Background
Sensorineural hearing loss is one of the most common congenital disorders, affecting at least 1 in 1,000 births [
1,
2]. Up to 39% of sensorineural deafness is associated with radiologically detectable inner ear malformations [
3‐
5]. Among those anomalies, complete labyrinthine aplasia (CLA), also known as "Michel aplasia," is reported to comprise 1% of cochlear bony abnormalites [
6]. CLA merits special consideration since it precludes cochlear implantation due to the lack of a spiral ganglion and cochleovestibular nerve fibers [
7‐
10]. CLA has been reported in association with other anomalies [
11,
12] including thalidomide embryopathy [
11], microtia and microdontia [
13]. Until recently, the etiology of CLA was unknown except for cases associated with thalidomide exposure [
11].
Mendelian inheritance was initially suggested for some cases of CLA, but no genetic linkage data were reported [
14,
15]. Tekin and co-authors subsequently described an autosomal recessive deafness phenotype, now referred to as LAMM syndrome (OMIM 610706), comprising CLA, microtia and microdontia [
9]. LAMM syndrome co-segregated with recessive mutations of the gene encoding fibroblast growth factor 3 (FGF3; OMIM 164950) on chromosome 11q13.2-q13.3 [
9,
10]. A total of five Turkish families and one Saudi family segregating LAMM syndrome and recessive
FGF3 mutations have been reported to date [
7,
9,
10]. All of the affected individuals had fully penetrant LAMM syndrome with CLA, microdontia, and type I microtia in which the auricle has a dysmorphic helix and antihelix but different parts of the auricle are still recognizable. In addition, temporal bone CT images of 14 affected members of the previously reported families revealed complete labyrinthine aplasia, although a cystic vestibulum was detected once [
7,
9,
10]. Ramsebner et al. (2009) recently reported a missense
FGF3 mutation (p.R95W) associated with less severe phenotypes, clinically distinct from those of LAMM syndrome [
16]. However, it was not clear whether this mild phenotype in the Somali family is due to the mutation, genetic background or both.
Mice deficient for FGF3 do not model LAMM syndrome but rather have abnormal inner ears with variable penetrance and expressivity on a uniform genetic background [
17‐
19]. In mice,
Fgf3 is expressed in the otic vesicle and in the adjacent hindbrain during otic placode induction and subsequent early inner ear morphogenesis [
18,
20]. Loss-of-function data from mice suggest that FGF3 affects molecular patterning of the dorsal otocyst and dorsal otic gene expression that is induced by WNT signals, but it does not critically influence ventrally expressed otic genes important for cochlear development [
18].
Here we report three Pakistani families co-segregating profound recessive deafness with mutant alleles of
FGF3. In one family, a recessive p.R95W mutation was associated with a variable inner ear and auricular phenotype,, similar to the reported mild phenotype of this allele [
16]. Unlike in the Somali family, significantly milder dental phenotypes were also observed in this Pakistani family. Therefore, we conclude that the phenotypic spectrum of recessive
FGF3 mutations range from fully penetrant LAMM syndrome to deafness with residual inner ear structures and absent or minimal syndromic features. We also suggest that p.R95W might exert a semi-dominant effect upon the function of
FGF3.
Discussion
Recessive
FGF3 mutations associate with a broad spectrum of inner ear and craniofacial phenotypes that show inter- and intrafamilial variability. Our study confirms that at least some recessive mutations of
FGF3 may not always result in fully penetrant LAMM syndrome. Furthermore, we noted a less prominent dental phenotype in subjects with p.R95W, in contrast to the severe dental phenotype reported from the Somali family [
16]. Taken together, our results show that the manifestations of recessive
FGF3 mutations can involve nearly non-syndromic deafness with variable inner ear structural development. In this regard, mutations of either
LRTOMT [
23] or
FGF3 should be considered when incompletely characterized nonsyndromic deafness is found to be linked to genetic markers of the
DFNB63 locus on chromosome 11q 13.2 -q13.3.
The less severe phenotype associated with p.R95W segregating in Pakistani family PKDF817 is probably not due solely to genetic background, since the milder phenotype of homozygous p.R95W individuals has also been reported in an unrelated family from Somalia. The amino acid substitution of p.R95W may have a milder pathogenic effect as compared with other mutations of FGF. Our molecular modeling results support the possibility of less severe pathogenic effects of this allele since this variant is predicted to affect neither FGFR2b receptor nor heparan sulfate binding sites. Alone or in combination with a less severe pathogenic potential of p.R95W, there might be a differential effect of mutant alleles of FGF3 on different target organs. While these molecular models provide a plausible rationale for the effects of the mutations identified here, there remains the caveat that available structural information does not account for the role that receptor glycosylation might play in FGF interaction. In addition, phenotypic variability within and between the affected individuals in these two families also indicates a role for environment, stochastic events, genetic background, or a combination of these influences. In affected members of family PKDF817, we could not find a variant in FGF10, a potential candidate for a modifier of FGF3. To address the presence of modifiers, if any, larger families segregating a milder phenotype and p.R95W would be required for linkage analyses.
Our identification of a mild to moderate conductive hearing loss in heterozygous carriers raises the question of whether p.R95W exerts a semi-dominant pathogenic effect upon the function of FGF3. These data warrant careful interpretation, since we were unable to obtain information about the status of the tympanic membrane and ossicles in carriers. We initially reasoned that this may reflect the same pathogenic mechanism underlying autosomal dominant lacrimo-auriculo-dento-digital syndrome (LADD syndrome; OMIM 149730) caused by heterozygous mutations of
FGFR2b [
33]. FGF3 is known to bind to FGFR2b with high affinity [
34,
35]. Some patients with LADD syndrome have been reported to show isolated mild conductive hearing loss [
36], although many patients manifest a mild to moderate mixed type hearing loss. Moreover, the dental and auricular phenotypes from LADD syndrome patients are very similar with those in LAMM syndrome, while another autosomal dominant oto-dental syndrome (MIM 166750) recently reported in association with heterozygous microdeletions affecting
FGF3 [
37] shows significantly different dental phenotypes. In this regard, this possible semi-dominant phenotype might be consistent with the hypothesis that p.R95W affects the interaction between FGF3 and FGFR2 [
16]. However, our molecular modeling results argue against this hypothesis, since p.R95 is located away from the FGFR2b binding surfaces of FGF3. Alternatively, an allele of a totally different gene accounts for the hearing loss in p.R95W heterozygous carriers in the inbred family PKDF817, as the Somali p.R95W heterozygotes were reported not to have hearing loss [
16] The resolution of this question will require careful molecular and clinical examination of additional carriers of this allele and other mutations of
FGF3. The inner ear phenotypic variability within families is consistent with the highly variable inner ear phenotype of
Fgf3 knockout mice on a uniform genetic background [
18,
19] and functional redundancy of
Fgf3 and
Fgf10 in otic vesicle formation [
17,
38].
The options for auditory rehabilitation of LAMM patients with CLA are limited to vibrotactile hearing devices or brainstem implants. The presence of an osseous IAC and a cochleovestibular nerve has never been described in association with homozygous FGF3 mutations before this study. The presence of a cochleovestibular nerve and a cochlear remnant is a significant consideration for cochlear implantation candidacy. Because of intrafamilial variability, some patients with FGF3-related hearing loss are potential candidates for cochlear implantation, even if CLA has already been documented in other affected family members.
Acknowledgements
We thank the members of the study families for their participation, Mayya N. Kawar, Erich Boger, and Barbara P. Zwiesler for technical assistance, and Meghan Drummond and Steven Raft for critical review of the manuscript. This study was supported by National Institute on Deafness and Other Communication Disorders intramural research funds DC00039-14 and DC00060-07. Part of the work in Pakistan was supported by the Higher Education Commission (HEC), Islamabad, Pakistan; EMRO/WHO-COMSTECH and Ministry of Science and Technology (MoST), Islamabad, Pakistan; the International Center for Genetic Engineering and Biotechnology, Trieste, Italy under project CRP/PAK08-01 (Contract no. 08/009). Work at Cincinnati Children's Hospital Research Foundation is supported by intramural research funds and the NIDCD research grant (R00-DC009287-03).
Competing interests
The authors declare that they have no competing interests.
Authors' contributions
SR, ZMA, SNK, IN, US and BYC carried out the molecular genetic studies and participated in the sequence alignment. RSH carried out the molecular modeling. JAB, AJG and BYC carried out the interpretation of clinical data. SR, ZMA and BYC participated in the design of the study. SR, ZMA and BYC conceived of the study and drafted the manuscript. SR, AJG and TBF supervised this study and critically revised the manuscript for important intellectual content. All authors read and approved the final manuscript.