Two Novel Frameshift Mutations in the GLI3 Gene Underlie Non-Syndromic Polydactyly in Chinese Families.

Objective: Polydactyly is characterized by multiple distinct heterogeneous phenotypes, the etiologies of which involve several genes. This study aimed to explore the genetic defects and further clarify the molecular mechanism of polydactyly in several Chinese families. Methods: Three families with diverse phenotypes of non-syndromic polydactyly were analyzed: two were cases of familial disease, whereas one was sporadic. PCR and Sanger sequencing were used to screen for pathogenic mutations in two known disease-associated genes, GLI3 and HOXD13, while bioinformatic analyses predicted the pathogenicity of the identified variants. Reverse transcription PCR was used to analyze the splicing effect of an intronic variant. Results: Two novel heterozygous frameshift mutations (c.4478delG/p.S1493Tfs*18; c.846_c.847insC/p.R283Qfs*21) were identified in the GLI3 gene from two of the pedigrees. Both c.4478delG and c.846_c.847insC were later confirmed in affected and unaffected members and normal controls, to truncate and disrupt the integrity of the GLI3 protein, reduce its level of expression, and disrupt its biological function through nonsense-mediated mRNA decay (NMD). In addition, a deep intron mutation (c.125-47 C>A) was detected in the GLI3 gene from the sporadic case, however, both bioinformatics analysis (HSF, splice AI, and CBS) and RT-PCR indicated that the variant c.125-47 C>A had minimal if any impact on splicing of the GLI3 gene. Conclusion: Two newly identified heterozygous frameshift mutations in the GLI3 gene were detected in two families with non-syndromic polydactyly, further extending the mutational spectrum of the GLI3 gene in non-syndromic polydactyly. Moreover, our study further expanded the phenotypic spectrum of non-syndromic polydactyly.

Transitional ankle fracture management using a new joystick technique.

PURPOSE: To investigate the feasibility and short-term clinical efficacy of the arthrography-assisted joystick technique for the treatment of adolescent transitional ankle fracture. METHODS: A retrospective analysis was performed in 10 adolescent transitional ankle fracture patients treated with the arthrography-assisted joystick technology. There were 7 male patients and 3 female patients with a mean age of 12.80 ± 1.81 years (range: 10-16 years). All cases were closed fractures, and the operation was performed after the failure of manual reduction [X-ray or computed tomography (CT) showed that the fracture gap was > 2 mm]. After the operation, X-ray or CT examination was performed to assess reduction of the fracture. Ankle function and fracture healing were evaluated by assessing the imaging indexes and evaluating the American Orthopedic Foot and Ankle Society (AOFAS) ankle score system. RESULTS: All patients were followed up for an average period of 12.00 ± 3.40 months (range: 8-20 months). The operation time was 40.50 ± 16.90 min (range: 25-80 min), the number of intraoperative fluoroscopy sessions was 18.70 ± 5.91 (range: 10-30 sessions), and the intraoperative blood loss was 5.90 ± 3.38 ml (range: 2-10 ml). X-ray examination showed that all cases achieved bone healing. The fracture healing time was 11.00 ± 2.45 weeks (range: 8-16 weeks). Four cases showed early closure of the epiphysis. The AOFAS scoring system evaluation results of these 10 pediatric cases were excellent. CONCLUSION: The arthrography-assisted joystick technique for the treatment of adolescent transitional ankle fracture offers advantages, such as minimal trauma, simple operation, ideal reduction effect, and the recent curative effect is satisfactory.

Do the Number, Size, and Position of Partially Threaded Screws Affect the Radiological Healing of Surgically Treated Displaced Femoral Neck Fractures? A Review of 136 Children.

Background and Objectives: The quantity, size, and position of implants might affect the fracture healing process of surgically treated displaced pediatric femoral neck fractures (PFNFs). The aim of this retrospective multicenter study was to evaluate the correlation between the time needed to achieve radiological union and the number, size, and location of the partially threaded cannulated screws (PTCSs) in children with displaced PFNFs. Materials and Methods: A retrospective review of 136 children (mean age: 10.6 ± 3.8 years) with displaced PFNFs treated by two (n = 103) or three (n = 33) PTCSs was carried out. Student's t-tests, one-way ANOVA, Cox regression analysis, and multiple linear regression analyses were performed to investigate the variables affecting the time needed to achieve radiological fracture healing according to the number, size, and position of PTCSs, as assessed on plain radiographs. Results: A total of 132 hips achieved union at an average of 3.2 ± 1.6 months after the initial surgery. The time needed to achieve union in the patients treated with two or three PTCSs was comparable (p = 0.36). Among the fractures treated by two PTCSs, the time needed to achieve union did not correlate with the size of the implant (p = 0.122), or with the angulation between the PTCSs on anterior-posterior (p = 0.257) and lateral radiographs (p = 0.547). The time needed to achieve union in the fractures that were fully compressed by the implants was similar to the partially compressed fractures (p = 0.08). Conclusions: The number, size, and position of the PTCSs do not affect the radiological healing in the children with displaced PFNFs treated surgically.

Genetically Predicted Milk Intake Increased Femoral Neck Bone Mineral Density in Women But Not in Men.

Background: Cow milk contains more calcium, magnesium, potassium, zinc, and phosphorus minerals. For a long time, people have believed that increasing milk intake is beneficial to increasing bone density. Many confounding factors can affect milk consumption, and thus the association described to date may not be causal. We explored the causal relationship between genetically predicted milk consumption and Bone Mineral Density (BMD) of the femoral neck and lumbar spine based on 53,236 individuals from 27 studies of European ancestry using the Mendelian randomization (MR) study. 32,961 individuals of European and East Asian ancestry were used for sensitivity analysis. Methods: A genetic instrument used for evaluating milk consumption is rs4988235, a locus located at 13,910 base pairs upstream of the LCT gene. A Mendelian randomization (MR) analysis was conducted to study the effect of selected single nucleotide polymorphisms (SNPs) and BMD. The summary-level data for BMD of the femoral neck and lumbar spine were obtained from two GWAS meta-analyses ['Data Release 2012' and 'Data Release 2015' in the GEnetic Factors for OSteoporosis Consortium (GEFOS)]. Results: we found that genetically predicted milk consumption was not associated with FN-BMD(OR 1.007; 95% CI 0.991-1.023; P = 0.385), LS-BMD(OR 1.003; 95% CI 0.983-1.024; P = 0.743) by performing a meta-analysis of several different cohort studies. High levels of genetically predicted milk intake were positively associated with increased FN-BMD in Women. The OR for each additional milk intake increasing allele was 1.032 (95%CI 1.005-1.059; P = 0.014). However, no causal relationship was found between milk consumption and FN-BMD in men (OR 0.996; 95% CI 0.964-1.029; P = 0.839). Genetically predicted milk consumption was not significantly associated with LS-BMD in women (OR 1.017; 95% CI 0.991-1.043; P = 0.198) and men (OR 1.011; 95% CI 0.978-1.045; P = 0.523). Conclusion: Our study found that women who consume more milk have a higher FN-BMD. When studying the effect of milk consumption on bone density in further studies, we need to pay more attention to women.

Causal Association Between Tea Consumption and Bone Health: A Mendelian Randomization Study.

Background: Much observational research reported that tea consumption decreases the risk of osteoarthritis (OA), rheumatoid arthritis (RA), and osteoporosis (OP) which are the three major bone disorders. However, the observed correlation is inconclusive. To determine the causal relationship between genetically predicted tea intake and OA, RA, and OP, we performed a two-sample Mendelian randomization (MR) study based on large samples. Methods: The European population's genome-wide association meta-analysis (GWAS) dataset identified SNPs associated with tea consumption was obtained from Neale Lab's analysis of UK Biobank data that comprised 349,376 participants of European ancestry. We extracted genetic data for knee OA (17,885 controls and 4,462 cases), hip OA (50,898 controls and 12,625 cases), and RA (43,923 controls and 14,361 cases) from the UK Biobank and OP cases (93083 controls and 1,175 cases) from FinnGen Data Freeze 2. A MR study was conducted to examine the effect of selected single nucleotide polymorphisms (SNPs) and OA, RA, and OP risk. Several sensitivity analyses were performed with weighted median and inverse-variance weighted methods for estimating the causal effects. Results: In this MR study, the genetically predicted per one cup increase of tea consumption was not associated with knee OA (OR 1.11,95% CI: 0.79-1.55) using IVW with random effect. Genetic predisposition to tea consumption was not associated with hip OA (OR: 1.20, 95% CI: 0.84-1.71), RA (OR: 1.24 95% CI: 0.81-1.91), and OP (OR: 1.11, 95% CI: 0.89, 1.39). Following the sensitivity analysis, there was no potential pleiotropy. Conclusion: According to our study, According to our study, there was no statistical power to confirm a causal relationship between tea consumption and the risk of knee OA, hip OA, RA, and OP.

Relationship Between Plasma Vitamin C and COVID-19 Susceptibility and Severity: A Two-Sample Mendelian Randomization Study.

Background: Considering the antioxidant function of Vitamin C, also called ascorbic acid, it is widely used against viral infections such as coronavirus disease (COVID-19) based on in vitro, observational, and ecological studies. Many confounding factors that can affect Vitamin C levels. Thus, the association described to date may not be causal. To determine the causal relationship between genetically predicted plasma Vitamin C and COVID-19 susceptibility and severity, we performed two-sample Mendelian randomization (MR) based on large samples. Methods: The summary-level data for Vitamin C was obtained from a GWAS meta-analysis, which included 52,018 individuals from four studies of European ancestry. Data for COVID-19 HGI results were obtained from the meta-analysis of 35 GWASs with more than 1,000,000 subjects of European ancestry, including 32,494 cases with COVID-19 susceptibility and 1,316,207 controls, 9,986 cases with COVID-19 hospitalization and 1,877,672 controls, and 5,101 cases with COVID-19 severe disease and 1,383,241 controls. Mendelian randomization (MR) analysis was conducted to examine the effect of selected single nucleotide polymorphisms and COVID-19 susceptibility, hospitalization, disease severity. Several sensitivity analyses were performed with inverse-variance weighted (random-effect model), inverse variance weighted (fixed-effect model), weighted median, and maximum likelihood methods for estimating the causal effects. Results: In this MR study, genetic predisposition to the levels of plasma Vitamin C was not associated with COVID-19 susceptibility (OR: 0.99, 95% CI: 0.84-1.17, P = 0.91), hospitalization (OR: 1.10, 95% CI: 0.71-1.71, P = 0.67) and severity (OR: 0.83, 95% CI: 0.43-1.59, P = 0.58). The association was consistent in complementary analyses. No potential heterogeneities and directional pleiotropies were observed for the analysis results. Conclusion: According to our study, no correlation was observed between plasma Vitamin C levels and COVID-19 susceptibility and severity. Further studies in different ethnics are necessary to explore the potential role and mechanisms of circulating serum Vitamin C levels on COVID-19.

A Novel Intronic Splicing Mutation in the EXT2 Gene of a Chinese Family with Multiple Osteochondroma.

Background: Multiple osteochondroma (MO), an autosomal dominant genetic disease, is caused by heterozygous mutations in the EXT1 and EXT2 genes. Approximately 80% of pathogenic mutations are nonsense/missense mutations, small indels, and splicing mutations. Splicing mutations, particularly at the 3' and 5' splice sites, disrupt normal mRNA processing and cause exon skipping or aberrant splicing, ultimately resulting in protein truncation and loss of function. Methods: Polymerase chain reaction (PCR) and Sanger sequencing were applied to detect subtle mutations in a Chinese family with MO, the pathogenicity of a splicing variant was predicted by bioinformatics and further verified using a minigene splicing assay. Results: A novel and heterozygous splicing mutation, c.626 + 2_626 + 5delTAGG, was identified in the EXT2 gene of the proband and the father by PCR and Sanger sequencing, whereas the unaffected mother and brother had wild-type alleles at the same site. Bioinformatics predicted that the 5' splicing site of exon 3 in the EXT2 gene was destroyed due to this mutation. A hybrid minigene splicing assay (HMSA) indicated that the mutation disturbed the normal splicing of the EXT2 gene mRNA and led to a deletion of 79 bp at the 5' end of exon 3, which resulted in aberrant splicing of exon 3 and introduced an earlier stop codon in the EXT2 gene. Conclusion: A novel splicing mutation was identified that produced the MO phenotype through aberrant splicing in a Chinese family. This observation, expands our knowledge of the spectrum of molecular pathogenic mechanisms leading to aberrant mRNA splicing.

miR-335-5P contributes to human osteoarthritis by targeting HBP1.

MicroRNA (miR)-335-5P has the ability to regulate chondrogenic differentiation and promote chondrogenesis in mouse mesenchymal stem cells. It is also abnormally elevated in human osteoarthritic chondrocytes. However, the biological function of miR-335-5P in osteoarthritis (OA) is not well understood. The present study investigated the mechanism of miR-335-5P in the pathogenesis of OA. To investigate the effect of miR-335-5P on the pathogenesis of OA in vitro, a miR-335-5P mimic and inhibitor were transfected into chondrocytes. Cell Counting kit-8 assay and flow cytometry were used to observe the effects of miR-335-5P on chondrocyte apoptosis and the expression of cartilage-specific genes, such as aggrecan, collagen II, matrix metalloproteinase 13 and collagen X, were detected by reverse transcription-quantitative PCR and western blot analysis. Moreover, the current study assessed whether HMG-box transcription factor 1 (HBP1) is a novel target of miR-335-5P with dual luciferase reporter assays. Finally, a rescue experiment was used to prove the regulation between miR-335-5P and HBP1. The results revealed that HBP1 was a novel target of miR-335-5P, and that miR-335-5P mediated the apoptosis of chondrocytes and changes in cartilage-specific genes via targeting HBP1. Overall, the present study revealed that miR-335-5P mediated the development of OA by targeting the HBP1 gene and promoting chondrocyte apoptosis. These data suggested that miR-335-5P may be used to develop novel early-stage diagnostic and therapeutic strategies for OA.

Application of percutaneous vertebroplasty and percutaneous kyphoplasty in treating Kümmell's patients with different stages and postural correction status.

Objectives: To compare the effects of percutaneous vertebroplasty (PVP) or percutaneous kyphoplasty (PKP) in treating Kümmell's disease (KD) patients.Methods: This prospective cohort study involved 19 KD patients (20 involved vertebrae). The surgical selection was dependent on clinical stages and postural correction status. The status of vertebral reduction, amount of cement injection, and amount of cement leakage during the surgery were recorded. The anterior vertebral height and kyphotic angle were measured by X-ray scan. The degree of low back pain and the quality of life were assessed by the visual analog scale and Oswestry disability index, respectively.Results: On the third-day post-operation and the last follow-up, anterior vertebral height was increased, kyphosis angle was reduced, low back pain was relieved, and life quality was improved in both groups compared with pre-operation. There was no significant difference between the two groups in cement amount, cement leakage rate, vertebral height correction, and kyphotic correction angle. The nerve function of stage III patients with spinal cord injury was improved from a Frankel grade D to E.Expert Opinion: PVP and PKP could be used in treating KD patients, and the surgical selection could be dependent on the clinical stages and status of postural correction.Conclusion: PVP and PKP could be used in treating KD patients, and the surgical selection could be dependent on the clinical stages and status of postural correction.

Comparison between the Pavlik harness and the Tübingen hip flexion splint for the early treatment of developmental dysplasia of the hip.

The Pavlik harness and the Tübingen hip flexion splint (Tübingen hip flexion splint) are two effective options for the early management of patients younger than 6 months of age with developmental dysplasia of the hip (DDH). The main objective of this study was to evaluate the clinical and radiological outcomes of patients younger than 6 months of age with type IIb to IV DDH managed by Pavlik harness or Tübingen hip flexion splint. The Pavlik harness and Tübingen hip flexion splint groups were comparable regarding the affected side (P = 0.09), Graf grade (P = 0.635), and age at initial treatment (P = 0.77). Overall, failure rates were 12 and 33% in Pavlik harness (4/33 hips) and Tübingen hip flexion splint groups (14/43 hips), respectively (P = 0.038). No cases of avascular necrosis (AVN) were found in either group. In the Tübingen hip flexion splint group, the failure rate was significantly higher in bilateral cases (66.6%; P = 0.004), in severe forms (Graf grade IV hips; P ≤ 0.0001), and in patients with lower age at initial treatment (67.7 ± 39.3 days; P = 0.005). The average follow-up time was 30.35 ± 3.58 months (range: 24-36). At the last follow-up visit, no statistically significant differences were found between the Pavlik harness and Tübingen hip flexion splint groups regarding the acetabular index (t = 0.632; P = 0.53) or center-edge angle (Z = -0.303; P = 0.762). Our study showed that both the brace treatments for DDH in children younger than 6 months of age were effective and well tolerated. However, Tübingen hip flexion splint should not be used in patients with severe forms of DDH (Graf grade IV hips).

Clinical, radiographic and pedobarographic analysis of skeletally immature patients with surgically treated distal metaphyseal fractures of the tibia: is concomitant fixation of the fibula necessary?

This study evaluated the outcomes of distal tibia metaphyseal fractures (DTMFs) managed surgically and compared the outcomes of patients with and without associated fibula fracture fixation. Thirty-two consecutive patients (14 males; mean age at the time of injury: 7.8 years) with closed displaced DTMFs, with (22 patients; group A) or without associated fibula fractures fixation (10 patients; group B), were included. Besides standard radiographic measurements, the following static and dynamic pedobarographic parameters were evaluated: foot total static plantar pressure percentage (PP%), static plantar pressure percentage of the forefoot (PP%) and of the rear foot (PP%), landing sequence of the metatarsals during contact with the ground (MT), and impulse percentage of the metatarsal heads (MT%) and the medial and lateral heel (MH% and LH%). All patients were followed for at least 2 years (range: 2-4.5 years). Functional outcomes were excellent to good in all but two patients (93.8%) at the last follow-up visit using the Johner-Wruths criteria'. Radiographic measurements, PP%, PP%, PP%, MT%, MH% and LH% were not significantly different between two groups as well as between injured and uninjured side of patients within the same group (P > 0.05). Abnormal MT sequence was found in 40.9% of group A (9/22) and in 40% of group B patients (4/10) (P > 0.05). Stabilization of associated fibula fractures did not significantly impact the clinical, radiographic and pedobarographic outcomes of the children with displaced DTMFs who were surgically treated. Level of evidence: Level III.

Free Fracture Fragment Predicts Poorer Outcomes in Adolescents With Delbet II Femoral Neck Fracture.

BACKGROUND: This retrospective multicenter study investigated whether the presence of free fracture fragments is associated with the clinical outcome of adolescents with Delbet II femoral neck fracture (FNF). MATERIALS AND METHODS: The study population comprised 74 adolescents with Delbet II FNF, with an average age of 13.31 y (range, 10-17 y). There were 19 and 55 patients, respectively, with and without (control) free fracture fragments. The patients underwent open (n = 41) or closed (n = 33) fracture reduction and then cannulated screw internal fixation. Postoperative functional evaluation was performed using Ratliff's criteria, and complications such as femoral head necrosis, premature epiphyseal closure, and coxa vara were recorded. RESULTS: All patients achieved bony union during the mean follow-up period of 15.6 mo (range, 12-78 mo). By Ratliff's criteria, the clinical outcomes of the patients with free fracture fragment (three excellent, eight good, eight poor) were significantly worse compared with the control group (26 excellent, 20 good, and nine poor). The rate of complications (femoral head necrosis, premature epiphyseal closure, and coxa vara) of patients with free fracture fragments was significantly higher than that of the control patients. CONCLUSIONS: In adolescents with Delbet II FNF, the presence of free fracture fragments is associated with poorer clinical outcomes. Delbet II FNF in adolescents with free fracture fragment may be a special clinical entity with a poorer outcome.

Influence of bone mineral density and hip geometry on the different types of hip fracture.

The aim of this study was to assess the influence of bone mineral density and hip geometry on the fragility fracture of femoral neck and trochanteric region. There were 95 menopausal females of age ≥ 50 years with fragility fracture of hip, including 55 cases of femoral neck fracture and 40 cases of trochanteric fracture. Another 63 non-fractured females with normal bone mineral density (BMD) were chosen as control. BMD, hip axis length, neck-shaft angle and structural parameters including cross surface area, cortical thickness and buckling ratio were detected and compared. Compared with control group, the patients with femoral neck fracture or trochanteric fractures had significantly lower BMD of femoral neck, as well as lower cross surface area and cortical thickness and higher buckling ratio in femoral neck and trochanteric region. There were no significant differences of BMD and structural parameters in the femoral neck fracture group and intertrochanteric fracture group. Hip axis length and neck-shaft angle were not significantly different among three groups. The significant changes of BMD and proximal femur geometry were present in the fragility fracture of femoral neck and trochanteric region. The different types of hip fractures cannot be explained by these changes.

[Surgical options for delayed osteoporotic vertebral collapse].

OBJECTIVE: To investigate the surgical options and clinical effects of delayed osteoporotic vertebral collapse. METHODS: From May 2010 to October 2014, 19 patients (20 vertebrae) with delayed osteoporotic vertebral collapse(Kümmell's disease) were enrolled in this study. There were 7 males and 12 females, aged from 65 to 87 years old with a mean of (73.5±5.62) years. According to Li staging system of Kümmell's disease, 3 cases were stage II, 13 cases (14 vertebrae) were stage III without spinal cord injury, 3 cases were stage III with spinal cord injury. Patients were respectively treated with percutaneous vertebroplasty(PVP) or percutaneous kyphoplasty(PKP) on the basis of the degree of postural reduction during operation. Injected cement volume, cement leakage, vertebral height restoration and local kyphotic reduction were observed. Visual analogue scale (VAS) and Oswestry Disability Index(ODI) were respectively used to assess the pain and function before and after operation. Frankel grade were used to evaluate neurological status. RESULTS: Seven vertebrae with satisfactory postural reduction were treated with PVP, 13 vertebrae with unsatisfactory postural reduction were treated with PKP, 3 patients with spinal cord injury were treated with decompression and posterior short segment fixation at the same time. All patients were followed up from 10 to 48 months with an average of 21.2 months. Cement leakage occurred in 4 cases with no symptom, 1 cases in PVP group and 3 cases in PKP group, there was no significant difference between two groups(P=0.561). The priming volume of cement was (6.40±0.94) ml in PVP group and (5.46±1.09) ml in PKP group (P>0.05). Three days after operation vs preoperation, the vertebral height restoration and kyphotic improvement was(31.71±11.35)%, (9.79±4.64)° in PVP group and (24.77±8.51)%, (8.15±2.97)° in PKP. There was no significant difference between two groups(P>0.05). Three days after operation, VAS of low back pain and ODI in all patients were improved than preoperative data(P<0.05), but there was no significant difference between two groups or between postoperative at 3 d and final follow up(P>0.05). Nerve function of 3 patients underwent decompression and fixation from Frankel D to E. CONCLUSIONS: According to Li staging system and the degree of introperative postural reduction, individualized surgical treatment for Kümmell's disease can obtain good clinical results. Bad postural reduction during operation maybe a risk factor of cement leakage.