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Article ; Online: A flow-cytometry-based protocol using diverse cell types for detecting autoantibodies from human plasma and serum samples.

Wong, Andrew Kam Ho / Kulpa, Deanna A / Silvestri, Guido / Maier, Cheryl L

STAR protocols

2021 Volume 2, Issue 4, Page(s) 100924

Abstract: ... of this protocol, please refer to Wong et al. (2021). ...

Abstract	Here, we describe a protocol for cell-based detection of autoantibodies from human plasma and serum samples using a standard flow cytometer. The protocol allows detection of autoantibodies against a wide array of extracellular antigens. Antigen coverage is limited to the cell types tested, and researchers will need to further determine if autoantibody-positive samples correlate with cytotoxic or clinical outcomes. This protocol is less expensive and faster to perform when compared to protein microarrays and requires no prior knowledge of potential targets. For complete details on the use and execution of this protocol, please refer to Wong et al. (2021).
MeSH term(s)	Adult ; Aged ; Aged, 80 and over ; Autoantibodies/blood ; Cell Culture Techniques ; Cell Line ; Female ; Flow Cytometry/methods ; Fluorescent Dyes ; Humans ; Male ; Middle Aged ; Young Adult
Chemical Substances	Autoantibodies ; Fluorescent Dyes
Language	English
Publishing date	2021-10-29
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ISSN	2666-1667
ISSN (online)	2666-1667
DOI	10.1016/j.xpro.2021.100924
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Article ; Online: A flow-cytometry-based protocol using diverse cell types for detecting autoantibodies from human plasma and serum samples

Andrew Kam Ho Wong / Deanna A. Kulpa / Guido Silvestri / Cheryl L. Maier

STAR Protocols, Vol 2, Iss 4, Pp 100924- (2021)

2021

Abstract: ... of this protocol, please refer to Wong et al. (2021). ...

Abstract	Summary: Here, we describe a protocol for cell-based detection of autoantibodies from human plasma and serum samples using a standard flow cytometer. The protocol allows detection of autoantibodies against a wide array of extracellular antigens. Antigen coverage is limited to the cell types tested, and researchers will need to further determine if autoantibody-positive samples correlate with cytotoxic or clinical outcomes. This protocol is less expensive and faster to perform when compared to protein microarrays and requires no prior knowledge of potential targets.For complete details on the use and execution of this protocol, please refer to Wong et al. (2021).
Keywords	Antibody ; Cell Biology ; Cell-based Assays ; Flow Cytometry/Mass Cytometry ; Health Sciences ; Science (General) ; Q1-390
Language	English
Publishing date	2021-12-01T00:00:00Z
Publisher	Elsevier
Document type	Article ; Online
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Article ; Online: Broad auto-reactive IgM responses are common in critically ill patients, including those with COVID-19.

Wong, Andrew Kam Ho / Woodhouse, Isaac / Schneider, Frank / Kulpa, Deanna A / Silvestri, Guido / Maier, Cheryl L

Cell reports. Medicine

2021 Volume 2, Issue 6, Page(s) 100321

Abstract: The pathogenesis of severe coronavirus disease 2019 (COVID-19) remains poorly understood. While several studies suggest that immune dysregulation plays a central role, the key mediators of this process are yet to be defined. Here, we demonstrate that ... ...

Abstract	The pathogenesis of severe coronavirus disease 2019 (COVID-19) remains poorly understood. While several studies suggest that immune dysregulation plays a central role, the key mediators of this process are yet to be defined. Here, we demonstrate that plasma from a high proportion (93%) of critically ill COVID-19 patients, but not healthy controls, contains broadly auto-reactive immunoglobulin M (IgM) and less frequently auto-reactive IgG or IgA. Importantly, these auto-IgMs preferentially recognize primary human lung cells
MeSH term(s)	Autoantibodies/blood ; Autoantibodies/immunology ; COVID-19/immunology ; COVID-19/pathology ; COVID-19/virology ; Cell Line ; Complement C4/metabolism ; Critical Illness ; Humans ; Immunoglobulin M/blood ; Immunoglobulin M/immunology ; Intensive Care Units ; Lung/metabolism ; Protein Array Analysis ; Proteome/analysis ; SARS-CoV-2/isolation & purification
Chemical Substances	Autoantibodies ; Complement C4 ; Immunoglobulin M ; Proteome
Language	English
Publishing date	2021-05-28
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ISSN	2666-3791
ISSN (online)	2666-3791
DOI	10.1016/j.xcrm.2021.100321
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Article ; Online: Acute postoperative endophthalmitis after cataract operation: result of early vitrectomy within 24 hours of presentation.

Iu, Lawrence Pui Leung / Chan, Ho Yan / Li, Gabriel Ka Hin / Ho, Mary / Mak, Andrew Chun Yue / Wong, Posey Po Yin / Kam, Ka Wai / Chen, Li Jia / Brelen, Marten / Young, Alvin Lerrmann

Eye (London, England)

2022 Volume 37, Issue 11, Page(s) 2344–2350

Abstract: Objectives: To evaluate result of early pars plana vitrectomy (PPV) within 24 hours of presentation for acute postoperative endophthalmitis after cataract operation, and to determine factors that predict visual outcome.: Methods: Consecutive patients ...

Abstract	Objectives: To evaluate result of early pars plana vitrectomy (PPV) within 24 hours of presentation for acute postoperative endophthalmitis after cataract operation, and to determine factors that predict visual outcome. Methods: Consecutive patients who developed acute postoperative endophthalmitis within 6 weeks after cataract operation were reviewed. Patients were divided into two groups for analysis: (1) those receiving PPV within 24 hours of presentation (early PPV group), and (2) those receiving initial intravitreal antibiotics only without PPV within 24 hours of presentation (IVA group). Results: Out of 41,411 cataract operations, 22 eyes developed acute postoperative endophthalmitis. Presenting VA was hand-movement or worse in 72.7%. The most common organisms were Staphylococcus (40.9%), Streptococcus (13.6%) and Enterococcus (13.6%). 22.7% of eyes had good final VA ≥ 20/30 and 27.3% had poor final VA < 20/400. Early PPV group had significantly lower rate of requiring additional treatments to control infection (25% versus 80%, P = 0.030), higher rate of retinal detachment (25% versus 0%, P = 0.221) and similar final logMAR VA (1.08 ± 1.08 versus 0.80 ± 0.80, P = 0.489) compared to IVA. Multivariate linear regression analysis showed that worse final VA was significantly associated with Streptococcus (ß = 1.92, P = 0.007) and retinal detachment (ß = 1.72, P = 0.005) but not with early PPV (P = 0.225). Conclusion: Early PPV was superior to initial intravitreal antibiotics alone as it required fewer additional treatments to control infection. Visual outcome was similar between early PPV and initial intravitreal antibiotics alone despite high number of poor presenting VA of light-perception in early PPV group. Streptococcal infection and retinal detachment were major poor prognostic factors for vision.
MeSH term(s)	Humans ; Vitrectomy ; Retinal Detachment/surgery ; Postoperative Complications/surgery ; Endophthalmitis/therapy ; Anti-Bacterial Agents ; Cataract ; Retrospective Studies ; Treatment Outcome
Chemical Substances	Anti-Bacterial Agents
Language	English
Publishing date	2022-12-13
Publishing country	England
Document type	Journal Article
ZDB-ID	91001-6
ISSN	1476-5454 ; 0950-222X
ISSN (online)	1476-5454
ISSN	0950-222X
DOI	10.1038/s41433-022-02347-1
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Article ; Online: Broad auto-reactive IgM responses are common in critically ill patients, including those with COVID-19

Andrew Kam Ho Wong / Isaac Woodhouse / Frank Schneider / Deanna A. Kulpa / Guido Silvestri / Cheryl L. Maier

Cell Reports Medicine, Vol 2, Iss 6, Pp 100321- (2021)

2021

Abstract: Summary: The pathogenesis of severe coronavirus disease 2019 (COVID-19) remains poorly understood. While several studies suggest that immune dysregulation plays a central role, the key mediators of this process are yet to be defined. Here, we demonstrate ...

Abstract	Summary: The pathogenesis of severe coronavirus disease 2019 (COVID-19) remains poorly understood. While several studies suggest that immune dysregulation plays a central role, the key mediators of this process are yet to be defined. Here, we demonstrate that plasma from a high proportion (93%) of critically ill COVID-19 patients, but not healthy controls, contains broadly auto-reactive immunoglobulin M (IgM) and less frequently auto-reactive IgG or IgA. Importantly, these auto-IgMs preferentially recognize primary human lung cells in vitro, including pulmonary endothelial and epithelial cells. By using a combination of flow cytometry, analytical proteome microarray technology, and lactose dehydrogenase (LDH)-release cytotoxicity assays, we identify high-affinity, complement-fixing, auto-reactive IgM directed against 260 candidate autoantigens, including numerous molecules preferentially expressed on the cellular membranes of pulmonary, vascular, gastrointestinal, and renal tissues. These findings suggest that broad IgM-mediated autoimmune reactivity may be involved in the pathogenesis of severe COVID-19, thereby identifying a potential target for therapeutic interventions.
Keywords	Medicine (General) ; R5-920
Subject code	610
Language	English
Publishing date	2021-06-01T00:00:00Z
Publisher	Elsevier
Document type	Article ; Online
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Article ; Online: The flexible N-terminal motif of uL11 unique to eukaryotic ribosomes interacts with P-complex and facilitates protein translation.

Yang, Lei / Lee, Ka-Ming / Yu, Conny Wing-Heng / Imai, Hirotatsu / Choi, Andrew Kwok-Ho / Banfield, David K / Ito, Kosuke / Uchiumi, Toshio / Wong, Kam-Bo

Nucleic acids research

2022 Volume 50, Issue 9, Page(s) 5335–5348

Abstract: Eukaryotic uL11 contains a conserved MPPKFDP motif at the N-terminus that is not found in archaeal and bacterial homologs. Here, we determined the solution structure of human uL11 by NMR spectroscopy and characterized its backbone dynamics by 15N-1H ... ...

Abstract	Eukaryotic uL11 contains a conserved MPPKFDP motif at the N-terminus that is not found in archaeal and bacterial homologs. Here, we determined the solution structure of human uL11 by NMR spectroscopy and characterized its backbone dynamics by 15N-1H relaxation experiments. We showed that these N-terminal residues are unstructured and flexible. Structural comparison with ribosome-bound uL11 suggests that the linker region between the N-terminal domain and C-terminal domain of human uL11 is intrinsically disordered and only becomes structured when bound to the ribosomes. Mutagenesis studies show that the N-terminal conserved MPPKFDP motif is involved in interacting with the P-complex and its extended protuberant domain of uL10 in vitro. Truncation of the MPPKFDP motif also reduced the poly-phenylalanine synthesis in both hybrid ribosome and yeast mutagenesis studies. In addition, G→A/P substitutions to the conserved GPLG motif of helix-1 reduced poly-phenylalanine synthesis to 9-32% in yeast ribosomes. We propose that the flexible N-terminal residues of uL11, which could extend up to ∼25 Å from the N-terminal domain of uL11, can form transient interactions with the uL10 that help to fetch and fix it into a position ready for recruiting the incoming translation factors and facilitate protein synthesis.
MeSH term(s)	Eukaryotic Cells/metabolism ; Humans ; Phenylalanine/metabolism ; Protein Biosynthesis ; Ribosomes/metabolism ; Saccharomyces cerevisiae/genetics
Chemical Substances	Phenylalanine (47E5O17Y3R)
Language	English
Publishing date	2022-05-11
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
ZDB-ID	186809-3
ISSN	1362-4962 ; 1362-4954 ; 0301-5610 ; 0305-1048
ISSN (online)	1362-4962 ; 1362-4954
ISSN	0301-5610 ; 0305-1048
DOI	10.1093/nar/gkac292
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Article ; Online: Modular Lentiviral Vectors for Highly Efficient Transgene Expression in Resting Immune Cells.

Fichter, Christina / Aggarwal, Anupriya / Wong, Andrew Kam Ho / McAllery, Samantha / Mathivanan, Vennila / Hao, Bailey / MacRae, Hugh / Churchill, Melissa J / Gorry, Paul R / Roche, Michael / Gray, Lachlan R / Turville, Stuart

Viruses

2021 Volume 13, Issue 6

Abstract: Gene/cell therapies are promising strategies for the many presently incurable diseases. A key step in this process is the efficient delivery of genes and gene-editing enzymes to many cell types that may be resistant to lentiviral vector transduction. ... ...

Abstract	Gene/cell therapies are promising strategies for the many presently incurable diseases. A key step in this process is the efficient delivery of genes and gene-editing enzymes to many cell types that may be resistant to lentiviral vector transduction. Herein we describe tuning of a lentiviral gene therapy platform to focus on genetic modifications of resting CD4
MeSH term(s)	Animals ; CD4-Positive T-Lymphocytes/immunology ; CD4-Positive T-Lymphocytes/metabolism ; Flow Cytometry ; Gene Expression ; Gene Transfer Techniques ; Genetic Vectors/genetics ; Genotype ; Humans ; Lentivirus/genetics ; Resting Phase, Cell Cycle ; T-Lymphocytes/metabolism ; Transduction, Genetic ; Transgenes
Language	English
Publishing date	2021-06-18
Publishing country	Switzerland
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	2516098-9
ISSN	1999-4915 ; 1999-4915
ISSN (online)	1999-4915
ISSN	1999-4915
DOI	10.3390/v13061170
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Article ; Online: Structural and Mutagenesis Studies Evince the Role of the Extended Protuberant Domain of Ribosomal Protein uL10 in Protein Translation.

Choi, Kwok-Ho Andrew / Yang, Lei / Lee, Ka-Ming / Yu, Conny Wing-Heng / Banfield, David K / Ito, Kosuke / Uchiumi, Toshio / Wong, Kam-Bo

Biochemistry

2019 Volume 58, Issue 36, Page(s) 3744–3754

Abstract: The lateral stalk of ribosomes constitutes the GTPase-associated center and is responsible for recruiting translation factors to the ribosomes. The eukaryotic stalk contains a P-complex, in which one molecule of uL10 (formerly known as P0) protein binds ... ...

Abstract	The lateral stalk of ribosomes constitutes the GTPase-associated center and is responsible for recruiting translation factors to the ribosomes. The eukaryotic stalk contains a P-complex, in which one molecule of uL10 (formerly known as P0) protein binds two copies of P1/P2 heterodimers. Unlike bacterial uL10, eukaryotic uL10 has an extended protuberant (uL10ext) domain inserted into the N-terminal RNA-binding domain. Here, we determined the solution structure of the extended protuberant domain of
MeSH term(s)	Amino Acid Sequence ; Animals ; Bombyx/chemistry ; Escherichia coli/genetics ; Gene Knockout Techniques ; Mutagenesis, Site-Directed ; Mutation ; Nuclear Magnetic Resonance, Biomolecular ; Protein Biosynthesis ; Protein Domains ; Ribosomal Proteins/chemistry ; Ribosomal Proteins/genetics ; Ribosomes/chemistry ; Saccharomyces cerevisiae/genetics ; Sequence Alignment
Chemical Substances	Ribosomal Proteins
Language	English
Publishing date	2019-08-22
Publishing country	United States
Document type	Journal Article ; Research Support, Non-U.S. Gov't
ZDB-ID	1108-3
ISSN	1520-4995 ; 0006-2960
ISSN (online)	1520-4995
ISSN	0006-2960
DOI	10.1021/acs.biochem.9b00528
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Zs.A 217: Show issues

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Article ; Online: CD8

Statzu, Maura / Jin, Wang / Fray, Emily J / Wong, Andrew Kam Ho / Kumar, Mithra R / Ferrer, Elizabeth / Docken, Steffen S / Pinkevych, Mykola / McBrien, Julia B / Fennessey, Christine M / Keele, Brandon F / Liang, Shan / Harper, Justin L / Mutascio, Simona / Franchitti, Lavinia / Wang, Hong / Cicetti, Davide / Bosinger, Steven E / Carnathan, Diane G /

Vanderford, Thomas H / Margolis, David M / Garcia-Martinez, J Victor / Chahroudi, Ann / Paiardini, Mirko / Siliciano, Janet / Davenport, Miles P / Kulpa, Deanna A / Siliciano, Robert S / Silvestri, Guido

Nature microbiology

2023 Volume 8, Issue 2, Page(s) 299–308

Abstract: Persistence of the human immunodeficiency virus type-1 (HIV-1) latent reservoir in infected individuals remains a problem despite fully suppressive antiretroviral therapy (ART). While reservoir formation begins during acute infection, the mechanisms ... ...

Abstract	Persistence of the human immunodeficiency virus type-1 (HIV-1) latent reservoir in infected individuals remains a problem despite fully suppressive antiretroviral therapy (ART). While reservoir formation begins during acute infection, the mechanisms responsible for its establishment remain unclear. CD8
MeSH term(s)	Animals ; Humans ; Simian Immunodeficiency Virus/genetics ; Simian Acquired Immunodeficiency Syndrome/drug therapy ; CD8-Positive T-Lymphocytes ; Anti-Retroviral Agents/therapeutic use ; Macaca mulatta ; HIV Infections/drug therapy
Chemical Substances	Anti-Retroviral Agents
Language	English
Publishing date	2023-01-23
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural
ISSN	2058-5276
ISSN (online)	2058-5276
DOI	10.1038/s41564-022-01311-9
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Article ; Online: Development and validation of an imaging and clinical scoring system to predict early mortality in spontaneous ruptured hepatocellular carcinoma treated with transarterial embolization.

Lee, Kam-Ho / Tse, Man-Lap Donald / Law, Martin / Cheng, Andrew Kai-Chun / Wong, Ho-Yuen Frank / Yu, Man-Leung / Li, Yan-Lin / Ho, Yuen-Chi / Chu, Ferdinand / Lam, Wendy Wai-Man

Abdominal radiology (New York)

2019 Volume 44, Issue 3, Page(s) 903–911

Abstract: Purpose: To develop and validate a scoring system using a combination of imaging and clinical parameters to predict 30-day mortality in ruptured HCC (rHCC) patients after transarterial embolization (TAE).: Methods: 98 consecutive patients with rHCC ... ...

Abstract	Purpose: To develop and validate a scoring system using a combination of imaging and clinical parameters to predict 30-day mortality in ruptured HCC (rHCC) patients after transarterial embolization (TAE). Methods: 98 consecutive patients with rHCC who underwent abdominal CT and subsequent TAE between January 2007 and December 2016 were retrospectively reviewed. The CT scans were reviewed by two radiologists blinded to the patient outcome. Clinical parameters including serum bilirubin, albumin, INR, creatinine, and hemoglobin were recorded. Independent risk factors for 30-day mortality after TAE were identified using multivariate binary logistic regression, for development of a scoring system. The scoring system was then validated in 20 patients between January 2017 and May 2018. Results: In the development cohort, bilobar tumor distribution (OR = 29.6), clinical parameters of bilirubin > 2.5 mg/dL (OR = 5.9), and albumin < 30 g/L (OR = 4.1) were independent predictors for 30-day mortality. A 6-point score was derived and yielded area-under-the-receiver-operating-characteristic-curve (AUC) of 0.904. A score ≥ 4 resulted in sensitivity of 80.5% and specificity of 91.2% for 30-day mortality. In the validation cohort, AUC for 30-day mortality was 0.939. A score ≥ 4 resulted in sensitivity of 81.2% and specificity of 88.9%. In both development and validation cohorts, the proposed scoring system was better than biochemical components of Child-Pugh score and serum bilirubin to predict 30-day mortality. Conclusion: Imaging and clinical parameters can be combined into a scoring system to accurately predict 30-day mortality after TAE in rHCC patients. The score may help identify and counsel high-risk patients.
MeSH term(s)	Aged ; Carcinoma, Hepatocellular/diagnostic imaging ; Carcinoma, Hepatocellular/mortality ; Carcinoma, Hepatocellular/therapy ; Cohort Studies ; Embolization, Therapeutic/methods ; Female ; Humans ; Liver/diagnostic imaging ; Liver Neoplasms/diagnostic imaging ; Liver Neoplasms/mortality ; Liver Neoplasms/therapy ; Male ; Middle Aged ; Predictive Value of Tests ; Reproducibility of Results ; Retrospective Studies ; Risk Assessment ; Rupture, Spontaneous ; Tomography, X-Ray Computed/methods ; Treatment Outcome
Language	English
Publishing date	2019-01-10
Publishing country	United States
Document type	Journal Article ; Validation Study
ZDB-ID	2839786-1
ISSN	2366-0058 ; 2366-004X
ISSN (online)	2366-0058
ISSN	2366-004X
DOI	10.1007/s00261-019-01895-7
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