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1.
Clin Lab Med ; 39(4): 591-607, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31668272

RESUMO

Flow cytometry is an incredibly powerful diagnostic tool in the evaluation of primary and secondary immune deficiencies. Assay design and setup involves a methodological consideration of specimen collection, marker and fluorochrome selection, antibody titration, instrumentation, compensation, gating, reference range development, and cross validation. Commonly used analyses for lymphocytes are the lymphocyte subset, T-cell subset, B-cell and T-cell naive/memory, double-negative T-cell, and plasmablast panels. Flow cytometry has direct clinical applicability to the workup of severe forms of primary immune deficiency disorders and is used diagnostically and for therapeutic monitoring in the context of secondary immune deficiency disorders.

2.
Int J Surg Pathol ; 25(5): 406-413, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28351195

RESUMO

OBJECTIVE: Tonsil surgical biopsy or excision is a very common procedure. However, there exist no consensus guidelines for the pathologic handling of tonsil specimens; gross and/or microscopic evaluation may be used. Diagnosis of tonsillar hematologic malignancy requires histology, immunohistochemistry and/or flow cytometry. Data regarding the utility of flow cytometry in tonsillar tissues are limited. We assessed our experience with flow cytometry for tonsil diagnosis with regard to accuracy and use patterns at a tertiary academic medical center. METHODS: We retrospectively analyzed all surgically biopsied or excised tonsil specimens that underwent flow cytometry evaluation from August 2011 to March 2014. Patient clinical information, intraoperative frozen section, histology, immunohistochemistry, and flow cytometry diagnoses were recorded. RESULTS: The study included 154 tonsil specimens from 89 females and 65 males. Patients averaged 27.4 years old (range 2-87 years); 73 were pediatric. Both histology and flow cytometry were benign for 148 patients (96.1%). Hematolymphoid malignancy was diagnosed in 6 adults by histology/immunohistochemistry: diffuse large B-cell lymphoma (2), small B-cell lymphoma (2), concomitant follicular lymphoma and histiocytic sarcoma (1), and extraosseous plasmacytoma (1). Flow cytometry identified abnormal populations in 5 of 6 cases, and detected clonal populations in 2 reactive follicular hyperplasia cases. CONCLUSION: Tonsillar hematolymphoid malignancy is uncommon, and flow cytometry was less accurate than histology/immunohistochemistry for its diagnosis. Despite the rarity of tonsillar lymphoma in children, nearly half of study patients were pediatric. Intraoperative frozen section diagnosis showed excellent sensitivity for malignancy, and could be used to effectively triage cases for flow cytometry evaluation.


Assuntos
Citometria de Fluxo/estatística & dados numéricos , Neoplasias Hematológicas/patologia , Tonsila Palatina/patologia , Doenças Raras/patologia , Neoplasias Tonsilares/patologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biópsia , Criança , Pré-Escolar , Feminino , Citometria de Fluxo/economia , Secções Congeladas , Neoplasias Hematológicas/cirurgia , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Tonsila Palatina/cirurgia , Doenças Raras/cirurgia , Estudos Retrospectivos , Neoplasias Tonsilares/cirurgia , Tonsilectomia , Adulto Jovem
3.
Am J Hematol ; 89(10): 978-84, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25042070

RESUMO

Flow cytometry (FCM) is an adjunct study to routine analysis of cerebrospinal fluid (CSF) to investigate for involvement by a hematologic malignancy. However, in our experience, FCM only infrequently detects abnormalities in CSF. To help optimize resources without forfeiting clinically important data, we sought to determine evidence-based indications and criteria for performing FCM on CSF. FCM results of 316 consecutive CSF specimens were retrospectively reviewed and correlated with clinical history, total nucleated cell (TNC) counts, and results of concurrent cytologic review. Of 255 samples adequate for analysis, 54% were from patients with a prior history of hematologic malignancy, of which 12% (17 cases) were abnormal by FCM. Corresponding TNC counts among samples with abnormal FCM ranged from 0-1050 cells/µL, and only 44% showed abnormal morphology on concurrent cytology. Of the remaining 46% of samples from patients with no known history of hematologic malignancy who had CSF sampling for neurological indications, only one (1%) was abnormal by FCM. This specimen had an elevated TNC count (39 cells/µL) but lacked clearly abnormal findings on concurrent cytology. These results support the use of CSF FCM only in patients with a history of hematologic malignancy or, in the absence of such a history, in samples showing pleocytosis. If these criteria were applied to the current cohort using a TNC count cut-off of >5 cells/µL, 23% of samples would have been deferred from testing, resulting in decreased cost, improved efficiency, and reduction in the need for unnecessary testing without a negative impact on clinical care.


Assuntos
Citometria de Fluxo , Neoplasias Hematológicas/líquido cefalorraquidiano , Neoplasias Hematológicas/diagnóstico , Contagem de Células/métodos , Feminino , Humanos , Masculino , Estudos Retrospectivos
4.
J Bone Miner Res ; 29(6): 1380-6, 2014 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-24420643

RESUMO

Cells of the osteoblast lineage play an important role in regulating the hematopoietic stem cell (HSC) niche and early B-cell development in animal models, perhaps via parathyroid hormone (PTH)-dependent mechanisms. There are few human clinical studies investigating this phenomenon. We studied the impact of long-term daily teriparatide (PTH 1-34) treatment on cells of the hematopoietic lineage in postmenopausal women. Twenty-three postmenopausal women at high risk of fracture received teriparatide 20 mcg sc daily for 24 months as part of a prospective longitudinal trial. Whole blood measurements were obtained at baseline, 3, 6, 12, and 18 months. Flow cytometry was performed to identify hematopoietic subpopulations, including HSCs (CD34+/CD45(moderate); ISHAGE protocol) and early transitional B cells (CD19+, CD27-, IgD+, CD24[hi], CD38[hi]). Serial measurements of spine and hip bone mineral density (BMD) as well as serum P1NP, osteocalcin, and CTX were also performed. The average age of study subjects was 64 ± 5 years. We found that teriparatide treatment led to an early increase in circulating HSC number of 40% ± 14% (p = 0.004) by month 3, which persisted to month 18 before returning to near baseline by 24 months. There were no significant changes in transitional B cells or total B cells over the course of the study period. In addition, there were no differences in complete blood count profiles as quantified by standard automated flow cytometry. Interestingly, the peak increase in HSC number was inversely associated with increases in bone markers and spine BMD. Daily teriparatide treatment for osteoporosis increases circulating HSCs by 3 to 6 months in postmenopausal women. This may represent a proliferation of marrow HSCs or increased peripheral HSC mobilization. This clinical study establishes the importance of PTH in the regulation of the HSC niche within humans. © 2014 American Society for Bone and Mineral Research.


Assuntos
Células-Tronco Hematopoéticas/citologia , Pós-Menopausa/efeitos dos fármacos , Teriparatida/farmacologia , Linfócitos B/citologia , Linfócitos B/efeitos dos fármacos , Biomarcadores/metabolismo , Densidade Óssea/efeitos dos fármacos , Osso e Ossos/efeitos dos fármacos , Osso e Ossos/metabolismo , Contagem de Células , Linhagem da Célula/efeitos dos fármacos , Feminino , Células-Tronco Hematopoéticas/efeitos dos fármacos , Humanos , Pessoa de Meia-Idade , Linfócitos T/citologia , Linfócitos T/efeitos dos fármacos
6.
Cytometry B Clin Cytom ; 84(3): 143-8, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23341189

RESUMO

BACKGROUND: B-cell lymphomas with concurrent translocations of MYC and BCL2 or BCL6, also known as "double-hit" lymphomas (DHL), are rare malignancies characterized by aggressive clinical behavior and poor prognosis. Previous reports suggest that decreased CD20 and/or CD19 expression by flow cytometry is relatively common in DHL and may help to identify cases requiring additional cytogenetic analysis. METHODS: We conducted a retrospective analysis of 26 cases of DHL, and compared their flow cytometric characteristics to cases of Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL). Cases were analyzed by four-color flow cytometry, and bivariate dot-plots were reviewed for light scatter characteristics, CD19, CD20, CD45, and surface light chain. RESULTS: Relatively few DHL cases showed dim expression of CD19 or CD20, and statistically significant differences were found only in the frequency of dim CD19 expression between DHL and BL or DLBCL. Although concomitant dim CD19 and CD20 expression was exclusive to DHL, it was present in only a minority of cases. CONCLUSIONS: We conclude that although a subset of DHL expresses aberrant levels of CD19 and/or CD20 by flow cytometry, these findings are of limited utility in identifying cases requiring cytogenetic analysis due to their low frequency. Until more sensitive pathologic parameters can be identified and validated, the decision to perform cytogenetic analysis should rest on a combination of clinical, morphologic, and immunophenotypic features suggestive of high-grade, aggressive disease.


Assuntos
Linfoma de Burkitt/diagnóstico , Proteínas de Ligação a DNA/análise , Citometria de Fluxo/normas , Linfoma Difuso de Grandes Células B/diagnóstico , Proteínas Proto-Oncogênicas c-bcl-2/análise , Proteínas Proto-Oncogênicas c-myc/análise , Idoso , Antígenos CD19/análise , Antígenos CD19/genética , Antígenos CD20/análise , Antígenos CD20/genética , Linfoma de Burkitt/genética , Análise Citogenética , Proteínas de Ligação a DNA/genética , Diagnóstico Diferencial , Feminino , Expressão Gênica , Humanos , Imunofenotipagem , Linfoma Difuso de Grandes Células B/genética , Masculino , Pessoa de Meia-Idade , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-6 , Proteínas Proto-Oncogênicas c-myc/genética , Estudos Retrospectivos
7.
Transfusion ; 53(8): 1763-71, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23145947

RESUMO

BACKGROUND: Although platelets (PLTs) are known to express ABH antigens, the extent of expression is different on PLTs compared with red blood cells and the relationship between PLT ABH expression and genotype has not been thoroughly investigated. STUDY DESIGN AND METHODS: We measured blood group H and A antigens on PLTs from 100 normal volunteers using fluorescent-conjugated reagents and flow cytometry. Individuals were also genotyped at the ABO locus using a commercially available genotyping system. RESULTS: Expression of A and H antigen varied widely on PLTs from different individuals. Among group A and AB persons, H antigen expression was significantly greater than A antigen (p < 0.0001). The ratio of H-to-A antigen expression varied more than 100-fold in a predictable fashion according to genotype with values lowest in A1/A1 < A1/O < A2/A2 < A2/O. H and A antigen expression was unaffected by secretor status. The proportion of PLTs with high A expression also varied directly according to genotype. CONCLUSIONS: Blood group A and H antigen expression on PLTs varies in a predictable fashion according to genotype. Flow cytometry and genotyping identify individuals who strongly express A antigens, a finding that may be relevant to clinical PLT transfusion across ABO barriers.


Assuntos
Sistema do Grupo Sanguíneo ABO/genética , Plaquetas/imunologia , Genótipo , Sistema do Grupo Sanguíneo ABO/metabolismo , Biomarcadores/sangue , Plaquetas/metabolismo , Citometria de Fluxo , Técnicas de Genotipagem , Humanos , Fenótipo
8.
J Infect Dis ; 204(8): 1227-36, 2011 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-21917896

RESUMO

BACKGROUND: Pro-inflammatory monocytes/macrophages may contribute to increased atherosclerosis in human immunodeficiency virus (HIV)-infected patients. We investigate--to our knowledge, for the first time--sCD163 and other markers of monocyte activation in relationship to atherosclerotic plaque in HIV-infected patients. METHODS: One hundred two HIV-infected and 41 HIV-seronegative men with equivalent cardiovascular risk factors and without history of coronary artery disease were prospectively recruited and underwent computed tomography coronary angiography. RESULTS: sCD163 levels and presence of plaque were significantly higher among antiretroviral-treated subjects with undetectable HIV RNA levels, compared with seronegative controls (1172 ± 646 vs. 883 ± 561 ng/mL [P = .02] for sCD163 and 61% vs. 39% [P = .03] for presence of plaque). After adjusting for age, race, lipids, blood pressure, glucose, smoking, sCD14, and HIV infection, sCD163 remained independently associated with noncalcified plaque (P = .008). Among HIV-infected patients, sCD163 was associated with coronary segments with noncalcified plaque (r = 0.21; P = .04), but not with calcium score. In contrast, markers of generalized inflammation, including C-reactive protein level, and D-dimer were not associated with sCD163 or plaque among HIV-infected patients. CONCLUSIONS: sCD163, a monocyte/macrophage activation marker, is increased in association with noncalcified coronary plaque in men with chronic HIV infection and low or undetectable viremia. These data suggest a potentially important role of chronic monocyte/macrophage activation in the development of noncalcified vulnerable plaque. CLINICAL TRIAL REGISTRATION: NCT00455793.


Assuntos
Antígenos CD/imunologia , Antígenos de Diferenciação Mielomonocítica/imunologia , Infecções por HIV/imunologia , HIV/imunologia , Ativação de Macrófagos/imunologia , Macrófagos/imunologia , Placa Aterosclerótica/virologia , Receptores de Superfície Celular/imunologia , Adolescente , Adulto , Fármacos Anti-HIV/efeitos adversos , Fármacos Anti-HIV/uso terapêutico , Antígenos CD/sangue , Antígenos de Diferenciação Mielomonocítica/sangue , Quimiocina CCL2/sangue , Quimiocina CCL2/imunologia , Angiografia Coronária , Citometria de Fluxo , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Humanos , Interleucina-6/sangue , Interleucina-6/imunologia , Receptores de Lipopolissacarídeos/sangue , Receptores de Lipopolissacarídeos/imunologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Osteopontina/sangue , Osteopontina/imunologia , Placa Aterosclerótica/imunologia , Estudos Prospectivos , Receptores de Superfície Celular/sangue , Estatísticas não Paramétricas , Adulto Jovem
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