phagocytosis of granules from disrupted mast cells

19
PHAGOCYTOSIS OF GRANULES FROM DISRUPTED MAST CELLS1 DOUGLAS E. SMITH AND YEVETTE S. LEWIS Division of Biological and Medical Research, Argonne Natioiial Laboratory, Lemont, Illinois TWENTY-FOUR FIGURES In the connective tissue throughout the mammalian or- ganisni the tissue mast cell is characteristically round or spindle-shaped and is filled with large cytoplasmic granules that stain metachromatically with toluidine blue. Occasion- ally, however, there occur apparently abnornial mast cells which show singly, or in combination, conglomeration of meta- chromatic granules, colorless or metachromatic vacuoles, or alterations in shape. Such cells make up 1-2% of the total mast cell population of untreated animals (Smith and Lewis, '541, 54b, '55) and become significantly greater in number in 1,cptone shock (Wilander, '3S), stimulation of urticaria pig- mentosa lesions (Drennan, '31) , and treatment with x-rays (Smith and Lewis, '53, '54a), nitrogen mustard (Schoch and Glick, '%), bacterial pyrog-ciis (Stuart, '51), cort,isone (Asboe- IIaiisen, '52 ; Cavallcro and Braccini, '51 ; Smith and Iiewis, '<j4h, 'Xj), aiid ACT11 (Smith and Lewis, '54b, '5.5). Recently Higginbotham et al. ('56) reported that the fibro- blasts of the connective tissue selectively take up and dispose of the cytoplasmic granules shed from mast cells disrupted by various treatments. Tb seemed to us that such fibroblasts closely resemble certain of the above-mentioned abnormal mast ce11s. It appeared possible to us, moreover, that some or all of the mast cells designated hitherto as abnormal or degen- 'This work was performed under the auspices of the U. S. Atomic Energy Coniniission. 93

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Page 1: Phagocytosis of granules from disrupted mast cells

PHAGOCYTOSIS O F GRANULES FROM DISRUPTED

MAST CELLS1

DOUGLAS E. SMITH AND YEVETTE S. LEWIS Division of Biological and Medical Research, Argonne Natioiial Laboratory,

Lemont, Illinois

TWENTY-FOUR FIGURES

I n the connective tissue throughout the mammalian or- ganisni the tissue mast cell is characteristically round or spindle-shaped and is filled with large cytoplasmic granules that stain metachromatically with toluidine blue. Occasion- ally, however, there occur apparently abnornial mast cells which show singly, or in combination, conglomeration of meta- chromatic granules, colorless or metachromatic vacuoles, or alterations in shape. Such cells make up 1-2% of the total mast cell population of untreated animals (Smith and Lewis, '541, 54b, '55) and become significantly greater in number in 1,cptone shock (Wilander, '3S), stimulation of urticaria pig- mentosa lesions (Drennan, '31) , and treatment with x-rays (Smith and Lewis, '53, '54a), nitrogen mustard (Schoch and Glick, '%), bacterial pyrog-ciis (Stuart, '51), cort,isone (Asboe- IIaiisen, '52 ; Cavallcro and Braccini, '51 ; Smith and Iiewis, '<j4h, 'Xj), aiid ACT11 (Smith and Lewis, '54b, '5.5).

Recently Higginbotham et al. ('56) reported that the fibro- blasts of the connective tissue selectively take up and dispose of the cytoplasmic granules shed from mast cells disrupted by various treatments. Tb seemed to us that such fibroblasts closely resemble certain of the above-mentioned abnormal mast ce11s. It appeared possible to us, moreover, that some o r all of the mast cells designated hitherto as abnormal o r degen-

'This work was performed under the auspices of the U. S. Atomic Energy Coniniission.

93

Page 2: Phagocytosis of granules from disrupted mast cells

94 DOUGLAS E. SMITH AND YEVETTE S. LEWIS

erated might be in reality fibroblasts or other cells which have ingested cytoplasmic granules from disrupted mast cells. The present paper is concerned with a microscopic examina- tion of (a) the fate of the cytoplasmic granules shed from mesenteric mast cells broken up as a result of exposure to dis- tilled water and of (b) abnormal mast cells in several tissues from normal and treated animals.

METHODS

The mesentery of the Sprague-Dawley rat (male and fe- male, weighing approximately 200 gm) was employed as the site for studying the fate of the cytoplasmic granules released from disrupted mast cells. Break-up of the mast cells was brought about by the method of Fawcett ('54) whereby distilled water is injected intraperitoneally into the unanesthe- tized animal. Rats were sacrificed by ether anesthesia at var- ious times (table 1) after the administration of distilled water. Whole mounts of mesentery were prepared as previously de- scribed (Smith and Lewis, '53). The tissues were fixed in 80% ethyl alcohol and stained with toluidine blue (.025% in 50% ethyl alcohol) or May-Grunwald stain. The preparations were examined under the light microscope at magnifications up to 1000 diameters. A similar study was carried out on fixed whole mounts of mesentery from 150-gm male Sprague-Dawley rats treated with 48/80.2 Twenty milliliters of Tyrode's solu- tion containing 1 mg of 48/80 was injected intraperitoneally, and portions of mesentery were taken from animals sacrificed at 10 minutes, 2, 6 and 24 hours after injection.

In addition to observations on fixed tissue, a microscopic study was carried out in. vivo on the mesenteries of rats in- jected intraperitoneally with distilled water. See table 1 for the schedule of the observations. For this study the animals were anesthetized with Nembutal R (sodium pentobarbital) and a loop of intestine was withdrawn through a mid-ventral incision. The rat was then placed on its side on a holder con- ' A polymer of N-methylhomoanisylamine and formaldehyde, supplied through

the courtesy of Dr. Edwin J. de Beer of Wellcome Research Laboratories.

Page 3: Phagocytosis of granules from disrupted mast cells

TABLE 1

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15

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12

34

57

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20

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Page 4: Phagocytosis of granules from disrupted mast cells

96 DOUGLAS E. SMITH AND YEVETTE S. LEWIS

taining a small lucite (methyl methacrylate) block over which the mesentery w7as laid in such a manner that it was transil- luminated on the microscope stage. Oxygenated Tyrode’s solution a t 37°C was allowed to drip continually on the mes- entery. Vital staining of these preparations was readily ac- complished by adding toluidine blue to the Tyrode’s solution. Observations were made at magnifications up to 900 dia- nieters with water immersion objectives, and the phenomena were recorded by time-lapse cinematography and still photo- graphy.

hlicroscopic examination was made of the abnormal mast cells in whole mounts of fixed and stained mesentery and skin of the Sprague-Dawley rat (200 gm) and mesentery, skin and cheek pouch of the Syrian hamster (100 gm) at various times after s-irradiation or administration of ACTH o r cortisone, Similar observations were made in untreated rats and liam- sters. The s-irradiations consisted of single total-body ex- posures to 600 r or 1200 r to rats, and 600 r, 715 r or 1200 r to hamsters. The radiation factors were 250 Bv, 15 ma, 0.; nim Cu, and 3.0 mm Bakelite filters, 26.7 ern target distance, 1.5 mm Cn half-value layer and 215-225 r per minute. Cortisone was administered subcutaneously or intramuscularly daily for 1-4 days in dosages of 200 nig/kg. ACTH ~ - a 7 injected subcutaneously for 1-4 days in dosages of 100 or 400 I.U./kg.

T n a few experiments in vico examination., wcre made of mesentcry of rats csposed to s-irradiation o r treated with cortisone or ACTH.

RES1T~T-S

Fired and s t c( in e (7 7 i cs P+Z t P r 71 o f rci t .s i i z j cc t c d in f ra p e r i- t o n e a1 1 i j 1 o i t h cl is ti1 1 (2 (1 u‘c, t-. Tithin 15 minutes after the administration of distilled water, the limitjng boundaries of many mast cells are no longer intact and metachromatic granules are seen free in the surrounding tissue. Some of these mast cells are completely disrupted and their granules occupy a space 11h to 3 times that of the volume of an intact mast cell; the remainder show the loss of a fern granules into the surrounding tissue. With the passage of time after in-

Page 5: Phagocytosis of granules from disrupted mast cells

PHAGOCYTOSIS O F MAST CELLS 97

jection more and more mast cells are found in the disrupted state until at 1-2 hours most of them are broken up and the mesentery is filled with mast cell granules. No changes are found in cells other than mast cells. No changes are apparent in the staining properties of the disrupted mast cells and their scattered granules except for the presence of an oc- casional enlarged, pink granule.

The fate of the freed mast cell granule is apparent as early as 15-30 minutes after distilled water injection. At this time metachromatic granules of the same size and shape as the cytoplasmic granules of the mast cell are found in macrophages, fibroblasts and leucocytes in the vicinity of the broken mast cells. As time passes and the number of disrupted mast cells increases in the preparations so does the number of macrophages, fibroblasts and leucocytes that contain meta- chromatic material. At 6-7 hours leucocytes are present throughout the tissue in impressive numbers (up to 200 and more per 0.0625 mm2 as compared with 0-5 per 0.0625 mm2 at 15-30 minutes). Many of the leucocytes contain no meta- chromatic granules. The distribution of macrophages and fibroblasts is not uniform throughout the tissue. Macro- phages are found in large numbers and fibroblasts are few near blood vessels, whereas fibroblasts are more numerous away from the blood vessels. Many of these cells contain metachromatic material. By 7 hours very few metachromatic granules are found free in the tissues, and at 24 hours no metachromatic material is found outside of fibroblasts, macro- phages and leucocytes. Many of these cells contain such large numbers of metachromatic granules at tv7o hours after treat- ment with distilled water that their nuclei are obscured, and they thus resemble intact mast cells. Others show few meta- chromatic granules in their cytoplasm even at 24 hours. Illustrations of metachromatic material in fibroblasts, macro- phages and leucocytes are found in figures 1-16.

Alterations in the ingested metachromatic material are ob- vious as early as two hours after treatment. I n some macro- phages, fibroblasts and leucocytes one now finds no discrete

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98 DOUGLAS E. SMITH AND YEVETTE S. LEWIS

granules but rather metachromatic vacuoles, some of which stain light red, others, purple. An individual cell may contain from one to dozens of such vacuoles. By 7 hours more than half of the macrophages, fibroblasts and leucocytes have at least some of their metachromatic material in this state, with the remainder in the form of discrete granules. At the end of 24 hours most of the cells have practically all of their metachromatic material in the form of vacuoles. Thereafter progressively fewer cells are found to contain metachromatic material in any form. By 4-5 days no such cells are found and the mesentery has a normal appearance except for a com- plete absence of mast cells and an increased number of eosin- ophils. Eosinophils are found in increasing numbers up to 24 hours after treatment and are still present in elevated numbers even at the end of one week. The eosinophils do not contain metachromatic inclusions.

After the intraperitoneal injection of 48/80, large numbers of cytoplasmic granules were released from the mast cells. These granules were ingested and disposed of by macro- phages, fibroblasts and leucocytes in the same fashion as those released by treatment with distilled water.

Mesentery--in. vivo study. Within 10 minutes after the intraperitoneal injection of 20 ml of distilled water, many of the mast cells are disrupted and their granules are scattered in the surrounding tissue. Here the granules show Brownian movement and freely bounce about. At this time macro- phages, fibroblasts and white blood cells containing some metachromatic granules can be found. As more mast cells break up, macrophages which were previously sessile and unnoticed become motile and evident. They move toward broken mast cells and engulf the granules with long proto- plasmic extensions. Leucocytes also make their way toward the loose granules by amoeboid movement. They do not sur- round the free mast cell granules with long pseudopodia but rather show a marked bubbling of the cell surface when in contact with the free granules; shortly thereafter the mast cell granule is seen within the leucocytc. Lcucocytes are

Page 7: Phagocytosis of granules from disrupted mast cells

PHAGOCYTOSIS O F MAST CELLS 99

frequently seen to traverse the walls of small blood vessels, and they are accumulated in large numbers in the tissue along- side the vessel walls at 2 4 hours after injection of distilled water. By 6-7 hours large numbers of leucocytes are visible at considerable distances from blood vessels throughout the mesentery. Fibroblasts show neither ambulatory nor surface activity. Free mast cell granules appear to make their way to fibroblasts by Brownian movement. After bouncing against the limiting surface of the fibroblast for a few minutes, the granules penetrate it and are seen within the cytoplasm of the fibroblast. The morphology and staining properties o€ the mast cell granule do not appear to be changed upon their entrance into the cytoplasm of the macrophage, fibroblast or leucocyte. Once inside these cells, the granules continue to show Brownian movement within the bounds of vacuoles. The vacuoles vary in size and in the number of granules which they contain. Later the granule (s) may disappear leaving the vacuole filled with material which appears uniformly purple. The phenomena observed after injections of 10 ml of distilled water were essentially the same as those above, ex- cept that some cells lose only a few granules and appear intact thereafter while other mast cells seem completely unaff ectcd by the treatment. By the end of 24 hours when all of the free mast cell granules have been ingested, no further activity is evident and the living preparations thereafter closely re- semble the fixed mesenteries of the same time period. 11- lustrations of typical findings in the living preparations are found in figures 17-24.

Fixed and staified tissues from untreated rats and ham- sters. Normal skin, mesentery and cheek pouch clearly show macrophages and fibroblasts which contain metachromatic granules and vacuoles. These cells are similar in appearance to those seen in the fixed and stained preparations of mesen- tery exposed to distilled water. One o r two such cells occur for every 100 mast cells. Cells indistinguishable from mast cells but containing conglomerations of granules and meta- chromatic vacuoles were found but rarely. Leucocytes con-

Page 8: Phagocytosis of granules from disrupted mast cells

100 DOUGLAS E. SMITH AND YEVETTE S. LEWIS

taining metachromatic material are not found in these preparations.

Fixed and staimed tissues from rats and hanzsters treated with z-radiation, cortisone and ACTB. The skin, mesentery and cheek pouch of animals subjected to x-irradiation or in- jection with cortisone or ACTH showed large numbers of macrophages and fibroblasts containing metachromatic gran- ules and vacuoles. These cells were similar to those found in the fixed and stained mesenteries exposed to distilled water. The number of such cells varied with duration and severity of treatment. I n the most striking instances the number of macrophages and fibroblasts containing metachromatic ma- terial equalled or even exceeded the number of intact mast cells found. As with the same tissues from untreated rats and hamsters, cells indistinguishable from normal mast cells except for conglomerations of granules and metachromatic vacuoles were found occasionally. Leucocytes with or without metachromatic material were seldom found. The i m vivo ob- servations on the mesentery of x-irradiated and cortisone and ACTH rats were in essential agreement with those on fixed preparations.

DISCUSSION

The finding in both living and fixed mesentery that the intraperitoneal injection of distilled water completely dis- rupts the tissue mast cells but causes no conspicuous effects in other cells agrees with the report of Fawcett ('54, '55) on fixed tissue. The observation that fibroblasts, macrophages and leucocytes ingest mast cell granules immediately after treatment with distilled water further indicates that general cellular damage does not follow this procedure.

The present finding of increased tissue eosinophils after distilled water treatment is at variance with the report of a decrease by Fawcett ( '55). It is possible that this discrepancy is due to the different strains of rats employed in the t ~ r o studies.

Page 9: Phagocytosis of granules from disrupted mast cells

PHAGOCYTOSIS O F MAST CELLS 101

Both the living and fixed mesenteries of the rat clearly show that fibroblasts, macrophages and leucocytes ingest and dispose of the cytoplasmic granules released from mast cells disrupted by distilled water. Prior reports have suggested that macrophages alone (Pawcett, '54) or fibroblasts alone (Higginbotham, '56) perform this phagocytic action, and, to the best of our knowledge, leucocytes containing meta- chromatic material have not been reported.

The differential distribution of fibroblasts and macro- phages with respect to their location near or away from blood vessels agrees with previous findings (Maximow and Bloom, '52). This differential distribution may account for the failure of prior workers to note that both macrophages and fibroblasts take up loose mast cell granules.

The findings after the injection of 48/80 indicate that the debris of mast cells disrupted by different procedures is phag- ocytized in the same fashion.

The macrophages and fibroblasts containing cytoplasmic granules from mast cells disrupted by distilled water or by 48/80 are grossly quite similar in appearance to the "abnor- mal" mast cells previously described as occurring in small numbers in normal tissue (Smith and Lewis, '54b, '55) and in quantity after treatment with ACTH (Smith and Lewis, '54b, '55) cortisone ( Asboe-Hansen, '52 ; Cavallero and Braccini, '51; Smith and Lewis, '54b, '55) o r x-radiation (Smith and Lewis, '53, '54a). In the present study, examination of normal and treated (ACTH, cortisone and x-irradiated tissue revealed that macrophages and fibroblasts that contain meta- chromatic granules and vacuoles occur with frequencies sim- ilar to those described for "abnormal" mast cells in prior studies. Only rarely does there appear a cell which contains conglomerated metachromatic granules or metachromatic vacuoles but which in other respects is a typical mast cell. Thus practically all of the cells previously designated as "abnormal" or degenerated mast cells (Wilander, '38 ; Cav- allero and Braccini, '51 ; Drennan, '51 ; Stuart, '51 ; Schoch and Glick, '53 ; Smith and Lewis, '53, '54a, '54b, '55) may well be

Page 10: Phagocytosis of granules from disrupted mast cells

102 DOUGLAS E. SMITH AND YEVETTE S. LEWIS

macrophages, fibroblasts and leucocytes that contain meta- chromatic material. Previous failures to identify these cells correctly probably resulted from the limitations of the meta- chromatic stains employed. The cellular detail brought out by the May-Grunwald stain used in the present study makes possible more accurate identification of the various cell types.

It was previously considered (Wilander, ’38 ; Cavallero and Braccini, ’51 ; Drennan, ’51 ; Stuart, ’51 ; Schoch and Glick, ’53; Smith and Lewis, ’53, ’54a, ’54b, ’55) that the “abnormal” mast cells reflected a series of degenerative changes that oc- cur spontaneously only rarely but with great frequency after certain treatments, the final step in degeneration being break up of the cell with release of the cell contents into the sur- rounding tissue. The present findings indicate that while such a degenerative change occurs, the usual process, which is seen only occasionally in untreated animals but frequently after the aforementioned treatments, is one of disruption of the mast cells with the freeing of their granules into the surround- ing tissue where macrophages, fibroblasts and leucocytes quickly ingest them. Since one is concerned with disruption of mast cells in both the past and the present interpretations, the proper identification of the cells does not require basic revision of conclusions drawn from experiments (Wilander, ’38 ; Cavallero and Braccini, ’51 ; Drcnnan, ’51 ; Stuart, ’51 ; Schoch and Glick, ’53 ; Smith and Lewis, ’53, ’54a, 54b, ’55) in which these cells were not properly identified. Since the remnants of a single disrupted mast cell may be taken up by more than one macrophage or fibroblast, however, the prior accountings (Smith and Lewis, ’54a, ’54b, ’55) in terms of “abnormal” mast cells would tend to give a somewhat exag- gerated idea of the number of mast cells being destroyed hy treatment. Although it would appear that absolute quantita- tion of mast cell destruction as a result of treatment is not possible from a measure of the increase in macrophages and fibroblasts and leucocytes in a preparation, it is certain that one can employ such counts to give qualitative and perhaps rough quantitative information on the effectiveness of cxperi-

Page 11: Phagocytosis of granules from disrupted mast cells

PHAGOCYTOSIS O F MAST CELLS 103

mental procedures. Failing to account f o r phagocytes con- taining metachromatic material, i.e., including them as mast cells, obviously leads to errors in interpreting data involving changes in mast cell number. It is possible that the increase in mast cell reported by Bensley ('52) and by Schiirer ('46) after histamine and heparin injection respectively are ex- amples of such a misinterpretation. Higginbotham et al. ( '56) have called attention to the possibility that such errors arise from failure to account for fibroblasts which have ingested inetachromatic material. Earlier claims (Devitt et al., '54) that similar treatments produce no change in mast cell num- ber may be explained in somewhat the same way. I n these studies phagocytes containing metachromatic material were not identified and cells of the "abnormal" type were con- sidered to be artifacts of handling and counted as normal mast cells.

The functional significance of the disruption of mast cells by various treatments is not clear. I n general, such disruption with release of granules into the surrounding material has been considered to constitute secretion (Wilander, '38 ; Cav- allero and Braccini, '51 ; Drennan, '51 ; Stuart, '51 ; Schoch and Glick, '53; Smith and Lewis, '53, '54a, '54b, '55). Indeed, large amounts of histamine are released into the peritoneal fluid after injection of the rat with histamine liberators and distilled water (Fawcett, '54). On the other hand, solutions of toluidine blue and protamine sulphate cause similar release of histamine without morphological changes in the mast cells of the mesentery (Smith, '58). I t is possible that the latter is the usual method of release of material (secretion) from the mast cell and that rupture of the cell with release of granules occurs only when marked changes occur in its environment.

SUMMARY

Microscopic examination was made of the fate of the cyto- plasmic granules shed from mesenteric mast cells disrupted by intraperitoneal injection of distilled water into the rat. Both fixed and living preparations of mesentery were employed.

Page 12: Phagocytosis of granules from disrupted mast cells

104 DOUGLAS E. SMITH AND YEVETTE S. LEWlS

I n addition microscopic examinations were carried out on the “abnorma1” or degenerated mast cells of several tissues from control and treated (x-radiation, ACTH, o r cortisone) rats and hamsters.

Treatment with distilled water completely disrupted the mast cells of the mesentery, while other cells appeared to be undamaged. Fibroblasts, macrophages and leucocytes were observed to take up and digest the shed mast cell granules. These phagocytes were quite similar in appearance to the “abnormal’ o r degenerated mast cells previously described as occurring in normal and treated animals.

Mast cell granules released as a result of 48/80 administra- tion met the same fate as those freed by distilled water treat- mcnt.

The examination of “abnormal” mast cells in control and treated animals indicated that most of these were macro- phages and fibroblasts that contained nietachromatic material.

The results are discussed in relation to the question of the phagocytosis of disrupted mast cells and of the significance of the “abnormal” or degenerated mast cell.

LITERAT URF CITED

ASBOE-HANSEN, G. 1952 The mast cell a n object of cortisone action on con-

BENSLEY, S. H. 1952 On the origin of mast cells. Anat. Rec., 122: 310. CAVALLERO, C., AND C. BRACCINI 1951 Effect of cortisone on the mast cells

of the rat. Proc. SOC. Exp. Biol. Med., 78: 141-143. DEVITT, J. E., P. B. SAMUELS, w. J. PIROZYNSKI AND D. R. WEBSTER 1954

Morphology of tissue mast cells; the frequency of art ifacts and the influence of certain biologic agents. Am. J. Pathol., 3 0 : 391401.

DRENNAN, J. M. 1951 The mast cells in urticaria pigmentosa. J. Pathol. Bac- teriol., 63: 513-520.

FAWCETT, D. W. 1954 Cytological and pl~~lrniacological observations on the re- lease of histamine by mast cells. J. Exp. Med., 100: 217-224.

1955 An experimental study of mast cell degranulation and re- generation. Anat. Rec., 121: 29.

HIGGINBOTHAN, R. D., T. F. DOUGIIERTY AND W. S. S. SEE 1956 Fa te of shed mast cell granules. Proc. Soe. Exp. Biol. Med., 92: 256-261.

MAXIMOW, A. A., AND W. BLOOM 1952 Textbook of Histology. Sixth Ed. p. 57. W. B. Saunders, Philadelphia.

nective tissues. Proc. SOC. Exp. Biol. Med., 80: 677-679.

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PIIAGOCYTOSIS O F MAST CELLS 105

SCHOCII, E. P., AXD D. GLICK 1953 The effect of cold stress, ACTH, cortisollc, pyroyeri and nitrogen mustard 011 tissue mast cells in the skin and subcutaneous tissues of the rat. J. Invest. Dermatol., 20: 119-131.

S C H ~ E R , W. 1946 Speicheruiig voii Heparin in den Zelleri des Iieticuloeiido- thcls. Helv. Mecl. Aeta, 13: 161-171.

S~IITII, I). E. 1958 The nature of tlic secretory activity of the mast cell, Amer. J. Physiol., 293: 573-575.

~ M I ' I T I , 1). E., AND Y. s. LEWIS 1953 Effects of total-body x-irradiation on the tissue mast cell. Proe. Soc. Exp. Biol. Med., 6'2: 208-212.

-~ 1954:~ Iiiflueiiec of total-body x-irradiation upon tissue iuast cell iiuiribcr. Ibid., X5 I 306-307.

1954b Iiiflueiice of hypophysis mid adrrrinl cortex upoil tissue mast crll of the rat. Ibitl., 87: 515-518.

1955 Tiiflueiice of hypophysis and of adreii:rl cortex upon tissuc mast cell of thc hamster. Ibid., 88: 631-634.

STliART, E. G. C'oiiiieetive tissuc in:~.st-cell response to b:rctcrinl pyrogrii, ovalbumin aiid cortisone. Anat. Rec., 109: 351.

~VILASDER, 0. 1938 Stnclicii iiber Heparin. Skand. Arch. Pllysiol., Xl : 1-89, Suppl. 15.

1951

Page 14: Phagocytosis of granules from disrupted mast cells

PLATE 1

EXPLANATION OF FIGURES

Pibro1)lasts f rom fixed :ind staiiied whole niounts of iiieseiitery of the rat after iiitr;ipeiitoiieal iiijectioii of distilled wxter. M:iy-Gruiiwilcl stain. X 1900.

1

2

3-4

5

G

i-8

A few mast cell granules i n n cell. Xotc the vacuole co~itniiiiiig a gr:iiiulc.

Maiiy mast cell granules iii a cell. Note tha t seine vacuoles coiitaiii several granules.

I ~ r g c x accuuiulatioiis of mast cell gr;riiulcs iii cells. K o t e sinnll eongloiiic~rn- tioris of granules.

Kote large conylomeratioiis of granules.

A cell aliriost coinpletely filled with nietncliromatic material.

Exaiiiplcs of dissolution of iiictaeliroiiintic material. Vacuoles show varyiiig degrees of mctachromasi:~ (pink t o red).

Page 15: Phagocytosis of granules from disrupted mast cells
Page 16: Phagocytosis of granules from disrupted mast cells

PLATE 2

EXPLASAI’ION O F FIGURES

Mncroplinges :iiid leucocgtes f rom fixed and stained mliole mounts of mesentery of the rat aftcr intraperitouenl iiljeCtioii of tlistilled water. May-Griinmnld stain. X 1900.

9 A niacropli:igc coiitaiiiiiig iiinst cell grauules. Note the vacuole coiit:iiiiing a granule.

h m:icropliage coiitaiiiiiig iii:iiig inast cell granules.

J1:icropliages coutniiiiiig co~iglonic~r:itio~~s of metaclironintic granules and nietaclironiatic vacuoles.

Kcutropliils coiitainiiig iii:ist ccll granules.

A nionoc>yte coiit:iiiiing ni:ist ccll granules.

1,yinplloc.ytcs eoiitaiuiiig mast cell granules.

10

11-12

13-14

15

16

108

Page 17: Phagocytosis of granules from disrupted mast cells

109

Page 18: Phagocytosis of granules from disrupted mast cells

PLATE 3

EXPLANATION OF FIGURES

Fibroblasts, uiacropliagcs atid leucocytcs in living m e s c n t e r ~ of r a t a f te r intrii- peritoncwl iiijrctiou of d i s t i l l d water. Toluidine blur stain. X 1000.

17-18 Fibroblasts mhicli 1i:rve taken up free niast cell grnuules. K o t c tllc vacuoles about t h e granules.

Fi1)rol)l:ists sliowiug ~iict:~rliroiiiatic inaterial in the process of dissolution. The vacuoles exliibit varying degrws of inetacliroinasia.

bfacroplinges eontainiug met:ichromntic granules aud vacuoles.

A lcucocpte wliicli has just engulfed a niast cell ~ K L I I U ~ C . S o t ( ’ tllr pro- top1:rsiiiie vxtensions of the cell.

A lcucocpte contaiiiiiig niast cell granules.

19-20

21-22

23

24

110

Page 19: Phagocytosis of granules from disrupted mast cells

I’IIA(:OCYTOSIS O F MAST CELLS u . F,. shl1’PII ASD Y. S. LEWIS

111