Electron Microscopic Study of the Mouse Leukemia Virus ... · C3Hf Parents1 DOROTHY G. FELDMAN,...

ICANCER RESEARCH 27 Part 1, 1792-1804, October 1967]

Electron Microscopic Study of the Mouse Leukemia Virus (Gross) inOrgans of Mouse Embryos from Virus-injected and NormalC3Hf Parents1

DOROTHY G. FELDMAN, YOLANDE DREYFUSS,AND LUDWIK GROSS

From the Cancer Research Unit, Veterans Administration Hospital, Bronx, New York 10468

SUMMARY

Ultrathin sections of thymus, spleen, liver, bone marrow, andkidney from embryos of 10 virus-injected and 9 normal nonin-jected C3Hf female mice were examined in the electron microscope. Leukemia virus particles were present in small numbersin thymus, spleen, liver, and bone marrow of embryos from bothvirus-injected and normal control parents. They appeared inapproximately the same amounts and with similar frequencyregardless of whether the embryos were removed from virus-injected or from normal control female mice. Particles were observed budding from lymphocytes, lymphoblasts, and epithelialcells in thymus, from erythroblasts and hemocytoblasts in liverand spleen, and from hemocytoblasts in bone marrow. The virusparticles most frequently observed were either budding or of thedoughnut-type;2 particles containing nucleoids2 appeared in only

one specimen of embryo thymus from normal noninjected parents. Leukemia virus particles were not observed in kidney tissuesfrom any of the embryos examined.

Examination of embryos from 4 Ak mice and from a normalnoninjected BALB/c mouse revealed the presence of virus particles in thymus, spleen, and liver tissues. Thus far, particles werenot observed in organs from embryos of 3 virus-injected or 3normal noninjected Sprague-Dawley rats.

INTRODUCTION

The mouse leukemia virus is transmitted under natural lifeconditions from one generation to another directly through theembryos. This was first demonstrated by Gross (10), and later

'Aided in part by grants from the Damon Runyon MemorialFund and the American Cancer Society.

2We have preferred using descriptive terms indicating the morphology of observed virus particles rather than arbitrary designation by alphabet letters. According to classification terminologysuggested by W. Bernhard (The Detection and Study of TumorViruses with the Electron Microscope. Cancer Res, 20: 712-727,1960), and more recently modified (Classification of OncogenicUNA Viruses. J. Nati. Cancer Inst., 37: 395-397, 19GG),the term"doughnut-type" particle employed in our study corresponds to"A" particle or "immature C" particle, and the term "particle-containing nucleoid" employed in our study corresponds to"mature C" particle.

Received February 9, 1967;accepted May 29, 1967.

extended in additional studies and discussed in subsequent publications (11, 14-16).

Since the mouse leukemia virus is transmitted through theembryos, it was of interest to examine electron microscopicallyorgans of mouse embryos presumably carrying the virus, for thepresence of characteristic virus particles. Previous electron microscopic studies reported the appearance of virus particles in embryos of Ak strain mice naturally infected with the mouse leukemia virus (4). Experiments reported in this study revealed thepresence of particles having the typical morphology of the mouseleukemia virus in organs of embryos from high-leukemic strainAk mice, as well as in embryos of strain C3Hf mice carrying themouse leukemia virus (Gross) as a result of experimental inoculation. However, rather unexpectedly, similar particles in comparable numbers were also found in a control series in normalembryos removed from healthy, noninjected C3Hf mice

MATERIALS AND METHODS

Animals

Embryos Removed from Virus-injected C3Hf Mice. Sixfemale mice of strain C3Hf, raised in our laboratory, were inoculated intra]Â¡eritoneally when less than 6 days old with passageA mouse leukemia (Gross) virus filtrate. When 3-4 weeks old, theinjected female mice were mated to their brothers. Five of the 6virus-injected females were mated to their virus-injected brothers.Four of these females developed symptoms of leukemia at thetime they were sacrificed at 2.5-3 months of age, and one had nosymptoms of leukemia when sacrificed at 2 months of age. Onevirus-injected female was mated to a noninjected brother and wasleukemic when sacrificed at 2.5 months of age. In addition, 4C3Hf females were inoculated with the virus filtrate at the age of3-5 weeks and were mated to their virus-injected brothers about2 weeks after virus inoculation. The virus-injected females in thisgroup had no symptoms of leukemia when they were sacrificedat 2.5-3 months of age. All injected females were sacrificed during

midpregnancy, or close to term, and their embryos were removedfor electron microscopic studies.

Embryos Removed from Ak Females. Four Ak femalesfrom our colony were mated to their brothers; they were sacrificedwhen pregnant, at ages varying from 2.5 to 6 months; their embryos were removed and used for electron microscopic studies.

Embryos Removed from Normal Control C3Hf Mice. Ina control series, 9 C3Hf female mice were mated to their brothers

1792 CANCER RESEARCH VOL. 27

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Leukemia Virus Particles in Mouse Embryos

as soon as they reached sexual maturity; none of these mice wasinjected with the mouse leukemia virus. The females were in goodhealth when they were sacrificed at the age of 2.5-5.5 months

during midpregnancy or close to term; their embryos were removed and used for electron microscopic studies.

Methods

Removal of Organs from Embryos. The pregnant femalemice were sacrificed by ether inhalation. The embryos were removed; fragments of thymus, liver, spleen, bone marrow,3 and

kidney tissues were then carefully excised under low magnification.4

Tissue fragments were also obtained from comparable organsof the pregnant female mice from which the embryos were removed in order to compare the results obtained in the study oÃembryo organs with those of their mothers.

Processing of Specimen for Electron Microscopy. Thetissue fragments were immediately fixed in 1% phosphate-buffered osmic acid on ice for 1-1.5 hours. The specimens were dehydrated in successive changes of 50-100';, ethanol, immersed in

propylene oxide, and embedded in Epon. Tissues were sectionedwith a diamond knife using a Porter-Blum microtome convertedinto a thermal advance model and placed on uncoated 300 meshcopper grids. Sections were lightly coated with carbon, stainedwith uranyl acetate and lead hydroxide (5, 18), and examinedin an RCA EMU-3F, or more recently in an RCA EMU-3Gelectron microscope at 50 kv.

RESULTS

Presence of Leukemia Virus Particles in Various Organs

of Embryos Examined

Leukemia virus particles appeared in thymus, spleen, liver,and bone marrow of embryos from virus-injected parents, andnormal noninjected parents with a similar high degree of frequency (Table 1). Particles were present in thymus of embryosfrom all 10 virus-injected mice and from 8 normal noninjectedmice. The spleen of embryos from 4 out of 5 virus-injected miceand from 8 out of 9 noninjected mice contained particles. Particles appeared in liver of embryos from all 7 virus-injected miceand from 4 out of 6 noninjeeted mice. Examination of embryobone marrow revealed the presence of particles in specimensfrom 1 out of 2 virus-injected mice, and from all 3 normal non-injected mice. The frequency of particles observed in embryothymus, spleen, liver, and bone marrow was apparently not

3Bone marrow of the embryos was not a solid mass of hemato-poietic cells which conici be readily dissected out for electronmicroscopy, but consisted rather of a bone marrow cavity containing cartilaginous and bone elements interspersed with mesen-chymal and blood cells. Therefore, the entire embryo femur wasexcised, processed, embedded, and sectioned. The sections werethen scanned in the electron microscope for foci of hematopoieticcells.

4Because several of the pregnant female mice were not sacrificed

close to term, some of the embryo organs were too small to bereadily located. Therefore, not all organs from every embryostudied were removed for electron microscopic examination.

influenced by whether or not the mother was virus-injected non-leukemic, leukemic, or normal noninjected when sacrificed. Noleukemia virus particles were found in any of the embryo kidneys examined.

Quantity and Type of Leukemia Virus Particles

Leukemia virus particles were found in approximately thesame quantity in thymus, spleen, liver, and bone marrow fromembryos of virus-injected parents and normal noninjected parents(Table 1). Budding and doughnut-type particles were the formsmost frequently observed in the specimens examined. Whenpresent, budding and doughnut-type particles were few in number. Particles containing nucleoids appeared in only one specimen examined, in embryo thymus from normal noninjected parents. In this sample, the particles with nucleoids occurred inmoderate numbers.

The morphology and size of the budding, doughnut-type particles, and particles containing nucleoids present in C3Hf embryoswere similar to particles described in C3Hf adult mice with passage A virus-induced leukemia (1, 3, 6, 17). In addition, smallerdoughnut-type particles5 previously described (6) within the

endoplasmic reticulum of cells from normal noninjected andvirus-injected C3Hf mice and from Ak mice, were found insmall numbers in thymus, spleen, liver, bone marrow, and kidneyfrom embryos of virus-injected and noninjected normal C3Hfparents.

Cells from which Leukemia Virus Particles Form

The cells from which leukemia virus particles appeared tobud were the same in tissues from embryos of virus-injected andnormal noninjected parents. Particles formed from lymphocytes,lymphoblasts, and epithelial cells in the thymus, from erythro-blasts and hemocytoblasts in spleen and liver, and from hemo-cytoblasts in bone marrow.

Thymus. The embryo thymuses studied appeared to becomposed of lymphoblasts (Lb), and lymphocytes (Lc), andepithelial cells (E) (Figs. 1-7). Leukemia virus particles were

observed budding from the cell membrane of lymphocytes andlymphoblasts, and from vacuolar membranes of epithelialcells. Figs. 1-7 are electron micrographs of embryo thymus,Fig. 1 from virus-injected parents and Figs. 2-7 from normalnoninjected parents. Higher magnifications of the outlined areasof Figs. 1 and 2 (Figs, lo, 2a) illustrate virus particles (arrows)budding from a lymphoblast (Fig. lo) and a lymphocyte (Fig.2a). Low-power micrographs of epithelial cells demonstratingthe presence of vacuoles (v) appear in Figs. 3 and 4. Enlargements of the outlined areas of Fig. 3 (Fig. 3a, 6), and Fig. 5illustrate virus particles (arrows) budding from vacuolar membranes. Doughnut-type particles (d) and particles with nucleoids (n) lying free within vacuoles are shown in Figs. 3o, 4, 6,and 7.

Spleen. The structure of the embryo spleens studied consisted

s"Smaller doughnut-type" particles have been designated"intracysternal A" particles according to classification termi

nology recently suggested (Classification of Oncogenic UNAViruses. J. Nati. Cancer Inst.,37: 395-397, 1900).

OCTOBER 1967 17Ã•M



Dorothy G. Feldman, Yolande Dreyfuss, and Ludwik Gross

TABLE 1Electron Microscopic Study of Embryos from Virus-injected and Normal Noninjected CSHf Parents

MouseNo.Virus-injected

leuke-micmothers1234'5Virus-injected

non-leukemicmothers678!Â»10Normal

non-injectedmothers111213141516171819EmbryosThymusE++++++++++++++++++"+SpleenE+++0++++0+++++LiverE+++++++0+0+++Bone

marrowE0++++KidneyE000000000MothersThymusL"riiiiNNNNNE+

+ +'+

+ +d+

+d+

++''++"-H+

*+

"++

+<*+

<+++000++SpleenLIIIINNNNNNNNNNNNNNNE+

+"+

"+

+"+

"+

+"++

d0+d++d000000000LiverLNNIINNNNNNNNNNNNNNNE00++d000000000000000Bone

marrowLINNNE+

+"+

+"+

+<<0+0+"0+000000KidneyLNINNNNNNNNNNNNNE0+*00000000000000

" E, electron microscopy: approximate average number of particles per grid square using 300 mesh grids: 0, no particles observed;+ , less than 5 (few); + + , 5-10 (moderate); + + + , more than 10 (abundant).

bL, light microscopy; I, infiltration; N, no infiltration.eOnly the mother received injection.dParticles containing nucleoids observed.

of a meshwork of mesenchyme within which foci of erythropoiesisand myelopoiesis were present; lymphoid cells were not evident.Fiji. 8 is a low-power electron micrograph demonstrating matureand immature granulocytes (G), hemocytoblast (H), reticulo-cyte (ffi), and mesenchymal cells (.I/). In Fig. 9 groups of platelets (P) and an erythroblast (Eb) appear, and in Fig. 10 anarea of mesenchymal cells (M), and collagen (C) near the periphery (lower right) of the spleen are shown. Virus particleswere found budding from erythroblasts (arrows, Figs. 11, 13;normal noninjeeted and virus-injected parents, respectively), andfrom hemocytoblasts as shown in Fig. 12a (arrow, higher magnification of outlined area of Fig. 12, virus-injected parents)and Fig. 14 (arrow, normal noninjected parents).

Liver. Besides parenchyma! and endothelial cells present inadult liver, embryo livers studied also contained various hema-topoietic cells. Figs. 15-18, and Fig. 20 are low-magnificationelectron micrographs of embryo liver illustrating areas of liverparenchyma (L, Fig. 17), hemocytoblasts (//, Figs. 17, 20),erythroblasts (Eb, Figs. 15, 17, 18, 20), granulocytes (G, Fig.

15), and megakaryocytes (.17, Fig. 16). As in embryo spleen,virus particles were observed budding from hemocytoblasts(right arrow, Fig. 20a, enlargement of outlined area of Fig. 20,virus-injected parents) and erythroblasts (left arrow, Fig. 20o;arrow, Fig. 19, virus-injected parents). A doughnut-type particle(d) proximal to the cell membrane of an erythroblast in liver ofembryo from normal noninjected parents appears in Fig. 18a(enlargement of outlined area oÃFig. 18).

Bone Marrow. Areas of hematopoietic cells (Figs. 21-23)consisted of mature and immature granulocytes (G), megakaryo-blasts (.1/6), hemocytoblasts (H), and occasionally erythroblastswithin a reticulum of mesenchymal cells (M). Thus far, in embryo bone marrow, budding of particles was observed from thecell membrane of hemocytoblasts only (arrow, Fig. 22, virus-injected parents; arrow, Fig. 23a, enlargement of outlined areaof Fig. 23, normal noninjected parents).

Kidney. No typical leukemia virus particles were evident inany of the kidney specimens studied from the embryos of virus-injected or normal noninjected parents. However, smaller




Leukemia Virus Particles in Mouse Embryos

doughnut-type particles, previously described (6), were present

in a sample of kidney of embryo from normal parents (arrow,Fig. 24a, enlargement of Fig. 24).

Comparison of Quantity and Type of Leukemia VirusParticles in Embryos and Their Mothers

Virus-Injected Mothers. The quantity and type of leukemiavirus particles found in tissues of the virus-injected female micefrom which the embryos were removed had no apparent influence upon the quantity and type of particles observed in theirrespective embryos. Thymus, spleen, and bone marrow fromvirus-injected leukemic mothers generally contained a moderateto abundant number of virus particles (Table 1). Also in eachspecimen examined, particles with nucleoids were present.

In thymus, spleen, and bone marrow from virus-injected non-leukemic mothers, few to moderate numbers of virus particlesand occasionally no particles were observed (Table 1). Particleswith nucleoids often appeared, but not as frequently as in comparable organs from leukemic mothers.

In thymus, spleen, and bone marrow from embryos of bothvirus-injected nonleukemic and leukemic mothers, virus particles api>eared with about the same frequency as in comparableorgans from the mothers. However, when particles were observedin the embryo, they were few in number. In addition, only budding and doughnut-type particles were present; no particles withnuc'leoids were thus far observed. The relatively larger numberof virus particles present in leukemic mothers than in virus-injected nonleukemic mothers apparently did not result in anincrease in the number of particles observed in embryos fromleukemic mothers.

No leukemia virus particles were observed in liver from virus-injected nonleukemic mothers, while a few particles appeared inliver from 2 out of 5 leukemic mothers. In one of these specimens,particles with nucleoids were found. In both samples, virusparticles appeared to bud from infiltrated cells, and in one, particles were also observed budding from liver parenchymal cellsand Kupffer cells.

However, in embryos from virus-injected parents, a fewdoughnut-type particles and particles budding from erythro-blasts and hemocytoblasts in zones of hematopoiesis were observed in every specimen of liver examined.

Among the kidneys examined from virus-injected mothers, afew virus particles appeared in one specimen from a leukemicmother (Table 1). However, no virus particles were observed inkidney of embryos from virus-injected mothers.

Normal Noiiinjected Parents. Leukemia virus particlesappeared with slightly greater frequency in thymus from normalembryos than from normal noninjected mothers. In all 8 embryothymuses examined virus particles were present, whereas thymusfrom 6 out of 9 mothers contained particles (Table 1). Whenvirus particles appeared in thymus from embryos or their mothers, they were few in number, except for one embryo thymuswhich contained a moderate number of particles. In addition,the particles observed were generally either budding or doughnut-type; particles containing nucleoids appeared in thymus fromone embryo and one mother only.

Of the specimens of liver, spleen, and bone marrow examinedfrom noninjected mothers, 1 or 2 budding and doughnut-type

particles were found in one sample of bone marrow; virus particles were not observed in liver and spleen. However, in the embryos from these mothers, particles frequently api>carcd in thespecimens of spleen, liver, and bone marrow examined.

No virus particles were observed in kidney tissue from eithernormal mothers or their embryos.

Ak Mouse Embryos. Ak embryos from 4 pregnant nonleukemic females were also studied. Examination of thymus, spleen,and liver tissues from these embryos revealed the presence ofleukemia virus particles. The frequency, quantity, and tyj>e ofparticles observed, as well as the cells forming the particles, weresimilar to C3Hf embryos.

Other Animals. Studies of embryos from other strains ofmice and also from rats are now in process in our laboratory. Theresults thus far observed are as follows:

BALB/c Mouse Embryo. Embryo organs from one normalnoninjected BALB/c mouse were examined in the electron microscope. Characteristic budding and doughnut-type particleswere observed in small numbers in thymus, liver, spleen, andbone marrow of the embryo.

Sprague-Dawley Rat Embryos. Embryos from 3 virus-injectedand 3 normal noninjected Sprague-Dawley rats were also studied.Thus far, examination of thymus, spleen, bone marrow, and liverfrom embryos of virus-injected parents, and thymus and liverfrom embryos of normal noninjected parents failed to reveal thepresence of leukemia virus particles.

DISCUSSION

The results of this study have shown that leukemia virusparticles are present in embryos from Ak strain mice, and alsoin embryos from both virus-injected and normal noninjectedC3Hf mice. The presence of virus particles in embryos from Akmice which normally carry the virus or in embryos from C3Hfmice injected with the virus was not surprising. The appearanceof virus particles in embryos from normal noninjected C3Hfmice was not unexpected either, since C3Hf strain mice areknown to carry a latent leukemogenic virus (12-14).

In previous studies of organs from virus-injected adult C3Hfmice, we found numerous virus particles in a large variety oforgans (6, 7, 9). However, in normal noninjected adult C3Hfmice few virus particles were observed and in a limited number oforgans, i.e., in thymus (2, 7), and very occasionally in bone marrow (7). On the basis of these experiments, we would have expected to see less virus particles in fewer organs in embryos fromnormal C3HI mice as compared with embryos from virus-injectedC3Hf mice. Contrary to such expectations, particles in embryoorgans from normal noninjected and virus-injected C3Hf mitÂ«were similar in quantity, frequency, and organ distribution.

There was also a similar quantity of virus particles observedin all C3HI embryos, whether from virus-injected nonleukemic,leukemic, or normal noninjected mothers. This was of particularinterest since organs from leukemic mothers contained considerably more virus particles than those from injected nonleukemicmothers, and the least amount of particles was found in organsfrom normal noninjected mothers. Apparently, the number ofvirus partides ]Â¡resentin the mothers had no influence on thequantity observed in their respective embryos.

In thymus from both normal noninjected adult C3Hf mice

OCTOBER 1967 1795



Dorothy G. Feldman, } olande Drey fuss, and Ludwik Gross

and C3Hf embryos, virus particles appeared to bud from lymphocytes, lymphoblasts, and epithelial cells. Although virusparticles were rarely found in bone marrow of normal adult C3Hfmice (7), in embryo bone marrow particles often appeared budding from hemocytoblasts. Thus far, virus particles were notobserved in liver and spleen from normal adult C3Hf mice (7).However, virus partielles forming from erythroblasts and hemocytoblasts of liver and spleen occurred not only in the embryosstudied, but also in noninjected mice at least until one week ofage (D. G. Feldiran and L. Gross, unpublished data). The association hetween virus \ article formation and hematopoiesis in theembryo and infant mouse is striking and warrants further studyto determine whether there is any relationship between these twophenomena.

In virus-injected adult C3Hf mice, leukemia virus particleswere observed not only in thymus, spleen, liver, and bone marrow, but in a variety of other organs such as lymph node, mammary and salivary glands, kidney, lung, pancreas (6, 7, 9), and inthe male arid female genital systems (8). In a recent study of thedigestive system, virus particles also appeared to form fromepithelial cells of both the small and large intestines (D. G.Fcldman and L. Gross, unpublished data).

REFERENCES1. Bernhard, W., and Gross, L. PrÃ©sencede particules d'aspect

vi rusai dans les tissus tumoraux de souris atteintes de leucÃ©mies induites. Compt. Rend., Â¿48:100-103, 1959.

2. de Harven, 10. Virus Particles in the Thymus of Conventionaland Germ-free Mice. J. Exptl. Med., ISO: Ã•57-SO?,1904.

3. Dmochowski, L., and Grey, C. E. Subcellular Structures ofPossible Viral Origin in Some Mammalian Tumors. Ann. N. Y.Acad. Sci., 68; 559-G15, 1957.

4. Dmochowski, L., Grey, C. E., Padgett, F., and Sykes, J. A.Studies on the Structure of the Mammary Tumor-inducingVirus (Bittner) and of Leukemia Virus (Gross). In: Viruses,Nucleic Acids, and Cancer. 17th Ann. Symp. on FundamentalCancer Research, Univ. of Texas, M. 1). Anderson IIosp. andTumor Inst., pp. 85-121. Baltimore: The Williams & WilkinsCo., 1903.

5. Feldman, 1). G. A Method of Staining Thin Sections withLead Hydroxide for Precipitate-free Sections. J. Cell Biol.,15: 592-595, 1962.

0. Feldman, D. G., and Gross, L. Electron Microscopic Study ofthe Mouse Leukemia Virus (Gross), and of Tissues from Micewith Virus-induced Leukemia. Cancer Res., 24: 1700-1783,1904.

7. Feldman, D. G., and Gross, L. Electron Microscopic Study ofthe Distribution of the Mouse Leukemia Virus (Gross) inOrgans of Mice and Rats with Virus-induced Leukemia. Cancer Res., m: 412-426, 1966.

8. Feldman, D. G., and Gross, L. Electron Microscopic Study ofthe Mouse Leukemia Virus (Gross) in Genital Organs of Virus-injected C3Hf Mice and of Ak Mice. Cancer Res., Â«7:1513-1527, 1967.

9. Feldman, D. G., Gross, L., and Dreyfuss, Y. Electron Microscopic Study of the Passage A Mouse Leukemia Virus inMammary Glands of Pregnant, Virus-injected, C3H(f) Mice.Cancer Hes., 23: 1604-1007, 1963.

10. Gross, L. Pathogenic Properties, and "Vertical" Transmis

sion of the Mouse Leukemia Agent. Proc. Soc. Exptl. Biol.Med., 78: 342-348, 1951.

11. Gross, L. Viral (Egg-Borne) Etiology of Mouse Leukemia.Filtered Extracts from Leukemic C58 Mice, Causing Leukemia(or Parotid Tumors) after Inoculation into Newborn C57Brown or C3H Mice. Cancer, 9: 778-791, 1956.

12. Gross, L. Attempt to Recover Filterable Agent from X-rayInduced Leukemia. Acta Haematol., 19: 353-361, 1958.

13. Gross, L. Serial Cell-free Passage of a Radiation-activated

Mouse Leukemia Agent. Proc. Soc. Exptl. Biol. Med.. 100:102-105, 1959.

14. Gross, L. Oncogenic Viruses, p. 393. New York, Oxford, London, Paris: Pergamon Press, 1961.

15. Gross, L. Pathogenic Potency and Host Range of the MouseLeukemia Virus. Acta Haematol., 29: 1-15, 1963.

16. Gross, L., and Dreyfuss, Y. How is the Mouse Leukemia VirusTransmitted from Host to Host Under Natural Life Conditions? In: Carcinogenesis, a Broad Critique. 20th Ann. Symp.on Fundamental Cancer Research. Univ. of Texas, M. D.Anderson Hosp. and Tumor Inst., Baltimore: The Williams& Wilkins Co., pp. 9-21.

17. Parsons, D. F. Structure of the Gross Leukemia Virus. J.Nati. Cancer Inst., 30: 509-583, 1963.

18. Watson, M. L. Staining of Tissue Sections for Electron Microscopy with Heavy Metals. J. Biophys. Biochem. Cytol.,4: 475-478, 1958.

FIG. 1. Section through thymus of embryo from virus-injected parents containing lymphoblasts (Lb), lymphocytes (Lc), and epithelialcells (E). X 12,400. a, higher magnification of the outlined area from Fig. 1 demonstrating a leukemia virus particle (arrow) buddingfrom a lymphoblast. X 42,800.

FIG. 2. Area of thymus from embryo of normal noninjected parents containing lymphoblasts (Lb), lymphocytes (Lc), and epithelialcells (E). X 9,200. a, higher magnification of the outlined area of Fig. 2 showing a leukemia virus particle (arrow) budding from alymphocyte. X 84,800.




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OCTOBER 1967



Dorothy G. Feldman, Yolande Dreyfuss, and Ludwik Gross

FIG. 3. Epithelial cell (E) from embryo thymus of normal uoiiinjected parents containing several vacuoles (v). X 20,650.a, b, highermagnifications of the outlined areas of Fig. 3. Present are virus particles budding (arrows) from vacuolar membranes, and doughnut -type particles (rf) and particles (n) with nucleoids lying free within the vacuoles. X 62,000.

FIG. 4. Epithelial cell from embryo thymus of normal noninjected parents containing several vacuoles (v) and particles (n) with nucleoids situated along the vacuolar membranes. X 23,600.

FIGS. 5-7. Portions of vacuoles from epithelial cells of embryo thymus from normal noninjected parents. In Fig. 5 a virus particle(arrow) appears to bud from a vacuolar membrane; in Fig. 6 a doughnut-type particle (d) and in Fig. 7 particles (n) with nucleoids aresituated within the vacuoles. Figs. 5, 6, X 106,750;Fig. 7, X 84,800.

FIGS.8-10. Sections of embryo spleen from virus-injected parents demonstrating mature and immature granulocytes (G), reticulo-cytes (K), hemocytoblasts (H), erythroblasts (Eb), platelets (P), mesenchymal cells (M), and collagen (C). Fig. 8, X 6,300; Fig. 9,X 9,200; Fig. 10, X 8,050.

FIG. 11. Erythroblast of embryo spleen from normal noninjected parents illustrating a virus particle (arrow) budding from the cellmembrane. X 54,000.

FIG. 12. Hemocytoblast from embryo spleen of virus-injected parents. X 13,760.a, higher magnification of the outlined area in Fig.12 showing a virus particle (arrow) budding from the cell membrane. X 67,840.

FIG. 13. Part of an erythroblast from embry.) spleen of virus-injected parents demonstrating a virus particle (arrow) budding fromthe cell membrane. X 62,000.

FIG. 14. Hemocytoblast from embryo spleen of normal noninjected parents; a virus particle (arrow) is apparently budding from thecell membrane. X 33,600.

FIGS. 15-18, 20. Areas of embryo liver from normal noninjected parents (Figs. 15-18) and virus-injected parents (Fig. 20) containinggranulocytes (G), erythroblasts (Eb), megakaryocytes (M), hemocytoblasts (H), and liver parenchyma (L). Fig. 15, X 7,360; Fig. 16,X 13,760;Fig. 17, X 7,360; Fig. 18, X 13,760;Fig. 20, X 12,400.

FIG. 18a. Higher magnification of the outlined area of Fig. 18 demonstrating a doughnut-type particle (</)appearing proximal to thecell membrane of an erythroblast. X 42,800.

FIG. 20a. Higher magnification of the outlined area in Fig. 20 illustrating a virus particle (arrow, lefl) budding from an erythroblast,and a virus particle (arrow, right) budding from a hemocytoblasl. X 42,800.

FIG. 19. Part of an erythroblast from embryo liver of virus-injected parents; a virus particle (arrow) appears to be budding from thecell membrane. X 42,800.

FIG. 21. Area of embryo bone marrow from normal noninjected parents containing granulocytes (G), megakaryoblast (Mb), and partof a mesenchymal cell (M). X 7,200.

FIG. 22. Hemocytoblast of embryo bone marrow from virus-injected parents illustrating a virus particle (arrow) budding from thecell membrane. X 33,600.

FIG. 23. Hemocytoblast of embryo bone marrow from normal noninjected parents. X 12,900.a, higher magnification of outlined areafrom Fig. 23 showing a virus particle (arrow) budding from the cell membrane. X 62,000.

FIG. 24. Section through embryo kidney from normal noninjected parents. X 12,400.a, higher magnification of the outlined area ofFig. 24 showing a smaller doughnut-type particle (arrow) within the endoplasmic reticulum. X 42,800.




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1967;27:1792-1804. Cancer Res Dorothy G. Feldman, Yolande Dreyfuss and Ludwik Gross Normal C3Hf Parents(Gross) in Organs of Mouse Embryos from Virus-injected and Electron Microscopic Study of the Mouse Leukemia Virus

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Electron Microscopic Study of the Mouse Leukemia Virus ... · C3Hf Parents1 DOROTHY G. FELDMAN,...

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