Compressive Myelopathy Secondary to TRPV4 Skeletal Dysplasia: Spondylometaphyseal Dysplasia, Kozlowski Type

Abstract

Transient receptor potential vanilloid 4 channel ( TRPV4 ) gene mutations have been described in skeletal system and peripheral nervous system pathology. The case described here is a 9-year-old male child patient, born to a nonconsanguineous marriage with normal birth history who had difficulty in walking and stiffness of joints for the last 7 years, and progressive weakness of all four limbs and urine incontinence for 1 year following falls. Physical examination showed below-average weight and height and short trunk. Musculoskeletal examination revealed bony prominence bilaterally in the knee joints and contractures in knee and elbow joints with brachydactyly; muscle tone was increased, with brisk deep tendon reflexes. Skeletal survey showed platyspondyly with anterior beaking with metaphyseal dysplasia. Magnetic resonance imaging of the spine revealed atlantoaxial instability with hyperintense signal changes at a cervicomedullary junction and upper cervical cord with thinning and spinal canal stenosis suggestive of compressive myelopathy with platyspondyly and anterior beaking of the spine at cervical, thoracic and lumbar vertebrae. Exome sequencing revealed a heterozygous de novo variant c.2389G > A in exon 15 of TRPV4 , which results in the amino acid substitution p.Glu797Lys in the encoded protein. The characteristics observed indicated spondylometaphyseal dysplasia, Kozlowski type (SMD-K). The child underwent surgical intervention for compressive myelopathy by reduction of atlantoaxial dislocation with C1 lateral mass and C2 pars fusion using rib graft and fixation using screws and rods. To conclude, for any child presenting with progressive kyphoscoliosis, short stature, platyspondyly, and metaphyseal changes, a diagnosis of SMD-K should be considered and the patient and family should be advised to avoid spinal injuries.

Introduction

Transient receptor potential vanilloid 4 channel (TRPV4) belongs to a calcium-permeable nonselective cation channel involved in many different cell functions. ¹ Mutations in TRPV4 have been shown to cause autosomal dominant diseases in the skeletal and peripheral nervous systems. ² Skeletal dysplasia caused by TRPV4 mutations includes brachyolmia (BO; OMIM 113500), spondylometaphyseal dysplasia, Kozlowski type (SMD-K; OMIM 184252), metatropic dysplasia (MD; OMIM 156530), parastremmatic dysplasia (OMIM 168400), and spondyloepimetaphyseal dysplasia, Maroteaux type (OMIM 184095). ^{3 4 5}

Kozlowski et al described a form of spondylometaphyseal dysplasia which is characterized by short stature, with unique radiologic changes that occur in the metaphysis of the distal femur, femoral neck and trochanteric area, and generalized platyspondyly. ⁶ The exact prevalence of the disease is not known. Here, we report an Indian case of SMD-K with a TRPV4 mutation.

Background

The transient receptor potential (TRP) superfamily of ion channels is implicated in sensing a variety of physical and chemical stimuli in almost the entire body. Although TRP channels have many roles in cellular function, TRPV4 has a critical role in the skeletal system, and is expressed in several tissues such as esophagus, kidney, salivary gland skin, spleen, testis, appendix, thyroid, and urinary bladder ⁷ including cartilage and bone. ⁸ In bone, TRPV4 plays an important part in bone homeostasis by regulation of osteoclast differentiation. Mice without TRPV4 have greater bone mass because of impaired bone resorption. ⁹ TRPV4 in cartilage has been shown to play an important role in chondrocyte mechanotransduction and osmosensation. ^{10 11} In chondrocytes, blocking the TRPV4 aborts an anabolic response to mechanical loads, while activating TRPV4 mimics mechanical loading. ¹¹

Clinical Manifestations

TRPV4-associated skeletal dysplasia has variable phenotype. ² The spectrum of TRPV4-associated skeletal dysplasia ranges from lethal MD to mild anauxetic dysplasia (AD) BO, as shown in Table 1 .

Table 1. Comparison of various subtypes of TRPV4 skeletal dysplasia .

Findings	Mild		Intermediate		Severe
	Familial digital brachydactyly	AD brachyolmia	SMD-K	SED-M	Parastremmatic dysplasia	Metatropic dysplasia
Hands/feet	Normal at birth; progressive swelling and arthropathy	Clinodactyly	Brachydactyly, hypoplastic and delayed carpal bones ossification	Brachydactyly	Joint contractures	Brachydactyly, delayed carpal ossification
Spine	Normal	Scoliosis ± , kyphosis; platyspondyly	Platyspondyly, overfaced pedicles		Significant kyphoscoliosis, pedicles	Platyspondyly, overfaced pedicles
Long bones	NA	Minimal metaphyseal changes; short femoral neck	Mild metaphyseal changes; genu varum	Mild-to-moderate metaphyseal changes; genu varum	Severe metaphyseal changes severe limb deformity; joint contractures	Dumbbell-shaped long bones, epiphyseal dysplasia and prominent joints
Pelvis	Normal	NA	Square, short, flared iliac wings; flat, irregular acetabulae; coxa vara	Champagne-glass configuration of pelvic inlet	Halberd-shaped pelvis; supra-acetabular notches
Other	Average height; early-childhood onset	Mild short stature; limbs unaffected; good physical function	Short-trunk short-stature dwarfism; broad chest; early-childhood onset waddling gait	Short-trunk short-stature dwarfism	Significant short-trunk short-stature dwarfism	Lethal prenatally or at birth, short-limb short-stature dwarfism

Abbreviations: AD, autosomal dominant; NA, not available; SED-M, spondyloepiphyseal dysplasia, Maroteaux type; SMD-K, spondylometaphyseal dysplasia, Kozlowski type.

Diagnosis

The skeletal dysplasia is usually recognized by age of onset, severity, clinical features, and skeletal survey. Table 1 shows a comparison of various skeletal dysplasia caused by TRPV4 mutations. Rock et al ³ showed that the molecular pathogenic basis of TRPV4-pathies is a gain of function characterized by increased constitutive activity of the Ca ²⁺ channel, and the elevated channel activation are due to variety of mechanisms. It is worthwhile to try to find valid genotype–phenotype correlations as there are TRPV4 conditions with considerable phenotypic overlap and blurred boundaries with SMD-K since all cases have radiographic evidence of metaphyseal involvement. ¹²

Management

There is no curative treatment for TRPV4- associated skeletal dysplasia. Injuries to the spine, such as in contact sports, should be avoided as they can cause compressive myelopathy. Supportive therapy and treatment of complications should be made available. Early diagnosis helps avoid circumstances that might increase complications. The prognosis depends on the severity of the disease. Life expectancy is not usually affected unless there are respiratory complications.

Case

History

A 9-year-old boy born to a nonconsanguineous marriage with normal birth history was brought with difficulty in walking and stiffness of joints for the last 7 years and a history of self-fall at 8 years of age followed by progressive weakness of all four limbs and urinary incontinence. Developmentally, the child started walking by 1 year, but with an abnormal gait, running by 2 years, climbing up and down stairs by 3 years, and riding a tricycle by 4 years. He had difficulty in activities of daily living such as eating, dressing, and brushing of teeth because stiffness of joints. Social and language milestones were normal. The child suffered injury due to fall after slipping, and falling on the floor a year earlier, the individual developed difficulty in getting up from squatting and supine positions, and required assistance all the time. He also developed difficulty in walking and was able to walk only with support. He also developed urinary incontinence. The mother also noticed abnormal gait in the patient while walking in the last 5 months: twisting of the right knee and bending while walking. Currently, the child is nonambulatory. No other family members are affected as shown in Fig. 1 .

Fig. 1.

The pedigree shows that the index patient is the only child born out of a nonconsanguineous marriage. None of the other family members affected.

Examinations

The weight was 28 kg (World Health Organization Z: −1.75) and height was 115 cm (Z scoreless than −3 standard deviation) suggesting short stature. The upper and lower segment ratio was 51/62 (0.82) and arm span of 125 cm, which is more than the height, is suggestive of a short trunk. Head size was 50 cm and no corneal clouding noted. Musculoskeletal examination revealed bony prominence in bilateral knee joints, and bilateral contractures of knee and elbow. Muscle tone was increased, with brisk deep tendon reflexes with a power of 3/5 on Medical Research Council grading. Fig. 2A shows small hands, mild flexion bilaterally at the elbows; Fig. 2B , before the onset of weakness and after the fall torticollis of neck, scoliosis of spine, genu valgus; and Fig. 2C , the child after surgery, wheelchair bound with weakness of all four limbs. However, a photograph of the child before surgery is not shown. Fig. 3A shows short stubby and clawed hands; Fig. 3B , prominence at knee joints; and Fig. 3C , in supine posture, brachydactyly and flexion at knee joints. Informed consent was obtained from the parents and ethical clearance was obtained from institutional ethical committee (No-IGICH/ACA/EC-45/2020-21).

Fig. 2.

Clinical photographs, before development of weakness around 6 years of age brachydactyly (A), after development of weakness showing contractures of knee, and elbow joints with kyphoscoliosis (B), and after surgery showing child is wheelchair bound and on tracheostomy tube (C).

Fig. 3.

Hands showing brachydactyly (A), contractures of elbow and knee lower limbs (B), and kyphoscoliosis of trunk (C).

Investigations

Complete hemogram showed hemoglobin of 12.9 g/dL, white blood cell count of 9,250 cells/mm ³ , and platelet count of 286,000. Serum creatine phosphokinase level was 83 U/L, calcium was 10.4 mg/dL, alkaline phosphatase was 193 U/L, and phosphorus was 4.5mg/dL. Nerve conduction studies are normal in all four limbs. The hearing evaluation was normal. The immunoglobulin levels, CD4, and CD8 counts were normal.

Radiological Investigations

Skeletal survey was suggestive of skeletal dysplasia as described later. Skull radiograph shows prognathism ( Fig. 4A ) and atlantoaxial subluxation ( Fig. 4A ). Frontal pelvic radiograph shows squaring of the iliac bone, small greater sciatic notch, flat acetabular roof, short femoral neck, champagne-glass configuration of the pelvic inlet and coxa vara ( Fig. 4B ). Widened irregular metaphysis with oblique angulation was seen bilaterally in the upper end of the bilateral tibia ( Fig. 4C ). Broadened lower end of the femur and upper end of the tibia were seen. Genu valgum was seen bilaterally ( Fig. 4C ). Frontal radiograph of both hands shows appearance of seven carpal bones with poor ossification; widened, irregular, and oblique metaphysis in the distal end of ulna and radius, short metacarpal and phalanges ( Fig. 5A ). Frontal ( Fig. 5B ) and lateral ( Fig. 5C ) radiographs of thoracic and lumbar spine show multiple platyspondyly with anterior beaking. In Fig. 6 , frontal and oblique radiographs of both feet and ankles show metaphyseal widening and irregularity in distal end of tibia with poorly ossified tarsal bones with loss of ankle mortise. Fig. 7 shows magnetic resonance imaging (MRI) of the spine with atlantoaxial instability ( Fig. 7A ) hyperintense signal at cervicomedullary junction and upper cervical cord with thinning and spinal canal stenosis suggesting compressive myelopathy ( Fig. 7A ). There is platyspondyly involving C4 to C7 cervical and T1 thoracic vertebral bodies ( Fig. 7B ). MRI of the dorsolumbar spine shows platyspondyly and anterior beaking in lower dorsal vertebral bodies ( Fig. 7C ). Serial axial MRI T2 fast spin-echo sections at craniovertebral junction level (left to right) at atlanto-occipital ( Fig. 8A ), atlantoaxial ( Fig. 8B ), and C2 vertebral body ( Fig. 8C ) show rotational subluxation of atlantoaxial joint with posterolateral displacement of dens causing compression on spinal cord. There is thinning of spinal cord and hyperintense signal at cervicomedullary junction and upper cervical cord ( Fig. 8A–C ).

Fig. 4.

Lateral skull radiograph shows prognathism (black arrow), atlantoaxial subluxation (thin white arrow) (A), frontal pelvic radiograph showing squaring of iliac bone (thick white arrow), small greater sciatic notch (thin white arrow), flat acetabular roof (white arrowhead), short femoral neck (thin black arrow), champagne-glass configuration of pelvic inlet and coxa vara (B), widened irregular metaphysis with oblique angulation in upper end of bilateral tibia (thin blue arrow). Broadened lower end of femur and upper end of tibia. Genu valgum seen bilaterally (C).

Fig. 5.

Frontal radiograph of bilateral hand shows appearance of seven carpal bones with poor ossification. Widened, irregular, and oblique metaphysis in distal end of ulna and radius (thin black arrow). Short metacarpal and phalanges (thick white arrows) (A), frontal and lateral radiographs of thoracic and lumbar spine show multiple platyspondyly (black arrow in B) with anterior beaking (black arrow in C).

Fig. 6.

(A) Frontal and oblique radiographs of bilateral foot and ankle show metaphyseal widening and irregularity in distal end of tibia (white arrows). Poorly ossified tarsal bones with loss of ankle mortise (B).

Fig. 7.

MRI T2 FSE sagittal sections and T1 FSE sagittal sections showing atlantoaxial instability (thin white arrow). T2 FSE hyperintense signal at cervicomedullary junction and upper cervical cord with thinning and spinal canal stenosis suggesting compressive myelopathy (thick white arrow) (A). Platyspondyly involving C4 to C7 cervical and T1 thoracic vertebral bodies (arrow heads) (b). MRI T2 FSE sagittal, STIR sagittal images of dorsolumbar spine showing platyspondyly (thin white arrow) and anterior beaking (thick white arrow) in lower dorsal vertebral bodies (C). FSE, fast spin-echo; MRI, magnetic resonance imaging; STIR, short tau inversion recovery.

Fig. 8.

Serial axial MRI T2 FSE sections at craniovertebral junction level (left to right) at atlantooccipital (A), atlanto axial (B), and C2 vertebral body (C) level. Rotational subluxation of atlantoaxial joint with posterolateral displacement of dens causing compression on spinal cord (thin white arrow). Thinning of spinal cord and hyperintense signal at cervicomedullary junction and upper cervical cord (thick white arrow).

Genetic Testing

The molecular analysis of the genomic DNA from the index case was performed using captured exons and sequencing using illumina chemistry. The obtained sequence tags were assembled using the reference sequence hg19. The calculation of the mismatch was performed using GATK pipeline. Exome sequencing was done, which showed a heterozygous variant c.2389G > A in exon 15 of the TRPV4 gene corresponding to amino acid substitution p. Glu797Lys in the protein. Fig. 9 shows organization of the genetic locus TRPV4. Sanger sequencing in the parents showed absence of mutation suggesting it to be de novo variant.

Fig. 9.

Organization of the genetic locus TRPV4. (A) The TRPV4 is located on chromosome 12 on negative strand. The mRNA reference sequence of 16 exons and isoforms are indicated. (B) The protein of 871 amino acid polypeptide sequence and the domains on the membrane protein are indicated as derived from protein databases. The spectrum of mutations is indicated in different colors to indicate different clinical conditions. (C) The protein translation of the CCDS sequence of TRPV4 indicating wildtype gene and E797L missense mutation in exon 15 of the gene. (D) The protein TRPV4 and the protein domains constituting the channel and domain with C-terminal mutation. (E) The blind spot with the 3D protein structure in C terminal and indicating the AIM domain near CBD domain. (F) The 3D structure constructed as per PDB ID: 6PVO and red dots indicate the predicted position in the extracellular domain of the protein. TRPV4, transient receptor potential vanilloid 4 channel.

Management

The patient underwent surgical intervention for compressive myelopathy by reduction of atlantoaxial dislocation with C1 lateral mass and C2 pars fusion using rib graft and fixation using screws and rods. There was improvement of weakness in all four limbs.

Discussion

The patient was a 9-year-old male child with short stature and short trunk type, quadriparesis, neurogenic bladder, and radiological features showing involvement of spine and metaphysis of long bones. The conditions considered for a differential diagnosis were mucopolysaccharidosis (MPS) type IV and skeletal dysplasia SMD-K, MD, BO 3, spondylometaphyseal dysplasia axial type (SMDAX). The main difference between SMD-K and MD is the absence of dumbbell-shaped long bones and halberd pelvis in the former. ¹³ BO 3 is characterized by the presence of mainly severe kyphoscoliosis and irregular flattened cervical vertebrae, but in this case, there is involvement of both axial and appendicular skeleton. ³ SMDAX patients mainly have predominant ocular complaints. ¹⁴ We also considered MPS type IV; however, the child does not have corneal clouding and hepatosplenomegaly, hence MPS type IV was ruled out. Platyspondyly with anterior beaking, metaphyseal changes, and delayed bone age suggest SMD-K as the likely diagnosis. It is usually recognized on skeletal survey. There were no immunological abnormalities noted in our case compared with case reported by Majhi et al. ¹⁵

The role of E797L mutation in the CA ²⁺ channel and calmodulin (CaM) response is investigated by researchers in Xenopus egg. The need for patient-derived cells or heterologous expression of domain expression of E497L in HEK 293 cells is needed for understanding membrane potential and Ca ²⁺ imaging in response to agonist and CaM in the media. However, we have described the experimental details which shed light on the role of C-terminal residues upstream of CaM and their role in channel activity. TRPV4 is identified as channel proteins activated by cell swelling. ^{16 17 18} It also can be activated by other influences including temperature and acidic pH. ¹⁹ The mutations in E797L are known to cause skeletal dysplasia (SD). ^{2 5 20} In comparison with wildtype TRPV4, P799L mutations yielded much longer basal current which represents constituent open channel without any additional activation. The Ca ²⁺ imaging experiments show intracellular concentration after application of phorbol 12,13-didecanoate -PDD C-terminal Ca(2+)-CaM binding domain (CBD) which cause higher response than wildtype TRPV4 transfected cells. ^{20 21} Loukin et al (2015) examined the E297L and P799L located immediate upstream CBD. They increase the basal activity. Channels examined have reduced response to Ca ²⁺ –CaM. Deletion of 10 residues upstream of CaM (Δ795–804) results in strong constitutive activity and complete lack of Ca ²⁺ –CaM response. It proposed a region immediately upstream of CBD in autoinhibitory domain that maintains closed state, though electrostatic interactions and adjacent detachable Ca ²⁺ strictly interfere with this autoinhibition. Thus, TRPV4 E799L causes constitutive leakage.

There are two reports of cases of SMD-K from India. Nampoothiri et al ²² reported a total of 514 skeletal dysplasias of which one case was of SMD-K. The second report was by Uttarilli et al, ²³ who reported a total of 557 individuals with confirmed skeletal dysplasia of whom four cases were SMD-K. Odontoid hypoplasia has been described in SMD-K, but compressive myelopathy has not been reported.

The variant c.2389G > A in exon 15 of the TRPV4 gene causes the amino acid substitution glutamate to lysine at position 797. This change is in a well-established functional domain without benign variation ( Fig. 9 ). This variant is labeled “damaging” by multiple computational tools including PolyPhen2, SIFT, and Fathmm. The variant is reported in ClinVar as pathogenic. Krakow et al ⁴ reported the variant as P799L in patients with SMD-K. The variant can be classified as likely pathogenic by American College of Medical Genetics criteria.

The human genome data suggest that these thermosensitive TRPV channels are highly variable and carry possible deleterious mutations in human population. The analysis of single nucleotide polymorphism database in TRPV4 has shown 1,759 mutations in the gene and 376 mutations in the coding gene of TRPV4 . TRPV4 had much fewer harmful mutations (∼25) suggesting that mutations in TRPV4 as such may not be tolerable. This is in agreement with many mutations in TRPV4 which have been linked with the developmental disorders commonly termed as “channelopathies.” TRPV4, a Pro-19-Ser change is associated with hyponatremia. Similarly, Thr-89-Ile, Lys-197-Arg, and Leu-199-Phe mutations in TRPV4 individually lead to MD. The spectral mutations in TRPV4 gene which is known to manifest as a group of disorders, namely, BO, SMDK, MD, CMT Type C, distal SMA, skeletal dysplasia, motor neuropathy, distal arthropathy, brachydactyly, and AD BO. ^{5 24 25 26 27}

Detailed musculoskeletal examination, recording of height, arm span and upper segment to lower segment ratio, and skeletal survey are useful in children with myelopathy. Genetic testing does not differentiate subtypes of TRPV4 skeletal dysplasia and neuropathies as the same genotype causes the different phenotypes. On the basis of age of onset, clinical features, and skeletal survey, we can differentiate different subtypes of TRPV4 skeletal dysplasia. The detailed classification of genotypic–phenotypic presentation in clinical cases with TRPV4 mutations is represented in Fig. 9B .

Conclusion

For any child with quadriparesis and neurogenic bladder secondary to compressive myelopathy, diagnosis of SMD-K should be considered in addition to MPS type IV if there is disproportionate short stature. Skeletal survey gives important clues for diagnosis. One should advise avoiding spinal injuries as they precipitate or aggravate compression of the spinal cord. Early recognition of weakness is important for early surgical intervention.