On the Origin of the Connective-tissue Ground- substance ... · of connective-tissue formation is a...

On the Origin of the Connective-tissue Ground-substance in the Chick Embryo.

By

George A. Baitsell,

Osborn Zoological Laboratory, Yale University, New Haven, Conn.

With Plates 44r-7.

SZILY (26) demonstrated in the early development of. the chickembryo, as well as in certain other species, a cell-free, fibrous,connective-tissue ground-substance which was quite generallypresent, filling the various cavities of the embryonic body.This investigator was of the opinion that this material wasi n t r a c e l l u l a r in its origin, arising, as he believed, by atransformation of cell processes from any of the surroundingcells, without regard to the particular germ-layer to whichthey belonged, but previous to the advent of the inesenchymecells.

In a previous communication (5) I have presented resultswhich demonstrate, I believe, that in the Amphibia the processof connective-tissue formation is a purely i n t e r c e l l u l a rone in which, f i r s t , a homogeneous intercellular ground-substance is formed, apparently as a result of the secretoryactivities of the surrounding embryonic cells, and, second,this secreted material is gradually transformed into the varioustypes of connective tissue.

In view of the earlier work of Szily, as noted above, I feltit necessary to re-investigate the development of connectivetissue in the chick, in order to see if, in my opinion, the processas there shown presented evidence contrary to that obtainedfrom the study of the amphibian tissues. It may be stated

N O . -~G Q q

572 GEORGE A. BAITSELL

at this time that the results obtained in the present investiga-tion are in harmony with the intercellular theory of connec-tive-tissue development, and therefore in agreement withmy previous work on the development of connective tissue inthe Amphibia (5).

MATERIAL AND METHODS.

Access has been had to a large collection of serial sectionsof chick embryos ranging from 16-90 hours incubation. Thismaterial was preserved in sublimate-acetic (saturated aqueoussolution of mercuric chloride with 5 per cent, glacial acetic),sectioned in paraffin either at 8/A or at 10/x and stained withDelafield's haematoxylin and orange 6. For comparison aconsiderable number of embryos were preserved in Zenker'sfluid (with 5 per cent, glacial acetic) and stained with theMallory connective-tissue stain. In general it has been foundthat the sublimate-acetic fluid followed by the Delafield staingives, on the whole, the most satisfactory results for chicktissues.

In addition to a study of the prepared material^ considerableemphasis has been placed upon results obtained from thedissection of living chick embryos. The method employed inthis phase of the investigation is briefly given below (p. 579)in connexion with the description of results obtained.

STUDY OF THE PREPARED MATERIAL.

In fig. 1, PL 44, is shown a portion of a transverse sectionthrough an 11-somite embryo incubated 29 hours. Thesection is taken in the region of the 9th somite. It will be notedfrom the figure that at this stage a considerable area of theintercellular ground-substance is present surrounding thenotochord and,, in general, filling the spaces below the somitesand lateral to the medullary cord. The surrounding cells arequite regular, rounded bodies which are devoid of projectingcell processes. There can be no question in such cases but thatthe ground-substance has arisen as an intercellular secretion

CONNECTIVE-TISSUE GHOUND-SUBSTANCE 578

of certain surrounding cells and not from any direct trans-formation of cytoplasm. In this embryo a considerable degreeof fibrillation is present in the ground-substance. A study ofmaterial obtained from a number of embryos shows that thedegree of fibrillation varies considerably, although there isa very definite increase to be noted in the older embryos.Thus in some cases the ground-substance is apparently homo-geneous, while in other embryos of the same age considerablefibrillation is present. I am of the opinion that this conditionis due, at least in part, to the varying action of the preservingfluid. However, the point to be emphasized, which is veryevident in this figure, is that the fibrillation unquestionablyarises by changes which occur in the ground-substance.

A somewhat later stage is shown in fig. 2, PI. 44, which isdrawn from a transverse section of a 14-somite embryo atabout the 36th hour of incubation. The section is also takenthrough the region of the 9th somite. Compared with thecondition shown in fig. 1, PL 44, the ground-substance enclosingthe notochord and filling the various spaces is more dense, andthe fibrillation is also considerably more advanced. The distinc-tion- between the areas around the notochord filled with thefibrillated ground-substance and the empty cavities of theneural tube and the aorta is very sharp. It will be noted ina few cases that certain of the mesodermal cells lying in theperipheral layer of the somites have lost their spherical shapeand assumed a more or less spindle shape with definite processesextending out into the ground-substance. Sections of thisembryo taken anterior to the one here figured show, in general,more of the peripheral cells which are beginning a definitemigration into the ground-substance. On the other hand, insections of this embryo taken posterior to the one shown thetransformation of the peripheral cells is less advanced thanshown in this figure.

In fig. 3, PL 45, we have a portion of a transverse sectionthrough the tail region of an 18-somite embryo, incubatedabout 42 hours. The section shown in this figure is taken aconsiderable distance to the right of the median line as seen

Q q 2


under the microscope, and exhibits the separation of thesplanchnic and somatic layers of the mesoderm to form theextra-embryonic coelom on that side of the embryo. In thisfigure it is desired to call particular attention to the denselayer of heavily fibrillated ground-substance lying between themesoderm and ectoderm, and between the mesoderm andendoderm, thus uniting all the layers and forming a compactwhole. As in the previous figure the contrast between theempty coelomic cavity and the completely filled areas betweenthe mesoderm and the other two layers is very marked. Themigration of the mesoderm cells into the ground-substancehas not begun as yet in the region figured.

In fig. 4, PL 45, is shown a portion of a frontal section througha 27-somite embryo of 48 hours incubation. This section istaken at the level of the notochord, and gives a very clearpicture of the relations between the ground-substance, noto-chord, and mesodermal somites. The figure gives added con-firmation to that noted previously in the transverse sections,namely the presence of the large area of ground-substancesurrounding the notochord and extending around the somites.In this region, at the stage shown, this material is cell-free,but it can be seen that long processes from several of the meso-derm cells are beginning to extend considerable distances into it.Some of these cell processes in the ground-substance are drawnout quite fine, but it is possible to distinguish between thecytoplasmic process and the fibres present in the ground-substance. There can be no doubt that the two are inde-pendent structures.

Fig. 5, PL 46, is a portion of a transverse section through theregion of the 21st somite of a 27-somite embryo, incubated48 hours. The portion figured lies ventral to. the neural tubeand shows a section of the notochord. This region is of parti-cular interest at this stage because some of the peripheralmesoderm cells from the pair of somites are migrating into thecell-free area of the ground-substance. A comparison of thisfigure with figs. 1 and 2 shows the changes that are takingplace. It is evident that these cells in their movements from

CONNECTIVE-TISSUE GROUND-SUBSTANCE 575

the cell groups, just as was previously shown in the case ofthe frog embryo (5), make use of the ground-substance as asupporting substratum. Their shape has undergone consider-able modification from that of the earlier condition, and longprocesses have formed which extend into the ground-substancefor considerable distances. Emphasis should be laid upon thefact that the cytoplasm of the migrating mesoderm cells, evenwhen drawn out to a very fine process, can be differentiatedfrom the surrounding ground-substance, so that it is possibleto be certain of the independence of the two materials.

A condition such as is shown with particular clearness infig. 5, PI. 46, gives striking evidence that the movements ofthese embryonic cells in the chick embryo are governed bythe same factors as are the living cells in tissue cultures. Inthe latter condition it has been conclusively demonstrated byL. Loeb, Harrison, and others that cells are positivelystereotropic, and in order for their migration to take placeaway from the cell groups it is necessary that a supportingmaterial of some sort be provided.

If, for example, the cells in a tissue culture are suspendedin a liquid medium so that they are not in contact with anysupporting material, they will show no tendency to leave thecell groups and move out into the liquid. On the other hand,if the cells are surrounded by a more or less solid medium suchas a blood or plasma clot, or if in a liquid medium a fibrillarmaterial, such as a filament from a spider web, is provided thecells in contact with the supporting material will, in general,begin to migrate away from the mass of tissue and out into thesurrounding areas. Furthermore, in such a movement, whenthe cells make use of the fibrillar materials, it is clear that theirshape represents an adjustment to the environment. Thuswhen they are quiescent in the group they remain as sphericalbodies, but when they are undergoing active movement theybecome spindle shaped, with the long axis parallel to thedirection they are moving. This fact is shown to particularadvantage in cases where the cells are moving along a singlefibre, such as in a spider web (13).


The study of connective-tissue development, both in thefrog and the chick, goes to show clearly that the behaviourof the embryonic cells in the developing animal with regard tomigration is the same as in the tissue cultures. It is necessarythat the cells in the embryo have a supporting material inorder to enable them to leave the cell groups and migrate intoand through other regions of the embryo. Such a material isprovided in the embryo from the earliest stages by the ground-substance which arises as a cell secretion; structureless atfirst, but variously modified later as the cells wander through it.

In fig. 6, PI. 46, is shown a portion of a transverse sectionthrough an embryo of 36 somites at the 72nd hour of incuba-tion. The section is through the region of the 21st somite.At the top of the figure a portion of the neural tube is shown.Lying below the latter a transverse section of the notochordmay be noted. The latter is surrounded by the fibrillar inter-cellular material which has arisen by a modification of thesecreted ground-substance and which is now infiltrated withgreat numbers of mesenehyme cells. A comparison of thisfigure with those previously described, particularly fig. 5,PL 46, demonstrates very clearly the marked changes whichhave taken place in this region of the embryo, due to theactive median migration of the mesenehyme cells from theregion of the somite on each side.

Mall in his study of the development of connective tissue inthe tadpole and pig (20) shows a stage similar to fig. 6, PI. 46,but interprets it differently. He holds, in accordance withthe exoplasmic theory of connective-tissue formation, that thefibrillar material, such as is shown in fig. 6, PI. 46, arises froma syncytium of the mesenehyme cells and therefore that itis really a modified exoplasm of the cells, intercellular inposition, but since it arises, as he believes, by a transformationof the peripheral cytoplasm, it is really intracellular in itsformation. When the fact is taken into account that theforerunner of the cell-containing, fibrillar material shown infig. 6, PI. 46, is the secreted, cell-free ground-substance whichmay be found generally present throughout the embryo, it is


clear that the formation of this material must be the resultof an intercellular action.

It is interesting to note in connexion with the so-calledmesenchyme syncytia postulated by the exoplasmic theory,certain studies of W. H. Lewis, who, working with mesenchymetissue of chick embryos in tissue cultures (18), shows in a numberof cases that cells, which to all appearances were completelyfused to form a definite syncytium, were really separate entitieswhich retained their individuality at all times. What appearedto be a syncytium was in reality only a group of cells in closeapposition to each other. In a later paper (19) Lewis says heis convinced that the mesenchyme cells of chicks do not formsyncytia in the tissue cultures, and he thinks it highly probablethat the same thing is true with regard both to the embryosof birds and of mammals. Whether or not it is true that themesenchyme cells never form syncytia, it can be demonstratedthat a basic, more or less, fibrillated ground-substance is presentin the embryo p r e v i o u s to any mesenchymal syncytium.The latter, therefore, if such occurs, is not responsible for theformation of the ground-substance.

In considering the discrepancy between the results of Szily(26) and those here recorded, the chief difference appears tobe due to the failure of Szily to recognize the earliest stagesin the formation of the ground-substance. According to thepresent findings, the ' zellfreie faserige Stiitzgewebe ' of Szilyis not the primary stage, but previous to this condition thereis present in the chick embryo a homogeneous ground-substancewhich gradually becomes fibrillated. The fibres which formare purely intercellular, having no direct connexion with cellsbut arising by a direct transformation of the ground-substance.

Cond i t i on of t h e G r o u n d - s u b s t a n c e in t h eHeart.—The early developmental stages of the chick heartconstitute probably the most satisfactory region of the embryofor demonstrating the presence of a large amount of the cell-free ground-substance. Szily (26) clearly showed the presenceof this material in the heart. Also Davis at the 1924 meetingof the American Association of Anatomists (8) presented

578 GEOUGE A. BAITSBLL

results showing that a homogeneous transparent material,which he terms ' cardiac jelly ', is present in the living heartof 48- to 72-hour chick embryos. He sectioned the livingheart in Locke's solution and found that a fine probe ' in-sinuated between the myocardium and endocardium meetswith resistance, and separation of the two layers is accom-plished with great difficulty '. He reported, however, that hewas unable to demonstrate this material microscopically eitherin fresh preparations or in material which had been preservedand sectioned. He concludes ' that the substance between themyocardium and endocardium is a homogeneous transparentjelly '.

I have found no difficulty in confirming these observationsof Davis on the presence of this material in the living hearts,and also in demonstrating it microscopically as shown previouslyby Szily. In figs. 7, 8, and 9, PL 47, are shown three sectionsthrough portions of the developing heart as different stages.Fig. 7, PI. 47, shows an early stage taken from a 27-hourembryo. In this figure a cell-free layer of ground-substance ofconsiderable thickness is to be noted lying between the endo-cardium and myocardium. In part, this material even underthe magnification appears homogeneous, in part it is fibrillar.Since at this stage the cells of both the myocardium andendocardium are definite rounded bodies, it is possible to saywith certainty that they are not in direct connexion with thefibres. It is clear, therefore, that the fibrillation arises bya change in the character of the ground-substance. The relativeamount of fibrillation varies somewhat in different preparations,and, as has been previously noted, this may be due in somedegree at least to the action of the killing fluid.

In fig. 8, PI. 47, a section is shown through a portion of theheart of a 42-hour embryo. In comparison with the earlierstage, sections through the heart of an embryo of this stageconsistently show, as evidenced in the figure, a marked increasein the density and in the fibrillation of the ground-substance.Here, again, it is clear, as in the earlier stage, that the develop-ment of the fibres is due to a change in the character of the inter-


cellular substance, and not to the transformation of cellprocesses or any other direct cell connexion.

In fig. 9, PI. 47, a section is shown through a portion of theheart of a 72-hour embryo. Somewhat previous to thismesenchyme cells have begun a migration into the intercellularground-substance. To the left of the figure a considerablenumber of these more or less spindle-shaped cells are shownlying in the ground-substance and apparently stretching outalong the fibres. To the right of the figure a portion of theground-substance is shown in which considerable areas arestill cell-free.

STUDY OF THE INTERCELLULAR GROUND-SUBSTANCE IN

LIVING CHICK EMBRYOS.

• It is possible to demonstrate the presence of a transparent,gelatinous material in living chick embryos from very earlystages just as it is in frog embryos (4). The method employedhas been as follows. The egg is opened in a dish of warm normalsaline. Then the blastodermic area with the embryo is cutfrom the yolk and floated into a Syracuse dish with some of thesame fluid. In some cases dishes have been used which con-tained a layer of paraffin to which the blastoderm could bepinned in order to keep it in position. In other cases theblastoderm has been held in position by means of lead weights.The examination of the embryos had been done under a Zeissbinocular microscope using either the intense direct illuminationof a small arc lamp, or the indirect illumination from the mirror.Dissection of the embryo has been accomplished by the use ofglass needles drawn to the desired fineness over a small gasflame according to the methods used by Chambers (7) formicrodissection needles. Very fine scissors have also beenused. By these methods it has been found possible to carryout almost any dissection desired on the various stages.

The earliest stage dissected has been that of the primitivestreak, ranging from 15-18 hours incubation. The dissectionof such an embryo permits the demonstration of an inter-


cellular ground-substance in various ways. Thus it will befound by careful manipulation with fine needles that theectoderm in the embryonic area is adherent to an underlyingtransparent material between the ectoderm and mesoderm sothat an attempt to remove the former meets with resistance.It is apparent from a study of both living and prepared materialthat the mesoderm cells are separated from the ectodermexcept possibly in the immediate vicinity of the primitivestreak. If transverse sections of the embryo through the primi-tive streak are made, and the sections manipulated with needlesit will be found that the ectoderm, mesoderm, and endodermtend to keep their relative positions, and any attempt tocompress or extend meets with resistance. Furthermore, whenthe pressure or tension is released the germ-layers tend at onceto assume the original position. In separating groups of theembryonic cells, it will be found that they are adherent toeach other. This is apparently due not to an adhesion of thelimiting cell membranes but to the presence of a transparentintercellular material, small areas of which under favourableconditions can be detected in various regions between the cells.

An embryo of 48-hours incubation is one of the best stagesfor demonstration of the ground-substance in the livingembryo, just as it is in the prepared material. An embryo atthis age is large enough to permit a comparatively easy dissec-tion. Transverse sections at various levels afford clear demon-strations of the presence of a homogeneous ground-substancefilling the spaces between the germ-layers and in various othercavities of the embryonic body. The results from such anexamination confirm those obtained from the study of theprepared material. Thus if one examines the cut edge ofa transverse section of a living embryo taken posterior to thecervical flexure at about the 15th somite, as shown in fig. 10,PI. 47, it is possible by means of fine glass needles to demon-strate the presence of the jelly in the area surrounding thenotochord, and also in the space between ectoderm and meso-derm. Any attempt to alter the position of the notochordmeets with resistance, as does also an attempt to strip away the


ectoderm. On the other hand, the needles can easily be insertedinto the extra-embryonic coelomic cavities between the somaticand splanchnic layers of the mesoderm or into the neural canal,all of which are apparently entirely free from any enclosedmaterial. Furthermore, just as in the earlier stages anyattempt to alter the position of the germ-layers or othorstructures in an embryo of this stage meets with resistance,so one must conclude from such an examination that theintercellular ground-substance is of quite general occurrencethroughout the embryonic body.

DISCUSSION.

The above results obtained from the study of both theprepared and living material present a very clear picture as tothe origin of the connective tissues in the chick. The primaryfact is the presence of an intercellular ground-substancethroughout the embryo from the earliest stages of development,which arises as a secretion from the cells of the various germ-layers. In some cases it appears that the mesoderm cells arenot concerned in the formation of the ground-substance, forareas of the material may be found which are situated a con-siderable distance from the cells of this layer. On the otherhand, in the development of the heart, it is apparent that theabundant formation of the ground-substance must be due tothe mesoderm cells.

The ground-substance in the early stages appears almoststructureless save for a slight fibrillation. Because of this factit is not easy of detection in the prepared material even underfavourable conditions of staining, but it can be demonstratedby dissection of living embryos. The degree of fibrillation,although subject to some variation in embryos of the samestage, shows a definite increase in the older embryos, and it canbe demonstrated that this phenomenon is due to changes whichtake place in the ground-substance and therefore indepen-dent of direct cellular connexion. There has been no questionfor some time but that in the later stages the development of


connective tissue took place by an intercellular action. Sinceit is also apparent that the same condition is true for theearlier stages as well, it must be concluded that connective-tissue formation is purely an intercellular process.

In connexion with the connective-tissue problem attentionshould be called to a noteworthy contribution by Harrison (14)obtained from his recent study of the development of thebalancer in Amblystoma, which, as he shows, affords idealmaterial for a study of connective-tissue development. Theresults presented afford a conclusive demonstration of thepresence of an intercellular, secreted ground-substance andthe gradual transformation, by changes in the material itself, ofa peripheral region to form a basement membrane. In thebalancer it is clear that the undifferentiated ground-substancearises as an intercellular formation of the mesoderm cells.The peripheral basement membrane which arises from thismaterial is ' composed of a dense felt of reticulum fibres 'and is formed under the influence of the surrounding epithelialcells. As to the type of influence exerted by the epithelialcells upon the ground-substance in order to form the basementmembrane, Harrison ' naturally thinks of an enzyme actionwhich condenses or coagulates the diffuse intercellular ground-substance transforming it into a fibrillar tissue which givesalmost the same chemical reactions as reticulum '.

' The balancer membrane thus affords a clear proof thatconnective-tissue fibres take origin in an amorphous ground-substance, independently of any direct action on the part ofthe mesenchyme cells. The fibres composing the membraneare neither intracellular nor are they formed in any particularoutside layer which could be called exoplasmic, unless thewhole of the intercellular ground-substance be designated assuch.' It is difficult to see how a more conclusive result couldbe obtained, or one which was more in harmony with theposition I have maintained for several years.

I am of the opinion that considering the results which arenow available the question of the origin of connective tissuemust be regarded as settled in favour of the intercellular

CONNECTIVE-TISSUE GEOUND-SUBSTANCE 583

theory. The next step is the definite linking up by conclusivechemical studies of the process of connective-tissue formationin the embryo with the process of wound healing as well asvarious other pathological conditions in the mature animal inwhich, in some cases at least, the transformed plasma clotplays such an important role. In this connexion it may beworth while to quote a paragraph from a previous paper (5).

' From the morphological standpoint the results of the presentstudy indicate that the formation of connective tissue in theamphibian embryo is similar to the process which takes placein transformation of the plasma clot. The intercellular ground-substance of developing connective tissue may therefore becompared in its morphology to the plasma clot. This ground-substance when first formed appears homogeneous or witha fine fibrillation. The process of transformation into a fibroustissue is a progressive one. The fibrillation increases, bundlesof fibres are formed, and in time the entire ground-substance,which at first showed such a high degree of homogeneity,becomes transformed into a fibrous tissue. It is indicatedthat this transformation occurs as the results of the introduc-tion of mechanical factors in the embryo. These factors maybe due to certain lines of tension in the embryo correspondingto the inherent polarity of the organism or, just as in theplasma clot, the movements of the cells through the ground-substance may introduce mechanical factors which aid in thetransformation of the ground-substance into a fibrous tissue.The cells, however, are to be regarded primarily as assimilativeand secretory agents, chiefly concerned in the formation of theundifferentiated ground-substance.'

The fact that the primary factor in connective-tissue forma-tion is a secrete'd intercellular ground-substance, which issecondarily invaded by mesenchyme cells and modified invarious ways, necessarily leads to an application of this know-ledge to certain features of development other than connective-tissue formation. In the first place, taking into account thefact (p. 575) that cells require a supporting material whenmoving away from cell groups, it is apparent that the presence


of the secreted, gelatinous ground-substance in an embryofrom the earliest stages is the s ine qua nonfor the migra-tion of isolated embryonic cells away from the cell groups andthe development of mesenchyme throughout the body. Forexample, as is well known in the chick embryo, the mesodermis first differentiated along the primitive streak. Prom thisregion the cells migrate laterally and anteriorly between theectoderm and endoderm. Now from the dissection of theliving embryo it can be demonstrated that the space betweenthe ectoderm and endoderm even at this very early stagecontains a gelatinous supporting material, the primitiveground-substance. The conclusion appears warranted thata type of cellular movement, such as is exhibited by the meso-derm cells at this stage in which isolated cells move away fromthe cell groups, is dependent in the embryo, just as it is intissue cultures, upon the presence of the secreted ground-substance.

Another striking example in the embryo of the relationsbetween the ground-substance and cell movement may benoted in the outgrowth of the nerve-fibre from the neuroblastas demonstrated by Harrison in tissue cultures (12). In thistype of movement the cell-body retains its position in the wallof the neural tube, but gradually form a process which extendsa considerable distance from the cell-body as a nerve-fibre.

Harrison in a later paper (13) says : ' With regard to themovements of the growing nerve-fibre the evidence . . . isnot quite so varied, but it is sufficient to warrant the con-clusion that also this protoplasm is stereotropic. No freeoutgrowth of nerves in a fluid medium has ever been observed,while such solids as the fibrin clot and smooth glass surfacesserve readily to support them, as do the surfaces of the largercell-masses and the interstitial protoplasmic network inside theembryo.'

In connexion with the normal growth of the nerve-fibrein the embryo an examination of the figures of Held as givenin his monograph on the development of nerves in the verte-brates (16) is very instructive in that they show many instances


in which the growing nerve-fibres are apparently utilizing amore or less fibrillated ground-substance in their extensionthrough the embryonic .spaces.

Finally, it should be noted that the early and abundantsecretion of a ground-substance by the embryonic cells may bean important factor in the growth of the embryo. In otherwords, it is possible that, at certain stages of development, thegrowth of an embryo may not be due so much to an increasein the number of cells formed by cell division as to an increasein the amount of the secreted ground-substance. If, forexample, one compares a chick embryo in the primitive streak-stage with a 48-hour embryo, a striking feature is the com-paratively great increase in the amount of the ground-substancepresent. On this basis one may postulate in development ofthe chick a series of cycles in which a period of cell divisionin certain regions of an embryo will be followed in turn bya period of inactivity as regards cell division, but a period inwhich the assimilative and secretory processes of the cellsattain their maximum activity thus resulting in an abundantformation of the ground-substance. The latter is at onceutilized as a supporting material and as the basic substance,under the influence of the invading cells, for the formation ofconnective tissue and various other structures.

SUMMARY.

1. In the chick, just as has been previously shown in theamphibian embryo, the forerunner of the connective tissues isa transparent, gelatinous, cell-free ground-substance which, ingeneral, pervades the embryonic body from very early stagesof development.

2. "This ground-substance can be demonstrated in variousregions of the body previous to the appearance of the mesen-chyme cells. It is evidently formed as a secretion of the cellsof the various germ-layers. The data at hand indicate nopossibility that it arises either from a syncytium or by a directtransformation of the cytoplasm.

586 GEORGE A. BAITSBLL

3. In the early stages the ground-substance appears, forthe most part, homogeneous, but with some fibrillation. Inthe later stages a progressive increase in the fibrillation isnoted. It is possible to show that the development of fibresin the ground-substance is due to changes in the ground-substance itself, and not to a direct transformation of cytoplasm.

4. The formation of the ground-substance is followed by theinvasion of the mesenchyme cells which, using it as a supportingmaterial, apparently in the same way that cells utilize theplasma clot in tissue cultures, move through and modify it invarious ways.

5. Emphasis is laid upon the fact that the presence of theground-substance through the embryo, together with the knownpositive stereotropism of cells as exhibited in tissue cultures,throws light upon certain features of early development,particularly cell movements and increase in body size.

BIBLIOGRAPHY.

1. Baitsell, G. A.—" The origin and structure of a fibrous tissue whichappears in living cultures of adult frog tissues ", ' Journ. Exp.Med.', vol. 21, 1915.

2. " The origin and structure of a fibrous tissue formed in woundhealing ", ibid., vol. 23, 1916.

3. " A study of the clotting of the plasma of frog's blood and thetransformation of the clot into a fibrous tissue ", ' Amer. Journ.Physiol.', vol. 44, 1917.

4. " Observations on the connective-tissue ground-substance inliving amphibian embryos ", ' Proc. Soc. Exper. Biol. and Med.',vol. 17, 1920.

5. " A study of the development of connective tissue in amphibia ",' Amer. Journ. Anat.', vol. 28, 1921.

6. " The development of connective tissue in the chick embryo ",' Proc. Amer. Assoc. Anats.', p. 194, ' Anat. Rec.', vol. 27, 1924.

7. Chambers, R.—"New apparatus and methods for the dissection andinjection of living cells " , ' Anat. Rec.', vol. 24, 1922.

8. Davis, C. L.—" Cardiac jelly in the chick embryo ", ibid., vol. 27, 1924.9. Flemming, W.—" t)ber die Entwickelung der collagenen Bindegewebs-

fibrillen bei Amphibien und Saugetieren ", ' Arch. f. Anat. u. Phys.,Anat. Abt.\ Bd. 21, 1897.


10. Hansen, Fr. C. C.—" Uber die Genese einiger Bindegewebsgrundsub-stanzen " , ' Anat. Anz.', Bd. 16, 1899.

11. " Untersuchungen iiber die Gruppe der Bindesubstanzen. I. DerHyalinknorpel ", ' Anat. Hefte ', Bd. 27, 1899.

12. Harrison, R. G.—" The outgrowth, of the nerve-fibre as a mode ofprotoplasmic movement", ' Journ. Exp. Zool.', vol. 9, 1910.

13. "The reaction of embryonic cells to solid structures", ibid.,vol. 17, 1914.

14. " The development of the balancer in Amblystoma, studied bythe method of transplantation and in relation to the connective-tissue problem ", ibid., vol. 41, 1925.

15. Heidenhain, M.—' Plasma und Zelle ', Abt. 1, Jena, I, 1907.16. Held, H.—' Die Entwicklung des Nervengewebes bei den Wirbel-

tieren', Leipzig, 1909.17. Hertzler, A. E.—" The development of fibrous tissues in peritoneal

adhesions ", ' Anat. Rec.', vol. 9, 1915.18. Lewis, W. H.—" Is mesenchyme a syncytium ? " ibid., vol. 23, 1923.19. " The adhesive quality of cells ", ibid.20. Mall, F. P.—" On the development of the connective tissues from the

connective-tissue syncytium ", ' Amer. Journ. Anat.', vol. 1, 1902.21. Matsumoto, S.—" Contribution to the study of epithelium movement.

The corneal epithelium of the frog in tissue culture " , ' Journ. Exp.Zool.', vol. 26, 1918.

22. Merkel, Fr.—" Betrachtungen iiber die Entwickelung des Binde-gewebes " , ' Anat. Hefte ', Bd. 38, 1908.

23. Nageotte, J.—" Les substances conjonctives sont des coagulumsalbuminoides, &c.", ' Compt.-rend. de la Soc. de Biol.', vol. 79,1916.

24. " Croissance, modelage et m6tamorphisme de la trame fibrineusedans les caillots cruoriques ", ' Compt.-rend. de l'Acad. des Sci.',vol. 170, 1920.

25. Studnicka, F. K.—" Uber einige Grundsubstanzgewebe ", ' Anat.Anz.', Bd. 31, 1907.

26. Szily, Al. v.—" Uber das Entstehen eines fibrillaren Stiitzgewebes imEmbryo und dessen Verhaltnis zur Glaskorperfrage", ' Anat.Hefte ', Bd. 35, 1908.

NO. 276


DESCRIPTION OF PLATES 44, 45, 46, AND 47.

ABBREVIATIONS (which apply to all figures).

AOE., aorta ; ECT., ectoderm ; END., endoderm ; M.C., mesoderm cellMES., mesoderm ; NC, notochord; N.T., neural tube ; G.S., ground-sub-stance ; E.E.C, extra-embiyonic ooelom; EN., endocardium; MYO.,myocardium.

PLATE 44.

Fig. 1.—Portion of a transverse section of a 29-hour, 11-somite chickembryo in the region of the 9th somite, x 634. The notochord is shownsurrounded by a more or less fibrillated, cell-free area of the ground-substance which extends laterally on each side under the somites anddorsally between the neural tube and the somite.

Pig. 2.—Portion of a transverse section of a 36-hour, 14-somite chickembryo in the region of the 9th somite, x 317. As compared with fig. 1the ground-substance shows a greater density and fibrillation. It ispractically cell-free, but certain cells in the peripheral layer of the somitescan be seen to be assuming a spindle-shape and to extend out into theground-substance.

PLATE 45.

Kg. 3.—Portion of a transverse section of a 42-hour, 18-somite chickembryo in the tail region. x317. The figure is taken a considerabledistance to one side of the mid-line and shows a portion of the extra-embryonic coelomic cavity of that side. Between the ectoderm and themesoderm and between the mesoderm and the endoderm the space is filledwith a dense fibrillated ground-substance which stands out in markedcontrast to the empty space of the coelomic cavity.

Fig. 4.—Portion of a frontal section of a 48-hour, 27-somite chick embryoat the level of the notochord. x 317. The notochord is shown embedded inconsiderable area of dense fibrillated cell-free ground-substance into whicha considerable number of the mesodermal cells from the somites arebeginning to extend long processes. It is possible to differentiate betweenthe fibres in the ground-substance and the cell processes and to be sure thatthe two structures are independent.

PLATE 46.

Fig. 5.—Portion of a transverse section of a 48-hour, 27-somite chickembryo in the region of the 21st somite, x 634. In this figure the begin-ning of the migration of the mesodermal cells into the area of the ground-substance surrounding the notochord is to be seen.

rCONNECTIVE-TISSUE GROUND-SUBSTANCE 589

Fig. 6.—Portion of the transverse section of a 72-hour, 36-somite chickembryo in the region of the 21st somite, x 634. This region is the sameas in fig. 5, and shows a considerably later stage in the migration of themeaodermal cells into the ground-substance surrounding the notochord.

PLATE 47.

Tig. 7.—Transverse section through the developing heart in a 27-hourchick embryo. x317. A considerable area of more or less fibrillatedground-substance is shown lying between the endocardium and myocar-dium.

Pig. 8.—Transverse section through the heart of a 42-hour chick embryo.X317. The figure shows a large area of dense and fibrillated cell-freeground-substance lying between the endocardium and myocardium.

Fig. 9.—Transverse section through the heart of a 72-hour chick embryo.X634. To the left of the figure is shown the invasion of the fibrillatedground-substance by the mesoderm cells, and to the right a considerablearea of the cell-free ground-substance.

Fig. 10.—A transverse section through a living 48-hour chick embryoin the region of the 15th somite, just posterior to the cervical flexure.X15 ± . In such a section the presence of the gelatinous ground-substancecan be detected in the area surrounding the notochord and lying betweenthe ectoderm and mesoderm and between the mesoderm and endoderm.These areas filled with the ground-substance are clearly differentiated fromempty cavities such as those of the extra-embryonic coelom and of theneural tube.

R r 2

QvuarlJour-nMUr. Sci. Vol.69MS.Pl.4.4>

NT.SOM.

AOR.

NT. ECT.

SOM.

G.S.

Quart.Journ.Micr.Sd. Vol.69.N.S.Pl.45.

ECT.

G.S

G.S

END

SOM. G.S G.S. SOM.

Qwcurt. Journ. Micr.Sci. Vol.69.N.S.Pl.46.

NC.

G.S.

N.T.

NC.

Quart Joum.Micr.Sd. Vol.69.MSPl. 4>7

MYO.

G.S.

M Y O .

GS.

8

G.S.

On the Origin of the Connective-tissue Ground- substance ... · of connective-tissue formation is a...

Documents

Transcript of On the Origin of the Connective-tissue Ground- substance ... · of connective-tissue formation is a...