Histologic criteria for age estimation of the developing human dentition

Oral pathology American

Academy

of Oral

Pathology

Donald Kerr, Editor

Histologic criteria for age estimation of the developing human dentition P. E. B. Calonius, Dr. Odont., Helsinki, l?inland, Martin Lulain, D.D.,O., M.P.H., Baltimore, Md., and Prank Stout, D.D.X., M.X., Baltimore, Md. II DEPARTMENT

OF PATHOLOGY,

OF PATHOLOGY,

SCHOOL

UNIVERSITY

OF DENTISTRY,

OF

USIVERSITY

HELSINKI,

AND

DEPARTMES?

OF MARYLAND

F

orensic odontology as a branch of forensic medicine has been used extensively for identification in a variety of medicolegal situations. An important part of this process is the identification of dead persons. In this procedure age estimation plays a significant role. Various anatomic features are useful in determining chronologie age ; among these are the characteristic skeletal changes and the conditions of the dentition associated with growth and aging. Although chronologic age is related to aging, ,Gustafsonl points out that there is a broad range of normal and for this reason it is usually advantageous to utilize as many features as possible in establishing age. Age estimation based on the adult dentition has been used widely, although the difficulty of estimating age from the condition of the teeth after the age of 14 years has led to GustafsonF’ method in which six different features are scored and the total is compared with a chart. This is a useful and accurate tcchnique. When other anatomic features, such as skeletal changes, are combined with it, one can attain considerable accuracy in age estimation. No definite system exists for estimating the chronologic age of fetuses and infants on the basis of the developing dcntition. There have been few studies of age estimation during intrauterine life and the period of life before the eruption of teeth. The fundamental data of the tlcveloping dentition are based on early studies by Logan and Kronfeld” and Scholl I* This study was supported in part by Grant No. FR05317-07 of Dental Research, Bethesda, Md. Presented at the twenty-second annual meeting of the Pathology, Phoenix, Ariz., April 16, 1968.

from American

the National Academy

Institute of

Oral

869

870

Calonius,

Lunin,

and Stout

Oral June,

Surg. 1970

and Massler.4 However, the material on which the data were based was limited. In Kronfeld’s study, for instance, there were, on the average, only two cases used to establish the development for each chronologic year from birth to 15 years of age.3 Recently, Gustafson and Koch1 constructed a graph relating mineralization to age and Stacks described a gravimetric method for early age estimation. The purpose of the present study was to record and describe a number of criteria useful in age estimation prior to birth and in infancy using histologic sections of the jaws. MATERIALS

AND

METHODS

The material used in arriving at criteria for age estimation consisted of ninety-two cases from three sources: 1. Twenty-one cases from the Carnegie Institution of Washington, Department of Embryology. These cases were mainly from early intrauterine life. Seventeen of the cases represented ages between 7 and 13 weeks. The ages in this period were calculated according to the method of Boving.‘j There were variations in fixation, decalcification technique, section thickness, cutting direction, and staining. 2. Eight cases from the University of Maryland. These cases represented ages from the eighth to the seventeenth weeks of intrauterine life. The specimens were fixed in neutral formalin, decalcified in formic acid and citrate buffer, embedded in paraffin, stained with hematoxylin and eosin or mucicarmine, and cut in sections 5 microns thick. 3. Sixty-three cases from the Children’s Hospital, University of Helsinki. These consisted of full-term children who died of acute diseases during the first 3 years of life. Complete clinical and autopsy reports were available for this study. The most common diagnoses were bronchopneumonia, acute gastritis, and other acute conditions of short duration. The specimens were taken from the left side of the posterior maxilla, from the cuspid-to-molar area, fixed in neutral formalin, decalcified with EDTA, sectioned serially in sections 5 to 7 microns thick, and stained with hematoxylin and eosin or by the van Gieson technique. Signs of development of bone, tooth, and accessory salivary glands were observed. The amount of hard tissue formed in the crowns, the maturation of the enamel, the root formation, and the topography of the tooth germs were all studied and recorded. For convenience, the phenomena observed were divided into three main groups : 1. Gross anatomic characteristics. These can be used for early intrauterine life prior to dental development. During the fifth or sixth week of embryonic life the primordia of the upper and lower jaws are seen. By the seventh week the lower and upper jaws are formed except for the midline zone. Despite their usefulness, the gross anatomic characteristics are not reported in this study because they are better observed grossly or radiographically than histologically.

Volume Number

Age estimation

29 6

Table I. Some oral histologic

criteria

Age (in weeks)*

useful

of developing

human

for age estimation,

dentitio?l

871

fetal period

Characteristics Early bone formation Invagination of dental lamina of deciduous central incisors and first molars; vestibular lamina formation Development of palatal salivary ducts and glands begins All deciduous teeth at least in bud stage All deciduous teeth at least in early or late bell stage Palatal salivary glands well developed and functioning Dentine matrix formation in upper central incisor begins Calcification is started in upper central incisors and first deciduous molars Calcification begins in upper deciduous cuspid; first upper permanent molar in cap stage The fist and second deciduous molars in occ~lus:~l view a~ equal in size Bone formation in upper jaw is prominent ; complete disappearance of Meckel’s cartilage

7 7 to 8 8 to 12 T3 14 13 to 14 14 to 15 1x “2 26

*Fetal ages were determined by using The ages may vary by 1 week for the first weeks, and about 4 weeks at term.8

standard tables of 17 weeks, 2 weeks

crown-rump length and age. by 25 weeks: 3 weeks at 33

Table II. Some oral histologic criteria useful for age estimation, postnatal Age Birth

to 5 days

2 to 3 months 6 to 7 months 8 to 10 months 2 years

4 to 6 months

2 years

6 months

Characteristics Coalescence of the cusps of maxillary first deciduous molar but the crown is not formed to the dentinocemental junction Slight loss of enamel matrix of maxillary first deciduous molar in decalcified sections Complete loss of enamel matrix of first upper deciduous molar in decalcified sections Slight loss of enamel matrix of maxillary second deciduous molar in decalcified sections; root formation under way in first deciduous molars Complete loss of enamel matrix of maxillary first permanent molars in decalcified sections Calcification of maxillary first premolars

2. Developmental characteristics. While development is a series of graduill changes, some stages can be noted in bone formation, early tooth formation, salivary gland development, mineralization of bone and teeth, and enamcxl maturation. Some of these stages are listed as histologic criteria for age est,inr;ltion in Tables I and II. 3. Topographic characteristics. These are important for orientation ant1 interpretation of findings. In the present study these chara.cteristics were usefIll in the interpretation of the autopsy material. The relative size and relationship between adjacent tooth germs and the relationship of the glandular and adipose tissue in the palate were used to identify teeth and establish anteropostcrior relationships. DISCUSSION It is generally agreed that the external features of the face begin to take shape between the sixth and eighth weeks and t.hat ossification centers in both

Fig. 2. Dental development of a human fetus in the thirteenth week. The maxillary first deciduous molars and the mandibular second deciduous molars are seen in the bell stage. Note the wide bony sockets and Meckel’s cartilage in the mandible. (Carnegie Collection No. 172. By Courtesy of the Carnegie Institution of Washington. Original magnification, x8.)

Fig. 3. Transverse section of a human mandible in the twenty-sixth well developed. Meckel’s cartilage is absent. (Carnegie Collection No. Carnegie Institution of Washington. Original magnification, x8.)

week. The mandible is 2969. By courtesy of the

Volume Number

29 6

Age estimation

of develop&g

human

dentition

873

the maxilla and the mandible are first observable at about this time.‘;-g The observations in this study confirm these findings. Ossification is gradual, and few clear-cut stages can be established during bone development. One clear-cut, stage appears to be the complete disappearance, in the twenty-sixth week, of Meckel’s cartilage (Figs. 1 and 2). In early ossification the bony crypt is wide and open on the occlusal and facial aspects. As bone formation continues, the crypt narrows and boric forms over the facial aspect of the tooth bud (F’igs. 1 and 2). Although tooth development is also a series of gradual changes, it is possible to est,ablish some observable chronologic stages because previous authors have amply defined and described the bud, cap, and bell stages. They also noted that at any time during development of the dentition different teeth have progressed to different stages73 8 For practical purposes, it is necessary to observe serial sections of more than one tooth for adequate age estimation. In our study serial sections were used to observe bud, cap, and bell stages and the earliest sign of tooth mineralization. In addition, the stage of devclopwas ment of minor salivary glands, based on the study by Stout and Lunin,‘” used to confirm age estimation. Using histologic criteria, we were able to confirm the chronology of tooth calcification reported by Christensen and Kraus,‘l Kraus and JordanI and Boller.13 These investigators reported on early mineralization as determined by gross methods in which early calcification and other features are shown 1~111. many histologic features are missing. Age estimation is relatively difficult during the period from birth to t.hc time the teeth begin to erupt. In our study the principal histologic criteria developed for age estimation during this time is the degree of maturation of

Fig. 3. A section of a human maxillary a 2M-month-old infant. In this section the magnification, x20.)

first deciduous organic enamel

molar matrix

from an autopsy specimen of is largely intact. (Original

074

Fig. specimen is largely

Calonius,

Lunin,

and &out

Oral June,

4. A section of a maxillary first deciduous molar of a B-month-old infant. was handled in the same manner as that seen in Fig. 3, the organic absent as a result of maturation.

Fig. 5. A section of a maxillary second deciduous section most of the enamel organic matrix is retained. seen in Fig. 4. (Original magnification, x17.)

molar of a ‘I-month-old Compare with the first

Burg. 1976

Although the enamel ma ttrix

infant. In this deciduous m olar

Volume Number

29 6

Age estimation

Fig. 6. The first and second maxillary autopsy specimen from a U&month-old infant. absent, while a large portion of the enamel magnific :ation, x9.)

lost.

Fig. 7. A section Root formation

of a first maxillary is almost complete.

of developing

human

dent&Son

875

deciduous molars in sagittal section. This is an Note that the enamel matrix of the first molar is matrix remains in the second molar. iOriginal

molar of a 2%month-old (Original magnification,

child. x9.)

The

enamel

matrix

is

876

Calonius,

Lunin,

and Stout

Oral June,

Surg. 1970

the enamel. Although the precise nature of maturation is not known, it appears that during maturation the protein of the matrix undergoes a marked change. There is an increase in its solubility, possibly by partial breakdown due to enzymatic degradation or the thixotropic properties of the protein.14 Recently Bergman and Hammerstrom, using radiographic and histochemical methods, described the maturation of enamel and the accompanying decrease in demonstrable proteins. The maturation appeared to progress from the oeclusal to the cervical region. Whatever the change, there remains in decalcified sections of mature enamel organic matrix a residue of only 0.2 per cent insoluble protein.16 Although it is subject to biologic variability, we found that the amount of enamel matrix remains a useful guide in age estimation (Figs. 3 to 7). SUMMARY Criteria have been developed for forensic odontologic age estimation using histologic techniques. Serial sections of ninety-two cases from 7 weeks in utero to 3 years of age were examined and useful histologic characteristics in bone, tooth, and salivary gland formation were tabulated. The authors are indebted to Dr. James ology, Carnegie Institution of Washington, B. G. Boving of the same institution for

D. Ebert, Director of the Department for facilities and materials provided his interest and help throughout the

of Embryand to Dr. work.

REFERENCES

1. Gustafson, G.: Forensic Odontology, New York, 1966, American Elsevier Publishing Company, Inc. 2. Gustafson, G.: Age Determination on Teeth, J. Am. Dent. Ass. 41: 45-54, 1950. 3. Logan, W. H. G., and Kronfeld, R.: Development of the Human Jaws and Surrounding Structures From Birth to the Age of Fifteen Years, J. Am. Dent. Ass. 20: 379-427, 1933. The Development of the Human Dentition, J. Am. Dent. 4. Schour, I., and Massler, M.: Ass. 28: 1153-1160, 1941. 5. Stack, M. V.: Forensic Estimation of Age in Infancy by Gravimetric Observations on the Developing Dentition, J. Forensic Sei. Sot. 1: 49-59, 1960. 6. Boving, B. G.: Anatomy of Reproduction. In Greenhill, J. P.: Obstetrics, Philadelphia, 1965, W. B. Saunders Company, pp. S-23. 7. Orban, B.: Oral Histology and Embryology, St. Louis, 1944, The C. V. Mosby Company. 8. Provenza, D. V.: Oral Histology, Inheritance and Development, Philadelphia, 1964, J. B. Lionincott Comnanv. 9. Woo, J.~~Ossification~and Growth of the Human Maxilla, Premaxilla and Palate Bone, Anat. Rec. 105: 737-761, 1949. 10. Stout. F., and Lunin, M.: The Development and Function of Minor Salivary Glands of the Palate, Program and Abstracts, 45th General Meeting, I.A.D.R., 1967,” p. 101. 11. Christensen, G. J., and Kraus, B. S.: Initial Calcification of the Human Permanent First Molar, J. Dent. Res. 44: 1338-1342, 1965. 12. Kraus, B. S., and Jordan, R. E.: The Human Dentition Before Birth, Philadelphia, 1965, Lea $ Febiger. 13. Boller, R. J.: Fetal Morphogenesis of the Human Dentition, J. Dent. Child. 31: 6797, 1964. J. H.: Maturation of Enamel. In Miles, A. E. W.: Structural and Chemical 14. Allan, Organization of Teeth, New York, 1967, Academic Press, Inc. 15. Bergman, G., and Hammarstrom, L.: Research on Calcified Dental Tissues. The Value of Combinimr Different Methods. Calcif. Tissue Res. 2: SUDD. 12. 1968. M. VI Mineral and Protein Levels in Enamel Fro~Human, Monkey and Rat 16. Stack, Molars, Odont. Rev. (Malmii) 8: 89-93, 1957.