A quality improvement assessment of multiple, concurrent flow cytometry analyses at a tertiary care center.

INTRODUCTION: The utility of flow cytometry (FC) in diagnosis and staging of hematologic malignancy is controversial. Often, multiple specimens from the same patient are processed concurrently for FC analyses, alongside tissue for histomorphologic diagnosis. METHODS: To assess the diagnostic utility of multiple, concurrent FC analyses, a 10-year retrospective review of cases with ≥2 concurrent specimens (from the same patient) submitted for FC was conducted. Light microscopic (LM) diagnoses were compared to FC findings, and the contribution of FC results to final diagnoses was examined. RESULTS: Of 4058 specimens (predominantly lymph nodes, bone marrows, and oropharyngeal tissues) submitted for FC analyses, 129 (3.2%) represented cases with multiple (average: 2.19) concurrent FC analyses. All were accompanied by tissues and/or aspirates for LM examination. In 115 (89.1%) cases, multiple FC analyses were performed prior to morphologic examination. In 87.0% of those cases, ≥1 FC result(s) aligned with LM findings. In 15 (13.0%) cases where FC results differed from morphologic diagnoses, 86.7% (13/15) failed to detect an abnormal cell population by FC in the presence of a hematologic malignancy by LM. In one case (0.9%), FC detected a lymphoma, without morphologic evidence by LM. CONCLUSIONS: Overall, multipart FC failed to demonstrate a significant contribution in initial diagnoses of hematologic malignancies compared with analysis of a single specimen.

Deafferentation-induced caspase-3 activation and DNA fragmentation in chick cochlear nucleus neurons.

Cochlea removal severs peripheral processes of cochlear ganglion cells and permanently abolishes afferent input to nucleus magnocellularis (NM) neurons. Deafferented chick NM neurons undergo a series of morphologic and metabolic changes, which ultimately trigger the death of 20%-40% of neurons. Previous studies suggested that this cell specific death involves activation of the intrinsic apoptotic pathway, including increased presence of cytochrome c and active caspase-9 in the cytoplasm of deafferented NM neurons. Interestingly, however, both markers were detected pan-neuronally, in both degenerating and surviving NM neurons [Wilkinson BL, Elam JS, Fadool DA, Hyson RL (2003) Afferent regulation of cytochrome-c and active caspase-9 in the avian cochlear nucleus. Neuroscience 120:1071-1079]. Here, we provide evidence for the increased appearance of late apoptotic indicators and describe novel characteristics of cell death in deafferented auditory neurons. Young broiler chickens were subjected to unilateral cochlea removal, and brainstem sections through NM were reacted for active caspase-3 and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL). Caspase-3 activation is observed in the cytoplasm of both dying and surviving deafferented NM neurons 24 h to 7 days following cochlea removal, suggesting that caspase-3, usually considered an "executioner" of apoptotic death, may also function as a "modulator" of death. In addition, we find that TUNEL labeling of degraded DNA is observed in deafferented NM. In contrast to upstream apoptotic markers, however, TUNEL labeling is restricted to a subpopulation of deafferented neurons. Twelve hours following cochlea removal, TUNEL labeling is observed as punctate accumulations within nuclei. Twenty-four hours following cochlea removal, TUNEL accumulates diffusely throughout neuronal cytoplasm in those neurons likely to die. This cytoplasmic TUNEL labeling may implicate mitochondrial nucleic acid degradation in the selective death of some deafferented NM neurons. Our study examines the subcellular distributions of two prominent apoptotic mediators, active caspase-3 and TUNEL, relative to known histochemical markers, in deafferented NM; provides new insight into the apoptotic mechanism of cell death; and proposes a role for mitochondrial DNA in deafferentation-induced cell death.