196321: 509-512

DOROTHEA ZUCKER-FRANKLIN

Lymphocytic Leukemia
Virus-like Particles in the Lymphocytes of a Patient with Chronic

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Copyright 2011 by The American Society of Hematology; all rights reserved.
Hematology, 2021 L St, NW, Suite 900, Washington DC 20036.
Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly by the American Society of
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Virus-like Particles in the Lymphocytes of a Patient
with Chronic Lymphocytic Leukemia
By DOROTHEA ZUCKER-FRANKLIN
I HE VIRAL ETIOLOGY of avian leukemia has been recognized for more
than 40 years while that of several types of murine leukemia has been
established within the last decade. So far, little evidence has been forth-
coming to suggest that similar agents may play a role in the human counter-
part of the disease. Virus-like particles have been described in vacuoles with-
in the cells of a few cases of acute human leukemia.5 However, such parti-
des have never been seen directly in the cytoplasm of blood cells freshly
removed from a human host afflicted with diseases of either neoplastic or
infectious nature. For this reason, the following observations are reported.
Lymphocytes were isolated on three separate occasions from the peripheral
blood of a patient with chronic lymphocytic leukemia. The specimens were
obtained at monthly intervals during which the white cell count decreased
from 240,000 to 8,000 without therapy. The diagnosis had been established
about 20 years previously, and the patient received intermittent splenic
radiation and Chiorambusil but was given no treatment during the 2 years
preceding the collection of specimens. W ithin 20 minutes of their isolation,
the lymphocytes were fixed in 1 per cent isotonic osmium tetroxide. They
were dehydrated in increasing concentrations of alcohol and embedded in
Epon 812.6 Thin sections were stained with isranyl acetate and/or lead
hydroxide and examined in a Siemens Elmiskop 1 electron microscope. About
half the cells exhibited some cytopathic changes consisting of clumped, un-
evenly distributed nuclear protein, an increase in nucleolar masses, and an
increase in the number of small vacuoles and RNP particles in the cytoplasm.
In 15-20 per cent of the cells, parallel rows of particles were observed in the
cytoplasm ( fig. la ) . Each row of particles was separated from the adjacent
one by a membrane about 80 A in diameter and several j in length. The center
rows of such particle aggregates were often indistinct or missing, suggesting
a cylindrical or tube-like structure sectioned longitudinally. This impression
was supported by the appearance of the aggregates in the cross-section il-
lustrated in figure ic. Here the particles are seen arranged in concentric
circles. Each circle of particles was again separated from the next by a
membrane. The central core consisted of a membrane-surrounded vacuole
which, at times, also contained electron-dense particles of varying size. Fila-
ments running at right angles to the long membranes separating individual
From the Department of Anatomy, New York University School of M edicine, New
York City.
The investigation was supported by Grant 2G-297 from the National Institutes of Health.
The support of Dr. I. Rhodin, in whose department these studies were carried out, is
gratefully acknowledged.
Submitted July 27, 1962; accepted for publication Sept. 10, 1962.
509
BLOOD, VOL. 21, No. 4 (APRIL), 1963
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DOROTHEA ZUCKER-FRANKLIN 510
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For personal use only. on May 17, 2014. by guest bloodjournal.hematologylibrary.org From
VUIUS-LIKE PARTICLES I N L Y M PH O C Y T E S 511
particles were also resolved on occasion ( see model fig. lb ) . In addition to
these organized arrays, particles have been observed lying singly or in groups
dispersed throughout the cytoplasm. In this case they resembled RNP particles,
though at high magnification budding from cytoplasmic filaments was sug-
gested.
The exact structure of the particles has not yet been determined and awaits
their isolation in purified form. They are most likely spherical with an irregular
outline, and a diameter of 80 to 120 A . It has not yet been possible to resolve
the particle aggregates with the light microscope utilizing Wrights and
Giemsa, PAS, Sudan Black, or Feulgen stains. Phase microscopy of living or
Epon-embedded sectioned cells also yielded negative results. Therefore, an
estimate of the percentage of cells involved is difficult at this time. It is also
too early to identify the particles with any known virus or to attribute to
them an etiologic role in the patients lymphocytic leukemia. The longitudinal
section of the particle aggregates resemble those published by Rifkind et al.7
of Echo 9 virus-infected tissue culture cells. The patient did not show clinical
evidence of such an infection and viral agglutination studies, kindly performed
by Dr. Millian at the New York City Board of Health, were negative. Tissue
culture studies are still in progress.
The electron-microscopic appearance of the particle aggregates also bore no
resemblance to the paracrystalline structures or sticks reported by Bernard
et al.8 in the cells of a patient with long-standing lymphocytic leukemia. The
authors suggested that the structures, which were fibrillar, represented an ab-
normal secretion of the cell. It is quite conceivable that the arrays of particles
reported here could also be the product of abnormal cellular metabolism.
However, intercurrent viral infections in patients with chronic lymphocytic
leukemia are common, and the role of the lymphocyte in such infections is
still obscure. It is noteworthy that the particles were not observed in any
peripheral blood cells other than lymphocytes. Though the observation tempts
to speculation, further interpretation is not warranted until the chemical and
biological nature of the isolated particles has been unequivocally established.
SUMMARY
Lymphocytes from the peripheral blood of a patient with chronic lympho-
cytic leukemia were isolated and subjected to electron microscopy. Aggregates
of virus-like particles measuring 80 to 120 A in diameter were observed as
organized structures in the cytoplasm of the cells. The true nature of the
particles remains to be established.
SuMM .mo IN INTERLINGUA
Lymphocytos ab le sanguine peripheric de un patiente con chronic leucemia
lymphocytic esseva isolate e subjicite a microscopia electronic. Aggregatos
de particulas virus-simile de 80 a 120 A in diametro esseva observate como
organisate structuras in le cytoplasma del cellulas. Le ver natura del particulas
remane a establir.
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512 DO R O T H E A z u C K E R -FR A N K L I N
REFERENCES
1 . Dmochowski, L., and Grey, C. E.: Sub-
cellular structures of possibile viral
origin in some mammalian tumors.
Ann. New York Acad. Sc. 68:559,
1957.
2. Beard, J. W. : Etiology of avian leukosis.
Ann. New York Acad. Sc. 68:473,
1957.
3 . Oberling, C., and Chevallier, P. : Sur 1-
origine infectieuse des leuc#{ 233} mies.
Proc. 3rd Internat. Soc. Haematol.,
1950, p. 241.
4. Dmochowski, L., and Grey, C. E. : Stud-
ies on submicroscopic structure of leu-
kemias of known or suspected viral
origin: A review. Blood 13: 1017, 1958.
5. -: Viruses & tumors in the light of
electron microscope studies : A review.
Cancer Res. 20:977, 1960.
6. Li ft, J. H. : Improvements in epoxy resin
embedding methods. J. Biophys. Bio-
chem. Cytol. 9:409, 1961.
7. Rifkind, R. A.. Godman, G. C., Howe,
C., Morgan, C., and Rose, M. M.:
Echo 9 virus in tissue culture observed
by light and electron microscopy.
Virology 12:331, 1960.
8. Bernard, J., Bessis, M., Soulier, J. P., and
Thiery, J. P. : Etude au microscope
#{ 233} lectronique dune leuc#{ 233} mie lymphoide
avec crystaux intracytoplasmique. Rev.
h#{ 233} mat. 14:227, 1959.
Dor othea Zucker -Fr ankl i n, M.D., Assi stant Attendi ng Physi -
ci an, Medi cal Di vLsi an (Hematol ogy), Montefi or e Hospi tal ;
Fel l ow i n El ectr onmi cr oscopy, Depar tmei U of Anatomy, New
Yor k Uni ver si ty School of Medi ci ne, New Yor k, N. Y.
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