some aspects of the fine structure of the reticulo-endothelial system; the cells which clear...

Zeitschrift fiir Zellforschung 89, 355--370 (1968)

Some Aspects of the Fine Structure of the Reticulo-Endothelial System;

the Cells Which Clear Colloids from the Blood Stream

IAN CARR *

Department of Human Biology and Anatomy, University of Sheffield, Sheffield, Yorks., England

Received March 18, 1968

Summary. The macrophages in various sites in the mouse have been examined with the electron microscope in normal animals, and in animals which had recently received intravenous colloids. There was a general structural similarity at all sites. The most obvious common feature was the presence of small granules which stain with toluidine blue. These were of two types. One, the smaller had a finely granular substructure and may represent the primary lysosome; the other, the larger was heterogeneous, contains banded probably fibrillar material with a repeating pattern of 35--50 A, and may represent the residual body.

Colloids given intravenously were cleared by phagocytosis by endothelial cells and endothelial macrophages, by leakage through patent intercellular junctions, and by phagocytosis by paravascular macrophages. Such clearance is commonly held to be a measure of reticuloendothelial function. This measure is therefore clearly dependent on several cellular factors.

The re t icu loendotheha l (RES) sys tem was def ined b y ASCHOFF in 1924 as composed of those cells which av id ly clear foreign ma te r i a l f rom the blood s t r eam; in th is na r row sense i t is made up of the macrophages . The def ini t ion is now often widened to include cells of the l y m p h o i d series and the whole of the immu- nological mechanism of the body, pe rhaps be t t e r descr ibed as the lymphore t i - cular sys tem. A vi r tue of the nar rower def ini t ion is t h a t the funct ion of the re t i cu loendotheha l sys tem as thus defined can be measured in a number of ways, n o t a b l y b y measur ing the ra te of clearance of in t r avenous ly in jec ted colloids f rom the blood s t ream (HALPERN et al., 1953). I t seems l ikely t h a t such clearance is a measure of the R E funct ion in an an imal over only a ve ry l imi ted t ime sequence, of the order of an hour or so. In i t i a l ly colloid is c leared f rom the blood by cells which happen to be in a posi t ion to do so; l a te r cells m a y migra te and there m a y be changes in funct ion of ind iv idua l cells, or even cell division, under the influence of the ac tua l colloid which is being used to es t imate funct ion. I t is of impor t ance in unde r s t and ing re t icu loendotheha l funct ion to know wha t is the s t ruc ture of the cells which no rma l ly clear colloids f rom the blood s t ream. There are m a n y accounts in the l i t e ra tu re of the s t ruc ture of macrophages a t ind iv idua l sites (e.g. MOVAT and FERNANDO, 1964; see CARI~, 1967a for fu r the r references). The only s t u d y of the R E S as an en t i t y in re la t ion to colloid c learance

* I am grateful to Prof. R. BARER for his advice and criticism, to Dr. G.A. MEEK for guidance on electron microscopy, and to Mrs. B. RoMAns and Miss M. Type for technical and photographic assistance.

This work was supported by grants from the M.R.C. and the University of Sheffield Tuberculosis Research Fund and by a grant to the Department from Unilever Ltd.

356 I. CAaR:

is that of MooR~ et al. (1961), who paid most attention to the period 24 hours after introductions of colloid. By this time the administered colloid has had an opportunity to modify the structure of the cells which have ingested it.

Previous reports from this laboratory (CARR, 1967, a and b, 1968) described the structure of the macrophages of the peritoneum of the mouse, and the changes which occur in these cells when they are stimulated, notably formation of lysosomes and increase in length of cytoplasmic processes. If the peritoneal macrophages are a fair sample of the RES, then it would be reasonable to extrapolate findings from the peritoneum, to the rest of the RES.

The present report is a study of the normal reticuloendothelial organs of the male white mouse, with particular relation to the cells in them which avidly take up injected colloids from the circulation. No account will be given of the topographical histology of the organs containing these cells. Emphasis will be laid on the features common to macrophages at all sites. The fine structure of circulating cells which take up colloid will be described in a further report.

Materials and Methods The animals used were male white mice 20--25 g in weight, of an inbred strain, bred in

the University of Sheffield Animal House. Tissues were examined from animals killed by cervical dislocation without prior treatment, and from animals which had received intravenous injections of various colloids - - 0.1~0.2 ml ferrivenin, thorotrast or undiluted Indian ink, (Gunther Wagner 1481 A). Others received a dose of 16 mg/100 g body weight carbon (Indian ink, Gunther Wagner 1481 A) made up in 1% gelatin as used for measuring carbon clearance. Animals were examined 10 min to 1 hour after injection. The tissues examined were: liver, spleen, axillary lymph node, femoral marrow, lung, skeletal muscle, subcutaneous tissue, adrenal, kidney, thymus, small intestine, large intestine, PETER'S patch, epididymal fat pad. A total of 18 animals was used.

Tissues were fixed in 3% glutaraldehyde in phosphate buffer, and post-osmicated in 2 % osmium tetroxide. Araldite blocks were sectioned and sections examined in an A. E. I. EM 6 B electron microscope, after staining with lead citrate.

Results

When toluidine blue stained thick sections of araldite blocks were examined with the light microscope, cells were identified which contained dense blue granules. They were seen both lining blood vessels and in the interstitial tissues of RE organs (Figs. 1 and 2). Evidence will be presented to support the view that these cells are avidly phagocytic, and that their granules have a characteristic structure.

As has been previously described by many authors, gaps exist between endothelial cells in RE organs. Through these gaps plasma presumably leaks under normal circumstances. After administration of colloids they leaked from the circulation through such gaps notably in liver and spleen, to a lesser extent in adrenal cortex and to minor degree in bone marrow. Colloids were phagocytosed by endothelial cells, notably in liver, but to a lesser degree in spleen and a much lesser extent in marrow. As will be pointed out many of these endothelial cells are in fact endothelial macrophages. Where leakage occurred, and where suitable paravascular macrophages existed these ingested very large amounts of colloid. This was marked in spleen. In the adrenal cortex leakage occurred but few

Fine Structure of the Reticulo-Endothelial System 357

macrophages were present so tha t an hour after administration much colloid lay in the interstitial spaces ; while in other sites, notably the testis a considerable number of macrophages were present, but no leakage occurred. In all of these organs small quantities of colloid were taken up by various types of cells including lymphocytes. These will be ignored. The pat tern of handling of colloids was similar with all of the colloids used. I t was notable tha t with the lower dose of carbon (that usually used in measuring carbon clearance) there was little leakage from vessels in the liver. In the spleen, however, even at this low dose level leakage from vessels and uptake by paravascular macrophages was marked.

All the material illustrated was fixed in glutaraldehyde and osmium and embedded in Araldite. The electron micrographs depict material further stained with lead citrate. Fig. 1. Light micrograph of 2 ~ araldite section of liver stained with toluidine blue. The attenuated endothelium of a sinusoid is visible in places. A Kupffer cell contains densely

staining granules of various sizes. Control animal. • 1,300 Fig. 2. Similar preparation of spleen. A large macrophage (outlined) contains numerous densely

staining granules. Control animal. • 1,300

1. Endothelial Phagocytosis Ingested material was seen in small or moderate quantities in flattened endo-

thelial cells in liver and spleen and to a much lesser extent in lymph node and bone marrow (Figs. 3, 4, 5); particles were seen apparently passing into flask- shaped caveolae a t the cell surface. These endothelial cells were relatively un- differentiated, tha t is they had little endoplasmic reticulum, few lysosomes and few mitochondria. They may be contrasted with the thicker more differentiated endothelial cells, notably in the liver, which are avidly phagocytic and have all the characteristics of macrophages.

2. Leakage ]rom Blood Vessels In control animals widely patent gaps between endothelial cells were identified

in only two sites in control animals, namely in liver and spleen. These became more obvious when occupied by colloid (Fig. 4), but probably neither more frequent nor wider. Only occasional and insignificant leaks were identified in

358 I. CARR:

Figs. 3--5


other organs, most often in bone marrow and PEYER'S patches. In the adrenal cortex, while no open gaps between endothelial cells were seen in control animals, an hour after administration of both carbon and thorotrast considerable quantities of colloid had leaked through patent gaps between endothelial cells and lay in the extravascular space. This was the only site in which there was evidence tha t the administration of the colloid caused increased vascular permeability. This is in contrast to the finding of ROWLEY (1963) who showed tha t thorotrast contained a factor (probably dextrin) potent in causing increased vascular permeability, probably mediated by mast cell damage.

3. Phagocytosis by Macrophages Nearly all of the cells which phagocytosed large quantities of colloid belonged

to the group normally possessing large granules staining with toluidine blue noted above. In the liver these were commonly endothelial in position, elsewhere notably in spleen they were paravascular.

Depending on their site they varied widely in size and shape. The endothelial cells in the area near the nucleus bulged into the lumen, while peripherally the cytoplasm became attenuated, often to the normal thickness of true endothelial cells. Short processes often projected into the lumen of the vessel.

Paravascular cells varied widely in size and shape. The commonest profile was elongated, up to 20 9 long by 4 ~ across with numerous cytoplasmic processes. In general the cytoplasmic structure of both endothelial and paravasculal macrophages was similar (Figs. 6 and 7). Phagocytic vacuoles 0.5--1 ~ in diameter were common in the cytoplasm, notably after injection of tracer particles but there were few micropinocytotic vesicles, or caveolae adjacent to the surface membrane. Granular endoplasmic reticulum was present in moderate amounts (Fig. 6). Aggregated free ribosomes (polyribosomes) are fairly frequent in the cytoplasm (Fig. 8). Numerous tubular and vesicular profiles scattered throughout the cytoplasm presumably represented agranular reticulum. "Alveola te" or " coa t ed" vesicles (RoTH and PORT],:R, 1964) were not commonly seen. Mitochon- dria showed as circular or ovoid profiles, rarely as elongated forms. Microtubules about 250A in diameter were common cytoplasmic components, often occurring in bundles (Fig. 9). Microtubules were much less commonly seen in endothelial macrophages, than in those in a paravascular site. This may be related to differences in motility. Also commonly found were microfibrils about 50A in diameter, similar to those first noted in peritoneal macrophages by DE PETRIS et al. (1963), (Fig. 10).

Fig. 3. Endothelium of capillary in lymph-node, showing carbon particles, administered intravenously one hour previously, being transported through the wall in membranous vesicles.

Carbon is present in the perivascular space. • 23,600 Fig. 4. Sinusoid in spleen of mouse 1 hour after intravenous administration of thorotrast. Thorotrast has been ingested by flattened endothelial cells lining sinusoids (L) and extravascular macrophages (M). The processes of lining cells (arrowed) may trap foreign particles.

• 6,800 Fig. 5. Detail from Fig. 4, to show leakage of thorotrast through a patent intercellular junction,

and uptake by both lining cell and pericyte. • 23,600

360 I. CARR:

Fig. 6. Liver of mouse one hour after intravenous injection of thorotrast. An endothelial maerophage (Kupffer cell) is intersected by deep invaginations, between which lies but a narrow isthmus of cytoplasm (arrowed). The cytoplasm contains numerous dense bodies, some small and homogeneous (A) and some heterogeneous (B). Thorotrast is present in large quantities along the cell membrane, in vacuoles (or invaginations) and in dense bodies. Despite the large amounts of thorotrast attached to the membrane of the macrophage, little is attached to the closely adjacent liver cell, which has however, ingested a little thorotrast.

x 17,500

The cells conta ined numerous dense bodies resembling lysosomes, which for convenience will be described as such. (Figs. 8, 9, 12). Two types were identif ied in macrophages at all sites. Some were homogeneous s t ructures showing some


Fig. 7. Field from spleen 1 hour after intravenous administration of thorotrast. M paravascular macrophage, P neutrophil polymorph, L lymphocyte. Thorotrast is conspicuously adherent to the cell membrane of the macrophage in very much greater degree than to the other cells. Invaginations of membrane suggest that ingestion was continuing at the time of sacrifice, The macrophage shows a considerable quantity of granular endoplasmic reticulum and free ribosomes, some aggregated. A heterogeneous dense body (A) does not show thoro-

trast while several homogeneous dense bodies (B) do contain thorotrast. )<23,600

var ia t ion in size and shape, bu t basically similar. These may be regarded as p r imary lysosomes; others were heterogeneous s t ructures in which f ragments

362 I. CARR:

Fig. 8. Cytoplasm of macrophage from lymph node showing numerous small dense bodies of homogeneous structure. At A and B the dense bodies are elongated and at C two small bodies are present within one membrane. They are arranged typically round the Golgi region of the cell (G). Numerous free ribosomes are present, often aggregated. There are several

"coated vesicles" present. •

of cytoplasmic debris could be recognised. The two groups were fairly dis t inct bu t in termediate forms were recoglfised.

The former, homogeneous dense bodies or p r imary lysosomes (Figs. l l , 12) ranged in size from 0.1 to 0.8 ~. There was no systematic var ia t ion in size or


Fig. 9. Cytoplasm of macrophage from spleen showing microtubules in longitudinal and cross-section. • 46,000

Fig. 10. Cytoplasm of macrophage from lymph node showing microfibrils in a focal aggregate. • 48,5OO

structure from one part of the RES to another. Most profiles were circular. The body was bounded by a membrane composed of two dense lines separated by a light zone some 40A across. Deep to this there frequently lay a less electron dense area and deep to this again, in lead stained specimens numerous dense granules falling into two populations, one 80--100A in diameter and the other about 30A in diameter. The images of these granules have been followed in through focus sets of micrographs. The granules were absent or much less dense in speeimes not stained with lead. I t is conceivable tha t the larger granules may represent aggregates of the smaller ones. Some of these dense bodies contained considerable amounts of ferritin, notably in the spleen, while others contained little or none (Fig. 11).

Apart from circular profiles, similar ovoid or sausage-shaped lysosomes were present; in some cases the shapes of these suggested tha t they were channels of the smooth (agranular) endoplasmie reticulum filled with lysosomal material (Fig. 8).

The heterogeneous dense bodies, (Figs. 13, 14) which might be reasonably regarded as residual bodies, were bound by a membrane of similar dimensions to tha t around the homogeneous bodies and contained a matrix with dense

364 I. CARR:

Fig. 11. Homogeneous dense body some 0.6 ~ in diameter from macrophage in spleen. The surrounding three layered membrane shows an intermediate electron-lucent layer ---40 A in diameter; deep to the membrane is a thicker electron lucent layer. The internal substance is fairly homogeneous, with one large dense area. There is however, a fine granularity. This

is the close-to-focns member of a through-focus series. • 140,000

Fig. 12. Detail from the same negative as Fig. I1. The fine granularity consists of densities of various sizes. The possibility cannot be excluded that there is a continuous size distribution, but densities of 80--100 A (encircled) and --~30 A (~) may be readily identified. A few smaller

densities ~-~15 A may be seen. (v) The latter are of doubtful significance. •

granules s imilar to t h a t found in the homogeneous bodies. I n add i t ion t hey conta ined larger dense aggregates 300 - -1500A in d iamete r , f ibri l- l ike s t ruc tures abou t 1 ~ long • 150A across, of ten in lead s ta ined sect ions showing per iodic band ing with a per iod of 3 5 - - 5 0 A (Figs. 13, 14, 15), and also dense bodies a b o u t 150A in d i ame te r which could represen t cross-sections of the above f ibri l - l ike s t ructures . I t is no t safe on the presen t evidence to claim t h a t these b a n d e d s t ruc tures are t rue fibrils. Clearly recognisable inges ted red blood cells also occurred no t ab ly in the spleen, bu t are no t ve ry common. Macrophages conforming in s t ruc ture to this general descr ip t ion were seen in a pa ravascu la r posi t ion in spleen, l y m p h node, test is , PEYE~'s pa tch , t h y m u s and bone marrow. A few such cells were seen in the extra .vascular spaces of the l iver.

W h e n colloids were given in t ravenous ly , par t ic les were seen wi thin an hour in pa ravascu la r macrophages in spleen and liver, and to a minor degree in bone


~'ig. 15

Fig. 13. Typical heterogeneous dense body about 1.8 ~ long from Kupffer cell, containing several dense areas round numerous elongated rod-like profiles frequently branching and interlacing. Small circular densities are of the right order of size to represent cross-sections

of these rod-like structures. • 46,000

Fig. 14. Heterogeneous dense body from Kupffer cell. Prominent rod-like structures are present which have a regular repeating pattern of densities (arrowed). Circular profiles are present

which could represent cross-sections of the rod-like structures. • 94,000

Fig. 15. Detail of 14. The rod-structures are some 90 A in transverse diameter. The dense lines occur about every 40 A. The dense lines are some 15 A and the light lines some 25 A thick.

• 202,000

marrow. Lit t le uptake was seen in lymph nodes or PEYER'S patches. I n spleen and liver the following changes were seen in cells after in t ravenous adminis t ra t ion of colloids.

1. A layer of colloid was seen apparen t ly adheren t to the surface of the macrophages. This behaviour was most notable with thoro t ras t (Figs. 6, 7).

2. Invag ina t ions of cytoplasm were filled with colloid. Numerous vesicles filled with colloid may either have been cross-sections of these, or ma y have been t rue vesicles, i ndependen t of the surface.

3. Colloid was rapidly segregated wi thin dense bodies of the homogeneous type. Colloid was no t so commonly seen in heterogeneous dense bodies (Figs. 6, 7).

366 I. CARR:

Discussion

The clearance of colloids from the circulation as displayed in these experiments is clearly dependent on many factors, among them the phagocytic properties of cells lining the blood vessels, the degree of leakage of the colloid from the circulation through patent intercellular junctions, and the avidity of paravascular macrophages.

I t appears likely that the behaviour of endothelium toward colloids in the blood stream passing it will vary, notably with the particle size and perhaps charge of the colloid, with the rate of flow, and with particle concentration. There was much confusion in the older literature about endothelial phagocytosis and it seems clear tha t much tha t was reported as endothelial phagocytosis was in fact leakage between patent endothelial cell junctions. The only circumstances where endothelial cell phagocytosis has been shown undoubtedly to occur in vivo, is after repeated doses of colloid (CoTRAN, 1965). In the present experiments, however, fairly marked phagoeytosis occurred in certain sites after single doses of colloid.

The profiles of phagocytosing cells in an endothelial site, seen in electron micrographs, were of two types. Some were flattened cells of true endothelial type, while others were cells protruding much further into the lumen of the vessel, and having the characteristic morphology of macrophages. The lat ter cells, endothelial macrophages, were particularly notable in the liver. I t cannot be certain without extensive serial section studies whether there are genuinely two types of lining cell in the vessels of the liver, or a single kind of cell, (see ATm~MAN, 1963; ROUILLEI~ and JF, ZEQUEL, 1963 for review). However, the clear demonstration by BUI~KEL and Low (1965) tha t the structure of the wall of the hepatic sinusoid varies from one point to another in the sinusoid, makes it very likely that there are two types of cell. Intermediate forms may and probably do exist. The mode of ingestion will differ between the two types of cells, or areas. In one area mobile flaps or fingers will t rap particles, elsewhere the particles will sink into the cell.

The classical view of the mechanism of reticulo-endothelial clearance of colloids was tha t it was due to the "pectic activity of the phagocytes of the reticulo-endothelial system lining the vascular bed which is being studied, the suspension of particles normally used being unable to pass through the vessel walls." (HALPV,~N et al., 1953). Leakage was not noted in the vessels of the liver by WIENEE et al. (1964) in their studies of reticuloendothelial depression. HAMPTON (1958) and CASr~EY-SMITH and READE (1965) on the other hand pointed out tha t there was leakage between the endothelial cells of the liver after the administration of various colloids.

In the present experiments, irrespective of the detailed morphology of the wall of the hepatic sinusoid, it is certain tha t the colloids used did not induce generalised increased vascular permeability. I t is clear that both in the case of high doses of colloids such as thorotrast and to a lesser, but still quite marked extent, in the case of low doses of carbon suspensions such as are used in quanti- tat ive estimates of colloid clearance, in at least the mouse, leakage through open


gaps between endothelial cells in spleen and liver is par t of the normal mechanism of colloid clearance. I t seems certain therefore that apart from serological factors, the cellular mechanisms of colloid clearance are complex, and may conceivably in RE stimulation and depression vary in an unrelated way. I t is interesting tha t in the present experiments in the mouse, clearance by the marrow does not seem very significant. This is in contrast to the findings of HUHN and STEIDLE (1967) in the rat.

I t seems tha t the cells which avidly ingest intravenously injected foreign material have a number of common structural features, by which they can be identified, without prior administration of foreign material - - in contrast to the view of Moo~E et al. (1961) that "Clearly anatomical location cannot be used to describe a cell as belonging to the RES, nor are there any cytological features tha t can consistently be used." The anatomical location which allows a cell to remove colloid from the blood stream is either an endothelial position, or proxi- mity to a patent intercellular junction. The cytological features which, taken together, suggest that a cell belongs to the RES are the presence of numerous small homogeneous dense bodies, or pr imary lysosomes, of characteristic structure, the presence of heterogeneous dense bodies and the presence of prominent microtubules and mierofibrils. In addition it appears likely tha t these cells have a component on the surface membrane which binds foreign materials preferentially.

The small homogeneous dense bodies are similar to those seen in normal and stimulated peritoneal macrophages; in the latter site some contain acid phos- phatase (CA~R, 1968). I t is likely that they are pr imary lysosomes and tha t images such as Fig. 8 represent stages in their formation. In spleen in particular they often contain ferritin; this is in keeping with the idea tha t they are cell products since the cell presumably synthesizes ferritin from ingested iron-containing material. A common feature in these bodies both in the peritoneum and elsewhere is the presence of dense granules some of about 100 A diameter and some much smaller. The nature of these is quite unknown. They are well seen after lead staining but are difficult to see without it; this suggests that they are not composed of iron. I t is at tractive to suppose that they are made up of aggregates of molecules of hydrolytic enzymes, but there is at present no good evidence for this view. The dense bodies are characteristic of macrophages at all sites and may be regarded as the specific macrophage granules. Similar granules have been noted in pulmonary macrophages (ScHuLZ, 1958).

0~o]~ and TSUKADA (1964) indicated the presence in low-power micrographs of "specific granules" in reticulum cells in spleen, which they considered to be analogous to the granules of polymorphs, but which they thought to be formed by pinocytosis. No information is available on the mechanism of formation of these granules in macrophages other than peritoneal macrophages. Here the view has been advanced (CARI~, 1968) tha t they, or significant parts of them are elaborated in the endoplasmie reticulum and segregated in the Golgi region. In the present study the morphological evidence suggests that maerophages at all sites have a common structural pattern, and that this pat tern is similar to that of the peritoneal macrophage. If this is so, it might be reasonable to assume the same general pat tern of granule formation.

25 Z. Zellforsch., Bd. 89

368 I. CARR:

The heterogeneous dense bodies probably represent residual bodies (see DE DUVE, 1963). Macrophages presumably ingest degenerate red cells and lymphoid cells in large numbers; but in this strain of animals, they are not commonly recognised as such. The striated structures seen in these bodies may well be truly fibrillar in form, but this cannot be stated with certainty from the present micrographs. Similar fibrillar structures have been demonstrated by FIELD and RAINE (1966) in the neurones of sheep affected by scrapie, in their case 150 A in diameter with a period of 50 A i.e. of apparently the same dimensions as found here. The authors suggested that their micrographs represented coiled filaments, possibly composed of DNA of mitochondrial origin. No evidence of a helical structure has been obtained in the present study.

More complex inclusions have been observed by ATHANASSIADES et al. (1965) in human macrophages in various pathological conditions. These were composed of smaller filaments some 33 A in diameter, arranged in complex concentric shells, and were thought to be possibly the remains of either mast cell granules or lipid membranes. No conclusion can be reached on the origin of the periodic structures seen in the present study, but they are found in macrophages in all the sites examined. I t is likely that they are derived from ingested material rather than a cell product.

The property of adsorbing colloidal particles, as a preliminary to their ingestion seems common to macrophages at all sites. This adhesion was noted in the amoeba (BRANDT and PAPPAS, 1960) and in cells in liver (HAMPTON, 1958) and lymph node (RSHLICH and T6RS, 1965). I t is now well known that the surface of most cells is covered to varying degrees by a carbohydrate-rich layer. Such a coat is well seen on the surface of peritoneal macrophages when stained by the uranium- osmium technique of NACrlMAS (1965). I t seems reasonable to ascribe the adhesive properties of macrophages elsewhere to a similarly prominent surface layer, though it is not clear whether there are any quantitative or qualitative characteristics of the maerophage cell coat which can account for the very marked adhesion of particles.

There are numerous variations in detailed structure between macrophages at different sites, none of them obviously significant. Significant functional differences have however been noted; for instance alveolar macrophages are more dependent on aerobic respiration than peritoneal cells (OREN et al., 1964). This difference is explainable though not necessarily explained by contamination of the population with great alveolar cells. There is a systematic variation in behaviour, both as to rate of cell division, and rate of spreading in culture when macrophages from various sites are explanted in culture (BENNETT, 1966).

I t has been previously reported that when peritoneal macrophages are stimulated, their processes become longer and they form more lysosomes, (CARR, 1967b). The variations that do occur in macrophages throughout the body, and the differences from the relatively homogeneous peritoneal population can be explained on the following hypothesis. When foreign material is ingested by a macrophage, it may to a varying degree stimulate the cell to produce primary lysosomes, which then fuse with the ingested material. The structure of the maerophage at any time will vary according to how great and how recent the stimulus was,


and according to how much of the s t imulan t mater ia l persists wi thin the cell.

The prominence of cytoplasmic processes may be re la ted to recent surface s t imu-

lat ion and of microtubules to moti l i ty . F ina l ly the overal l shape of the macrophage will conform to tissue pressures, and will be modified accordingly as the cell has

an endovascular or a paravascular position.

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370 I. CARR: Fine Structure of the Reticulo-Endothelial System

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Dr. IAN CARR Department of Human Biology and Anatomy University of Sheffield Sheffield, Yorks., England

some aspects of the fine structure of the reticulo-endothelial system; the cells which clear...

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