Phosphorus physiology of the marine cyanobacterium Trichodesmium

Submitted in partial fulfillment of the requirements for the degree of Doctor of Philosophy at the Massachusetts Institute of Technology and the Woods Hole Oceanographic Institution February 2010Primary producers play a critical role in the oceanic food chain and the global cycling of carbon. The marine diazotroph Trichodesmium is a major contributor to both primary production and nitrogen fixation in the tropical and subtropical oceans. These regions are often characterized by low phosphorus (P) concentrations, and P deficiency of Trichodesmium could limit growth, and constrain nitrogen fixation by this genus. Three key components of P metabolism in cyanobacteria include: inorganic phosphate (Pi) uptake, dissolved organic phosphorus (DOP) hydrolysis, and polyphosphate (polyP) biosynthesis and catabolism. These three pathways lay the foundation for the four chapters that make up the body of this thesis. Chapter two examines the relative importance of Pi and DOP in Trichodesmium in the Sargasso Sea by measuring uptake rates and uptake kinetics of 33Pi and DOP (using alpha labeled adenosine-5’-triphosphate 33P-ATP as a model P ester compound). The in situ uptake rates suggest that the contribution of P ester to total P uptake can be greater than 25% for Trichodesmium, and therefore P esters are important for supplementing Trichodesmium’s P demand. Chapter three confirms the presence of polyP in the Sargasso Sea populations. This is the first detection of polyP in an oligotrophic system and may indicate micro-scale variability in DIP or DOP supply. Chapter four examines the regulation of several genes involved in P scavenging in cultures of Trichodesmium erythraeum IMS101: two copies of a high affinity phosphate binding protein (pstS and sphX) and two putative alkaline phosphatases (phoA and phoX). Chapter five calibrates the expression of phoX to decreases in nitrogen fixation rate in Trichodesmium cultures allowing the use of phoX expression as a molecular indicator of P limitation of nitrogen fixation. Detection of phoX expression in the North Atlantic and South Pacific identified the extent and magnitude of P limitation in these regions. Taken together this thesis provides new insights into how Trichodesmium survives in low P environments, utilizes DOP, stores polyP, and the extent to which P can limit nitrogen fixation by this genus.I have been supported by the National Science Foundation (grants OCE-0549794 and CCF-424599), the Environmental Protection Agency (R-83041501-0) and the WHOI Academic Programs Office

Tumour occurrence in women with Turner syndrome: a narrative review and single-centre case series

Background: Population studies suggest cancer morbidity may be different in Turner syndrome (TS) compared to the background female population. However, significant variability is observed in cancer associations likely due to heterogeneity in patient cohorts. We explored the prevalence and patterns of cancer amongst a cohort of women with TS attending a dedicated TS clinic. Methods: Retrospective analysis of the patient database was performed to identify TS women who developed cancer. Population data (available before 2015) from the National Cancer Registration and Analysis Service database were used for comparison. Results: Out of 156 TS women, median age of 32 (range 18–73) years, 9 (5.8%) had a recorded cancer diagnosis. Types of cancers were, bilateral gonadoblastoma, type 1 gastric neuroendocrine tumour (NET), appendiceal-NET, gastrointestinal stromal tumour, plasma cell dyscrasia, synovial sarcoma, cervical cancer, medulloblastoma and aplastic anaemia. Median age at cancer diagnosis was 35 (range 7–58) years and two were detected incidentally. Five women had 45,X karyotype, three received growth hormone treatment and all except one received oestrogen replacement therapy. The cancer prevalence of the background age-matched female population was 4.4%. Conclusions: We confirm the previous observations that women with TS do not appear to be at overall increased risk of common malignancies. Our small cohort showed a spectrum of rare malignancies that are not typically associated with TS, except for a single patient with a gonadoblastoma. The slightly higher prevalence of cancer in our cohort might simply represent increased cancer prevalence in the background population, or might be related to small sample size and regular monitoring of these women due to TS per se

Intrathecal enzyme replacement for Hurler syndrome: biomarker association with neurocognitive outcomes.

PurposeAbnormalities in cerebrospinal fluid (CSF) have been reported in Hurler syndrome, a fatal neurodegenerative lysosomal disorder. While no biomarker has predicted neurocognitive response to treatment, one of these abnormalities, glycosaminoglycan nonreducing ends (NREs), holds promise to monitor therapeutic efficacy. A trial of intrathecal enzyme replacement therapy (ERT) added to standard treatment enabled tracking of CSF abnormalities, including NREs. We evaluated safety, biomarker response, and neurocognitive correlates of change.MethodsIn addition to intravenous ERT and hematopoietic cell transplantation, patients (N = 24) received intrathecal ERT at four peritransplant time points; CSF was evaluated at each point. Neurocognitive functioning was quantified at baseline, 1 year, and 2 years posttransplant. Changes in CSF biomarkers and neurocognitive function were evaluated for an association.ResultsOver treatment, there were significant decreases in CSF opening pressure, biomarkers of disease activity, and markers of inflammation. Percent decrease in NRE from pretreatment to final intrathecal dose posttransplant was positively associated with percent change in neurocognitive score from pretreatment to 2 years posttransplant.ConclusionIntrathecal ERT was safe and, in combination with standard treatment, was associated with reductions in CSF abnormalities. Critically, we report evidence of a link between a biomarker treatment response and neurocognitive outcome in Hurler syndrome

A noncanonical autophagy pathway restricts Toxoplasma gondii growth in a strain-specific manner in IFN-γ-activated human cells

ABSTRACT A core set of autophagy proteins is required for gamma interferon (IFN-γ)-mediated clearance of Toxoplasma gondii in the mouse because of their control of several downstream effectors, including immunity-related GTPases (IRGs) and guanylate-binding proteins (GBPs). However, these effectors are absent (i.e., IRGs) from or nonessential (i.e., GBPs) in IFN-γ-activated human cells, raising the question of how these cells control parasite replication. Here, we define a novel role for ubiquitination and recruitment of autophagy adaptors in the strain-specific control of T. gondii replication in IFN-γ-activated human cells. Vacuoles containing susceptible strains of T. gondii became ubiquitinated, recruited the adaptors p62 and NDP52, and were decorated with LC3. Parasites within LC3-positive vacuoles became enclosed in multiple layers of host membranes, resulting in stunting of parasite replication. However, LC3-positive T. gondii-containing vacuoles did not fuse with endosomes and lysosomes, indicating that this process is fundamentally different from xenophagy, a form of autophagy involved in the control of intracellular bacterial pathogens. Genetic knockout of ATG16L or ATG7 reverted the membrane encapsulation and restored parasite replication, indicating that core autophagy proteins involved in LC3 conjugation are important in the control of parasite growth. Despite a role for the core autophagy machinery in this process, upstream activation through Beclin 1 was not sufficient to enhance the ubiquitination of T. gondii-containing vacuoles, suggesting a lack of reliance on canonical autophagy. These findings demonstrate a new mechanism for IFN-γ-dependent control of T. gondii in human cells that depends on ubiquitination and core autophagy proteins that mediate membrane engulfment and restricted growth

Metabolic syndrome components and their response to lifestyle and metformin interventions are associated with differences in diabetes risk in persons with impaired glucose tolerance

AIMS: To determine the association of metabolic syndrome (MetS) and its components with diabetes risk in participants with impaired glucose tolerance (IGT), and whether intervention-related changes in MetS lead to differences in diabetes incidence. METHODS: We used the National Cholesterol Education Program/Adult Treatment Panel III (NCEP/ATP III) revised MetS definition at baseline and intervention-related changes of its components to predict incident diabetes using Cox models in 3234 Diabetes Prevention Program (DPP) participants with IGT over an average follow-up of 3.2 years. RESULTS: In an intention-to-treat analysis, the demographic-adjusted hazard ratios (95% confidence interval) for diabetes in those with MetS (vs. no MetS) at baseline were 1.7 (1.3-2.3), 1.7 (1.2-2.3) and 2.0 (1.3-3.0) for placebo, metformin and lifestyle groups, respectively. Higher levels of fasting plasma glucose and triglycerides at baseline were independently associated with increased risk of diabetes. Greater waist circumference (WC) was associated with higher risk in placebo and lifestyle groups, but not in the metformin group. In a multivariate model, favourable changes in WC (placebo and lifestyle) and high-density lipoprotein cholesterol (placebo and metformin) contributed to reduced diabetes risk. CONCLUSIONS: MetS and some of its components are associated with increased diabetes incidence in persons with IGT in a manner that differed according to DPP intervention. After hyperglycaemia, the most predictive factors for diabetes were baseline hypertriglyceridaemia and both baseline and lifestyle-associated changes in WC. Targeting these cardiometabolic risk factors may help to assess the benefits of interventions that reduce diabetes incidence

Metabolic syndrome components and their response to lifestyle and metformin interventions are associated with differences in diabetes risk in persons with impaired glucose tolerance

Aims To determine the association of metabolic syndrome (MetS) and its components with diabetes risk in participants with impaired glucose tolerance (IGT), and whether intervention-related changes in MetS lead to differences in diabetes incidence. Methods We used the National Cholesterol Education Program/Adult Treatment Panel III (NCEP/ATP III) revised MetS definition at baseline and intervention-related changes of its components to predict incident diabetes using Cox models in 3234 Diabetes Prevention Program (DPP) participants with IGT over an average follow-up of 3.2 years. Results In an intention-to-treat analysis, the demographic-adjusted hazard ratios (95% confidence interval) for diabetes in those with MetS (vs. no MetS) at baseline were 1.7 (1.3–2.3), 1.7 (1.2–2.3) and 2.0 (1.3–3.0) for placebo, metformin and lifestyle groups, respectively. Higher levels of fasting plasma glucose and triglycerides at baseline were independently associated with increased risk of diabetes. Greater waist circumference (WC) was associated with higher risk in placebo and lifestyle groups, but not in the metformin group. In a multivariate model, favourable changes in WC (placebo and lifestyle) and high-density lipoprotein cholesterol (placebo and metformin) contributed to reduced diabetes risk. Conclusions MetS and some of its components are associated with increased diabetes incidence in persons with IGT in a manner that differed according to DPP intervention. After hyperglycaemia, the most predictive factors for diabetes were baseline hypertriglyceridaemia and both baseline and lifestyle-associated changes in WC. Targeting these cardiometabolic risk factors may help to assess the benefits of interventions that reduce diabetes incidence

Recommendations for the management of MPS IVA: systematic evidence- and consensus-based guidance.

IntroductionMucopolysaccharidosis (MPS) IVA or Morquio A syndrome is an autosomal recessive lysosomal storage disorder (LSD) caused by deficiency of the N-acetylgalactosamine-6-sulfatase (GALNS) enzyme, which impairs lysosomal degradation of keratan sulphate and chondroitin-6-sulphate. The multiple clinical manifestations of MPS IVA present numerous challenges for management and necessitate the need for individualised treatment. Although treatment guidelines are available, the methodology used to develop this guidance has come under increased scrutiny. This programme was conducted to provide evidence-based, expert-agreed recommendations to optimise management of MPS IVA.MethodsTwenty six international healthcare professionals across multiple disciplines, with expertise in managing MPS IVA, and three patient advocates formed the Steering Committee (SC) and contributed to the development of this guidance. Representatives from six Patient Advocacy Groups (PAGs) were interviewed to gain insights on patient perspectives. A modified-Delphi methodology was used to demonstrate consensus among a wider group of healthcare professionals with experience managing patients with MPS IVA and the manuscript was evaluated against the validated Appraisal of Guidelines for Research and Evaluation (AGREE II) instrument by three independent reviewers.ResultsA total of 87 guidance statements were developed covering five domains: (1) general management principles; (2) recommended routine monitoring and assessments; (3) disease-modifying interventions (enzyme replacement therapy [ERT] and haematopoietic stem cell transplantation [HSCT]); (4) interventions to support respiratory and sleep disorders; (5) anaesthetics and surgical interventions (including spinal, limb, ophthalmic, cardio-thoracic and ear-nose-throat [ENT] surgeries). Consensus was reached on all statements after two rounds of voting. The overall guideline AGREE II assessment score obtained for the development of the guidance was 5.3/7 (where 1 represents the lowest quality and 7 represents the highest quality of guidance).ConclusionThis manuscript provides evidence- and consensus-based recommendations for the management of patients with MPS IVA and is for use by healthcare professionals that manage the holistic care of patients with the intention to improve clinical- and patient-reported outcomes and enhance patient quality of life. It is recognised that the guidance provided represents a point in time and further research is required to address current knowledge and evidence gaps

Recommendations for the management of MPS VI: systematic evidence- and consensus-based guidance.

IntroductionMucopolysaccharidosis (MPS) VI or Maroteaux-Lamy syndrome (253200) is an autosomal recessive lysosomal storage disorder caused by deficiency in N-acetylgalactosamine-4-sulfatase (arylsulfatase B). The heterogeneity and progressive nature of MPS VI necessitates a multidisciplinary team approach and there is a need for robust guidance to achieve optimal management. This programme was convened to develop evidence-based, expert-agreed recommendations for the general principles of management, routine monitoring requirements and the use of medical and surgical interventions in patients with MPS VI.Methods26 international healthcare professionals from various disciplines, all with expertise in managing MPS VI, and three patient advocates formed the Steering Committee group (SC) and contributed to the development of this guidance. Members from six Patient Advocacy Groups (PAGs) acted as advisors and attended interviews to ensure representation of the patient perspective. A modified-Delphi methodology was used to demonstrate consensus among a wider group of healthcare professionals with expertise and experience managing patients with MPS VI and the manuscript has been evaluated against the validated Appraisal of Guidelines for Research and Evaluation (AGREE II) instrument by three independent reviewers.ResultsA total of 93 guidance statements were developed covering five domains: (1) general management principles; (2) recommended routine monitoring and assessments; (3) enzyme replacement therapy (ERT) and hematopoietic stem cell transplantation (HSCT); (4) interventions to support respiratory and sleep disorders; (5) anaesthetics and surgical interventions. Consensus was reached on all statements after two rounds of voting. The greatest challenges faced by patients as relayed by consultation with PAGs were deficits in endurance, dexterity, hearing, vision and respiratory function. The overall guideline AGREE II assessment score obtained for the development of the guidance was 5.3/7 (where 1 represents the lowest quality and 7 represents the highest quality of guidance).ConclusionThis manuscript provides evidence- and consensus-based recommendations for the management of patients with MPS VI and is for use by healthcare professionals that manage the holistic care of patients with the intention to improve clinical- and patient-reported outcomes and enhance patient quality of life. It is recognised that the guidance provided represents a point in time and further research is required to address current knowledge and evidence gaps

Clinical trial of laronidase in Hurler syndrome after hematopoietic cell transplantation.

BackgroundMucopolysaccharidosis I (MPS IH) is a lysosomal storage disease treated with hematopoietic cell transplantation (HCT) because it stabilizes cognitive deterioration, but is insufficient to alleviate all somatic manifestations. Intravenous laronidase improves somatic burden in attenuated MPS I. It is unknown whether laronidase can improve somatic disease following HCT in MPS IH. The objective of this study was to evaluate the effects of laronidase on somatic outcomes of patients with MPS IH previously treated with HCT.MethodsThis 2-year open-label pilot study of laronidase included ten patients (age 5-13 years) who were at least 2 years post-HCT and donor engrafted. Outcomes were assessed semi-annually and compared to historic controls.ResultsThe two youngest participants had a statistically significant improvement in growth compared to controls. Development of persistent high-titer anti-drug antibodies (ADA) was associated with poorer 6-min walk test (6MWT) performance; when patients with high ADA titers were excluded, there was a significant improvement in the 6MWT in the remaining seven patients.ConclusionsLaronidase seemed to improve growth in participants <8 years old, and 6MWT performance in participants without ADA. Given the small number of patients treated in this pilot study, additional study is needed before definitive conclusions can be made