19 results on '"Ward HH"'
Search Results
2. Metabolic acidosis in a lactating woman induced by a deliberate ketogenic diet.
- Author
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Hong MJ, Schwartz LE, Ward HH, Morgan JC, and Jacoby JL
- Subjects
- Female, Humans, Infant, Lactation, Vomiting, Acidosis etiology, Diet, Ketogenic adverse effects, Ketosis etiology
- Abstract
Abstract: This article describes a rare case of lactation ketoacidosis in a patient who started a ketogenic diet while nursing an infant and toddler. The patient presented to the ED with a history of nausea, vomiting, and postural dizziness, and was found to have a significant metabolic acidosis and elevated lipase level. The metabolic changes induced in this patient could occur in anyone with high metabolic demands who also is on a strict ketogenic diet. The case highlights the importance of a dietary history in patients with unexplained metabolic derangements., (Copyright © 2021 American Academy of Physician Assistants.)
- Published
- 2021
- Full Text
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3. Citrobacter koseri meningitis with cerebral edema and pneumocephalus in a neonate.
- Author
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Ward HH, Lauber P, Laubach LT, Fishbein J, and Greenberg MR
- Abstract
Sometimes the only indicator of a serious infection in a neonate is a fever. Citrobacter koseri (C. koseri) has been reported to cause neonatal brain abscesses in the setting of meningitis. Although rare, pneumocephalus, secondary to C. koseri , carries a very high mortality. A 17-day-old male presented to the emergency department with a fever, decreased oral intake, and lethargy. The patient developed pneumocephalus and cerebral edema and was diagnosed with C. koseri meningitis, leading to death. This case demonstrates the presentation of C. koseri meningitis with pneumocephalus and cerebral edema in a neonate presenting with fever., (© 2020 The Authors. Published by Elsevier Inc. on behalf of University of Washington.)
- Published
- 2020
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4. Clinical and Demographic Parameters of Patients Treated Using a Sepsis Protocol.
- Author
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Ward HH, Kiernan EA, Deschler CL, Murillo SM, Karoly EA, Macfarlan JE, McCambridge MM, Richardson DM, Mackenzie RS, Greenberg MR, and Jacoby JL
- Subjects
- Aged, Aged, 80 and over, Female, Hospital Mortality, Hospitalization, Humans, Male, Middle Aged, Retrospective Studies, Sepsis epidemiology, Sepsis mortality, Sepsis therapy
- Abstract
Purpose: The purpose of this study was to investigate potential differences by sex in the demographic and clinical characteristics of patients treated utilizing a sepsis electronic bundle order set. Risk factors for in-hospital mortality were also assessed., Methods: Data on patients in whom the sepsis order set was initiated in the emergency department over a 16-month period were entered into the hospital database. Data were analyzed for differences by sex in demographic and clinical factors, treatment modalities, and in-hospital mortality. The Bonferroni correction was applied to account for multiple comparisons; α was set at 0.006 for sex differences., Findings: A total of 2204 patients were included. Male and female cohorts were similar with regard to a variety of demographic and clinical factors, including age, Emergency Severity Index (ESI) levels 1 and 2, time to disposition, appropriateness of antibiotics, and total fluids given by weight. The ESI is an assessment score ranging from 1 to 5 (1 is emergent). There were modest differences in the source of infection (genitourinary was 4% more common in women; P = 0.03) and mode of arrival (men were 4% more likely to arrive by ambulance; P = 0.03). These differences did not achieve our predefined α of 0.006 when the Bonferroni correction was applied. Factors associated with in-hospital mortality were advanced age, arrival by ambulance, and an ESI level of 1 or 2 (all, P < 0.01)., Implications: Women were more likely to have a genitourinary cause of sepsis and less likely to arrive by ambulance. Risk factors of in-hospital mortality were older age, arrival by ambulance, and an ESI level of 1 or 2, but not sex., (Copyright © 2019. Published by Elsevier Inc.)
- Published
- 2019
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5. Intermittent hypoxia exacerbates increased blood pressure in rats with chronic kidney disease.
- Author
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Riggs JL, Pace CE, Ward HH, Gonzalez Bosc LV, Rios L, Barrera A, and Kanagy NL
- Subjects
- Animals, Arterial Pressure physiology, Cardiovascular Diseases physiopathology, Kidney physiopathology, Kidney Function Tests, Male, Rats, Sprague-Dawley, Renal Insufficiency, Chronic complications, Blood Pressure physiology, Hypertension physiopathology, Hypoxia physiopathology, Renal Insufficiency, Chronic physiopathology
- Abstract
Kidney injury and sleep apnea (SA) are independent risk factors for hypertension. Exposing rats to intermittent hypoxia (IH) to simulate SA increases blood pressure whereas adenine feeding causes persistent kidney damage to model chronic kidney disease (CKD). We hypothesized that exposing CKD rats to IH would exacerbate the development of hypertension and renal failure. Male Sprague-Dawley rats were fed a 0.2% adenine diet or control diet (Control) until blood urea nitrogen was >120 mg/dl in adenine-fed rats (14 ± 4 days, mean ± SE). After 2 wk of recovery on normal chow, rats were exposed to IH (20 exposures/h of 5% O
2 -5% CO2 7 h/day) or control conditions (Air) for 6 wk. Mean arterial pressure (MAP) was monitored with telemeters, and plasma and urine samples were collected weekly to calculate creatinine clearance as an index of glomerular filtration rate (GFR). Prior to IH, adenine-fed rats had higher blood pressure than rats on control diet. IH treatment increased MAP in both groups, and after 6 wk, MAP levels in the CKD/IH rats were greater than those in the CKD/Air and Control/IH rats. MAP levels in the Control/Air rats were lower than those in the other three groups. Kidney histology revealed crystalline deposits, tubule dilation, and interstitial fibrosis in both CKD groups. IH caused no additional kidney damage. Plasma creatinine was similarly increased in both CKD groups throughout whereas IH alone increased plasma creatinine. IH increases blood pressure further in CKD rats without augmenting declines in GFR but appears to impair GFR in healthy rats. We speculate that treating SA might decrease hypertension development in CKD patients and protect renal function in SA patients.- Published
- 2018
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6. Variability of PD-L1 expression in mastocytosis.
- Author
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Hatch EW, Geeze MB, Martin C, Salama ME, Hartmann K, Eisenwort G, Blatt K, Valent P, Gotlib J, Lee JH, Chen L, Ward HH, Lidke DS, and George TI
- Subjects
- Adult, Aged, B7-H1 Antigen analysis, Bone Marrow pathology, Diagnosis, Differential, Female, Humans, Male, Mastocytosis diagnosis, Mastocytosis, Cutaneous diagnosis, Mastocytosis, Systemic diagnosis, Middle Aged, Neoplasms diagnosis, Programmed Cell Death 1 Receptor analysis, Young Adult, B7-H1 Antigen metabolism, Mastocytosis chemistry, Programmed Cell Death 1 Receptor metabolism
- Abstract
Mastocytosis is a rare disease with heterogeneous clinical manifestations and few effective therapies. Programmed death-1 (PD-1) and its ligands (PD-L1 and PD-L2) protect tissues from immune-mediated damage and permit tumors to evade immune destruction. Therapeutic antibodies against PD-1 and PD-L1 are effective in the treatment of a variety of neoplasms. In the present study, we sought to systematically analyze expression of PD-1 and PD-L1 in a large number of patients with mastocytosis using immunohistochemistry and multiplex fluorescence staining. PD-L1 showed membrane staining of neoplastic mast cells (MCs) in 77% of systemic mastocytosis (SM) cases including 3 of 3 patients with MC leukemia, 2 of 2 with aggressive SM, 1 of 2 with smoldering SM, 3 of 4 with indolent SM, and 9 of 12 with SM with an associated hematologic neoplasm (SM component only). Ninety-two percent (23 of 25) of cutaneous mastocytosis (CM) cases and 1 of 2 with myelomastocytic leukemia expressed PD-L1, with no expression found in 15 healthy/reactive marrows, 18 myelodysplastic syndromes (MDSs), 16 myeloproliferative neoplasms (MPNs), 5 MDS/MPNs, and 3 monoclonal MC activation syndromes. Variable PD-L1 expression was observed between and within samples, with PD-L1 staining of MCs ranging from 10% to 100% (mean, 50%). PD-1 dimly stained 4 of 27 CM cases (15%), with no expression in SM or other neoplasms tested; PD-1 staining of MCs ranged from 20% to 50% (mean, 27%). These results provide support for the expression of PD-L1 in SM and CM, and PD-1 expression in CM. These data support the exploration of agents with anti-PD-L1 activity in patients with advanced mastocytosis., (© 2018 by The American Society of Hematology.)
- Published
- 2018
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7. Transcriptome analysis reveals manifold mechanisms of cyst development in ADPKD.
- Author
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de Almeida RM, Clendenon SG, Richards WG, Boedigheimer M, Damore M, Rossetti S, Harris PC, Herbert BS, Xu WM, Wandinger-Ness A, Ward HH, Glazier JA, and Bacallao RL
- Subjects
- Adult, Case-Control Studies, Cell Line, Gene Expression Profiling, Gene Ontology, Humans, Male, Metabolic Networks and Pathways, Middle Aged, Polycystic Kidney, Autosomal Dominant pathology, TRPP Cation Channels genetics, TRPP Cation Channels metabolism, Polycystic Kidney, Autosomal Dominant metabolism, Transcriptome
- Abstract
Background: Autosomal dominant polycystic kidney disease (ADPKD) causes progressive loss of renal function in adults as a consequence of the accumulation of cysts. ADPKD is the most common genetic cause of end-stage renal disease. Mutations in polycystin-1 occur in 87% of cases of ADPKD and mutations in polycystin-2 are found in 12% of ADPKD patients. The complexity of ADPKD has hampered efforts to identify the mechanisms underlying its pathogenesis. No current FDA (Federal Drug Administration)-approved therapies ameliorate ADPKD progression., Results: We used the de Almeida laboratory's sensitive new transcriptogram method for whole-genome gene expression data analysis to analyze microarray data from cell lines developed from cell isolates of normal kidney and of both non-cystic nephrons and cysts from the kidney of a patient with ADPKD. We compared results obtained using standard Ingenuity Volcano plot analysis, Gene Set Enrichment Analysis (GSEA) and transcriptogram analysis. Transcriptogram analysis confirmed the findings of Ingenuity, GSEA, and published analysis of ADPKD kidney data and also identified multiple new expression changes in KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways related to cell growth, cell death, genetic information processing, nucleotide metabolism, signal transduction, immune response, response to stimulus, cellular processes, ion homeostasis and transport and cofactors, vitamins, amino acids, energy, carbohydrates, drugs, lipids, and glycans. Transcriptogram analysis also provides significance metrics which allow us to prioritize further study of these pathways., Conclusions: Transcriptogram analysis identifies novel pathways altered in ADPKD, providing new avenues to identify both ADPKD's mechanisms of pathogenesis and pharmaceutical targets to ameliorate the progression of the disease.
- Published
- 2016
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8. Dual-Purpose Bioreactors to Monitor Noninvasive Physical and Biochemical Markers of Kidney and Liver Scaffold Recellularization.
- Author
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Uzarski JS, Bijonowski BM, Wang B, Ward HH, Wandinger-Ness A, Miller WM, and Wertheim JA
- Subjects
- Animals, Cell Culture Techniques methods, Cells, Cultured, Humans, Male, Rats, Rats, Sprague-Dawley, Antigens, Differentiation biosynthesis, Bioreactors, Kidney chemistry, Liver chemistry, Tissue Scaffolds chemistry
- Abstract
Analysis of perfusion-based bioreactors for organ engineering and a detailed evaluation of physical and biochemical parameters that measure dynamic changes within maturing cell-laden scaffolds are critical components of ex vivo tissue development that remain understudied topics in the tissue and organ engineering literature. Intricately designed bioreactors that house developing tissue are critical to properly recapitulate the in vivo environment, deliver nutrients within perfused media, and monitor physiological parameters of tissue development. Herein, we provide an in-depth description and analysis of two dual-purpose perfusion bioreactors that improve upon current bioreactor designs and enable comparative analyses of ex vivo scaffold recellularization strategies and cell growth performance during long-term maintenance culture of engineered kidney or liver tissues. Both bioreactors are effective at maximizing cell seeding of small-animal organ scaffolds and maintaining cell survival in extended culture. We further demonstrate noninvasive monitoring capabilities for tracking dynamic changes within scaffolds as the native cellular component is removed during decellularization and model human cells are introduced into the scaffold during recellularization and proliferate in maintenance culture. We found that hydrodynamic pressure drop (ΔP) across the retained scaffold vasculature is a noninvasive measurement of scaffold integrity. We further show that ΔP, and thus resistance to fluid flow through the scaffold, decreases with cell loss during decellularization and correspondingly increases to near normal values for whole organs following recellularization of the kidney or liver scaffolds. Perfused media may be further sampled in real time to measure soluble biomarkers (e.g., resazurin, albumin, or kidney injury molecule-1) that indicate degree of cellular metabolic activity, synthetic function, or engraftment into the scaffold. Cell growth within bioreactors is validated for primary and immortalized cells, and the design of each bioreactor is scalable to accommodate any three-dimensional scaffold (e.g., synthetic or naturally derived matrix) that contains conduits for nutrient perfusion to deliver media to growing cells and monitor noninvasive parameters during scaffold repopulation, broadening the applicability of these bioreactor systems.
- Published
- 2015
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9. Epithelial Cell Repopulation and Preparation of Rodent Extracellular Matrix Scaffolds for Renal Tissue Development.
- Author
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Uzarski JS, Su J, Xie Y, Zhang ZJ, Ward HH, Wandinger-Ness A, Miller WM, and Wertheim JA
- Subjects
- Animals, Bioreactors, Male, Rats, Rats, Sprague-Dawley, Epithelial Cells cytology, Extracellular Matrix physiology, Kidney cytology, Tissue Engineering methods, Tissue Scaffolds
- Abstract
This protocol details the generation of acellular, yet biofunctional, renal extracellular matrix (ECM) scaffolds that are useful as small-scale model substrates for organ-scale tissue development. Sprague Dawley rat kidneys are cannulated by inserting a catheter into the renal artery and perfused with a series of low-concentration detergents (Triton X-100 and sodium dodecyl sulfate (SDS)) over 26 hr to derive intact, whole-kidney scaffolds with intact perfusable vasculature, glomeruli, and renal tubules. Following decellularization, the renal scaffold is placed inside a custom-designed perfusion bioreactor vessel, and the catheterized renal artery is connected to a perfusion circuit consisting of: a peristaltic pump; tubing; and optional probes for pH, dissolved oxygen, and pressure. After sterilizing the scaffold with peracetic acid and ethanol, and balancing the pH (7.4), the kidney scaffold is prepared for seeding via perfusion of culture medium within a large-capacity incubator maintained at 37 °C and 5% CO2. Forty million renal cortical tubular epithelial (RCTE) cells are injected through the renal artery, and rapidly perfused through the scaffold under high flow (25 ml/min) and pressure (~230 mmHg) for 15 min before reducing the flow to a physiological rate (4 ml/min). RCTE cells primarily populate the tubular ECM niche within the renal cortex, proliferate, and form tubular epithelial structures over seven days of perfusion culture. A 44 µM resazurin solution in culture medium is perfused through the kidney for 1 hr during medium exchanges to provide a fluorometric, redox-based metabolic assessment of cell viability and proliferation during tubulogenesis. The kidney perfusion bioreactor permits non-invasive sampling of medium for biochemical assessment, and multiple inlet ports allow alternative retrograde seeding through the renal vein or ureter. These protocols can be used to recellularize kidney scaffolds with a variety of cell types, including vascular endothelial, tubular epithelial, and stromal fibroblasts, for rapid evaluation within this system.
- Published
- 2015
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10. Optimization and critical evaluation of decellularization strategies to develop renal extracellular matrix scaffolds as biological templates for organ engineering and transplantation.
- Author
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Caralt M, Uzarski JS, Iacob S, Obergfell KP, Berg N, Bijonowski BM, Kiefer KM, Ward HH, Wandinger-Ness A, Miller WM, Zhang ZJ, Abecassis MM, and Wertheim JA
- Subjects
- Animals, Cell Differentiation, Cells, Cultured, Detergents pharmacology, Humans, Kidney Tubules blood supply, Kidney Tubules drug effects, Male, Perfusion, Rats, Rats, Sprague-Dawley, Endothelium, Vascular cytology, Extracellular Matrix physiology, Induced Pluripotent Stem Cells cytology, Kidney Tubules physiology, Organ Transplantation standards, Tissue Engineering, Tissue Scaffolds
- Abstract
The ability to generate patient-specific cells through induced pluripotent stem cell (iPSC) technology has encouraged development of three-dimensional extracellular matrix (ECM) scaffolds as bioactive substrates for cell differentiation with the long-range goal of bioengineering organs for transplantation. Perfusion decellularization uses the vasculature to remove resident cells, leaving an intact ECM template wherein new cells grow; however, a rigorous evaluative framework assessing ECM structural and biochemical quality is lacking. To address this, we developed histologic scoring systems to quantify fundamental characteristics of decellularized rodent kidneys: ECM structure (tubules, vessels, glomeruli) and cell removal. We also assessed growth factor retention--indicating matrix biofunctionality. These scoring systems evaluated three strategies developed to decellularize kidneys (1% Triton X-100, 1% Triton X-100/0.1% sodium dodecyl sulfate (SDS) and 0.02% Trypsin-0.05% EGTA/1% Triton X-100). Triton and Triton/SDS preserved renal microarchitecture and retained matrix-bound basic fibroblast growth factor and vascular endothelial growth factor. Trypsin caused structural deterioration and growth factor loss. Triton/SDS-decellularized scaffolds maintained 3 h of leak-free blood flow in a rodent transplantation model and supported repopulation with human iPSC-derived endothelial cells and tubular epithelial cells ex vivo. Taken together, we identify an optimal Triton/SDS-based decellularization strategy that produces a biomatrix that may ultimately serve as a rodent model for kidney bioengineering., (© Copyright 2014 The American Society of Transplantation and the American Society of Transplant Surgeons.)
- Published
- 2015
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11. OFD1 and flotillins are integral components of a ciliary signaling protein complex organized by polycystins in renal epithelia and odontoblasts.
- Author
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Jerman S, Ward HH, Lee R, Lopes CA, Fry AM, MacDougall M, and Wandinger-Ness A
- Subjects
- Adult, Cell Line, ErbB Receptors metabolism, Green Fluorescent Proteins metabolism, Humans, Kidney Tubules metabolism, Male, Multiprotein Complexes metabolism, Mutant Proteins metabolism, Protein Transport, Signal Transduction, Cilia metabolism, Epithelium metabolism, Kidney metabolism, Membrane Proteins metabolism, Odontoblasts metabolism, Proteins metabolism, TRPP Cation Channels metabolism
- Abstract
Mutation of the X-linked oral-facial-digital syndrome type 1 (OFD1) gene is embryonic lethal in males and results in craniofacial malformations and adult onset polycystic kidney disease in females. While the OFD1 protein localizes to centriolar satellites, centrosomes and basal bodies, its cellular function and how it relates to cystic kidney disease is largely unknown. Here, we demonstrate that OFD1 is assembled into a protein complex that is localized to the primary cilium and contains the epidermal growth factor receptor (EGFR) and domain organizing flotillin proteins. This protein complex, which has similarity to a basolateral adhesion domain formed during cell polarization, also contains the polycystin proteins that when mutant cause autosomal dominant polycystic kidney disease (ADPKD). Importantly, in human ADPKD cells where mutant polycystin-1 fails to localize to cilia, there is a concomitant loss of localization of polycystin-2, OFD1, EGFR and flotillin-1 to cilia. Together, these data suggest that polycystins are necessary for assembly of a novel flotillin-containing ciliary signaling complex and provide a molecular rationale for the common renal pathologies caused by OFD1 and PKD mutations.
- Published
- 2014
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12. Inversin modulates the cortical actin network during mitosis.
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Werner ME, Ward HH, Phillips CL, Miller C, Gattone VH, and Bacallao RL
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- Animals, Cell Migration Assays, HEK293 Cells, HeLa Cells, Humans, Kidney Cortex embryology, Mice, Mice, Knockout, Microscopy, Confocal, Transcription Factors genetics, Actins physiology, Kidney Cortex cytology, Mitosis physiology, Transcription Factors metabolism
- Abstract
Mutations in inversin cause nephronophthisis type II, an autosomal recessive form of polycystic kidney disease associated with situs inversus, dilatation, and kidney cyst formation. Since cyst formation may represent a planar polarity defect, we investigated whether inversin plays a role in cell division. In developing nephrons from inv-/- mouse embryos we observed heterogeneity of nuclear size, increased cell membrane perimeters, cells with double cilia, and increased frequency of binuclear cells. Depletion of inversin by siRNA in cultured mammalian cells leads to an increase in bi- or multinucleated cells. While spindle assembly, contractile ring formation, or furrow ingression appears normal in the absence of inversin, mitotic cell rounding and the underlying rearrangement of the cortical actin cytoskeleton are perturbed. We find that inversin loss causes extensive filopodia formation in both interphase and mitotic cells. These cells also fail to round up in metaphase. The resultant spindle positioning defects lead to asymmetric division plane formation and cell division. In a cell motility assay, fibroblasts isolated from inv-/- mouse embryos migrate at half the speed of wild-type fibroblasts. Together these data suggest that inversin is a regulator of cortical actin required for cell rounding and spindle positioning during mitosis. Furthermore, cell division defects resulting from improper spindle position and perturbed actin organization contribute to altered nephron morphogenesis in the absence of inversin.
- Published
- 2013
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13. High resolution 4-dimension imaging of metanephric embryonic kidney morphogenesis.
- Author
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Clendenon SG, Ward HH, Dunn KW, and Bacallao R
- Subjects
- Animals, Boron Compounds metabolism, Ceramides metabolism, Equipment Design, Fluorescent Dyes metabolism, Gestational Age, Image Processing, Computer-Assisted, Kidney metabolism, Mice, Morphogenesis, Organ Culture Techniques, Time Factors, Imaging, Three-Dimensional, Kidney embryology, Microscopy, Fluorescence, Multiphoton instrumentation
- Abstract
High-resolution three-dimensional imaging of fixed embryonic kidney tissues has advanced considerably in the past decade. Here we developed a new process for imaging whole metanephric organ culture at cell resolution in three dimensions over time. This technique combines the use of the newly available generation of infrared-optimized long working distance, high numerical aperture objectives and multiphoton fluorescence microscopy with a new system for vital staining of metanephric organ cultures with bodipy ceramide. This allows all cells in the organ culture to be visualized over time, enabling detailed observation of tissue morphogenesis. Thus, our method offers a powerful new approach for visualizing and understanding early events in renal development and for extending observations made in genetically manipulated models.
- Published
- 2013
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14. A telomerase immortalized human proximal tubule cell line with a truncation mutation (Q4004X) in polycystin-1.
- Author
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Herbert BS, Grimes BR, Xu WM, Werner M, Ward C, Rossetti S, Harris P, Bello-Reuss E, Ward HH, Miller C, Gattone VH 2nd, Phillips CL, Wandinger-Ness A, and Bacallao RL
- Subjects
- Flow Cytometry, Fluorescent Antibody Technique, Humans, Immunoblotting, Microscopy, Electron, Scanning, Microscopy, Electron, Transmission, Transduction, Genetic, Cell Line, Codon, Nonsense genetics, Kidney Tubules, Proximal cytology, Polycystic Kidney, Autosomal Dominant pathology, TRPP Cation Channels genetics, Telomerase metabolism
- Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is associated with a variety of cellular phenotypes in renal epithelial cells. Cystic epithelia are secretory as opposed to absorptive, have higher proliferation rates in cell culture and have some characteristics of epithelial to mesenchymal transitions. In this communication we describe a telomerase immortalized cell line that expresses proximal tubule markers and is derived from renal cysts of an ADPKD kidney. These cells have a single detectable truncating mutation (Q4004X) in polycystin-1. These cells make normal appearing but shorter cilia and fail to assemble polycystin-1 in the cilia, and less uncleaved polycystin-1 in membrane fractions. This cell line has been maintained in continuous passage for over 35 passages without going into senescence. Nephron segment specific markers suggest a proximal tubule origin for these cells and the cell line will be useful to study mechanistic details of cyst formation in proximal tubule cells.
- Published
- 2013
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15. Adult human CD133/1(+) kidney cells isolated from papilla integrate into developing kidney tubules.
- Author
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Ward HH, Romero E, Welford A, Pickett G, Bacallao R, Gattone VH 2nd, Ness SA, Wandinger-Ness A, and Roitbak T
- Subjects
- AC133 Antigen, Adult Stem Cells transplantation, Animals, Cell Differentiation, Cell Separation, Coculture Techniques, Colony-Forming Units Assay, Humans, Mice, Organ Culture Techniques, Polycystic Kidney, Autosomal Dominant therapy, Adult Stem Cells cytology, Adult Stem Cells immunology, Antigens, CD metabolism, Glycoproteins metabolism, Kidney Medulla cytology, Kidney Tubules cytology, Kidney Tubules growth & development, Peptides metabolism
- Abstract
Approximately 60,000 patients in the United States are waiting for a kidney transplant due to genetic, immunologic and environmentally caused kidney failure. Adult human renal stem cells could offer opportunities for autologous transplant and repair of damaged organs. Current data suggest that there are multiple progenitor types in the kidney with distinct localizations. In the present study, we characterize cells derived from human kidney papilla and show their capacity for tubulogenesis. In situ, nestin(+) and CD133/1(+) cells were found extensively intercalated between tubular epithelia in the loops of Henle of renal papilla, but not of the cortex. Populations of primary cells from the renal cortex and renal papilla were isolated by enzymatic digestion from human kidneys unsuited for transplant and immuno-enriched for CD133/1(+) cells. Isolated CD133/1(+) papillary cells were positive for nestin, as well as several human embryonic stem cell markers (SSEA4, Nanog, SOX2, and OCT4/POU5F1) and could be triggered to adopt tubular epithelial and neuronal-like phenotypes. Isolated papillary cells exhibited morphologic plasticity upon modulation of culture conditions and inhibition of asymmetric cell division. Labeled papillary cells readily associated with cortical tubular epithelia in co-culture and 3-dimensional collagen gel cultures. Heterologous organ culture demonstrated that CD133/1(+) progenitors from the papilla and cortex became integrated into developing kidney tubules. Tubular epithelia did not participate in tubulogenesis. Human renal papilla harbor cells with the hallmarks of adult kidney stem/progenitor cells that can be amplified and phenotypically modulated in culture while retaining the capacity to form new kidney tubules. This article is part of a Special Issue entitled: Polycystic Kidney Disease., (Copyright © 2011 Elsevier B.V. All rights reserved.)
- Published
- 2011
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16. Receptor protein tyrosine phosphatases are novel components of a polycystin complex.
- Author
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Boucher CA, Ward HH, Case RL, Thurston KS, Li X, Needham A, Romero E, Hyink D, Qamar S, Roitbak T, Powell S, Ward C, Wilson PD, Wandinger-Ness A, and Sandford RN
- Subjects
- Amino Acid Sequence, Animals, Cadherins chemistry, Cadherins metabolism, Cell Line, Cell Membrane chemistry, Humans, In Vitro Techniques, Kidney metabolism, Mice, Models, Molecular, Multiprotein Complexes chemistry, Mutagenesis, Site-Directed, Peptide Library, Polycystic Kidney, Autosomal Dominant genetics, Polycystic Kidney, Autosomal Dominant metabolism, Protein Interaction Domains and Motifs, Receptor-Like Protein Tyrosine Phosphatases genetics, Receptor-Like Protein Tyrosine Phosphatases metabolism, Receptor-Like Protein Tyrosine Phosphatases, Class 2 chemistry, Receptor-Like Protein Tyrosine Phosphatases, Class 2 genetics, Receptor-Like Protein Tyrosine Phosphatases, Class 2 metabolism, Receptor-Like Protein Tyrosine Phosphatases, Class 5 chemistry, Receptor-Like Protein Tyrosine Phosphatases, Class 5 genetics, Receptor-Like Protein Tyrosine Phosphatases, Class 5 metabolism, Recombinant Proteins chemistry, Recombinant Proteins genetics, Recombinant Proteins metabolism, Signal Transduction, TRPP Cation Channels genetics, TRPP Cation Channels metabolism, Transcription Factor AP-1 metabolism, Receptor-Like Protein Tyrosine Phosphatases chemistry, TRPP Cation Channels chemistry
- Abstract
Autosomal dominant polycystic kidney disease (ADPKD) is caused by mutation of PKD1 and PKD2 that encode polycystin-1 and polycystin-2. Polycystin-1 is tyrosine phosphorylated and modulates multiple signaling pathways including AP-1, and the identity of the phosphatases regulating polycystin-1 are previously uncharacterized. Here we identify members of the LAR protein tyrosine phosphatase (RPTP) superfamily as members of the polycystin-1complex mediated through extra- and intracellular interactions. The first extracellular PKD1 domain of polycystin-1 interacts with the first Ig domain of RPTPσ, while the polycystin-1 C-terminus of polycystin-1 interacts with the regulatory D2 phosphatase domain of RPTPγ. Additional homo- and heterotypic interactions between RPTPs recruit RPTPδ. The multimeric polycystin protein complex is found localised in cilia. RPTPσ and RPTPδ are also part of a polycystin-1/E-cadherin complex known to be important for early events in adherens junction stabilisation. The interaction between polycystin-1 and RPTPγ is disrupted in ADPKD cells, while RPTPσ and RPTPδ remain closely associated with E-cadherin, largely in an intracellular location. The polycystin-1 C-terminus is an in vitro substrate of RPTPγ, which dephosphorylates the c-Src phosphorylated Y4237 residue and activates AP1-mediated transcription. The data identify RPTPs as novel interacting partners of the polycystins both in cilia and at adhesion complexes and demonstrate RPTPγ phosphatase activity is central to the molecular mechanisms governing polycystin-dependent signaling. This article is part of a Special Issue entitled: Polycystic Kidney Disease., (Copyright © 2010 Elsevier B.V. All rights reserved.)
- Published
- 2011
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17. A conserved signal and GTPase complex are required for the ciliary transport of polycystin-1.
- Author
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Ward HH, Brown-Glaberman U, Wang J, Morita Y, Alper SL, Bedrick EJ, Gattone VH 2nd, Deretic D, and Wandinger-Ness A
- Subjects
- ADP-Ribosylation Factors genetics, ADP-Ribosylation Factors metabolism, Adaptor Proteins, Signal Transducing metabolism, Amino Acid Sequence, Animals, Cell Line, Dogs, GTPase-Activating Proteins metabolism, Gene Expression, Microtubule-Associated Proteins metabolism, Molecular Sequence Data, Primary Cell Culture, Protein Binding, Protein Interaction Domains and Motifs, RNA Interference, Recombinant Fusion Proteins metabolism, TRPP Cation Channels chemistry, rab GTP-Binding Proteins genetics, rab GTP-Binding Proteins metabolism, Cilia metabolism, Conserved Sequence, Multiprotein Complexes metabolism, Protein Sorting Signals, Protein Transport, TRPP Cation Channels metabolism
- Abstract
Primary cilia regulate epithelial differentiation and organ function. Failure of mutant polycystins to localize to cilia abolishes flow-stimulated calcium signaling and causes autosomal dominant polycystic kidney disease. We identify a conserved amino acid sequence, KVHPSST, in the C-terminus of polycystin-1 (PC1) that serves as a ciliary-targeting signal. PC1 binds a multimeric protein complex consisting of several GTPases (Arf4, Rab6, Rab11) and the GTPase-activating protein (GAP), ArfGAP with SH3 domain, ankyrin repeat and PH domain 1 (ASAP1) in the Golgi, which facilitates vesicle budding and Golgi exocytosis. A related N-terminal ciliary-targeting sequence in polycystin-2 similarly binds Arf4. Deletion of the extreme C-terminus of PC1 ablates Arf4 and ASAP1 binding and prevents ciliary localization of an integral membrane CD16.7-PC1 chimera. Interactions are confirmed for chimeric and endogenous proteins through quantitated in vitro and cell-based approaches. PC1 also complexes with Rab8; knockdown of trafficking regulators Arf4 or Rab8 functionally blocks CD16.7-PC1 trafficking to cilia. Mutations in rhodopsin disrupt a similar signal and cause retinitis pigmentosa, while Bardet-Biedl syndrome, primary open-angle glaucoma, and tumor cell invasiveness are linked to dysregulation of ASAP1 or Rab8 or its effectors. In this paper, we provide evidence for a conserved GTPase-dependent ciliary-trafficking mechanism that is shared between epithelia and neurons, and is essential in ciliary-trafficking and cell homeostasis.
- Published
- 2011
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18. Analysis of multiple Invs transcripts in mouse and MDCK cells.
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Ward HH, Wang J, and Phillips C
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Blotting, Northern, Cloning, Molecular, DNA, Complementary chemistry, DNA, Complementary genetics, Dogs, Gene Expression, Humans, Mice, Molecular Sequence Data, Nuclear Localization Signals, Protein Structure, Tertiary, Reverse Transcriptase Polymerase Chain Reaction, Sequence Homology, Amino Acid, Alternative Splicing, Exons genetics, Kidney metabolism, Transcription Factors chemistry, Transcription Factors genetics, Transcription Factors metabolism
- Abstract
Infantile nephronophthisis is associated with cystic kidneys, situs inversus, and INVS mutations. The function of the INVS product, inversin, is unknown but evidence suggests there are multiple inversin isoforms with differing molecular weights, cellular localization patterns, and binding partners. We used Northern blots, RT-PCR, and sequence analysis to identify alternative INVS transcripts. Northern blots probed with Invs cDNA detected four bands in normal mouse kidney. RT-PCR of mouse kidney RNA revealed Invs transcripts with skipping of exon 5, 11, or 13. We sequenced canine (MDCK-II cells) INVS and determined that the corresponding full-length protein shares identity with mouse (74%) and human (84%) inversin. Canine INVS produces a transcript that skips exon 12. Exon skips cause loss of inversin protein motifs, including ankyrin repeats, IQ domains, destruction boxes, and nuclear localization signals. Identification of INVS splice variants will help us determine which inversin protein motifs contribute to left-right asymmetry and kidney development.
- Published
- 2004
- Full Text
- View/download PDF
19. A businessman's view of occupational health and human values.
- Author
-
Ward HH
- Subjects
- Economic Competition, Humans, United States, United States Occupational Safety and Health Administration, Industry, Occupational Diseases, Social Values
- Published
- 1986
- Full Text
- View/download PDF
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