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Article ; Online: Characterization and CRISPR/Cas9-mediated genetic manipulation of neutrophils derived from Hoxb8-ER-immortalized myeloid progenitors.

Shannon, Jeffrey G / Hinnebusch, B Joseph

2023 Volume 114, Issue 1, Page(s) 42–52

Abstract: Neutrophils represent a first line of defense against a wide variety of microbial pathogens. Transduction with an estrogen receptor-Hoxb8 transcription factor fusion construct conditionally immortalizes myeloid progenitor cells (NeutPro) capable of ... ...

Abstract	Neutrophils represent a first line of defense against a wide variety of microbial pathogens. Transduction with an estrogen receptor-Hoxb8 transcription factor fusion construct conditionally immortalizes myeloid progenitor cells (NeutPro) capable of differentiation into neutrophils. This system has been very useful for generating large numbers of murine neutrophils for in vitro and in vivo studies. However, some questions remain as to how closely neutrophils derived from these immortalized progenitors reflect primary neutrophils. Here we describe our experience with NeutPro-derived neutrophils as it relates to our studies of Yersinia pestis pathogenesis. NeutPro neutrophils have circular or multilobed nuclei, similar to primary bone marrow neutrophils. Differentiation of neutrophils from NeutPro cells leads to increased expression of CD11b, GR1, CD62L, and Ly6G. However, the NeutPro neutrophils expressed lower levels of Ly6G than bone marrow neutrophils. NeutPro neutrophils produced reactive oxygen species at slightly lower levels than bone marrow neutrophils, and the 2 cell types phagocytosed and killed Y. pestis in vitro to a similar degree. To further demonstrate their utility, we used a nonviral method for nuclear delivery of CRISPR/Cas9 guide RNA complexes to delete genes of interest in NeutPro cells. In summary, we have found these cells to be morphologically and functionally equivalent to primary neutrophils and useful for in vitro assays related to studies of bacterial pathogenesis.
MeSH term(s)	Mice ; Animals ; Homeodomain Proteins/genetics ; Homeodomain Proteins/metabolism ; Neutrophils/metabolism ; Receptors, Estrogen/metabolism ; CRISPR-Cas Systems ; Cell Differentiation ; Myeloid Progenitor Cells
Chemical Substances	Homeodomain Proteins ; Receptors, Estrogen ; Hoxb8 protein, mouse
Language	English
Publishing date	2023-03-29
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Intramural
ZDB-ID	605722-6
ISSN	1938-3673 ; 0741-5400
ISSN (online)	1938-3673
ISSN	0741-5400
DOI	10.1093/jleuko/qiad036
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Article ; Online: Antibody Opsonization Enhances Early Interactions between Yersinia pestis and Neutrophils in the Skin and Draining Lymph Node in a Mouse Model of Bubonic Plague.

Shannon, Jeffrey G / Hinnebusch, B Joseph

Infection and immunity

2020 Volume 89, Issue 1

Abstract: Bubonic plague results ... ...

Abstract	Bubonic plague results when
MeSH term(s)	Animals ; Antibodies, Bacterial/immunology ; Antibody-Dependent Cell Cytotoxicity/immunology ; Complement System Proteins/immunology ; Complement System Proteins/metabolism ; Cytokines/metabolism ; Disease Models, Animal ; Host-Pathogen Interactions/immunology ; Immunity, Innate ; Lymph Nodes/immunology ; Lymph Nodes/metabolism ; Lymph Nodes/pathology ; Mice ; Neutrophils/immunology ; Neutrophils/metabolism ; Plague/etiology ; Plague/pathology ; Reactive Oxygen Species/metabolism ; Skin/immunology ; Skin/metabolism ; Skin/microbiology ; Skin/pathology ; Type III Secretion Systems/immunology ; Type III Secretion Systems/metabolism ; Yersinia pestis/immunology
Chemical Substances	Antibodies, Bacterial ; Cytokines ; Reactive Oxygen Species ; Type III Secretion Systems ; Complement System Proteins (9007-36-7)
Language	English
Publishing date	2020-12-15
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Intramural
ZDB-ID	218698-6
ISSN	1098-5522 ; 0019-9567
ISSN (online)	1098-5522
ISSN	0019-9567
DOI	10.1128/IAI.00061-20
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Zs.A 684: Show issues

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Article: Looking beyond year 1 in the molecular era of pediatric brain tumor diagnosis: confirmatory testing of germline variants found on tumor sequencing.

Greene, Brittany L / Stasi, Shannon M / Ting, Michelle A / Waligorski, Natalie / Cole, Bonnie L / Lockwood, Christina M / Paulson, Vera A / Buchan, Jillian G / Lee, Amy / Ojemann, Jeffrey G / Ellenbogen, Richard G / Stevens, Jeffrey / Leary, Sarah E S

Frontiers in oncology

2024 Volume 14, Page(s) 1338022

Abstract: Purpose: Somatic molecular profiling of pediatric brain tumors aids with the diagnosis and treatment of patients with a variety of high- and low-grade central nervous system neoplasms. Here, we report follow-up targeted germline evaluation for patients ... ...

Abstract	Purpose: Somatic molecular profiling of pediatric brain tumors aids with the diagnosis and treatment of patients with a variety of high- and low-grade central nervous system neoplasms. Here, we report follow-up targeted germline evaluation for patients with possible germline variants following tumor only testing in the initial year in which somatic molecular testing was implemented at a single institution. Patients and methods: Somatic testing was completed for all tumors of the central nervous system (CNS) undergoing diagnostic workup at Seattle Children's Hospital during the study period of November 2015 to November 2016. Sequencing was performed in a College of American Pathologists-accredited, Clinical Laboratory Improvements Amendments-certified laboratory using UW-OncoPlex™ assay (version 5), a DNA-based targeted next generation sequencing panel validated to detect genetic alterations in 262 cancer-related genes. We tracked subsequent clinical evaluation and testing on a subgroup of this cohort found to have potential germline variants of interest. Results: Molecular sequencing of 88 patients' tumors identified 31 patients with variants that warranted consideration of germline testing. To date, 19 (61%) patients have been tested. Testing confirmed germline variants for ten patients (31% of those identified for testing), one with two germline variants ( Conclusion: Clinically validated molecular profiling of pediatric brain tumors identifies patients who warrant further germline evaluation. Despite this, only a subset of these patients underwent the indicated confirmatory sequencing. Further work is needed to identify barriers and facilitators to this testing, including the role of genetic counseling and consideration of upfront paired somatic-germline testing.
Language	English
Publishing date	2024-03-06
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2649216-7
ISSN	2234-943X
ISSN	2234-943X
DOI	10.3389/fonc.2024.1338022
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Article ; Online: Intravital Confocal Microscopy of Dermal Innate Immune Responses to Flea-Transmitted Yersinia pestis.

Shannon, Jeffrey G / Hinnebusch, B Joseph

Methods in molecular biology (Clifton, N.J.)

2019 Volume 2010, Page(s) 57–68

Abstract: The technique known as intravital microscopy (IVM), when used in conjunction with transgenic mice expressing fluorescent proteins in various cell populations, is a powerful tool with the potential to provide new insights into host-pathogen interactions ... ...

Abstract	The technique known as intravital microscopy (IVM), when used in conjunction with transgenic mice expressing fluorescent proteins in various cell populations, is a powerful tool with the potential to provide new insights into host-pathogen interactions in infectious disease pathogenesis in vivo. Yersinia pestis, the causative agent of plague, is typically deposited in a host's skin during feeding of an infected flea. IVM has been used to characterize the innate immune response to Y. pestis in the skin and identify differences between the responses to needle-inoculated and flea-transmitted bacteria that would have been difficult, if not impossible, to detect by other means. Here we describe techniques used to image the neutrophil response to flea-transmitted Y. pestis in the dermis of live mice using conventional confocal microscopy.
MeSH term(s)	Animals ; Dermis/immunology ; Dermis/microbiology ; Disease Models, Animal ; Immunity, Innate ; Insect Vectors/microbiology ; Intravital Microscopy/methods ; Mice ; Microscopy, Confocal/methods ; Neutrophils/immunology ; Neutrophils/microbiology ; Plague/immunology ; Plague/microbiology ; Plague/transmission ; Siphonaptera/microbiology ; Yersinia pestis/immunology
Language	English
Publishing date	2019-05-02
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Intramural
ISSN	1940-6029
ISSN (online)	1940-6029
DOI	10.1007/978-1-4939-9541-7_5
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Article ; Online: Characterization of

Dudte, Sophia C / Hinnebusch, B Joseph / Shannon, Jeffrey G

Frontiers in cellular and infection microbiology

2017 Volume 7, Page(s) 358

Abstract: ... Yersinia ... ...

Abstract	Yersinia pestis
Language	English
Publishing date	2017
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2619676-1
ISSN	2235-2988 ; 2235-2988
ISSN (online)	2235-2988
ISSN	2235-2988
DOI	10.3389/fcimb.2017.00358
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Article ; Online: Year 1 in the Molecular Era of Pediatric Brain Tumor Diagnosis: Application of Universal Clinical Targeted Sequencing in an Unselected Cohort of Children.

Cole, Bonnie L / Lockwood, Christina M / Stasi, Shannon / Stevens, Jeffrey / Lee, Amy / Ojemann, Jeffrey G / Ellenbogen, Richard G / Leary, Sarah E S

JCO precision oncology

2022 Volume 2, Page(s) 1–13

Abstract: Purpose: Next-generation sequencing is gaining acceptance as a clinical tool to aid diagnosis and guide treatment of pediatric cancer. Prior pilot studies have evaluated the feasibility and utility of clinical genomic profiling in a subset of selected ... ...

Abstract	Purpose: Next-generation sequencing is gaining acceptance as a clinical tool to aid diagnosis and guide treatment of pediatric cancer. Prior pilot studies have evaluated the feasibility and utility of clinical genomic profiling in a subset of selected patients with brain tumors. Here, we report an unselected prospective cohort study to evaluate the clinical use of universal targeted sequencing in pediatric patients with brain tumors. Methods: We applied a universal sequencing protocol for all tumors of the CNS undergoing diagnostic workup at Seattle Children's Hospital during the study period of November 2015 to November 2016. All tumors were sequenced using the UW-OncoPlex platform, which is a multiplexed targeted deep gene sequencing panel that detects genetic alterations in 262 cancer-related genes performed in a College of American Pathologists-accredited Clinical Laboratory Improvements Amendments-certified laboratory. Results: Eighty-eight patients underwent diagnostic evaluation during the study period, of which 85 tumors (95%) yielded sufficient DNA for sequencing, including 59 newly diagnosed and 26 relapsed. Clinically relevant genetic alterations were identified in 68 of 85 patients (80%). Of these, 57 (67%) had disease-defining or disease-modifying mutations, 44 (52%) had potentially targetable mutations, and 31 (36%) had mutations requiring germline follow-up. As of the last follow-up, seven patients had been prescribed targeted agents on the basis of sequencing results, and nine had confirmed deleterious germline mutations. Conclusion: Clinically validated molecular profiling of pediatric brain tumors aids diagnosis and treatment of patients with a variety of high- and low-grade primary and relapsed pediatric brain tumors.
Language	English
Publishing date	2022-01-27
Publishing country	United States
Document type	Journal Article
ISSN	2473-4284
ISSN (online)	2473-4284
DOI	10.1200/PO.17.00151
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Article ; Online: Genotype-by-environment interactions and local adaptation shape selection in the US National Chip Processing Trial.

Agha, Husain I / Endelman, Jeffrey B / Chitwood-Brown, Jessica / Clough, Mark / Coombs, Joseph / De Jong, Walter S / Douches, David S / Higgins, Charles R / Holm, David G / Novy, Richard / Resende, Marcio F R / Sathuvalli, Vidyasagar / Thompson, Asunta L / Yencho, G Craig / Zotarelli, Lincoln / Shannon, Laura M

TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik

2024 Volume 137, Issue 5, Page(s) 99

Abstract: Key message: We find evidence of selection for local adaptation and extensive genotype-by-environment interaction in the potato National Chip Processing Trial (NCPT). We present a novel method for dissecting the interplay between selection, local ... ...

Abstract	Key message: We find evidence of selection for local adaptation and extensive genotype-by-environment interaction in the potato National Chip Processing Trial (NCPT). We present a novel method for dissecting the interplay between selection, local adaptation and environmental response in plant breeding schemes. Balancing local adaptation and the desire for widely adapted cultivars is challenging for plant breeders and makes genotype-by-environment interactions (GxE) an important target of selection. Selecting for GxE requires plant breeders to evaluate plants across multiple environments. One way breeders have accomplished this is to test advanced materials across many locations. Public potato breeders test advanced breeding material in the National Chip Processing Trial (NCPT), a public-private partnership where breeders from ten institutions submit advanced chip lines to be evaluated in up to ten locations across the country. These clones are genotyped and phenotyped for important agronomic traits. We used these data to interrogate the NCPT for GxE. Further, because breeders submitting clones to the NCPT select in a relatively small geographic range for the first 3 years of selection, we examined these data for evidence of incidental selection for local adaptation, and the alleles underlying it, using an environmental genome-wide association study (envGWAS). We found genomic regions associated with continuous environmental variables and discrete breeding programs, as well as regions of the genome potentially underlying GxE for yield.
MeSH term(s)	Gene-Environment Interaction ; Genome-Wide Association Study ; Plant Breeding ; Genotype ; Phenotype
Language	English
Publishing date	2024-04-10
Publishing country	Germany
Document type	Journal Article
ZDB-ID	2170-2
ISSN	1432-2242 ; 0040-5752
ISSN (online)	1432-2242
ISSN	0040-5752
DOI	10.1007/s00122-024-04610-3
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Zs.A 1692: Show issues

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Article ; Online: Monoaminergic mediation of hyperalgesic and analgesic descending control of nociception in mice.

Nemoto, Wataru / Kozak, Dalia / Sotocinal, Susana G / Tansley, Shannon / Bannister, Kirsty / Mogil, Jeffrey S

Pain

2022 Volume 164, Issue 5, Page(s) 1096–1105

Abstract: Abstract: Descending control of nociception (DCN; also known as conditioned pain modulation [CPM], the behavioral correlate of diffuse noxious inhibitory controls) is the phenomenon whereby pain inhibits pain in another part of the body and is the ... ...

Abstract	Abstract: Descending control of nociception (DCN; also known as conditioned pain modulation [CPM], the behavioral correlate of diffuse noxious inhibitory controls) is the phenomenon whereby pain inhibits pain in another part of the body and is the subject of increasing study because it may represent a biomarker of chronic pain. We recently discovered that pain modulation on the application of a DCN paradigm involving low-intensity test stimuli occurs in the direction of hyperalgesia in healthy mice and rats, whereas the use of high-intensity stimuli produces analgesia. To elucidate the physiological mechanisms underlying hyperalgesic DCN, we administered agonists and antagonists of norepinephrine (NE) and serotonin (5-HT) receptors, key neurochemical players in the production of analgesic DCN. We find that 3 different monoamine reuptake inhibitors-the NE-selective reboxetine, the 5-HT-selective fluoxetine, and the dual NE/5-HT agonist duloxetine-all abolish hyperalgesic DCN when administered into the spinal cord (but not systemically), with no effect on heat or mechanical pain sensitivity. The reversal by reboxetine of hyperalgesic DCN is mediated by α 2 -adrenergic receptors (ie, blocked by atipamezole), and the fluoxetine reversal is mediated by 5-HT 7 receptors (ie, blocked by SB269970). By contrast, analgesic DCN was found to be reversed by atipamezole and SB269970 themselves, with no effect of reboxetine or fluoxetine. Thus, hyperalgesic DCN seems to be the neurochemical opposite to analgesic DCN. These data further validate and help elucidate a preclinical paradigm that mimics dysfunctional CPM and thus may form the basis of translational experiments that aim to reveal preventative pharmacological strategies for individuals predisposed to persistent pain.
MeSH term(s)	Rats ; Mice ; Animals ; Hyperalgesia/drug therapy ; Fluoxetine/pharmacology ; Fluoxetine/therapeutic use ; Serotonin ; Reboxetine ; Nociception ; Rats, Sprague-Dawley ; Analgesics ; Norepinephrine/physiology ; Chronic Pain
Chemical Substances	Fluoxetine (01K63SUP8D) ; Serotonin (333DO1RDJY) ; Reboxetine (947S0YZ36I) ; Analgesics ; Norepinephrine (X4W3ENH1CV)
Language	English
Publishing date	2022-10-19
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	193153-2
ISSN	1872-6623 ; 0304-3959
ISSN (online)	1872-6623
ISSN	0304-3959
DOI	10.1097/j.pain.0000000000002806
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Article ; Online: The impact of imposed delay in elective pediatric neurosurgery: an informed hierarchy of need in the time of mass casualty crisis.

Ahluwalia, Ranbir / Rocque, Brandon G / Shannon, Chevis N / Blount, Jeffrey P

Child's nervous system : ChNS : official journal of the International Society for Pediatric Neurosurgery

2020 Volume 36, Issue 7, Page(s) 1347–1355

Abstract: SARS-CoV-2 COVID-19, coronavirus, has created unique challenges for the medical community after national guidelines called for the cancellation of all elective surgery. While there are clear cases of elective surgery (benign cranial cosmetic defect) and ... ...

Abstract	SARS-CoV-2 COVID-19, coronavirus, has created unique challenges for the medical community after national guidelines called for the cancellation of all elective surgery. While there are clear cases of elective surgery (benign cranial cosmetic defect) and emergency surgery (hemorrhage, fracture, trauma, etc.), there is an unchartered middle ground in pediatric neurosurgery. Children, unlike adults, have dynamic anatomy and are still developing neural networks. Delaying seemingly elective surgery can affect a child's already vulnerable health state by further impacting their neurocognitive development, neurologic functioning, and potential long-term health states. The purpose of this paper is to demonstrate that "elective" pediatric neurosurgery should be risk-stratified, and multi-institutional informed guidelines established.
MeSH term(s)	Betacoronavirus ; COVID-19 ; Child ; Coronavirus Infections/epidemiology ; Coronavirus Infections/surgery ; Elective Surgical Procedures/trends ; Health Services Needs and Demand/trends ; Humans ; Mass Casualty Incidents/prevention & control ; Neurosurgical Procedures/trends ; Pandemics ; Pneumonia, Viral/epidemiology ; Pneumonia, Viral/surgery ; SARS-CoV-2 ; Time-to-Treatment/trends
Keywords	covid19
Language	English
Publishing date	2020-05-20
Publishing country	Germany
Document type	Journal Article ; Review
ZDB-ID	605988-0
ISSN	1433-0350 ; 0302-2803 ; 0256-7040
ISSN (online)	1433-0350
ISSN	0302-2803 ; 0256-7040
DOI	10.1007/s00381-020-04671-x
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Zs.A 2036: Show issues

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Article ; Online: Disparities in preventative diabetic foot examination.

Fermawi, Sarah Ali / Tolson, Jeffrey P / Knapp, Shannon M / Marrero, David / Zhou, Wei / Armstrong, David G / Tan, Tze-Woei

Seminars in vascular surgery

2023 Volume 36, Issue 1, Page(s) 84–89

Abstract: The objective of this study was to assess the overall differences in the standard of preventive foot care for patients at risk of diabetic foot ulceration and to identify specific demographic factors affecting these health care practices, including race ... ...

Abstract	The objective of this study was to assess the overall differences in the standard of preventive foot care for patients at risk of diabetic foot ulceration and to identify specific demographic factors affecting these health care practices, including race and ethnicity. The National Health and Nutrition Examination Survey data for 2011 to 2018 were analyzed. Participants (20 years and older) with diabetes were categorized as White, Black, Hispanic, Asian, and others (including multiracial participants) based on self-reported race and ethnicity. The primary outcome was foot examination over the past year administered by a medical professional. Logistic regression was performed to examine the effects of race and ethnicity on the annual diabetic foot examination, controlling for age (65 years and older), gender, and health insurance status. Among the 2,836 participants included in the study (weighted percentage: 61.1% were White, 13.9% were Black, 15.1% were Hispanic, 5.4% were Asian, and 4.5% were other), 2,018 (weighted percentage: 71.6%) received annual diabetic foot examination over the past year. Hispanic participants (adjusted odds ratio [aOR] = 0.685; 95% CI, 0.52-0.90) were significantly less likely than White participants to receive an annual foot examination (Black participants: aOR = 1.11; 95% CI, 0.83-1.49; Asian participants: aOR = 0.80; 95% CI, 0.60-1.07; other participants: aOR = 0.66; 95% CI, 0.40-1.10). Factors associated with receipt of foot examination were age 65 years or older (aOR = 1.42; 95% CI, 1.05-1.92) and having health insurance (aOR = 3.02; 95% CI, 2.27-4.03). Our findings suggest that Hispanic adults with diabetes are receiving disproportionately lower rates of preventive foot care compared with their White counterparts. This significant variation in the standard of care for individuals with diabetes reflects the need to further identify factors driving the disparities in preventive foot care services among racial and ethnic minority groups.
MeSH term(s)	Adult ; Humans ; Diabetes Mellitus ; Diabetic Foot/diagnosis ; Diabetic Foot/prevention & control ; Ethnicity ; Healthcare Disparities ; Minority Groups ; Nutrition Surveys ; United States/epidemiology
Language	English
Publishing date	2023-01-06
Publishing country	United States
Document type	Journal Article
ZDB-ID	645131-7
ISSN	1558-4518 ; 0895-7967
ISSN (online)	1558-4518
ISSN	0895-7967
DOI	10.1053/j.semvascsurg.2023.01.001
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