Genetic correlation in otosclerosis

Genetic Correlation

in Otosclerosis

Surinder K. Singhal, DLO, MS, DipNBE, Sher Baj Singh Mann, MS, MNAMS, Usha Datta, MD, MNAMS, FIC[Path), Naresh K. Panda, MS, DipNBE, Ashok Kumar Gupta, DLO, MS Purpose: The aim of this study was to determine the relationship of HLAtyping in patients with otosclerosis. Materials and Methods: We used a prospective study in which HLA typing of 100 surgically confirmed otosclerosis patients were compared with age and sex matched normal individuals. Results: HLA-AS, HLA-AS, HLA-All, and HLA-B13 were found to be significantly higher (P < .05, .Ol , and .Ol, respectively). HLA-A9 and HLA-All were found to be higher (P < .Ol) in patients with a positive family history, indicating genetic heterogenicity. Conclusions: Higher values of HLA-AS, HLA-11, and HLA-B13 in patients with otosclerosis compared with normal individuals strongly suggests a genetic, HLA-related component. (Am J Otolaryngoll999;20:102-105. Copyright 0 1999 by W.B. Saunders Company)

Otosclerosis is a familial and genetically determined process inherited as an autosomal dominant penetrance. It is a disease of the otic capsule that is characterized by resorption and redeposition of bony tissue.l The disease is common in whites and is an important cause of deafness in the Indian subcontinent, Europe, the Middle East, and in North and South America. The histological form of otosclerosis is about 10 times more common than clinical otosclerosis. An estimated incidence of 0.5% has been reported in North America.2 In reviewing English literature, an association between otosclerosis and ABO blood grouping has been reported,3 whereas some workers have failed to establish such an association with a particular genotypea Toynbee was the first to observe a familial pattern of deafness from this disease in 1861, but later it was established that the mode of inheritance is autosomal dominant.5 A higher incidence of the disease in families and homo-

From the Departments of Otolatyngology and Head & Neck Surgery and the Department of Immunopathology, Postgraduate Institute of Medical Education and Research, Chandigarh 160 012, India. Address reprint requests to Sher Baj Singh Mann, MS, MNAMS, Head, Department of Otolaryngology and Head & Neck Surgery, Postgraduate Institute of Medical Education and Research, Chandigarh 160 012, India. Copyright 0 1999 by W.B. Saunders Company 0196-0709/99/2001-0005$10.00/0 102

American

Journal

of Otolaryngology,

Vol20,

zygotic twins” constitutes substantial evidence for the genetic nature of this disorder. The present study was undertaken to find the genetic correlation of otosclerosis with AB specificity of major histocompatibility complex antigen class I (MHC-Class I).

MATERIAL

AND METHODS

The present study was conducted in the Departments of Otolaryngology and Head & Neck Surgery and the Department of Immunopathology, Nehru Hospital, Postgraduate Institute of Medical Education and Research, Chandigarh where HLA typing of 100 cases of clinically diagnosed and surgically confirmed otosclerosis were compared with 100 age and sex matched normal subjects who showed no evidence of otosclerosis.

HLA Typing The standard microlymphocytotoxic assay as described by MacKintosh7 was carried out. Heparinized blood (10 mL) was collected and the mononuclear cells (MNC) were isolated over a ficoll/ isopaque gradient. The platelets were removed by slow centrifugation. Three washes to the MNC were given and adjusted to 2 X lo6 cells/ml. Sixty well tarasaki plates (Nunc Germany) were oiled and 1 FL of antisera of a known specificity (Biotest/Pelfreeze) were dispensed in each well. A minimum of four antisera of a specificity and 11 antisera of B specificity were tested (Table 1). Prepared MNC (1 FL) were added to each well and incubated at 22°C for 45 minutes to let the antigen-antibody reaction take place. Rabbit complement (5 pL) was added to each well, and after 890 No 2 (March-April),

1999:

pp 102-105

GENETICS

AND

TABLE 1.

OTOSCLEROSIS

HLA Antigens Patients

Otosclerotic HLA

in Controls

No. of Patients (n = 100)

Al A2 A3 A9 A10 All A24 A28 85 87 B8 812 813 815 817 827 B35 B40 844

103

12 25 2.5 54 14 44 6 2 46 16 16 14 8 10 22 8 20 16 4

compared

and

All No. of Controls (n = 100) 26 36 10’ 1st 16

20* 6 4 36 24 12 20 2 10 18 10 26 10 0

Relative

-

Risk

(RR) 0.38 0.56 2.84 5.34 0.85 3.14 1 .oo 0.49 1.51 0.60 1.40 0.65 4.26 1 .oo 1.28 0.78 0.71 1.71

*XT = 3.91, P< .05, RR, 2.4. tx: = 14.58, P < .Ol , RR, 3. *XT = 7.01, P < .Ol, RR, 2.2.

minutes at 22”C, 5 FL of eosin was added to assess the cell death. Cells were fixed with 5 FL of 37% formaldehyde, and cytotoxicity was assessed under an inverted microscope after the cells had settled Dead cells look dull after imbibing the dye, which confirms the presence of the antigen. Statistics The association of HLA antigen with otosclerosis was calculated by applying the chi-square (X2) test. Values were considered at P .05 level. Relative risk (RR) was calculated according to the following equation: RR=

P+ cp-xc+

where P = patient and C = control

RESULTS The patients with otosclerosis were between the ages of 17 and 48 years, out of which 68% were men and 32% were women. HLA typing shows the A3, A9, All, and B13 antigens to be significantly higher in patients with otosclerosis compared with the control group (P < .05,

.Ol, and .Ol, respectively) (Table 1). The RR was highest (5.34 times) for A9, followed by B-13 (4.26 times). The B13 antigen was present in 2% of the control group,

8% in patients

HLA-A9

to be significantly

and

higher in

male patients compared to HLA-A3, which was found to be significantly higher in female patients (Table 2). Thirty patients had a family history that was positive for otosclerosis. HLA-A9 and All were found to be significantly higher in patients with a positive family history (P < .Ol) compared to HLA-A3, which was found to be significantly

higher in patients with a negative

family history for otosclerosis (Table 3). There is a negative correlation between HLA-Al and disease because the RR is very low (0.38). Therefore, the chance of individuals with HLA-Al developing otosclerosis is low. Similarly, there is a negative correlation between HLA-Al among men.

and incidence

of disease

DISCUSSION

Otosclerosis can be due to hereditary, endocranial, biochemical, metabolic, vascular, infectious, or traumatic anomalies of the temporal bone. Otosclerosis is reported to present a familial pattern of deafness and, on compreTABLE 2.

(x20).

with

were found

HLA in Male

and Female

Patients

HLA Antigen

No. of Male Patients (n = 68)

No. of Female Patients (n = 32)

Al A2 A3 A9 A10 Alit Al3 A19 A24 A28 A32 85 B7 B8 812 813 815 B17 827 B35 B37 840 844

12 14 8 40 8 34 0 0 2 2 0 26 10 12 6 6 4 14 4 12 0 8 2

0 10 16 14 6 10 0 0 4 0 0 20 4 4 8 2 6 8 4 8 0 8 2

*xf = 3.93, P< tx: = 3.38, P<

.05, RR, 1.06. .05, RR, 1.08.

104

SINGHAL

TABLE 3. Versus

HLA in Patients Negative History

HLA Antigen Al A2 A3’ W A10 Ail* A24 A28 85 B7 B8 812 B13 B15 817 B27 B35 840 844 *x: = 21.05, tx: = 15.44, $x: = 18.23,

With

Male Patients (n = 68) 12 14 8 40 8 34 2 2 26 10 12 6 6 4 14 4 12 8 2

Positive

Family

History

Female Patients (n = 32)

0 10 16 14 6 10 4 0 20 4 4 8 2 6 8 4 8 8 2

P < .Ol , RR, 5. P < .Ol, RR, 0.08. P < .Ol , RR, 7.

hensive investigation show a simple autosoma1 dominance with a 25% penetrance of abnormal genes.8 The otosclerotic lesions are pleomorphic, varying from spongiotic bone with relatively large areas that are replaced by fibrous tissue to dense sclerotic bone. There are a few recognizable haversian systems, which are replaced by fibrous tissue in the spongiotic lesions and by interwoven bands of osseous tissue in the sclerotic lesions. The common characteristics of spongiosis, sclerosis, or both is total disorganization of the lesion that usually replaces the normal bone. Many diseases cluster in families for various reasons. Such inherited disorders are intrinsically more difficult to solve because genes cannot be observed under a microscope like bacteria. Many inherited traits are variable in their expression; thus, the genetic mechanism may be complex. In some instances, the genetic basis is predicted on its observed distribution in families. The present higher values of HLA-A3, HLAA9, HLA-All, and HLA-B13 in patients with otosclerosis are contrary to the observations made by Wauoff et a1,Qwho found HLA-B8 to be higher in these cases.Q We found HLA-B8 to be equally distributed in both the groups.

ET AL

Dahlqvist et al lo found HLA-BlO to be 10% and 19% in the patients group and the control group, respectively.lO The frequency of HLA-A9 and HLA-B13 (RR value 5.34 and 4.26, respectively) was significantly higher in the otosclerosis patients than in the control group, which suggests that the presence of those antigens may be related to an increased susceptibility to otosclerosis or its clinical outcome. This finding finding suggests that otosclerosis has a genetic HLA-related component. Similar findings have also been suggested by Gamir et alI1 We found a definite relationship between sex of individuals and HLA typing in patients with otosclerosis. HLA-A9 and HLA-All were found significantly increased among male patients (P < .Ol), compared with HLA-A3 (P < .05), which was higher among female patients. On reviewing the available English Literature, we could not come across any reference that shows higher values of HLA-A3 in women. It is well documented that women suffer from otosclerosis 1.6 times more often than males. Histologically, otosclerotic foci can be found in the temporal bones of women 1.9 times more often than in those of men. The hypothecated reason for this finding is the measles viral infection, which has been proved by elevated values of specific antimeasles IgG in the perilymph fluid of patients with otosclerosis.12 Higher values of HLA-A3 in women strongly suggests a HLA genetic linkage. Therefore, HLA typing of women during pregnancy can be more useful in identifying patients with increased HLA-A3 who are at risk of developing otosclerosis. Estrogen, a known stimulator of osteolytic activity, can play a further role in ossification of otospongeotic bone lesions. Although otosclerosis is considered to be an inherited disorder, there was no documented proof of its possible genetic linkage. Thirty percent of the patients in the present study had a genetic linkage and 30% had a positive family history. Significantly higher values of HLA-A9 and HLA-All (P < .Ol) in patients with a positive family history indicate that a genetic factor may be involved in the development of this disease. In otosclerotic patients with a positive family history, higher values of HLA-All, HLA-B35, and HLA-B14 has been reported. l3 Tissue typing of family members of such patients to evaluate the significance and

GENETICS

AND

OTOSCLEROSIS

HLA-All will be most useful. A positive correlation between a positive family history and HLA-All will highlight the importance of HLA typing in the absence of a reliable family history, enabling a close follow-up of the members of such families. The lack of similar association in the group without a positive family history indicates a possible genetic heterogeneity between the two groups. We could find no study in the English literature to suggest a relationship between sex and a positive family history in cases of otosclerosis, which has been highlighted in this study.

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105

3. Soifer NN: Otosclerosis: A review. Acta Otolaryngol Suppl (Stockh) 269:1-25,197O 4. Morrison AW: Genetic factors in otosclerosis. Ann R Co11 Surg Engl41:202-237,1967 5. Toynbee J: Pathological and surgical observations on the diseases of the ear. Med Chir Trans 24:190-211,186l 6. Larson A: Acta Otolaryngol Suppl (Stockh) 154:1, 1960 7. MacKintosh P: HLA typing, in Thompson RA ted): Technique in Clinical Immunology. London, Blackwell Science Publishers, 1981, pp 203-221 8. Larson A: Genetic problems in otosclerosis, in Schuknecht HF (ed): Otosclerosis. London, Churchill Livingstone, 1962, pp 109-117 9. Wauoff M, Chobaut JC, Raffoux C, et al: Systeme HLA et otospongiose J Fr Otorhinolaryngol28:299-301,1979 10. Dahlqvist A, Diamant H, Rantapaa D, et al: HLA antigens in patients with otosclerosis. Acta Otolaryngol IStockh~100:33-35.1985 11. damir MA, Mallea I, Mafrco J, et al: Human leukocvte antigen and otosclerosis. Acta Otorrinolarinaol Esn 4-7:26-28,1996 12. Arnold W, Niedermeyer HP, Altermatt HS, et al: Pathogenesis of otosclerosis. State of the art. Head Neck Oncol44:121-129,1996 13. Ginsberg IA, Hoffman SR, Stinziano GD, et al: Stapedectomy-In depth analysis of 2405 cases. Laryngoscope 88:1999-2016,1978