Micromelia of chicken embryos and newly hatched chicks caused by a nutritional deficiency.код для вставкиСкачать
MICROMELIA O F CHICKEN EMBRYOS AND NEWLY HA4TCHED CHICKS CAUSED BY A NUTRITIONAL D EF IC IENC Y W A L T E R LANDAU ER Storm Agricitllurcil Ezperinwnt Sloliov, Storrs, Connecticut FIVE FIGURES Certain types of deficient diets hare been found to cause the occurreace o f a peculiar deformity of chicken embryos and newly hatched chicks, characterized by shortness of the long bones of the extremities (micromelia) and, in a part o f the specimens, by ail abnormal shape of the head (Byerly, Titus, Ellis and Landauer, ' 3 5 ) . A short description of the gross a i d histological appearance of the bones in this new type of developmeiital abnormality is presented herewith. AISTERJAL AKD METHODS The material used consisted of about forty micromelic embryos which had died during the last days of development, and which had been taken from the unhatched eggs at the end of the incubation period ; of five newly hatched micromelie chicks ; of one pullet which was definitely micromelic at hatching time and had been kept on a normal diet for 3 months ; and, finally, of three embryos with only a moderate shortening of the long bones which were found in the eggs of chickens that had been kept on a diet less deficient than that which was responsible for the occurrence of the more extremely deformed embryos. F o r histological sindy, sections of all the long boiics of the extremities were made of five strikingly deformed embryos which had died a short time before hatching, of three newly 267 T H F 4 N A T O I I I C I L RECORD, l 0 T 1 64, NO 2, AAD SIPPLFMFNT A O 2 268 WALTER LAXDAUER hatched chirks, of the 3-month-old pullet, and of two embryos showing only a moderate degree of micromelia. Sections of thc same bones of aormal chicks were used for comparison. In addition, sections of ribs of abnormal and normal late embryos and newly hatched chicks were compared. Fig. 1 Abnormal einbryo of about 19 days of development. Extremities very short, head brachycephalic. E’ig. 2 Metaphyseal region of humerus of micromelic embryo. Age about 19 days. Cartilage calcification chiefly along periphery, few osteoblasts, abnormal periosteal bone. Fig. 3 Marrow cavity of ulna of iiiieroinelie embrgo. Age about 19 dajs. Typical network marrow. Fig. 4 Tibia of newly hatched chick with low degrec of microniclia. A h o r m a 1 periostwl hone of diaphpi6. E’ig. 5 Tibia of a pullet at the age of 3 months, niicromclic at hatchirig, raised on a normal diet. Periosteal hone of the diaphysis. Peripherally normal bone, towaril inside narrow layer of nbnormal bone in process of resorption. NUTRITIONAL MICROMELIA O F C H I C K S 269 GROSS MORPHOLOGY There is considerable variation in the expression of this deformity. The more striking abnormalities always seem to cause death of the embryos during about the last 3 days of incubation. These embryos appear to be normal in size of the body, or nearly so, but have strikingly short extremities (fig. 1). The head of these embryos frequently, but not always, shows an abnormal shape, being of a more brachycephalic type than normal. Even in embryos with extreme shortening of the legs and wings, however, the head may be entirely normal. The great majority of chicks which were hatched in these experiments did not show any abnormalities but there were a few with definite shortening of the extremities. I n sonde of the abnormal newly hatched chicks the degree of micromelia was about as extreme as in the abnormal embryos which died before hatching, but the head was always normal; in others, the extremities were less conspicuously shortened. Some such chicks were raised on a normal diet, and the only one of them examined histologically, still showed definite shortness of the long bones at the age of 3 months. Some measurements were taken of the length of several long bones, and these give an approximate picture of the retardation in growth of the bones. Since we had only a few specimens, however, in which age was well timed, only the average length of the bones in per cent of normal length is given here. I n each instance, the average of ten measurements of normal specimens served as a basis of comparison. These figures (table 1) show that all the long bones are strikingly-shortened. The bones of legs and wings Seem to be involved to about the same extent. On the other hand, in the legs the degree of reduction definitely increases in a proximo-distal direction. The decrease in relative length of the long bones from the twentieth day of embryonic development to the first day after hatching is due to a much smaller increase in length during this period in the abnormal embryos than in normal ones, Finally, for the one pullet which 'PEE ANATOMIOAL REOOED, VOL. 64, NO. 2, AND SUPPLEMENT NO. 2 270 WALTER LANDAUER had short extremities at hatching and was raised on a normal diet for 3 months, our figures show that the relative length of the long bones had more closely approached, but not reached, the normal level. There is a striking resemblance in gross appearance between these embryos with nutritiond micromelia and embryos with sporadic chondrodystrophy, described in an earlier report (Landauer, '27). I n both types of malformed embryos we find the extreme shortening of both pairs of extremities relative to body size; in both we find a shortened and malformed head in some of the specimens, but not in others. The ehondrodystrophic embryos almost always have a striking bendin the shaft of the tibia which is visible externally or can TABLE 1 Length of long bones of micromelic chicks in per cent of norm1 length N AGIE ~ 20-day embryos Newly hatched chicks 3-month pullet FEMUR _ 8 4 1 _ 76.8 75.4 87.9 TIBIA HUMERUS _ 75.3 70.7 83.8 ULNA ______ 71.5 69.7 80.7 77.9 70.4 86.2 77.9 75.0 88.1 be palpated; in the embryos described here, the tibia always appears to be straight. In embryos with nutritional micromelia the long bones of the wings are reduced in length to almost the same degree as those of the legs, while in chondrodystrophy the wing bones are much less affected than the leg bones. I n many cases, however, it would be difficult to distinguish the two types of malformed embryos from external appearance alone. Histologically, these two types of micromelia are quite distinct. HISTOLOGICAL OBSERVATIONS The long bones in late embryonic stages and ifi newly hat,ched chicks. The principle histological features of the different long bones of the extremities are fairly uniform, so that a general description will be sufficient. NUTRITIONAL MICROMELIA OF CHICKS 271 The epiphyseal ends of the bones tend to be somewhat wider than normal. The growing cartilage shows no conspicuous abnormalities. The marrow canals are fewer in number than normally and vascularisation of the epiphyses is less well developed. The zone of cartilage columns is irregular. Usually, this zone is narrower than normal, and the arrangement of the cells tends to be irregular although short and poorly formed columns are still present in most cases. The hypertrophic cartilage cells tend to be smaller than normal and have more matrix between each other. The most striking abnormalities, however, concern calcification and bone formation. Calcification of the metaphyseal cartilage normally is well advanced at this stage of development and extends across the whole width of the metaphysis. I n this material, on the contrary, calcification is confined to a narrow border at the periphery of the bones (fig. 2). There is no normal periosteal bone. I n its place we find a faintly staining tissue which appears to consist chiefly of cartilage matrix, containing much debris from disintegrated capsules of cartilage cells. I n the neighborhood of the metaphyseal region all transitional stages can be seen from hypertrophic cartilage cells to the tissue which takes the place of the periosteal bone. The capsules of the cartilage cells can be observed to fragment, and, later, to disappear entirely. Further away from the zone of hypertrophic cartilage, the cell fragments gradually diminish in size until only small particles of irregular shape are seen scattered through the matrix. I n some places fine fibrillar structures are found in the matrix which also appear to be remains of disintegrated cartilage capsules. Osteoblasts are few in number, and frequently they show an abnormal, elongated shape. Osteoclasts are very rare or absent. It is interesting that this abnormal tissue presents a trabecular structure somewhat similar to that of the periosteal bone, except that the intertrabecular spaces tend to be larger than those of periosteal bone of normal embryos. The marrow cells in these intertrabecular spaces are in large part replaced by fibrous connective tissue. I n the marrow cavity of ulna (fig. 272 WALTER LANDAUER 3) and femur and in parts of that of the tibia, the marrow is replaced by an irregular connective tissue network which contains few marrow cells (‘Ceriistmark’). Those embryos which show less extreme shortening of their extremities also exhibit less severe histological lesions in the long bones. The columns of cartilage cells show normal arrangement, and calcification extends across the whole width of the bones. In the metaphyseal region the periosteal bone exhibits osteoid character. Toward the center of the diaphysis the structure of the periosteal bone is similar to that which has been described fo r the embryos with more extreme shortening of the long bones (fig. 4):there is matrix containing numerous fine fibrillae and many fine granular inclusions (debris of degenerated cartilage cells); there are fewer bone cells than normally, and these cells are arranged irregularly; the intertrabecular spaces frequently are occupied by fine connective tissue fibrillae instead of marrow cells. The Eomg homes of a micromelic pullet at the age of 3 months. The various long bones show great differences in histological structure. The femur appears entirely normal. I n the tibia the shaft shows an outer layer of normal periosteal bone, staining intensely red. Toward the marrow cavity, along the whole length of the shaft, there is a layer of abnormal bone with all the peculiarities described for late embryonic stages (fig. 5 ) . This abnormal layer of periosteal bone stains bluish red instead of red (hemalum-eosin). There are very few, if any, typical bone cells. Large areas do not show any cells but only matrix containing granular and irregularly shaped debris ; in other places, degenerated cartilage cells can be seen. The abnormal and the normal periosteal bone are connected by transitional stages. These areas of transitional structure contain many osteoclasts, and it is obvious that transformation of abnormal into normal bone is taking place. I n the tarsometatarsus abnormal bone is found chiefly in the region of the distal metaphysis. The diaphysis contains only normal periosteal bone but there is a layer of dense connective tissue along the inside of the shaft which appears to have replaced NUTRITIONAL MICROHELIA O F CHICKS 273 some of the earlier abnormal bone. The humerus has an almost normal histological structure. There are large cavities along the inner periphery of the shaft which contain connective tissue and many osteoclasts, probably the last signs of a practically completed transformation of abnormal into normal bone. Radius and ulna still show small areas of abnormal bone inside of a normal layer of periosteal bone of the diaphysis. Ribs. The skeletal lesions of the abnormal embryos and newly hatched chicks are not confined to the long bones of the extremities. The ribs, for instance, exhibit changes of a similar nature to those reported for the long bones, though less extreme in degree. Calcification of the cartilage is much retarded and practically confined to a narrow peripheral zone, while in ribs of normal specimens of the same age calcification extends across the whole width of the ribs. Otherwise, the cartilage appears to be fairly normal. The periosteal bone of the ribs shows fewer bone cells than normally, and these cells exhibit irregularities in shape and arrangement. Toward the inside, parts of the layer of periosteal bone are still cartilaginous. The formation of endochondral bone is somewhat retarded, and, similar to the periosteal bone, the number of bone cells is reduced. There were small hemorrhages in the musculature between the ribs. I n contradistinction to our findings in cartilage bone of these embryos and chicks, the membrane bones of the skull appeared to be normal. DISCUSSION It is obvious from our observations that calcification and the formation of periosteal bone are the seat of the principle disturbances in the condition with which we are dealing. The changes in the growing cartilage, on the other hand, are relatively slight, and are likely to be of a secondary nature. Some of the features of this micromelia bear a definite resemblance to the symptoms of mammalian scurvy: the general osteoporosis ;the relative scarcity of osteoblasts ; the unorganized detritus in the tissue which takes the place of periosteal 274 WALTER U N D A U E R bone ; the occasional occurrence of a layer of connective tissue covering the inner surface of the abnormal periosteal bone ; the framework marrow, and what irregularities there are in the growing cartilage all are reminiscent of scurvy. In the costal musculature we could even at times observe small hemorrhages, though subperiosteal hemorrhages, most common in scurvy, have not been found. Sacchetto ( '23) described an experimental form of beri-beri in young chicks, occurring on a diet of polished rice o r polished barley. He did not find a disproportionate arrest in growth of the long bones, but the histological lesions in the bones apparently were somewhat similar to those found in our material, though much less extreme. Dam and Schonlieyder ( '34) produced in chicks a deficiency disease resembling mammalian scurvy with extensive hemorrhages and changes in the mucous membrane of the gizzard, but they did not describe the condition of the bones in their specimens. It is obvious, of course, that our material differs in various ways from the picture found in typical mammalian scurvy. What similarities there are, together with the fact that prompt healing takes place on realimentation, enable us to classify the symptoms as typical effects of a nutritional deficiency. On the other hand, interesting similarities exist between our micromelic chicks and the condition of abnormal human development known as osteogenesis imperfecta. I n both instances, we find a disproportionate shortness of the extremities, lack o r very deficient formation of the periosteal bone with granular debris in the tissue which takes the place of the normal bone, and relatively slight disturbances in the epiphyseal cartilage, depending in their extent upon the severity of disturbance of periosteal bone formation. Multiple fractures, the most characteristic feature of osteogenesis imperfecta, have not been found in our material. A comparison of the pictures found in specimens differing in degree of expression of this abnormality illustrates the intimate developmental correlations existing in growing bone. As long as formation of periosteal bone is taking place to a NUTRITIONAL MICROMELIA OF CHICKS 275 certain extent, we find that hypertrophic and columnar cartilage are nearly normal; but, if normal periosteal bone formation is suppressed entirely, the hypertrophic cartilage disintegrates and the cartilage columns become irregular. Conversely, on realimentation healing sets in with the formation of normal periosteal bone and is followed by the resorption of the degenerated cartilage cells. Finally, it should be pointed out that the abnormalities which have been described, seem to constitute the first instance in which a prenatal nutritional deficiency results in a disproportionate stunting of the extremities. SUMMARY Certain nutritional deficiencies in the diet of laying hens, as reported elsewhere, lead to the occurrence of malformed embryos and newly hatched chicks showing disproportionate shortness of the extremities, and, sometimes, abnormal shape of the head. Morphological and histological studies on such material brought out the following principle facts. 1. I n wings and legs the long bones are shortened to approximately the same extent. I n the legs, the degree of reduction in length of the long bones increases in proximo-distal direction. 2. The greater the shortening of the bones, the more abnormal is their histological structure. In the most extreme cases, the periosteal bone is replaced entirely by matrix containing fragments of degenerated cartilage cells. Osteoblasts are few in number. Calcification of the cartilage in the metaphyseal region is restricted to the periphery of the bones. The cartilage columns are low and irregular. The hypertrophic cartilage cells are smaller than normal. The marrow cavity in some of the bones contains network marrow. Abnormalities, less extreme in degree but similar in nature, are found in the ribs, but the membrane bones of the skull appear to be normal. 3. On realimentation, normal new periosteal bone is laid down and the abnormal bone is resorbed or transformed into normal bone. 276 WALTER LANDAUER ACKNOWLEDGMENT The material used in this study and all information concerning it s experimental production were obtained through the cooperation of the Bureau of Animal Industry, U. 8.Department of Agriculture. LITERATURE CITED BYERLY, T. C., H. W. TITUS, N. R. BLLTSAND W. LANDAUER1935 A new nutritional disease of the chick embryo, Proc. 800. Exp. Biol. and Med., vol. 32, pp. 1542-154G. DAM, H., AND F. SCHONREYDER 1934 A dedciency disease in chicks resembling scurvy. Biochem. J., vol. 28, pp. 1355-1359. DIETMCH,A. 1929 Die Entwicklungsstiirungen der Knochen. Handbuch der speziellen pathologischen Anatomie und Hiatologie. Bd. 9, S. 166-221, Berlin. L A N D AW. ~ 1927 Untersuehungen iiber Chondrodyatrophie. I. Archiv. f. Entwick.-mech., Bd 110, S. 195-278. SACCHETFO,I. 1923 Le alterazioni di sviluppo dello scheletro nel beri-beri sperimentale. Pathologica, vol. 15, pp. 515-521.