65
J. Anat. (1976), 121, 1, pp. 65-70 With 7 figures Printed in Great Britain
A presomite human embryo of Horizon VII R. E. REWELL AND R. G. HARRISON
Department ofPathology, The Women's Hospital, Liverpool, and Department of Anatomy, The University of Liverpool
(Accepted 20 April 1975) INTRODUCTION
The early presomite human embryo described in this paper showed some interesting features at variance with those of other embryos of a similar age, including certain slightly abnormal ones. It is proposed to designate the embryo 'Liverpool III,'. MATERIAL AND METHODS
A woman aged thirty years presented with a request for ligation of her uterine tubes as a contraceptive measure. She had given birth to five living children and had experienced three spontaneous abortions. The operation, together with a customary curettage of the uterus, was performed on 24 June. Her youngest child was then aged 7 months. It is not known when she started to menstruate again after his birth, but she recorded a normal menstrual period the first day of which was on 21 May, i.e. 34 days before operation. Unfortunately her coital history was not recorded, but, assuming a cycle length of 28 days, ovulation may be presumed to have taken place on or about 4 June and fertilization on or about 6 June, with a time gap between fertilization and curettage of about 18 days. The curettings were placed immediately in Bouin's fluid and left for about 7 hours. Pieces for section, chosen late the same afternoon, were transferred to formol-saline in a tissue-processing machine which ran overnight. They were blocked the next morning and sectioned and stained that afternoon. On microscopic examination the endometrial glands were seen to display a late progestational response; the arterioles showed no premenstrual dilatation. However, the progestational response was not as advanced as is usually demanded for the diagnosis of early pregnancy, nor was the formation of decidua beyond what is often seen in premenstrual specimens. One fragment of tissue was found to contain an early embryo. Serial sections were made at 7 um and stained with Weigert's haematoxylin and eosin. Fortunately, the entire embryo remained in the block and was so aligned that the embryo was cut in the transverse plane at an angle of 10 degrees caudal to the vertical. Unfortunately, however, the fixation (intended only for routine histological diagnosis) had caused shrinkage of tissues and some distortion of the embryo, so that the ectoderm of the embryonic plate had become buckled ventrally.
5
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66
R. E. REWELL AND R. G. HARRISON
,~~~~~~~~ 4' -
Sf
4''
1.4.
...
~
~ ~ 1.
A~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~A.
-'.
~
-~~~~~ I.
~
~
Fig. 1. A photomicrograph of the chorionic vesicle at its mid-point. The exocoelomic membrane is visible, and the villi are clearly quite primitive. The buckling of the ectoderm of the embryonic plate can also be seen. x 65. DESCRIPTION OF THE SPECIMEN
A complete section through the chorionic vesicle at its mid-point is shown in Figure 1. It measured 0-714 x 0 526 mm in its largest diameters. In the surrounding endometrium there was no necrosis of decidual cells, despite statements to the contrary in earlier descriptions of embryos of comparable age. The endometrial glands were compressed, but none were seen to have been broken open. Some Fig. 2. Photomicrograph of the decidua capsularis of the embryo. The periphery of the chorionic vesicle is visible in the lower part of the figure. x 120. Fig. 3. The ectoderm of the embryonic plate is seen to be buckled ventrally and the intraembryonic mesoderm is continuous with the extraembryonic mesoblast at the lateral margins of the embryonic disc. x 190. Fig. 4. Transverse section through the cephalic end of the embryo, showing narrowing of the amniotic cavity and a ventral proliferation of the ectoderm to form an eccentric nodule in contact with the subjacent endoderm. x 190. Fig. 5. Photomicrograph showing early branching of the chorionic villi. x 190. Fig. 6. Vacuolation of cytotrophoblast cells at the tip of a chorionic villus. x 770. Fig. 7. The erosion of the endothelial wall of a capillary by syncytiotrophoblast. x 490.
A presomite human embryo
67 I0
3
5-2
R. E. REWELL AND R. G. HARRISON 68 contained blood, though this could have been a consequence of the trauma of the curettage. A well-marked zone of lymphocytic infiltration was present in the decidua on the maternal side of the vesicle. The embryo was completely embedded, the decidua having closed over the implantation site (Fig. 2). At one point blood was visible within the tissues between the chorion and the uterine cavity, but as this was mainly at the edge of the implantation site it could well have been the result of trauma. Extraembryonic mesoderm (mesoblast) was present and formed a well-marked somatopleuric layer 0-072 mm thick, which extended into the villi. The mesoblast was limited centrally by an exocoelomic (Heuser's) membrane (Fig. 1). The endoderm of the embryonic disc measured 0392 mm in cephalo-caudal length, the ectoderm only 0238 mm. In other words, there was a 0077 mm overlap of the yolk sac at each end of the embryo. This was probably the result of the buckling of the ectoderm so that its margins were raised from the endoderm. Similarly, the endoderm had a maximum transverse diameter of 0090 mm, but the distortion of the ectoderm of the plate prevented accurate assessment of its width. The separation of the two germ layers revealed the presence of intraembryonic mesoderm between them, and this was clearly continuous with the extraembryonic mesoblast at the lateral margins of the disc (Fig. 3). A primitive streak was present and its cells could be seen to be continuous with intraembryonic mesoderm laterally. At its cephalic extremity, the amniotic cavity narrowed considerably (Fig. 4). At this point the ectoderm of the disc showed a proliferation ventrally, forming an eccentric nodule which was in close contact with the subjacent endoderm. At this point of contact the endoderm also showed a proliferation of cells into the lumen of the yolk sac. Villi were most noticeable at the embryonic pole of the chorionic vesicle; only a very few of them were beginning to branch (Figs. 1, 5). Differentiation into cytotrophoblast and syncytiotrophoblast had begun, this being most marked at the embryonic pole. Mitoses were not found in the syncytial nuclei, but where the differentiation into two layers was least advanced, formation of syncytium by fusion of cells could be discerned. The cells of the cytotrophoblast were sufficiently well differentiated to be vacuolated (Fig. 6). Syncytial buds were forming all over the surface of the chorion and syncytial streamers were obvious, especially at the embryonic pole. Endometrial capillaries at the embryonic pole were dilated to form sinusoids which communicated with lacunae formed in the syncytiotrophoblast. However, there was as yet no one vessel much larger than others to serve as the main channel for drainage of blood. The lacunar spaces at the embryonic pole intercommunicated. At one point the syncytiotrophoblast had eroded the wall of a capillary and bulged into its lumen (Fig. 7). It has long been known that trophoblast may be found in the maternal circulation later in pregnancy (Schmorl, 1893), but it was surprising to observe invasion of a capillary as early as this. No fibrin (or fibrinoid) could be detected between trophoblast and decidua in these haematoxylin and eosin sections. DISCUSSION
This embryo can be assigned to Horizon VII of Streeter, i.e. 'Branching villi, axis of germ disc defined' (Streeter, 1942). Comparison of its size, histological characteristics, menstrual and embryological age with others of the same Horizon
A presomite human embryo
69
reveals that there are considerable variations between this embryo and those already documented. Thus the embryo described by Ingalls (1918) was thought on clinical grounds to have been dated 17-18 days after fertilization. It was not so fully embedded as the present embryo, the maternal blood in the intervillous spaces varied greatly in amount between different areas and yet it was further advanced in having a better developed primitive streak. By contrast, the embryo described by Heuser (1932), also thought to be of 18 days development, was in some respects further developed than the Liverpool III embryo; a definite notochordal canal was present, for instance. Concerning younger embryos, that of Brewer (1938) was thought to be of 11-12 days development. It was not embedded completely, villi were present but unbranched, no allantois was found and the cells forming the yolk sac were grouped together in clusters. The primitive streak was confined to the five caudal sections made; indeed it was hardly a distinct structure, being represented merely by an intermingling of the cells of the two layers of the germ disc. In the embryo of Streeter (1926), thought to be of 10-11 days development, the trophoblast had already begun to erode maternal capillaries, but the trophoblastic lacunae were not yet flooded with blood. The 16 day embryo reported by Harrison & Jeffcoate (1953) showed an early stage of the primitive streak. The true (secondary) yolk sac appeared to have formed and the blastocoele was largely occupied by extraembryonic mesoblast. The present embryo, whilst similarly displaying a yolk sac, also had a very clearly developed exocoelomic membrane (Fig. 1). Yet an earlier embryo of 13} days (Lewis & Harrison, 1966) showed an elongated yolk sac duct in process of being constricted off from the embryo. These findings are not easy to reconcile with theories regarding disposal of the exocoelomic membrane by means of a yolk sac duct if these embryos belong to a uniform population with constant time relationships. The thickened nodule in the ectoderm of the embryonic plate in this embryo showed similarities with a thickening in the ectoderm of a pathological presomite embryo (Harrison, Jones & Jones, 1966). This latter specimen showed well-marked syncytiotrophoblastic streamers and a pronounced endometrial reaction of a degree comparable to that displayed in the present embryo but much greater than that seen in the embryos of Harrison & Jeffcoate (1953) and Lewis & Harrison (1966). Such findings suggest that the degree of development, and the possible presence of abnormal features, in such early embryos, may be related to the degree of endometrial reaction, which in turn may possibly be dependent on the extent of immunological conformity between maternal and embryonic tissues. SUMMARY
A presomite embryo of Horizon VII aged approximately 18 days is described and illustrated. It is compared with some other embryos of a similar age, and a considerable variation of histological characteristics within the same Horizon is noted.
70
R. E. REWELL AND R. G. HARRISON
REFERENCES BREWER, J. 1. (1938). A human embryo in the bilaminar blastodisc stage (the Edwards-Jones-Brewer ovum). Contributions to Embryology of the Carnegie Institute of Washington 27, 85-93. HARRISON, R. G. & JEFFCOATE, T. N. A. (1953). A presomite human embryo showing an early stage of the primitive streak. Journal ofAnatomy 87, 124-129. HARRISON, R. G., JONES, C. H. & JONES, E. P. (1966). A pathological presomite human embryo. Journal of Pathology and Bacteriology 92, 583-584. HEUSER, C. H. (1932). A presomite human embryo with a definite chorda canal. Contributions to Embryology ofthe Carnegie Institute of Washington 23, 251-267. INGALLS, N. W. (1918). A human embryo before the appearance of myotomes. Contributions to Embryology of the Carnegie Institute of Washington 7, 111-134. LEWIS, B. V. & HARRISON, R. G. (1966). A presomite human embryo showing a yolk-sac duct. Journal ofAnatomy 100, 389-396. SCHMORL, G. (1893). Pathologische-anatomische Untersuchungen iiber Puerperal-Eklampsie. Leipzig: Vogel. STREETER, G. L. (1926). The " Miller" ovum, the youngest normal human embryo thus far known. Contributions to Embryology of the Carnegie Institute of Washington 18, 31-48. STREETER, G. L. (1942). Developmental horizons in human embryos. Contributions to EmbrVology of the Carnegie Institute of Washington 30, 211-245.
J. Anat. (1976), 121, 1, pp. 65-70 With 7 figures Printed in Great Britain
A presomite human embryo of Horizon VII R. E. REWELL AND R. G. HARRISON
Department ofPathology, The Women's Hospital, Liverpool, and Department of Anatomy, The University of Liverpool
(Accepted 20 April 1975) INTRODUCTION
The early presomite human embryo described in this paper showed some interesting features at variance with those of other embryos of a similar age, including certain slightly abnormal ones. It is proposed to designate the embryo 'Liverpool III,'. MATERIAL AND METHODS
A woman aged thirty years presented with a request for ligation of her uterine tubes as a contraceptive measure. She had given birth to five living children and had experienced three spontaneous abortions. The operation, together with a customary curettage of the uterus, was performed on 24 June. Her youngest child was then aged 7 months. It is not known when she started to menstruate again after his birth, but she recorded a normal menstrual period the first day of which was on 21 May, i.e. 34 days before operation. Unfortunately her coital history was not recorded, but, assuming a cycle length of 28 days, ovulation may be presumed to have taken place on or about 4 June and fertilization on or about 6 June, with a time gap between fertilization and curettage of about 18 days. The curettings were placed immediately in Bouin's fluid and left for about 7 hours. Pieces for section, chosen late the same afternoon, were transferred to formol-saline in a tissue-processing machine which ran overnight. They were blocked the next morning and sectioned and stained that afternoon. On microscopic examination the endometrial glands were seen to display a late progestational response; the arterioles showed no premenstrual dilatation. However, the progestational response was not as advanced as is usually demanded for the diagnosis of early pregnancy, nor was the formation of decidua beyond what is often seen in premenstrual specimens. One fragment of tissue was found to contain an early embryo. Serial sections were made at 7 um and stained with Weigert's haematoxylin and eosin. Fortunately, the entire embryo remained in the block and was so aligned that the embryo was cut in the transverse plane at an angle of 10 degrees caudal to the vertical. Unfortunately, however, the fixation (intended only for routine histological diagnosis) had caused shrinkage of tissues and some distortion of the embryo, so that the ectoderm of the embryonic plate had become buckled ventrally.
5
ANA ItI
s - S ' ; ' @ * ' S s ; * - ' *$ > j S f X w , ~ I '4
66
R. E. REWELL AND R. G. HARRISON
,~~~~~~~~ 4' -
Sf
4''
1.4.
...
~
~ ~ 1.
A~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~A.
-'.
~
-~~~~~ I.
~
~
Fig. 1. A photomicrograph of the chorionic vesicle at its mid-point. The exocoelomic membrane is visible, and the villi are clearly quite primitive. The buckling of the ectoderm of the embryonic plate can also be seen. x 65. DESCRIPTION OF THE SPECIMEN
A complete section through the chorionic vesicle at its mid-point is shown in Figure 1. It measured 0-714 x 0 526 mm in its largest diameters. In the surrounding endometrium there was no necrosis of decidual cells, despite statements to the contrary in earlier descriptions of embryos of comparable age. The endometrial glands were compressed, but none were seen to have been broken open. Some Fig. 2. Photomicrograph of the decidua capsularis of the embryo. The periphery of the chorionic vesicle is visible in the lower part of the figure. x 120. Fig. 3. The ectoderm of the embryonic plate is seen to be buckled ventrally and the intraembryonic mesoderm is continuous with the extraembryonic mesoblast at the lateral margins of the embryonic disc. x 190. Fig. 4. Transverse section through the cephalic end of the embryo, showing narrowing of the amniotic cavity and a ventral proliferation of the ectoderm to form an eccentric nodule in contact with the subjacent endoderm. x 190. Fig. 5. Photomicrograph showing early branching of the chorionic villi. x 190. Fig. 6. Vacuolation of cytotrophoblast cells at the tip of a chorionic villus. x 770. Fig. 7. The erosion of the endothelial wall of a capillary by syncytiotrophoblast. x 490.
A presomite human embryo
67 I0
3
5-2
R. E. REWELL AND R. G. HARRISON 68 contained blood, though this could have been a consequence of the trauma of the curettage. A well-marked zone of lymphocytic infiltration was present in the decidua on the maternal side of the vesicle. The embryo was completely embedded, the decidua having closed over the implantation site (Fig. 2). At one point blood was visible within the tissues between the chorion and the uterine cavity, but as this was mainly at the edge of the implantation site it could well have been the result of trauma. Extraembryonic mesoderm (mesoblast) was present and formed a well-marked somatopleuric layer 0-072 mm thick, which extended into the villi. The mesoblast was limited centrally by an exocoelomic (Heuser's) membrane (Fig. 1). The endoderm of the embryonic disc measured 0392 mm in cephalo-caudal length, the ectoderm only 0238 mm. In other words, there was a 0077 mm overlap of the yolk sac at each end of the embryo. This was probably the result of the buckling of the ectoderm so that its margins were raised from the endoderm. Similarly, the endoderm had a maximum transverse diameter of 0090 mm, but the distortion of the ectoderm of the plate prevented accurate assessment of its width. The separation of the two germ layers revealed the presence of intraembryonic mesoderm between them, and this was clearly continuous with the extraembryonic mesoblast at the lateral margins of the disc (Fig. 3). A primitive streak was present and its cells could be seen to be continuous with intraembryonic mesoderm laterally. At its cephalic extremity, the amniotic cavity narrowed considerably (Fig. 4). At this point the ectoderm of the disc showed a proliferation ventrally, forming an eccentric nodule which was in close contact with the subjacent endoderm. At this point of contact the endoderm also showed a proliferation of cells into the lumen of the yolk sac. Villi were most noticeable at the embryonic pole of the chorionic vesicle; only a very few of them were beginning to branch (Figs. 1, 5). Differentiation into cytotrophoblast and syncytiotrophoblast had begun, this being most marked at the embryonic pole. Mitoses were not found in the syncytial nuclei, but where the differentiation into two layers was least advanced, formation of syncytium by fusion of cells could be discerned. The cells of the cytotrophoblast were sufficiently well differentiated to be vacuolated (Fig. 6). Syncytial buds were forming all over the surface of the chorion and syncytial streamers were obvious, especially at the embryonic pole. Endometrial capillaries at the embryonic pole were dilated to form sinusoids which communicated with lacunae formed in the syncytiotrophoblast. However, there was as yet no one vessel much larger than others to serve as the main channel for drainage of blood. The lacunar spaces at the embryonic pole intercommunicated. At one point the syncytiotrophoblast had eroded the wall of a capillary and bulged into its lumen (Fig. 7). It has long been known that trophoblast may be found in the maternal circulation later in pregnancy (Schmorl, 1893), but it was surprising to observe invasion of a capillary as early as this. No fibrin (or fibrinoid) could be detected between trophoblast and decidua in these haematoxylin and eosin sections. DISCUSSION
This embryo can be assigned to Horizon VII of Streeter, i.e. 'Branching villi, axis of germ disc defined' (Streeter, 1942). Comparison of its size, histological characteristics, menstrual and embryological age with others of the same Horizon
A presomite human embryo
69
reveals that there are considerable variations between this embryo and those already documented. Thus the embryo described by Ingalls (1918) was thought on clinical grounds to have been dated 17-18 days after fertilization. It was not so fully embedded as the present embryo, the maternal blood in the intervillous spaces varied greatly in amount between different areas and yet it was further advanced in having a better developed primitive streak. By contrast, the embryo described by Heuser (1932), also thought to be of 18 days development, was in some respects further developed than the Liverpool III embryo; a definite notochordal canal was present, for instance. Concerning younger embryos, that of Brewer (1938) was thought to be of 11-12 days development. It was not embedded completely, villi were present but unbranched, no allantois was found and the cells forming the yolk sac were grouped together in clusters. The primitive streak was confined to the five caudal sections made; indeed it was hardly a distinct structure, being represented merely by an intermingling of the cells of the two layers of the germ disc. In the embryo of Streeter (1926), thought to be of 10-11 days development, the trophoblast had already begun to erode maternal capillaries, but the trophoblastic lacunae were not yet flooded with blood. The 16 day embryo reported by Harrison & Jeffcoate (1953) showed an early stage of the primitive streak. The true (secondary) yolk sac appeared to have formed and the blastocoele was largely occupied by extraembryonic mesoblast. The present embryo, whilst similarly displaying a yolk sac, also had a very clearly developed exocoelomic membrane (Fig. 1). Yet an earlier embryo of 13} days (Lewis & Harrison, 1966) showed an elongated yolk sac duct in process of being constricted off from the embryo. These findings are not easy to reconcile with theories regarding disposal of the exocoelomic membrane by means of a yolk sac duct if these embryos belong to a uniform population with constant time relationships. The thickened nodule in the ectoderm of the embryonic plate in this embryo showed similarities with a thickening in the ectoderm of a pathological presomite embryo (Harrison, Jones & Jones, 1966). This latter specimen showed well-marked syncytiotrophoblastic streamers and a pronounced endometrial reaction of a degree comparable to that displayed in the present embryo but much greater than that seen in the embryos of Harrison & Jeffcoate (1953) and Lewis & Harrison (1966). Such findings suggest that the degree of development, and the possible presence of abnormal features, in such early embryos, may be related to the degree of endometrial reaction, which in turn may possibly be dependent on the extent of immunological conformity between maternal and embryonic tissues. SUMMARY
A presomite embryo of Horizon VII aged approximately 18 days is described and illustrated. It is compared with some other embryos of a similar age, and a considerable variation of histological characteristics within the same Horizon is noted.
70
R. E. REWELL AND R. G. HARRISON
REFERENCES BREWER, J. 1. (1938). A human embryo in the bilaminar blastodisc stage (the Edwards-Jones-Brewer ovum). Contributions to Embryology of the Carnegie Institute of Washington 27, 85-93. HARRISON, R. G. & JEFFCOATE, T. N. A. (1953). A presomite human embryo showing an early stage of the primitive streak. Journal ofAnatomy 87, 124-129. HARRISON, R. G., JONES, C. H. & JONES, E. P. (1966). A pathological presomite human embryo. Journal of Pathology and Bacteriology 92, 583-584. HEUSER, C. H. (1932). A presomite human embryo with a definite chorda canal. Contributions to Embryology ofthe Carnegie Institute of Washington 23, 251-267. INGALLS, N. W. (1918). A human embryo before the appearance of myotomes. Contributions to Embryology of the Carnegie Institute of Washington 7, 111-134. LEWIS, B. V. & HARRISON, R. G. (1966). A presomite human embryo showing a yolk-sac duct. Journal ofAnatomy 100, 389-396. SCHMORL, G. (1893). Pathologische-anatomische Untersuchungen iiber Puerperal-Eklampsie. Leipzig: Vogel. STREETER, G. L. (1926). The " Miller" ovum, the youngest normal human embryo thus far known. Contributions to Embryology of the Carnegie Institute of Washington 18, 31-48. STREETER, G. L. (1942). Developmental horizons in human embryos. Contributions to EmbrVology of the Carnegie Institute of Washington 30, 211-245.
