OPPGClinical manifestation Patient 1

Patient 1 (P1) was a 9-year-old girl born at full-term with healthy non-consanguineous parents. Her psychomotor development was normal. She was congenitally blind. The visual problems were noted at 3 months of age, showing right cataract and left microphthalmia in the ophthalmologic findings (Fig. 2a). At the age of 9, she had back pain and developed a kyphosis after falling. Radiologic examination showed significant systemic osteoporosis, increased dorsal kyphosis, and multiple vertebral compression fractures (Fig. 3a). BMD value of L1-4 was 0.363 g/cm2, which was significantly lower than the normal range for children of the same age and gender [21]. Results of biochemical parameters were mainly within the normal range, except for the deficiency of 25(OH)D (Table 1). Concurrently, ophthalmic reexamination showed that there was an absence of bilateral optic nerve.

Fig. 2

Ocular clinical features of patients with OPPG. a Cataract (right) and microphthalmia(left). b Enucleation of eyeball (right) and cataract (left)

Fig. 3

X-rays of the spine and pelvic of OPPG. a Generalized osteoporosis with fish mouth vertebra of patient 1 before treatment. b Improvement of lumbar vertebral body morphology and increased BMD of patient 1 after 3-year alendronate treatment. c Generalized osteoporosis with lumbar scoliosis of patient 2 before treatment and d after 6-month zoledronate treatment

Table 1 Baseline clinical features and bone mineral density in affected patients Patient 2

Patient 2 (P2) was the first child born to non-consanguineous parents. She was born blind and was diagnosed with right retinoblastoma and left retinal atrophy. At the age of 13, she underwent enucleation of her left eye due to tumor swelling and retinal perforation (Fig. 2b). She experienced chronic systemic bony pain and had her first nontraumatic fracture at the manubrium sterni at the age of 9. In the following years, many fractures occurred, including the ribs, humerus, thoracolumbar spine, hip, femur, tibia, and ankle, mainly involving long bones and vertebrae. When she was 16, she could not keep her upper body upright due to multiple vertebral compression fractures. She could stand unsupported but unable to walk independently and must rely on a wheelchair. In addition, she had torn ligaments in her knees and ankles and was also troubled by recurrent aseptic inflammation of her sacroiliac joint.

Besides, she has had frequent premature beats since childhood and developed into paroxysmal atrial fibrillation a few years later, and once had a cardiac arrest. However, no organic lesions were found by cardiac examination.

The results of biochemical parameters including serum calcium, phosphonium, ALP, PTH, and bone turnover markers were summarized in Table 1. Except vitamin D deficiency, all indicators were within the normal range. Radiologic examination showed thoracolumbar scoliosis, mild compression fractures of the local thoracic spine, and low pelvic BMD (Fig. 3c). The BMD of L1-4 was 0.630 g/cm2, and the Z-score value was − 4. Her parents and brother were healthy and had no vision or skeletal problems.

Identification of LRP5 variants

We verified the mutations in the patients and their family members if they agreed, and the results are presented in Figs. 1a, b and 6a. Sanger sequencing confirmed that P1 had a compound heterozygous mutation in exon 7 (c.1455G > T, p.Glu485Asp) and exon 8 (c.1708C > T, p.Arg570Trp) and P2 had a compound heterozygous mutation in exon 6 (c.1145C > T, p.Pro382Leu) and exon 23 (c.4830dupC, p.cys1611leufsx33). Meanwhile, some healthy relatives of the two patients carried a heterozygous mutation from their corresponding family.

Variant prediction and structural analysis

Multiple sequence alignment reveals that Glu485 is highly conserved in different species (Fig. 6a), and Glu485Asp is predicted as “disease-causing” by PolyPhen-2, MutationTaster, SIFT, and PROVEAN (Supplementary table). Furthermore, the three-dimensional structure homology modeling and visualization of the native and the three missense mutant proteins show that the three mutations have no apparent effect on protein structure. As for Pro382Leu, since no hydrogen bonds are formed before and after the mutation, there are no changes at least in the hydrogen bonds formed with neighboring atoms at the mutation site. However, as for Glu485Asp, our AlphaFold result shows there are changes in the length of the hydrogen bonds, while Rosetta result shows the number of hydrogen bonds changed from four in the native type to two in the mutant type. It also forms fewer hydrogen bonds at the site of the Arg570Trp in both modeling results (Fig. 6b and SI).

Treatment and follow-up

P1 received oral alendronate treatment at a dose of 70 mg/week from the age of 9 to 12. During the 3-year treatment, bone pain was significantly relieved. The baseline and follow-up serum levels of calcium, phosphorus, bone turnover markers, alkaline phosphatase, and PTH were summarized in Table 2. All of them were within in the normal level and showed no significant difference. Oral administration of 600 IU of vitamin D3 per day did not improve the state of vitamin D deficiency. The spinal X-ray after treatment showed significant improvement in the shape of the thoracolumbar spine (Fig. 3b), and there were no new fractures during the treatment. Furthermore, the BMD of L1-4, femur neck, and total hip was significantly increased by 107.2%, 70.0%, and 65.5%, respectively (Table 2).

Table 2 Pre- and post-treatment biochemical data and bone mineral density of two OPPG patients

As for P2, she was diagnosed as “osteoporosis” or “osteogenesis imperfecta” in other hospitals and took alendronate 70 mg per week at the age of 13. Six months later, she quit because of recurrence of fractures and no significant improvement in BMD. Last year, at the age of 27, she received 5 mg of zoledronic acid intravenously. Simultaneously, she took vitamin D3 800 IU and calcium 1000 mg per day. The clinical symptoms improved within half a year after injection. She could walk longer without a wheelchair. Although she fell down twice, she did not have a fracture, which was likely to happen in the past. BMD of L1-4, femoral neck, and total hip increased by 5.87%, 6.76%, and 4.15%, and β-CTX and osteocalcin decreased slightly after 6 months of treatment. Serum calcium, phosphorus, and parathyroid hormone levels were almost unchanged. Although the level of 25(OH)D increased, it was still below the normal range (Table 2).

No serious adverse drug reactions occurred in both patients during the treatment.

LRP5-HBMClinical manifestations Patient 3

Patient 3 (P3), a 32-year-old male, came to our department because of lumbodorsal pain. A torus palatinus in the center of the oral hard palate was detected by physical examination (Fig. 4a). X-ray examination revealed that the skull, thoracolumbar vertebrae, and pelvis bones were dense, and the cranial plate was significantly thickened (Fig. 5a). The Z-scores of BMD of the L1-4, femur neck, and total hip were + 10.5, + 11.1, and + 12.3, respectively (Table 1). The patients have no history of fractures, sensorimotor neuropathy, and visual or hearing impairment. The level of serum calcium, phosphonium, ALP, PTH, 25(OH)D, and bone turnover markers was all within the normal range (Table 1). His family members were all healthy.

Fig. 4

Clinical manifestations of patients with LRP5-HBM. a, b, c The torus palatinus in the center of the hard palate of 3 patients. d A slight depression in the right chest of patient 4

Fig. 5

X-rays of the skull, spine, humerus, and femur of two LRP5-HBM patients (a: patient 3; b: patient 4). Osteosclerosis of the skull with an enlarged sella turcica. Generalized osteosclerosis. Cortical thickening of the humerus and femur with a normal external shape

Patient 4

Patient 4 (P4) was a 6-year-old boy born at full-term who was found to have thoracic deformity at the age of 3. He was diagnosed as “Rickets” in the local hospital, but the symptoms did not improve after calcium and vitamin D supplementation. At the age of 6, the abnormal increased bone mineral density of ribs, sternum, thoracic vertebrae, and bilateral humeral metaphysis was identified by occasionally chest X-ray examination due to cough. In order to confirm the diagnosis, the child underwent further radiological examination. The examination found that the patient’s whole-body BMD increased and the skull plate thickened (Fig. 5b). The patient had normal tooth development and maxillary morphology. No jaw enlargement or palatal ring was found. The patient had a slight depression on the right side of the chest (Fig. 4d) and had no history of fracture.

Moreover, the value of BMD of L1-4, femoral neck, and total hip was 1.149 g/cm2, 1.327 g/cm2, and 1.321 g/cm2, respectively (Table 1). The biochemical indices including serum calcium and phosphonium, ALP, PTH, 25(OH)D, β-CTX, and OC were within the normal range (Table 1).

His parents were healthy and non-consanguineous.

Patients 5 and 6

Patients 5 (P5) and 6 (P6) were a mother-daughter pair, as reported in our previous study [18]. In brief, both of them had chronic lumbodorsal pain, an elongated mandible and torus palatinus (Fig. 4b and c), high bone mass, but no history of fractures. X-ray radiographs showed thickening of the cranial plates, an elongated mandible, cortical thickening of the long bones, and degenerative changes. The biochemical indices showed that they both had vitamin D deficiency. The serum ALP level of patient 6 was slightly higher than normal range, while β-CTX was much higher than the normal range (Table 1).

Identification of LRP5 variants

The LRP5 gene detection results of patients and their family members were shown in Figs. 1c, d, and e and 6a. By Sanger sequencing, the heterozygous c.724G > A (p.Ala242Thr) mutation was detected in P3, P5, and P6. And the heterozygous c.640G > A (p.Ala214Thr) mutation was detected in P4. We did not identify any pathogenic variants in family members other than the patients.

Fig. 6

Sanger sequencing and the three-dimensional modeling. a Multiple sequence alignments revealed that the glu485 residue in the LRP5 protein was highly conserved among species; Sanger sequencing of the LRP5 mutations: p.Glu485Asp (E485D, c.1455G > T) and p.Arg570Trp (R570W, c.1708C > T) in patient 1; p.Pro382Leu (P382L, c.1145C > T) and p.Cys1611LeufsX33 (c.4830dupC) in patient 2; p.Ala242Thr (A242T, c.724G > A) in patients 3, 5, and 6; p.Ala214Thr (A214T, c.640G > A) in patient 4. b Comparison of the three-dimensional modeling of wild and missense mutations’ local structures by AlphaFold; distribution of the six mutation sites on the LRP5 protein