Pelger-Huet anomaly (PHA) is a benign hematological anomaly that is characterized by impaired lobulation of neutrophils with a coarse nuclear chromatin. Skeletal abnormalities may accompany this anomaly. Autosomal recessive deafness-4 (DFNB4) with enlarged vestibular aqueduct (EVA) comprises a phenotypic spectrum of sensorineural hearing loss (SNHL). We report a case with SNHL, multiple skeletal anomalies including osteochondroma, developmental delay, and PHA. Molecular studies revealed a heterozygous pathogenic variant in the LBR gene and a homozygous likely pathogenic variant in the SLC26A4 gene. Due to these 2 variants, he was diagnosed with PHA and DFNB4 with EVA. If goiter develops, DFNB4 with EVA is named Pendred syndrome (PDS), so the patient will be followed up for this condition, and in the current literature, there is no case with PDS and PHA co-existence either. PHA may be accompanied by multiple skeletal abnormalities. In our case, there is also concomitance with osteochondroma. Although these are independent and distinct diagnoses, we present this case due to the concomitance of these situations.

Established Facts

  • Skeletal abnormalities may accompany Pelger-Huet anomaly.

  • If goiter develops, autosomal recessive deafness-4 with enlarged vestibular aqueduct is named Pendred syndrome.

Novel Insights

  • Osteochondroma may accompany Pelger-Huet anomaly.

  • This is the first case known to have concomitance of Pelger-Huet anomaly and autosomal recessive deafness with enlarged vestibular aqueduct.

Pelger-Huet anomaly (PHA) is a benign hematological anomaly that is characterized by impaired lobulation of neutrophils with a coarse nuclear chromatin. Its prevalence is 1/6,000 live births, and it is inherited as an autosomal dominant trait. Neutrophils are hypolobulated and usually have 2 lobes connected by a thin chromatin bridge. Due to this appearance, they have been compared to a pair of glasses without arms when viewed under a microscope. It results from a deficiency of neutrophil differentiation caused by mutations in the lamin B receptor (LBR) gene. The amount of LBR protein may determine the amount of segmentation, so that heterozygotes present with bilobed nuclei (dumbbell shape), whereas homozygotes show unsegmented (round or ovoid) nuclei [Hoffmann et al., 2002; Speeckaert et al., 2009]. Some PHA patients have additional abnormalities such as ventricular septal defect, macrocephaly, and mild skeletal anomalies [Gastearena et al., 1982].

Hearing loss (HL) is the most common sensory defect affecting approximately 360 million people worldwide [Stevens et al., 2013]. HL is categorized as nonsyndromic (70%) or syndromic (30%) depending on the absence or presence of anomalies in other organs [Pourahmadiyan et al., 2019]. Autosomal recessive deafness-4 (DFNB4) with enlarged vestibular aqueduct (EVA) comprises a phenotypic spectrum of sensorineural hearing loss (SNHL) that is reported as severe to profound with vestibular dysfunction and temporal bone abnormalities such as bilateral EVA with or without cochlear hypoplasia. In addition to the clinical features of DFNB4 with EVA, euthyroid goiter develops from late childhood to early adulthood in Pendred syndrome (PDS). PDS, which is an example of syndromic HL, results from biallelic variants in the SLC26A4 (solute carrier family 26, member 4) gene. PDS was first described in 2 members of a large family presenting with deafness and goiter by Pendred in 1896 [Wémeau and Kopp, 2017; Gettelfinger and Dahl, 2018]. EVA (Mondini deformity) demonstrated by auditory canal magnetic resonance imaging (MRI) can support the diagnosis [Smith et al., 2016]. PDS is caused by biallelic variants of SLC26A4. The SLC26A4 gene is located on chromosome 7q31 and consists of 21 exons that encode the anion carrier transmembrane protein called pendrin that is expressed in the thyroid, kidneys, and cochlea [Park et al., 2005]. More than 200 different mutations have been reported to cause autosomal recessive nonsyndromic HL since 1997 [Koohiyan, 2019]. Pathogenic variants in SLC26A4 can result in partial or complete loss of pendrin activity. In mice, pendrin has been shown to be required between embryonic day 16.5 and postnatal day 2 for normal hearing but not for maintenance of hearing [Choi et al., 2011]. Three pathogenic variants including p.Leu236Pro (26%), p.Thr416Pro (15%), and c.1001+1G>A (14%) are seen more frequently than other pathogenic variants in individuals of northern European descent. Among the Chinese, Japanese/Korean, and Pakistani populations c.919–2A>G, p.His723Arg, and p.Val239Asp are prevalent pathogenic variants, and p.Gln514Lys is common amongst the Spanish population [Tsukamoto et al., 2003].

We present a boy with heterozygous LBR and homozygous SLC26A4 variants. Due to these variants he was diagnosed with PHA and PDS/DFNB4 with EVA.

The patient is a 13-year-old boy of Turkish origin. He is the third of 4 children from a first-degree cross cousin marriage and his siblings are healthy. He had SNHL that required a cochlear implant at the age of 3 months. He was first referred to our pediatric genetic clinic at the age of 7 years due to hearing loss, walking disability, and dysmorphic facial appearance. He was noted to have cognitive impairment, low frontal hairline, prominent nose, big ears, scoliosis, and a wide-based gait. His fingers were thin and long, and examination of his lower extremities revealed medial deviation in the big toes. There was no pathology in cardiovascular and abdomen examinations. Anthropometric measurements (occipitofrontal circumference, weight, and height) were in the normal range according to Turkish growth charts. Complete blood count, comprehensive metabolic tests, creatine kinase and thyroid function tests were all normal. A peripheral blood smear revealed hypolobulated, bi-lobed nuclei in almost all neutrophils that were suggestive of PHA (Fig. 1). The other family members were also screened, and the father was found to have similar abnormalities in the neutrophils.

Fig. 1.

Peripheral blood smear revealing neutrophils with bi-lobed nuclei (×1,000, Wright stain).

Fig. 1.

Peripheral blood smear revealing neutrophils with bi-lobed nuclei (×1,000, Wright stain).

Close modal

Radiographic examination confirmed scoliosis of the vertebral column (Fig. 2). His bone age was consistent with his chronological age. MRI revealed osteochondroma located in the metaphysis of the proximal side of the left tibia (Fig. 3). Echocardiography and abdominal ultrasound were normal. In addition, temporal bone computed tomography (CT), cranial MRI, and thyroid ultrasound tests were performed to investigate temporal bone anomalies, goiter, and vestibular aqueduct enlargement. Thin-section temporal CT revealed bilateral enlargement in the vestibular aqueducts.

Fig. 2.

Anteroposterior view of our patient with thoracolumbar scoliosis.

Fig. 2.

Anteroposterior view of our patient with thoracolumbar scoliosis.

Close modal
Fig. 3.

Magnetic resonance imaging located osteochondroma in the left tibia.

Fig. 3.

Magnetic resonance imaging located osteochondroma in the left tibia.

Close modal

We performed several tests over a period of 6 years. Cytogenetic analysis revealed a normal male karyotype. A normal CGG repeat allele was observed in the Xq27.3 locus of the FMR1 gene. Array-comparative genomic hybridization analysis was normal. No pathogenic variant was detected in X-linked intellectual disability and mitochondrial next-generation sequencing (NGS) panels. A mutational analysis for GAA expansion of the FXN locus divulged normal GAA repeat.

We finally decided to perform an inherited panel test which includes 597 genes associated with mendelian disorders. It revealed a heterozygous, pathogenic, stop codon forming variant [(NM_002296.4) c.43C>T, p.(Arg15Ter)] in the second exon of the LBR gene (chr1:225,611,735; rs192681330), and a homozygous, likely pathogenic variant [(NM_000441.2) c.397T>A, p.(Ser133Thr)] in the fourth exon of the SLC26A4 gene (chr7:107,312,675; rs121908365). In the following studies, a heterozygous variant was identified for the LBR gene in the father in segregation analysis. His mother and father were carriers of the SLC26A4 variant.

Sample Collection and DNA Extraction

DNA extraction was done by the commercial QIAcube (Qiagen, Germany) according to extraction and purification protocols offered by the producer. The DNA was stored at −20°C until the NGS study.

Next-Generation Sequencing of 597 Genes

DNA quality and concentration measurements were performed by Qubit (Thermofisher, USA) for the preparation of DNA samples for the test kit (Celemics, South Korea). After adjustment of the proper DNA amount, the library preparation step was done. In order, steps of the targeted NGS test kit include: DNA fragmentation, purification, DNA end repair, purification, A-tailing, purification, adapter ligation, purification, indexing, purification, probe hybridization, targeted library selection by streptavidin beads, amplification of targeted library, and purification according to the recommendations of the producer. The library run was performed by the NGS platform (MiniSeq, Illumina, USA). The data delivered from the NGS platform were analyzed by software Genomize Seq (V.16.7) (Genomize, Turkey). In addition, the ClinVar database (https://www.ncbi.nlm.nih.gov/clinvar/) and Varsome web tool (https://varsome.com/) were also used. The coding regions (±10 bp intronic) of 597 genes were investigated. For the pathogenicity interpretation and classification of genetic variants, the scoring system of ACMG recommendations was used [Nykamp et al., 2017].

Our patient has a blended phenotype and is unusual in many aspects. In a study conducted by Posey et al. [2017], 4.9% of patients had a molecular diagnosis that involved 2 or more loci. Consanguity was reported in 36.4% of the patients with homozygous variants [Posey et al., 2017]. Our patient was a child of consanguineous parents. In the same study, the most common mode of inheritance was autosomal dominant + autosomal dominant, followed by autosomal dominant + autosomal recessive. Our patient had an autosomal dominant + autosomal recessive inheritance pattern. There is no known concomitance of both DFNB4/EVA with PHA and/or PHA with osteochondroma in the literature. We believe that our patient is of significant importance, given that he has the concomitance of the aforementioned diseases.

The variant in the LBR gene caused a nonsense mutation of the arginine amino acid, forming a nonfunctional, truncated protein. This arginine residue is fully conserved among the LBR proteins of humans [McGinnis and Madden, 2004]. In silico prediction programs suggest that this truncated protein is potentially harmful [Adzhubei et al., 2010]. The DANN score was 0.9973 and MutationTaster (www.mutationtaster.org) predicted the variant as “disease causing.” Moreover, 7 of the 9 estimators identified the variant as damaging.

There are many variants associated with PHA [Borovik et al., 2013]. A mutation in exon 2 of the LBR gene has previously been identified. In a study, a young patient was described with regressing severity of rhizomelic limb shortening and PHA associated with compound heterozygous variants in LBR, p.Arg586Ser and p.Arg15*. Unlike the patient in that study, our patient’s heterozygous variant is not compound [Carvalho et al., 2017]. Whether mutations in exon 2 or the concomitance of PHA and PDS cause worse clinical features is currently not known.

LBR abnormalities that result in PHA are loss-of-function variants [Olins et al., 2010a, b] and can be found in either the hydrophobic or nucleoplasmic domain. While normal cells show a high amount of LBR, heterozygotes have decreased amounts of LBR. Segmentation of granulocyte nuclei is associated with the amount of LBR protein present. Granulocyte nuclei of PHA heterozygotes have less nuclear segmentation than normal granulocyte nuclei, and have a bi-lobulated, dumbbell shape [Hoffmann et al., 2002]. In our patient and his father all neutrophils were bi-lobulated similar to the literature.

Some patients only have isolated PHA (non-segmented nuclei), whereas others have accompanying signs and symptoms such as cognitive impairment, seizures, cardiac defects, and skeletal deformities [Oosterwijk et al., 2003]. Skeletal anomalies in PHA include short stature, short upper limbs, short metacarpals, postaxial polydactyly, and kyphosis [Hoffmann et al., 2002]. Our patient was found to have osteochondroma. To the best of our knowledge no patient has been reported to have both PHA and osteochondroma.

Human SLC26 family members are involved in a range of key anion transport activities including Cl/HCO3, I/HCO3, and SO42–/HCO3 exchange; and are linked to various debilitating disorders, including PDS/DFNB4 with EVA [Dawson and Markovich, 2012]. SLC26A4 is a member of the solute carrier 26 gene family and overlaps SLC26A4-AS1 (SLC26A4 antisense RNA 1), located on the minus strand. The mRNA product is approximately 5 kb long, with an open reading frame of 2,343 bases producing the 780-amino-acid protein pendrin which functions as a chloride, iodide, bicarbonate, and formate transporter.

Our patient revealed a biallelic, likely pathogenic variant (c.397T>A, p.Ser133Thr) in the fourth exon of the SLC26A4 gene (chr7:107,312,675, rs121908365), different from the ones frequently seen in the literature for PDS/DFNB4 with EVA. Ser133Thr is located in transmembrane domain II and likely impairs the stability of the protein at the cell membrane level, as previously reported for mutations affecting other transmembrane domains [Fugazzola et al., 2002].

The identification and interpretation of temporal bone defects requires appropriate testing. Thin-slice CT as a routine CT of the temporal bones typically will not suffice. Bilateral wide vestibular aqueduct was detected in the patient on 1-millimeter CT slices.

In addition to deafness/hearing impairment, temporal bone abnormalities, goiter, and an abnormal organification of iodide, with or without hypothyroidism may also be present in PDS [Wémeau and Kopp, 2017; Mey et al., 2019]. Functional studies suggest that missense SLC26A4 pathogenic variants that retain residual iodide transport function are more likely to be associated with PDS/DFNB4 with EVA than with PDS [Scott et al., 2000]. Some studies suggest that goiter develops in only 50% of individuals with PDS [Wémeau and Kopp, 2017]. PDS also includes the development of euthyroid goiter in late childhood to early adulthood, so we will have to follow-up our patient for years for an accurate diagnosis. In the first tests of our patient, thyroid ultrasound and function tests were normal. Thus, we defined our patient as DFNB4 with EVA. In the literature, a p.Ser133Thr amino acid substitution was observed in a 24-year-old male patient with subclinical hypothyroidism, severe SNHL, and EVA and he was evaluated as PDS [Fugazzola et al., 2002].

In summary, our patient is the first known case with both DFNB4/EVA and PHA. In addition, although there are patients with mild skeletal abnormalities, there is no patient known to have osteochondroma. Our patient’s clinical features are severe, and we can not conclude if the clinical severity is due to the concomitance of PHA and DFNB4/EVA or the mutation of exon 2 or the nonsense mutation in transmembrane domain II in the LBR gene. In our patient, it is not clear whether osteochondroma is a third diagnosis or a manifestation of the skeletal anomalies seen in PHA. More studies are required to clarify the effects of LBR mutations and other combinations of mutations in this gene.

Samples from the patients were obtained in accordance with the Helsinki Declaration. The paper is exempt from ethical committee approval. Ethical approval was not required for this study in accordance with local/national guidelines. Written informed consent for genetic testing was obtained from the patient’s mother.

All authors declare that they have no conflict of interest.

There is no funding source for the study.

Project design: Ö.T., M.D.E., and Ö.G.B. Data collection and clinical evaluation: T.C., C.Y.U., and Ş.A. NGS analysis: M.D.E. Preparation of the manuscript: T.C., C.Y.U., Ş.A., Ö.T., Ö.G.B. All authors read and approved the manuscript.

The authors confirm that all relevant data are included in the article. Further inquiries can be directed to the corresponding author.

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