[ 331 ] THE LATE INTRAUTERINE AND POSTNATAL DEVELOP- MENT OF HUMAN RENAL GLOMERULI

Transcription

1 [ 331 ] THE LATE INTRAUTERINE AND POSTNATAL DEVELOP- MENT OF HUMAN RENAL GLOMERULI BY MORAG S. MACDONALD* AND JOHN L. EMERY Department of Pathology, The Children's Hospital, Sheffield When examining a child's kidney, one meets the difficulty that there is no available standard, other than weight, against which to assess its maturity. The present paper reports an attempt to work out the normal standards of histological maturity of renal glomeruli during the later stages of intrauterine life and throughout childhood. Since the beginning of the nineteenth century much work has been done on the structural development of the kidney, but most of it has been concerned with animals other than man. Herring (1898) gave a full review of the work published before During the present century Kittleson (1917) and Arataki (1925) have studied the albino rat kidney, in which the formation of new glomeruli is known to continue after birth. They found immature forms in animals up to 50 days old, which they considered to correspond with a human age of 8 years. It has been known for many years (Peter, 1909, 1927) that the human kidney is incompletely developed at birth but most of the work on the human kidney has been carried out on single cases or on groups of cases from limited periods of childhood. Waschetko (1914), from work on six cases aged 1 day to 5 weeks, concluded that the postnatal growth of the kidney is due to the increasing size and diameter of the tubules and glomeruli and not to the formation of new nephrons. Campos (1923) used the presence or absence of the nephrogenic zone as an indication of the degree of development of the kidney in a survey of its pathological changes in congenital syphilis, and compared his findings with those in a series of forty-three normal controls. He concluded that the persistence of the nephrogenic zone beyond full term is an indication of retardation of kidney development, but he quotes Aschoff (1911) as stating that the nephrogenic zone is present in the subcapsular layer of the kidney up to 3-6 months after birth. Tsuda (1943) described a histological survey of the foetal human kidney in which he recorded the number of layers of glomeruli seen in the cortex at four different stages of intrauterine life, and he compared the diameter of the peripheral glomeruli with that of the central ones. He stated that the first glomerulus appears when the embryo is 3 in. long. At 4-5 months, he found 3-5 layers of glomeruli; at 7-9 months 9-12 layers, and at the 10th month layers. He also stated that in all cases the central glomeruli have a greater diameter than the peripheral ones, in some cases twice as great, but never more than this. Gruenwald & Popper (1940) examined the kidneys of thirtynine children under 22 months of age. They were concerned with juxtamedullary glomeruli only, and classified them into four stages of maturity. They considered that all are mature by the age of 2 years, and could be taken as a good indication of the general maturity of the child. Potter & Thierstein (1943) surveyed 1000 kidneys * Working under a grant from the Medical Research Council.

2 332 Morag S. Macdonald and John L. Emery from infants dying in the perinatal period, and concluded that the presence or absence of the nephrogenic zone was closely related to the weight of the kidneys and to the infant's weight, and constituted a good measure of the maturity at birth of the infant. Roosen-Runge (1949) used a control series of twenty-two normal kidneys for comparison with a series from patients with early cerebral lesions. Eight of the controls were under 2 years old and fourteen were aged years. From these kidneys he concluded that the cuboidal epithelium has normally disappeared from the glomerular tuft by the age of 14 months. MATERIAL Kidneys for the present survey were obtained from necropsies carried out by the staff of the Department of Pathology, The Children's Hospitals, Sheffield, and included a large number of infants studied through the courtesy of Dr A. J. N. Warrack at the City General Hospital, Sheffield. The total number of kidneys available for sampling was 1800 and the ages ranged from 22 weeks gestation to 15J years. From this material, 235 cases were examined in this survey. SELECTION OF MATERIAL Selection was made initially by using the post-mortem record number of the case with tables of random numbers, but after 100 cases had been selected in this way it was found that the natural age incidence of mortality had weighted the series very heavily in favour of infants under 6 months. We therefore began to select cases of 6 months and over, in chronological order of death, from the post-mortem register. In this way approximately even numbers of cases were obtained above and below 6 months of age. Only normal kidneys were included in the survey. All sections showing recognized pathological changes were discarded. METHOD The kidneys were all formalin-fixed, the sections were cut from paraffin blocks and were stained either with haematoxylin and eosin, or Masson's trichrome technique. The glomeruli were classified in six stages of maturity and counts were made of the numbers of each stage present throughout the depth of the cortex. In order to do this fairly, only those sections showing the columnar arrangement of the glomeruli clearly were used, and ten columns were counted from each case. It was found that many routine sections were unsuitable for this purpose and in these cases fresh kidney blocks were cut in such a plane that the section included a complete papilla in longitudinal section with the corresponding cortex showing unbroken medullary rays running out to the subcapsular zone. One column was regarded as the area of cortex lying between two adjacent complete medullary rays and extending from the medulla to the capsule. The criteria used in classifying the glomeruli into six stages were as follows: and are illustrated in PI. 1. Stage I. The 'S '-shaped proglomerulus (or pronephron, as it is better called). Stage II. The glomerular tuft is recognizable as such, but it is mushroom-shaped

3 Postnatal development of human renal glomeruli 333 and possesses no true vascular pole. Bowman's capsule is crescentic in cross-section and is lined by cuboidal epithelium. Stage III. The glomerular tuft has a vascular pole, but it is not divided into lobules and is covered by a continuous layer of epithelium. Bowman's capsule is circular in section and is lined by cuboidal epithelium. Stage IV. The glomerular tuft has expanded and lobulation is present but its epithelial covering is still continuous and dips down into the clefts between the lobules. The epithelium of Bowman's capsule is being flattened. Stage V. Considerable growth of the capillary tuft has taken place and its epithelial covering is fragmented, being represented by groups of cells in broken pallisade. Stage VI. The capillary tuft is in adult form, the capillary spaces in it are easily seen and its epithelial covering has practically disappeared, being represented by only a few cells scattered singly over its surface. In addition to these six categories it was necessary to provide two others: one for abnormal involuted or scarred forms, and one for 'unclassifiable glomeruli', i.e. those in which the section included only a small group of cells from the surface of the glomerular tuft and those in which the tuft had been lost from the section, the glomerulus being represented by an empty Bowman's capsule. Each case in the survey had a separate working sheet on which the results of the ten column counts were recorded individually and then the total number of glomeruli in each of the eight categories was expressed as a percentage of the total count. It was found that the taking of careful blocks suitable for column counting and the searching of sections to find ten different perfect columns was so time-consuming as to limit the general usefulness of the method. For this reason the method was altered to that of battlement counts made on routine sections which included the full depth of the cortex. The results of these counts were also expressed as percentages of the eight different categories present in a total count of at least 100 glomeruli. At the time of counting, only the post-mortem number of the case was known, but after the count had been completed, the age, weight, crown-rump length, total kidney weight, cause of death and duration of illness were obtained from the hospital post-mortem records and written on the working sheet of each case. In children younger than 2 months at death the age was expressed as gestation time plus postnatal age, but in those dying 2 months or more after birth the postnatal age alone was used. REPRODUCIBILITY TEST In order to test the reliability of our results we carried out a reproducibility survey. This test was designed by Dr G. H. Jowett. He selected sixteen cases from the four age groups that showed the highest individual variation in the original counts and three sections were chosen from each of these cases. These sections were named fq, and y and on each of ft and y two columns were selected and marked. On sections a and, simple battlement counts were made, and on sections ft and y the marked columns were used for column descriptions and counts. Thus four counts were made on each case, i.e. two battlement and two column counts. The survey was arranged so that on each of four consecutive days, one of the four types of count was done on each of the sixteen cases and when the whole series had been completed

4 RL zo~t a)-~~~~c)o a S ~~ cq 00 -oo GP Co 9eq Iti to e o~~~~~~c~c v o o o v o m cog o Co Cd P C) c) c) 0 CO C0 C ca Cao t co Go gl5c > > oo. O. O~ t~ Cs~ ooc a = Q X : O H O un~ i ao Q- c~ ko m~~~~~~~~~~~~o.~~-cc Q oqccbs co 0~~~ c,-4t- C' Go0.5" c.i 0 ~o0 Coc ra >.5C? 00 <5 0 S40C aq I 111 ~~~~~~~~~~911 ~CO >0C, CXsXnt-C O SHOCiXOtOc?-~~~~~~~ C? 0C)o c t-cq c DO b toc Ca -0CaC~~ ~~~ c ~ ~ ~ c co -tt g0 a) -4- ce~~~~~

5 Postnatal development of human renal glomeruli 335 it was repeated in the same order. A Latin square was drawn giving the order of counting so that each type of count was preceded and followed by the other three types in turn. The results were recorded on a separate sheet for each case, and as each day's count was completed, the sheet was folded down so that it could not be read the following day. It was then put away in an envelope containing slides a, ft and y belonging to the same case. Each of us carried out this survey independently of the other, and neither set of results was seen until both surveys had been completed. RESULTS The data of the cases examined is presented in Tables 1 and 2, that in Table 1 being given according to age, and that in Table 2 according to crown-rump length. Text-fig. 1 shows the proportion of glomeruli of stages I and II related to the gestation age. It seems that the most primitive type of glomeruli constitute- about 10 % of the total before 30 weeks' gestation and that these glomeruli diminish rapidly in numbers during the next 10 weeks, constituting about 1 % at 40 weeks, i.e. normal Table 2. Analysis of glomerular counts according to crown-rump length (For particulars see Table 1.) 45 C.R. length (cm.) and over I * II * X9 0 III *8 6*9 4* *5 0 IV 58'4 50* * V * * III+IV+V * *7 100 VI * Total cases in each group 'full-term'. None were seen in normal kidneys over 46 weeks' gestation. Stage II, the slightly more organized form, which would, however, usually be called a proglomerulus, constitutes up to 5 % of glomeruli at 40 weeks' gestation. It seems that these normally disappear by the end of the second postnatal month. Their occasional occurrence beyond this age may represent an unrecognized abnormality. The incidence related to age, of what appear to be fully mature (stage VI) glomeruli is presented in Text-fig. 2. This shows two fairly definite features: first, there is a nearly constant small proportion of stage VI glomeruli present, even in the most immature infants; secondly, the glomeruli are not all fully mature until the age of 12 years, i.e. approaching puberty. Text-figs. 3 and 4 are maturity profiles at different ages, the younger age groups being represented in Text-fig. 3 and the older children in Text-fig. 4. These amplify the statements of the previous two figures, and illustrate also the general difference between the kidneys of children under one year and those over 2 years of age. In children of 44 weeks and under, the predominant glomerular type is stage IV. By 6 months after birth the predominance has shifted to stage V. This predominance 22 Anat. 93

6 336 Morag S. Macdonald and John L. Emery Stage n - - Stage II Weeks gestation Months after birth Text-fig. 1. Percentage of stages I and II present according to age. Gestation age is the period of gestation plus the postnatal age and is used in all infants ot a postnatal age of less than 2 months I- E 0 bo o 4). Eto be m C a Weeks gestation Months after birth Years Text-fig. 2. Percentage of stage VI glomeruli present according to age. There is a very small percentage of adult forms present in the most immature kidneys, but all glomeruli have reached adult form by 12 years of age.

7 of stage V appears to carry on throughout infancy to the age of 5 years, after which fully mature glomeruli predominate. 2 years, but glomeruli in which the epithelial covering of the tuft is present in the form of a fragmented pallisade (stage V) constitute 7 % of the total in the age group 9-12 years. The relationship between the crown-rump length of the infant and the maturity of the glomeruli is represented in Text-fig. 5, and in Table 2. The crown-rump measurement is used as it probably gives the best single general indication of maturity in an infant. In this series, glomeruli of stage I are no longer found at a crown-rump length of about 85 cm. and the maturation of stage II seems to follow the stage I pattern very closely. Concurrently the stage III glomeruli mature rapidly, giving a steeper disappearance curve than stages I and II, until they are absent at crown-rump length cm. Their subsequent reappearance in the months (p.n.) He.~~~~~~~~~~~~~~~~ I: Postnatal development of human renal glomeruli 337 Glomeruli of stages I and III are absent by 100_ Over 12 years 90 _ years. 60 a' ~ ~~ ~ ~ ~ ~ ~~~~,~~ 2-5 years a40- Fg ~ ~ a weeks (g1) F weeks (g) pori ~~~~~~~ Maturity grades Maturity grades 5 6 Fig. 3 Fig. 4 Text-fig. 3. The percentages of the six maturity grades of glomeruli present in three age groups below six postnatal (p n.) months. At 28 weeks from conception there is a high proportion of the most primitive glomeruli (g.) though the majority are at stage IV. Between 36 and 44 weeks from conception a much greater majority are at stage IV and the percentage of primitive forms has fallen markedly. At a postnatal age of 4-6 months the great majority of glomeruli have matured to stage V and the primitive forms have virtually disappeared. Text-fig. 4. The percentages of the six maturity grades of glomeruli present in three age groups above 2 years. Stages I, II and III have disappeared. At 2-5 years the majority of glomeruli are still at stage V though adult forms constitute a large minority. From 6 to 12 years the adult glomeruli form the great majority. From 12 years onwards, only adult forms are present. 22-2

8 338 Morag S. Macdonald and John L. Emery longest infants is an unexplained finding, but it may represent a group of abnormal involuting forms. Glomeruli of stage IV predominate throughout the whole period, but seem to diminish fairly rapidly from the 30 to 35 cm. group, being replaced by glomeruli of stage V. Glomeruli of the most mature form (stage VI) are present at all crown-rump lengths in this series and all are located in the juxta medullary area, often very close to the deepest layer of the arcuate vessels , 60 -,x 4l U 4' Stage 50- IVx bo~~~~~~~~~~~~~~~~~~~~~~i X /~~~~~~~ 10 VI C Crown-rump length (cm.) Text-fig. 5. The percentages of the six maturity grades of glomeruli present according to crownrump length. In general the three most primitive stages disappear at a crown-rump length of cm. While they are present, the majority of glomeruli are at stage IV, but from 35 cm. onwards the percentage of this stage diminishes rapidly and no longer constitutes the majority by the time the crown-rump length has reached cm. DISCUSSION The aim of this study was to achieve a picture of the normal histological maturity of renal glomeruli at different ages in infancy and childhood in order to be able to assess variation and diagnose particular renal immaturities. But the individual variation between cases, and, more than that, the intrinsic difficulty in attempting to put a subjective impression of histological form on to a statistical basis has, to some extent, defeated us. The reproducibility survey showed that one of us (M. S. M.) had subconsciously altered the standard of assessment during the survey, and that there was error in

9 Postnatal development of human renal glomeruli 339 the grading of glomeruli between adjacent categories. This basic error in assessment of individual glomeruli was, fortunately, largely offset by the number of kidneys examined and by the random selection of cases, so that the general pattern described and conclusions drawn from it are likely to be as near the truth as the personal factor inherent in our method will allow. The following pattern of development appears in the cases we have examined. Very primitive incompletely formed glomeruli (stage I) are not to be expected after the age of 44 weeks from conception, nor in infants with crown-rump length greater than cm. In kidneys in which stage I are present, the majority of glomeruli will be at stage IV, but after stage I has disappeared the majority will be at stage V. These remain in the majority during the first 5 years of life, but from 6 years onwards stage VI predominates, though it is not until after the age of 12 that all glomeruli may be expected to be of adult form. The number of fully mature glomeruli, sometimes found in the juxta-medullary region of kidneys still showing stage I, do not begin to increase rapidly until 3-5 years of age. The large foetal juxta-medullary glomeruli have been described a number of times and are the subject of a separate communication. The present survey has been carried out on a somewhat different basis from that of Potter & Thierstein (1943). They were seeking a histological tool for the assessment of general maturity, whereas we have attempted to establish normal histological standards of renal maturity throughout childhood. Nevertheless, the general conclusion we have reached from our observations on the newborn infants in our series agree with theirs, though we have found a scatter in the time of disappearance of proglomeruli sufficient to make us doubtful of its value as a criterion of maturity in an individual case. In the older children in our series we have found that full histological maturity is reached at a later age than has been generally believed. It seems therefore that the histological maturation of the kidney may be roughly divided into three phases: (1) The nephrogenic phase in which new glomeruli are being formed. This phase may continue until 44 weeks after conception, but is usually over by the time the infant has reached the 36th week of gestation or has attained a body length of cm. (2) A phase in which the full complement of glomeruli is present, but almost all are immature. This phase lasts until 3-5 years of age. (3) A phase of final maturation which appears to extend from 3 to 12 years. During this phase there is a steady increase in the proportion of fully mature glomeruli of adult form present in the kidney. SUMMARY A survey has been made of the histological maturity of the renal glomeruli of 235 children whose ages ranged from 26 weeks of gestation to 131 years. The most primitive glomeruli are not found after the 44th week from conception and are found only occasionally after the 36th week. Primitive forms may persist until 4-6 months of age.

10 340 Morag S. Macdonald and John L. Emery The majority of glomeruli are of mature type from 6 years onwards. It is not until after 12 years of age that all glomeruli are of adult form in all kidneys. A very wide individual variation in the rate of glomerular maturation was found. REFERENCES ARATAKI, M. (1925). On the post-natal growth of the kidney with special reference to the number and size of the glomeruli. Amer. J. Anat. 36, ASCHOFF (1911). (Quoted by Campos, 1923.) CAMPOS, E. S. (1923). Pathological changes in the kidney in congenital syphilis. Johns Hopk. Hosp. Bull. 34, GRUENWALD, P. & POPPER, H. (1940). The histogenesis and physiology of the renal glomerulus in early postnatal life in histological examinations. J. Urol. 43, HERRING, P. T. (1898). The development of the Malpighian bodies of the kidney and its relation to the pathological changes which occur in them. J. Path. Bact. 6, KITTLESON, J. A. (1917). The post natal growth of the kidney of the albino rat with observations on adult human kidney. Anat. Rec. 13, PETER, K. (1909). Die Nierenkanalchen des Menschen und einiger Saugetiere. In Untersuchungen fiber Bau und Entwicklung der Niere, No. 1. Jena: Gustav Fischer. PETER, K. (1927). Der Ausbau der Nierensubstanz. In Untersuchungen fiber Bau und Entwicklhng der Niere, no. 6. Jena: Gustav Fischer. POTTER, E. L. & THIERSTEIN, S. (1943). Glomerular development in the kidney as an index of foetal maturity. J. Pediat. 22, ROOSEN-RUNGE, E. C. (1949). Retardation of post-natal development of the kidney in persons with early cerebral lesions. Amer. J. Dis. Child. 77, TSUDA (1943). Histologic investigation of the foetal kidney. Jap. J. Obstet. Gynec. 17, WASCHETKO, N. P. (1914). Zur Frage von dem physiologischen Wachstum der Niere. Zbl. allg. Path. path. Anat. 25, EXPLANATION OF PLATE Fig. 1. Stage I glomerulus. Fig. 2. Stage II glomerulus. Fig. 3. Stage III glomerulus. Fig. 4. Stage IV glomerulus. Fig. 5. Stage V glomerulus. Fig. 6. Stage VI glomerulus. All glomeruli are at the same magnification (1/500).

11 Journal of Anatomy, Vol. 93, Part 3 Plate 1 L1.U 1 1 'A. :,.. `:... i- AM ;.... m vx. A AWL F., t -q *.1r "F MACDONALD AND EMERY-POSTNATAL DEVELOPMENT OF HUMAN RENAL GLOMERULI (Facing p. 340)