Hereditary gingival fibromatosis associated with growth hormone deficiency

British Journal 0

ofOraland

1990 The British

MariNofacial

Association

Surgery

(1990) 28,3

of Oral and Maxillofacial

Su

Hereditary gingival fibromatosis associated with growth hormone deficiency K. Oikarinen,

T. Salo, M-L. Kti%r, P. Lahtela, M. Altonen

Department of Oral and Maxillofacial Surgery, Department of Orthodontics, Institute of Dentistry, and Department of Pediatrics, University of Oulu, Finland

SUMMARY. A case report of gingival fibromatosis in association with growth hormone (GH) deficiency due to a lack of growth hormone releasing factor (GRF) is presented. The girl is the youngest member of a family of eight children, five of whom lack the same hormone and have or have had similar gingival enlargements. After the growth hormone deficiency had been diagnosed and hormone substitute administered the dental age of the girl presented came closer to that of her age and sex-matched controls but did not reach the corresponding values even though the teeth were exposed Ibyexcising the overgrown gingiva. Test fibroblasts cultured from the overgrown gingiva proliferated at a slower rate than those cultured from age-matched controls. Total RNA was extracted from the test and three control fibroblasts and examined by Northern hybridisation using cDNAs for proal(1) and proal(II1) chains. The amount of type I and III procollagen mRNAs were lower in the test fibroblasts as compared to the controls.

and Maxillofacial Surgery at the Institute of Dentistry, University of Oulu due to pronounced fibrous gingival overgrowth covering all the maxillary and mandibular incisors and canines except for the tips of the lower medial incisors (Fig. 1A). The overgrowth formed a thickened alveolar crest which was evident also in the molar region and caused narrowing of the palatal space (Fig. 1B). At the age of 8.5 years the growth hormone deficiency had been diagnosed by the clonidine test, and it was assumed to be of hypothalamic origin as there was a good response to growth hormone releasing factor (GFW) (Fig. 2). Four years after the first operation and 3 years after the commencement of hormone therapy, eruption of permanent teeth was almost complete even though palatal fibrous overgrowth was still evident (Fig. 3). A growth chart, including two measurements of bone age according to a method introduced by Greulich and Pyle (1959) is ‘shown in Figure 4. Up to the age of 6 years the girl’s growth had been within the normal limits, after which it had become retarded until the point at which growth hormok deficiency was diagnosed and GRF substitute initiated. Although her growth reached the lower limit of the average’ growth chart of Finnish girls, her bone age was still almost 4 years behind her chronological age. No signs of puberty were present by the age of 12 years.

INTRODUCTION Gingival

overgrowth

is a common

side

effect

of

several drugs such as phenytoin (Modeer et al., 1986), nifedipine (Lederman et al., 1984) and cyclosporine-A (Rateitschak-Pluss et al., 1983). It has also been shown to be associated with some syndromes (Gorlin et al. ,1976). Kilpinen et al. (1978) have described a Finnish family in which four out of eight children expressed various degrees of fibrous gingival overgrowth associated with delayed eruption of permanent teeth and retarded skeletal growth. Gingival overgrowth had been present in the preceeding generations on the mother’s side and was reported to have been associated with a lack of secondary sexual characteristics in some female members (Kilpinen et al., 1978). Endocrinological studies by Collan et al. (1982) concerning the affected members of this family suggested that the association of retarded growth with gingival overgrowth was coincidental. The etiology of the retardation of the growth of the members of this family has been re-examined and it has been shown that five children and the mother have growth hormone (GH) deficiency due to a shortage of growth hormone releasing factor (GRF). This paper sets out to describe the findings of the youngest member of the family who during the time of previous studies was newborn and had nol gingival changes.

MATERIAL,, AND METHODS Histological;sections taken from the overgrown gingiva were stained with haematoxylin-eosin and examined by light’microscope with magnificatidn X40. Radiographic determination of dental age at ages 7.9, 9.2, 10.0 and 11.8 years was performed by a system introduced by Demirjian. et al. (1973). Each

Case report

orthopantoqograph taken at the various ages was compared to 10 age-matched and sex-matched radiographs which served as controls.

An &year-old girl, the youngest in the eight kindred family, was first referred to the Department of Oral 335

336

British Journal

Fig. 1A - Clinical of the mandibular from the maxilla.

of Oral and Maxillofacial

Surgery

view of the girl’s dentition, as seen at the first visit. A fibrous overgrowth covered all the incisors except for the tips incisors, (B) a heavy palatal fibrous overgrowth of the alveolar crest reducing the palatal space is seen on a cast

serum growth krmone

20

,\ \

f i

i

10

therapy

; 30

clonldine

/_a

I 0

60

90

120

150

test

t,me

(m,n,

180

Fig. 2 - Growth hormone response in the clonidine test (150 ugiml iv) and response to growth hormone releasing factor (GRF) (3 ug/kg iv).

I 0

5

10

15

20 ,P%,

Fig. 4 - Growth chart of the girl. The time of growth hormone deficiency diagnosis and the commencement of the GRF substitution is denoted by an arrow. Bone age measured at chronological ages of 9 and 12 years is illustrated by Q.

Fig. 3 - Four years after the first operation the clinical view shows almost complete eruption of the front teeth.

Gingival fibroblasts (test cells) were cultured from the overgrown gingiva taken during the first operation. Control fibroblasts were obtained during surgi-

cal exposure of impacted canines of four similarly aged girls. Tissue samples were dissected with a sterile blade in PBS (Phosphate-buffered saline). Explants were plated on a 6 cm diameter Falcon dish with 5 ml of Dulbecco’s Modified Eagle’s Medium (DMEM) containing 10% fetal calf serum (FCS), 100 kg/ml penicillin and 100 ug/ml streptomycin and they were allowed to reach confluency before subculturing using routine methods. The cells were examined daily under inverted microscope to assess their fibroblastic nature: Only those cultures which did not contain epithelial cells were used for the cell

Hereditary gingival fibromatosis

function studies. Gingival fibroblasts were used for the proliferation studies between passages 3 to 5. The cells were plated in T-25 Falcon flasks at a concentration of 2 x lo4 cells/5 ml and the cells were allowed to attach in culture medium for 24 h at +37”C. Cell proliferation was assessed by trypsinisation and counting the number of cells at various intervals up to 16 days. The values given are the means (*SD) of triplicate samples. During the recovery period the cell culture medium was changed twice a week. Total RNA was extracted and purified using guanidium thiocyanate-phenol-chloroform as described by Chomczynski and Sacchi (1987). Equal amounts (15 ug) of RNA were fractioned on 1% agarose gels in the presence of 18% formaldehyde. The gel was stained with ethidium bromide to visualise the rRNAs and, after taking a photograph, the RNA was transferred to a nitrocellulose filter. Hybridisation was carried out at 42°C in 50% formamide using the nick, translated and 32p-labelled pro&(I) collagen, 12 cDNA probe (Pyeritz et al., 1984) and proal(II1) collagen, E6 cDNA probe (Loidl et al., 1984). The filters were washed under stringent conditions (2xSSC, 0.1% SDS at 65°C for up to 20 min), using standard protocols (Thomas, 1980). Autoradiographs and photographs were analysed by quantitative densitometry.

337

Fig. 5 -Histological section of the overgrown gingiva. A thickened epithelium with long rete pegs is seen. (H&E. Original magnification X40.)

RESULTS Histological sections taken from the overgrown gingiva showed a thickened epithelium with long, elongated rete pegs while no subepithelial inflammation was present (Fig. 5). The difference in the dental age of the girl presented to the controls was highest (2.2 years) at the second measurement at the age of 8.8 years after which the difference decreased and was 1.1 years at fourth measurement (at the age of 11.1 years, Fig. 6). Test fibroblasts proliferated at a slower level than the control cells throughout the cultivation period (Fig. 7). The relative mRNA amounts of both type I and type III collagen of the test cells were markedly decreased as compared to the control cells (Fig. 8).

DISCUSSION Gingival overgrowth can be associated with the use of several drugs (Rateitschak-Pluss et aE., 1983; Lederman et al., 1984; Modeer et al., 1986) or with various syndromes (Gorlin et aE., 1976), or can appear as an idiopathic condition (Horning et al., 1985). A Finnish family with fibrous gingival overgrowth and retarded skeletal growth has been described by Kilpinen et al. (1978). Earlier studies concerning this family could not confirm hormonal background to the condition as no change in the response t:o growth hormone therapy was noted (Collan et al., 1982). The discovery of hypothalamic regulation of the

pituitary release of growth hormone by Rivers et al. (1982) has nevertheless now enabled it to be shown that five members of this family are lacking this growth hormone releasing factor (GRF). The girl described here and one older girl also have a gonadotropin deficiency due to a lack of gonadotropin releasing factor. This indicates that members of this family have a hereditary lack of several hormones whilch may have influenced their skeletal, dental and sexual maturation. All the other children in this family have passed puberty, so that dental and skel.etal maturations after GRF substitution could be followed only in this youngest child. It has been confirmed that all those five members of this eight kindred family who have or have had gingival enlargement and delayed tooth eruption also have decreased secretion of growth hormone. The disturbed growth hormone secretion explains the delayed eruption to some extent, as it is known that many pituitary hormones, including growth hormone can affect the development of the teeth (Garn et al., 1965). The girl’s dental age was 2.0 years behind that of her age and sex-matched controls at the time when the treatment with the hormone substitute was started, even though her anterior teeth had been exposed 14 months earlier. This seemed to indicate that removing the mechanical obstruction did not accelerate the dental maturation in this case even though eruption of teeth progressed. After the hormone substitute had started the difference in dental

338 British Journal of Oral and Maxillofacial Surgery

Fig. 6 C Chronological and dental age of the girl with fibrous gingival overgrowth and mean dental ages of 10 age’and sexmatched control subjects, measured according to Demirjian et al., 1983. Treatment with hormone substitute (GRP) was started at the ‘age of 8.5 years.

0

0

3

7

10

14

16

Days of cultivation Fig. 7 - Proliferation of the test fibroblast and the mean proliferation of four control cell lines during 16 days cultivation period. Cell proliferation was assessed by trypsinisation and counting the number of cells with a haemocytometer after various intervals up to 16 days.

age between her and the controls narrowed to 1.1 years within 3 years. This accelerated dental maturation may have been due to the hormone therapy, although her bone age was still clearly retarded in relation to her chronological age. Delayed eruption of the permanent teeth after loss of the primary teeth could result in a fibrous gingiva due to irritation by masticatory functions, but palatal enlargement is not easily explained by the influence of chewing. Retarded skeletal growth caused by growth hormone deficiency also results in reduced size of the facial bones (Sarnat etal., 1988). Assuming that the development of soft tissues was not affected, the overgrown gingiva could have been the result of a normal-sized gingiva in retarded jaws. This theory is confronted, however, by the fact that no other connective tissue abnormality has been observed in the affected members of the family. Histologically, the appearance of the hereditary gingival enlargement resembled that described by Shirasuna et al. (1989) with thick epithelium having rete pegs emerging deep into the connective tissue. In contrast to drug-induced gingival overgrowth, no clear sub-

T

C

T

C

T

C

Fig. 8 - Hybridisation

of Northern blots containing total RNA extracted from test gingival fibroblasts (T) and one control gingival fibroblasts (C) with cDNA probes for human type I and III collagen mRNAs. Aliquotes of 15 ug of total RNA was electrophoresed under denaturing conditions. The gel was stained with ethidium bromide (panel A) to visualise the rRNAs (marked on the left) after which it was transferred to Schleicher-Schull nitrocellulose filter and hybridised with nick translated 32plabelled probes. The autoradiograph with type I collagen probe is shown in panel B and with type III collagen in panel C.

epithelial colonisation of inflammatory cells was noticed as is common for phenytoin-induced gingival overgrowth lesions (Shafer et al, 1983). Gingival fibroblasts cultured from the overgrown gingiva proliferated at a slower rate than their controls in a way similar to that shown with the fibroblasts taken from verapamil-induced gingival overgrowth (Pernu et al., 1989). Shirasuna et aE. (1989) studied two siblings with congenital gingival fibromatosis and found low growth activity of the fibroblasts, as we did, but in contrast the collagen production was increased in their cases. In our case we found a decrease in the level of mRNA for types I and III collagen. Both decreased proliferation and reduced mRNA levels of types I and III collagen indicate the retarded activity of these cells which might be associated with the deficiency or shortage of growth hormone. This result does not explain the mechanism of gingival fibromatosis in viva but clearly demonstrates that cells derived from affected gingiva are not produking two main components of connective tissue at normal rates. One of the explanations could be that in addition to decreased production of collagen and proliferation, the degradation of extracellular proteins is reduced which in turn causes enlargement of the gingival tissue. Decreased collagenase activity has been suspected as one possible mechanism of drug-induced gingival overgrowth (Keller & Bohr, 1988). ! In conclusion, it can be said that the girl,is suffering from several hormone secretion deficiencies which must be connected with the altered skeletal and dental development as well as with the altered function of gingival fibrdblasts.

Hereditarv gingival fibromatosis in type III procollagen. American Journal of Medical

Acknowledgement

Genetics, 19, 607.

This study was supported Research Foundation.

by a grant from Oulu University

Rateitschak-Pluss, E. M., Hefti, A., Liirtschen, R. & Thiel, G. (1983). Initial observation that cyclosporine-A induces gingival enlargement in man. Journal of Clinical Periodontology,

References Chomczynski, P. & Sac&i, N. (1987). Single-step method of RNA isolation by acid guanidium thiocyanate-phenolchloroform extraction. Analytical Biochemistry, 162, 156. Collan, Y., Ranta, H., Vartio, T., Perheentupa, J. & Raeste, A.-M. (1982). Histochemical and biochemical study of hereditary fibrous hyperplasia of the gingiva. Scandinavian Journal of Dental Research, 90,20.

Demirjian, A., Goldstein, H. & Tanner, J. M. (1973). A new system of dental age assessment. Human Biology, 45,211. Garn, S. M., Lewis, A. B. & Blizzard, R. M. (1965). Endocrine factors in dental development. Journal of Dental Research, 44,243.

Gorlin, R. J., Pindborg, J. J. & Cohen, M. M. (1976). (Syndromes of the Head and Neck, 2nd Ed., p. 329. New York: McGrawHill Book Co. Greulich, W. W. & Pyle, S. I. (1959). RadiographicAtlas of Skeletal Development of the Hand and Wrist. 2nd Ed. California, U.S.A.: Stanford University Press. Horning, G. M., Fisher, J. G., Barker, B. F., Killoy, ‘W. J. & Lbwe, J. W. (1985). Gingival fibromatosis with hyperhidrosis. A case report. Journal of Periodontology, 56, 344.

Keller, U. & Mohr, W. (1988). Die Cyclosporine-induzierte Gingivalhyperplasie. Morphologie und Ueberlegungen zur Pathogenese. Deutsche Zeitschrift fuer Kiefer- und Gesichtschirurgie, 12,262.

Kiluinen. E.. Raeste. A.-M. & Collan, Y. (1978). Hereditary L g&&al’hyperpl&ia and physical maturation. Scandinavian Jburnal of Dental Research, 86, 118.

Lederman, D., Lumeiman, H., Reuben, S. & Freedman, P. ($984) Gingival hyperplasia associated with nifedipine therapy. Oral Surgery, Oral Medicine, Oral Pathology, 51, 620.

Loidl, H., Brinker, J. M., May, M., Pihlajaniemi, T., Morrow, S., Rosenbloom, J. & Myers, J. C. (1984). Molecular cloning and carboxvl-propeptide analysis of human type III procoiagen. Nucleid kc& Research,.12,9383. Modeer. T.. Dahlliif. G. & Theorell, K. (1986). Oral health in non-insiitutionaitied epileptic children with special rkference io phenytoin medication. Community Dentistry &al Epidemiology,

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Pernut, H. E., Oikarinen, K., Hietanen, J. & Knuutila, M. (1989). Verapamil-induced gingival overgrowth - a clinical, histological and biochbmical approach. Journal of Oral

10,237.

Rivers, J., Speiss, J., Thorner, M. & Vale, W. (1982). Characterization of a growth hormone-releasing factor from a human pancreatic islet tumour. Nature, 300, 276. Sarnat, H., Kaplan, I., Pertzelan, A. & Laron, Z. (1988). Comparison of dental findings in patients with isolated growth hormone deficiency &eateb with human growth hormone (hGH) and in untreated patients with Laron-type dwarfism. Oral Surgery, Oral Medicine, Oral Pathology, 66, 581.

Shafer, W. G., Hine, M. K. & Levy, B. M. (1983). Textbook of Oral Pathology, 4th Ed., p. 786. Philadelphia: W. B. Saunders Co. Shirasuna, K., Okura, M. Watatani, K., Hayashido, Y., Saka, M. & Matsuya, T. (1989). Abnormal cellular property of ffibroblasts from congenital gingival fibromatosis. Journal of (Oral Pathology, 7,381.

Thomas, P. S. (1980). Hybridization of denaturated RNA and small DNA fragments transferred to nitrocellulose. Proceedings of National Academic Science USA, 77,520l.

The Authors KyGsti Oikarinen DDS, PhD Senior Lecturer Tuula Sale DDS, PhD Assistant Mikko Altonen MD, DDS, PhD

Professor Deuartment of Oral and Maxillofacial Surgerv Pia’Lahtela

DDS

Resident Department of Orthodontics Institute of Dentistry University of Oulu Aapistie 3 90220 Oulu Marja-Liisa

KBlr MD, PhD

Associate Chief Pediatrician Department of Pediatrics University of Oulu Finland Correspondence & requests for offprints to Dr. Kyijsti Oikarinen, Institute of Dentistry, Aapistie 3, SF-90220 Oulu, Finland

Pathology, 18,422.

Pyeritz, R. E., Stolle, C. A., Parfrey, N. A. & Myers, J. C. (1984). Ehlers-Danlos syndrome IV due to a novel defect

Paper received 15 September 1989 Accepted 30 November 1989