcobalamin(Me-B12); cyanocobalamin(CN-B12)) were identified in Mankai extracts, whereas no pseudo B12 was detected. A higher abundance of 16S-rRNA gene amplicon sequences associated with a genome containing a KEGG ortholog involved in microbial B12 metabolism were observed, compared with control bioreactors that lacked Mankai. Following the DIRECT-PLUS intervention (n = 294 participants; retention-rate = 89%; baseline B12 = 420.5 ± 187.8 pg/mL), serum B12 increased by 5.2% in control, 9.9% in Mediterranean, and 15.4% in Mankai-containing green Mediterranean/low-meat diets (p = 0.025 between extreme groups). CONCLUSIONS: Mankai plant contains bioactive B12 compounds and could serve as a B12 plant-based food source.

Multisector Approach to Improve Healthy Eating and Physical Activity Policies and Practices in Early Care and Education Programs: The National Early Care and Education Learning Collaboratives Project, 2013-2017.

PURPOSE AND OBJECTIVES: Embedding healthy eating and physical activity best practices in early care and education settings is important for instilling healthy behaviors early in life. A collaborative partnership between Nemours Children's Health System and the Centers for Disease Control and Prevention was created to implement the National Early Care and Education Learning Collaboratives Project (ECELC) in childcare settings in 10 states. We measured improvement at the program level by the self-reported number of best practices implemented related to healthy eating and physical activity. INTERVENTION APPROACH: The ECELC implemented a collaborative model with state-level partners (eg, child care resource and referral networks) and early care and education programs. Intervention components received by program directors and lead teachers included 1) self-assessment, 2) in-person learning and training sessions, 3) action planning and implementation, 4) technical assistance, and 5) post-reassessment. EVALUATION METHODS: A pre-post design assessed self-reported policies and practices related to breastfeeding and infant feeding, child nutrition, infant and child physical activity, screen time, and outdoor play and learning as measured by the validated Nutrition and Physical Activity Self-Assessment for Child Care (NAP SACC) best practices instrument. The sample included 1,173 early care and education programs. RESULTS: The number of best practices met for each of the 5 NAP SACC areas increased from pre-assessment to post-assessment approximately 6 months later and ranged from 1.5 to 4.7 best practices (P < .001). Almost all increases occurred regardless of participation in the Child and Adult Care Food Program, Quality Rating Improvement System, Head Start/Early Head Start, and/or accreditation status. IMPLICATIONS FOR PUBLIC HEALTH: The innovative and collaborative partnerships led to broad implementation of healthy eating and physical activity-based practices in early care and education settings. Development, implementation, and evaluation of policy and practice-based partnerships to promote healthy eating and physical activity among children attending early care and education programs may contribute to obesity prevention in the United States.

Accelerating efforts to prevent childhood obesity: spreading, scaling, and sustaining healthy eating and physical activity.

During the past decade, progress has been made in addressing childhood obesity through policy and practice changes that encourage increased physical activity and access to healthy food. With the implementation of these strategies, an understanding of what works to prevent childhood obesity is beginning to emerge. The task now is to consider how best to spread, scale, and sustain promising childhood obesity prevention strategies. In this article we examine a project led by Nemours, a children's health system, to address childhood obesity. We describe Nemours's conceptual approach to spreading, scaling, and sustaining a childhood obesity prevention intervention. We review a component of a Nemours initiative in Delaware that focused on early care and education settings and its expansion to other states through the National Early Care and Education Learning Collaborative to prevent childhood obesity. We also discuss lessons learned. Focusing on the spreading, scaling, and sustaining of promising strategies has the potential to increase the reach and impact of efforts in obesity prevention and help ensure their impact on population health.

Applying a 3.0 transformation framework to guide large-scale health system reform.

Implementation of the Affordable Care Act is unleashing historic new efforts aimed at reforming the US health system. Many important incremental improvements are under way, yet there is a growing recognition that more transformative changes are necessary if the health care system is to do a better job of optimizing population health. While the concept of the Triple Aim-dedicated to improving the experience of care, the health of populations, and lowering per capita costs of care-has been used to help health care providers and health care systems focus their efforts on costs, quality, and outcomes, it does not provide a roadmap for a new system. In this article we describe the 3.0 Transformation Framework we developed to stimulate thinking and support the planning and development of the new roadmap for the next generation of the US health care system. With a focus on optimizing population health over the life span, the framework suggests how a system designed to better manage chronic disease care could evolve into a system designed to enhance population health. We describe how the 3.0 Transformation Framework has been used and applied in national, state, and local settings, and we suggest potential next steps for its wider application and use.

Obesity prevention in the early care and education setting: successful initiatives across a spectrum of opportunities.

With an estimated 12.1% of children aged 2-5 years already obese, prevention efforts must target our youngest children. One of the best places to reach young children for such efforts is the early care and education setting (ECE). More than 11 million U.S. children spend an average of 30 hours per week in ECE facilities. Increased attention at the national, state, and community level on the ECE setting for early obesity prevention efforts has sparked a range of innovative efforts. To assist these efforts, CDC developed a technical assistance and training framework - the Spectrum of Opportunities for Obesity Prevention in the ECE setting - which also served as the organizing framework for the Weight of the Nation ECE track. Participants highlighted their efforts at national, state, and local levels pursuing opportunities on the Spectrum, the standards and best practices that had been the emphasis of their efforts, and common steps for developing, implementing, and evaluating initiatives. Strong leadership and collaboration among a broad group of stakeholders; systematic assessment of needs, opportunities and resources; funding sources; and training and professional development were reported to be integral for successful implementation of standards and best practices, and sustainability.

SILAC compatible strain of Pichia pastoris for expression of isotopically labeled protein standards and quantitative proteomics.

The methylotrophic yeast Pichia pastoris is a powerful eukaryotic platform for the production of heterologous protein. Recent publication of the P. pastoris genome has facilitated strain development toward biopharmaceutical and environmental science applications and has advanced the organism as a model system for the study of peroxisome biogenesis and methanol metabolism. Here we report the development of a P. pastoris arg-/lys- auxotrophic strain compatible with SILAC (stable isotope labeling by amino acids in cell culture) proteomic studies, which is capable of generating large quantities of isotopically labeled protein for mass spectrometry-based biomarker measurements. We demonstrate the utility of this strain to produce high purity human serum albumin uniformly labeled with isotopically heavy arginine and lysine. In addition, we demonstrate the first quantitative proteomic analysis of methanol metabolism in P. pastoris, reporting new evidence for a malate-aspartate NADH shuttle mechanism in the organism. This strain will be a useful model organism for the study of metabolism and peroxisome generation.

A statewide strategy to battle child obesity in Delaware.

In 2006, approximately 37 percent of Delaware's children were overweight or obese. To combat Delaware's childhood obesity epidemic, Nemours, a leading child health care provider, launched a statewide program to improve child health. The "social-ecological" strategy reaches beyond clinical encounters to promote better health and behavior at multiple levels. Early results show that the initiative halted the increase in the prevalence of overweight and obese children, since no statistically significant change occurred during the two-year span between administrations of the Delaware Survey on Children's Health. The initiative also spurred increased knowledge of healthy eating and awareness of the need for increased physical activity in school, child care, and primary care settings.

Proliferating cell nuclear antigen uses two distinct modes to move along DNA.

Proliferating cell nuclear antigen (PCNA) plays an important role in eukaryotic genomic maintenance by topologically binding DNA and recruiting replication and repair proteins. The ring-shaped protein forms a closed circle around double-stranded DNA and is able to move along the DNA in a random walk. The molecular nature of this diffusion process is poorly understood. We use single-molecule imaging to visualize the movement of individual, fluorescently labeled PCNA molecules along stretched DNA. Measurements of diffusional properties as a function of viscosity and protein size suggest that PCNA moves along DNA using two different sliding modes. Most of the time, the clamp moves while rotationally tracking the helical pitch of the DNA duplex. In a less frequently used second mode of diffusion, the movement of the protein is uncoupled from the helical pitch, and the clamp diffuses at much higher rates.

DNA damage tolerance: when it's OK to make mistakes.

Mutations can be beneficial under conditions in which genetic diversity is advantageous, such as somatic hypermutation and antibody generation, but they can also be lethal when they disrupt basic cellular processes or cause uncontrolled proliferation and cancer. Mutations arise from inaccurate processing of lesions generated by endogenous and exogenous DNA damaging agents, and the genome is particularly vulnerable to such damage during S phase. In this phase of the cell cycle, many lesions in the DNA template block replication. Such lesions must be bypassed in order to preserve fork stability and to ensure completion of DNA replication. Lesion bypass is carried out by a set of error-prone and error-free processes collectively referred to as DNA damage tolerance mechanisms. Here, we discuss how two types of DNA damage tolerance, translesion synthesis and template switching, are regulated at stalled replication forks by ubiquitination of PCNA, and the conditions under which they occur.

Beyond medical care: how health systems can address children's needs through health promotion strategies.

This paper describes an innovative approach that integrates community-based health promotion and disease prevention into a well-established pediatric medical care system. System components include a population-level focus in multiple service sectors, community coalitions, knowledge dissemination, and social marketing. The combination of these components is intended to bring about widespread changes in health/social policy and professional practice, which, in turn, should improve health behavior and outcomes. Early lessons are discussed.

Monoubiquitination of proliferating cell nuclear antigen induced by stalled replication requires uncoupling of DNA polymerase and mini-chromosome maintenance helicase activities.

Proliferating cell nuclear antigen (PCNA) is a homotrimeric, ring-shaped protein complex that functions as a processivity factor for DNA polymerases. Following genotoxic stress, PCNA is modified at a conserved site by either a single ubiquitin moiety or a polyubiquitin chain. These modifications are required to coordinate DNA damage tolerance processes with ongoing replication. The molecular mechanisms responsible for inducing PCNA ubiquitination are not well understood. Using Xenopus egg extracts, we show that ultraviolet radiation and aphidicolin treatment induce the mono- and diubiquitination of PCNA. PCNA ubiquitination is replication-dependent and coincides with activation of the ataxia telangiectasia mutated and Rad3-related (ATR)-dependent DNA damage checkpoint pathway. However, loss of ATR signaling by depletion of the ATR-interacting protein (ATRIP) or Rad1, a component of the 911 checkpoint clamp, does not impair PCNA ubiquitination. Primed single-stranded DNA generated by uncoupling of mini-chromosome maintenance helicase and DNA polymerase activities has been shown previously to be necessary for ATR activation. Here we show that PCNA ubiquitination also requires uncoupling of helicase and polymerase activities. We further demonstrate that replicating single-stranded DNA, which mimics the structure produced upon uncoupling, is sufficient to induce PCNA monoubiquitination. Our results suggest that PCNA ubiquitination and ATR activation are two independent events that occur in response to a common single-stranded DNA intermediate generated by functional uncoupling of mini-chromosome maintenance (MCM) helicase and DNA polymerase activities.

Applying lessons learned in communities to programs and policies at the federal level.

As solutions to the problems of the uninsured are debated, there are lessons to be learned from community-based initiatives. Such efforts can provide information on different models as well as key political lessons. Defining the specific role that community efforts play is also critical. Actively involving community stakeholders of such community initiatives in health care policy debates will result in more workable policies.

Crystal structure of a human cyclin-dependent kinase 6 complex with a flavonol inhibitor, fisetin.

Cyclin-dependent kinases (CDKs) play a central role in cell cycle control, apoptosis, transcription, and neuronal functions. They are important targets for the design of drugs with antimitotic or antineurodegenerative effects. CDK4 and CDK6 form a subfamily among the CDKs in mammalian cells, as defined by sequence similarities. Compared to CDK2 and CDK5, structural information on CDK4 and CDK6 is sparse. We describe here the crystal structure of human CDK6 in complex with a viral cyclin and a flavonol inhibitor, fisetin. Fisetin binds to the active form of CDK6, forming hydrogen bonds with the side chains of residues in the binding pocket that undergo large conformational changes during CDK activation by cyclin binding. The 4-keto group and the 3-hydroxyl group of fisetin are hydrogen bonded with the backbone in the hinge region between the N-terminal and C-terminal kinase domain, as has been observed for many CDK inhibitors. However, CDK2 and HCK kinase in complex with other flavone inhibitors such as quercetin and flavopiridol showed a different binding mode with the inhibitor rotated by about 180 degrees. The structural information of the CDK6-fisetin complex is correlated with the binding affinities of different flavone inhibitors for CDK6. This complex structure is the first description of an inhibitor complex with a kinase from the CDK4/6 subfamily and can provide a basis for selecting and designing inhibitor compounds with higher affinities and specificities.

Americans' views about the adequacy of health care for children and the elderly.

Recent attempts to increase health coverage for specific populations incrementally have been more successful than efforts to dramatically reconfigure the health care system. We present findings from a survey to assess support for programs for children compared with those for the elderly, as well as the public's desire to prioritize whether the needs of one should be addressed over the needs of the other. Americans believe that the health care needs of both children and the elderly are not being met, and there is clear and widespread support for a government role in ensuring adequate health care.

The State Children's Health Insurance Program: successes, shortcomings, and challenges.

This paper examines successes and shortcomings of the State Children's Health Insurance Program (SCHIP). SCHIP is a source of coverage for millions of children, improving their access to health care and sparking innovation in program design and improvements in Medicaid. However, SCHIP adds to the complexity of the insurance system and introduces new inequities in access to insurance; it is imperfectly targeting eligible children who are uninsured; and its financing is problematic because of the block-grant funding structure and use of SCHIP funds to cover adults. These issues need to be addressed during the SCHIP reauthorization process.