Cant1 Affects Cartilage Proteoglycan Properties: Aggrecan and Decorin Characterization in a Mouse Model of Desbuquois Dysplasia Type 1.

Desbuquois dysplasia type 1 (DBQD1) is a recessive chondrodysplasia caused by mutations in the CANT1 gene, encoding for the Golgi Calcium-Activated Nucleotidase 1 (CANT1). The enzyme hydrolyzes UDP, the by-product of glycosyltransferase reactions, but it might play other roles in different cell types. Using a Cant1 knock-out mouse, we demonstrated that CANT1 is crucial for glycosaminoglycan (GAG) synthesis; however, its impact on the biochemical properties of cartilage proteoglycans remains unknown. Thus, in this work, we characterized decorin and aggrecan from primary chondrocyte cultures and cartilage biopsies of mutant mice at post-natal day 4 by Western blots and further investigated their distribution in the cartilage extracellular matrix (ECM) by immunohistochemistry. We demonstrated that the GAG synthesis defect caused by CANT1 impairment led to the synthesis and secretion of proteoglycans with shorter GAG chains compared with wild-type animals. However, this alteration did not result in the synthesis and secretion of decorin and aggrecan in the unglycanated form. Interestingly, the defect was not cartilage-specific since also skin decorin showed a reduced hydrodynamic size. Finally, immunohistochemical studies in epiphyseal sections of mutant mice demonstrated that the proteoglycan structural defect moderately affected decorin distribution in the ECM.

Clinical and Genetic Insights into Desbuquois Dysplasia: Review of 111 Case Reports.

Skeletal disorders encompass a wide array of conditions, many of which are associated with short stature. Among these, Desbuquois dysplasia is a rare but severe condition characterized by profound dwarfism, distinct facial features, joint hypermobility with multiple dislocations, and unique vertebral and metaphyseal anomalies. Desbuquois dysplasia is inherited in an autosomal recessive manner, with both the DBQD1 (MIM 251450) and DBQD2 (MIM 615777) forms resulting from biallelic mutations. Specifically, DBQD1 is associated with homozygous or compound heterozygous mutations in the CANT1 gene, while DBQD2 can result from mutations in either the CANT1 or XYLT1 genes. This review synthesizes the findings of 111 published case reports, including 54 cases of DBQD1, 39 cases of DBQD2, and 14 cases of the Kim variant (DDKV). Patients in this cohort had a median birth weight of 2505 g, a median length of 40 cm, and a median occipitofrontal circumference of 33 cm. The review highlights the phenotypic variations across Desbuquois dysplasia subtypes, particularly in facial characteristics, joint dislocations, and bone deformities. Genetic analyses revealed a considerable diversity in mutations, with over 35% of cases involving missense mutations, primarily affecting the CANT1 gene. Additionally, approximately 60% of patients had a history of parental consanguinity, indicating a potential genetic predisposition in certain populations. The identified mutations included deletions, insertions, and nucleotide substitutions, many of which resulted in premature stop codons and the production of truncated, likely nonfunctional proteins. These findings underscore the genetic and clinical complexity of Desbuquois dysplasia, highlighting the importance of early diagnosis and the potential for personalized therapeutic approaches. Continued research is essential to uncover the underlying mechanisms of this disorder and improve outcomes for affected individuals through targeted treatments.

[Surgical treatment of polydactyly]. / Chirurgische behandeling van polydactylie.

This clinical lesson addresses the treatment options for polydactyly, emphasizing the lack of consensus in the Netherlands regarding the timing and method of intervention. The study aims to provide evidence-based recommendations for the management of post-axial polydactyly type B. Two cases are presented, each illustrating different approaches to surgical intervention for post-axial polydactyly type B in a 1-year-old boy (Patient A) and a newborn girl (Patient B). Patient A undergoes surgical removal of an extra digit under general anesthesia after waiting for a year, while Patient B undergoes prompt surgical removal under local anesthesia. Both procedures are successful with no complications, demonstrating positive outcomes for early surgical intervention under local anesthesia. The study advocates for revising outdated national guidelines, recommending surgical removal under local anesthesia within the first three months after birth for post-axial polydactyly type B. Delaying intervention increases stress, risks, and costs without apparent benefits. This clinical lesson calls for optimizing care for children with post-axial polydactyly type B through guideline updates.

The First Patient with Tibial Hemimelia-Polysyndactyly-Triphalangeal Thumb Syndrome Caused by De Novo c.423+4916 T>C ZRS Variant: A Case Report.

Genetic variants in the zone of polarizing activity regulatory sequence (ZRS) that induce ectopic expression of the SHH gene have been associated with different ZRS-related phenotypes. We report the first patient with a de novo variant, c.423+4916 T>C, in ZRS (previously classified as a variant of uncertain significance) that causes tibial hemimelia-polysyndactyly-triphalangeal thumb syndrome (THPTTS). A two-month-old male patient presented with bilateral preaxial polydactyly, triphalangeal thumb, and tibial agenesis and was heterozygous for the variant c.423+4916T>C (neither of his parents was a carrier). The findings obtained from the family study were sufficient to reclassify the variant from "uncertain significance" to "likely pathogenic" according to three criteria from the American College of Medical Genetics and Genomics guidelines, as follows: (1) absence of gnomAD, (2) confirmation of paternity and maternity, and (3) strong phenotype-genotype association. In ZRS-associated syndromes, a wide clinical spectrum has been observed, ranging from polydactyly to THPTTS; our patient has the most severe and rare phenotype. We did not perform functional assays. However, the c.423+4916T>C variant is flanked by three variants, which have been proven not only to cause the phenotype but also to increase the expression of SHH. Through all this data gathering, we consider the c.423+4916T>C variant to be causative of THPTTS.

Cancer risk in individuals with polydactyly: a Swedish population-based cohort study.

BACKGROUND: Polydactyly is a feature of several cancer predisposition syndromes (CPS), however, cancer risk in individuals with polydactyly is largely unknown. METHODS: We performed a matched cohort study using data from Swedish national registers. We included 6694 individuals with polydactyly, born in Sweden between 1970-2017. Polydactyly was categorised as thumb polydactyly, finger polydactyly, polydactyly+ (additional birth defects and/or intellectual disability) or isolated polydactyly. Each exposed individual was matched to 50 comparisons by sex, birth year and birth county. Associations were estimated through Cox proportional hazard models. FINDINGS: An increased childhood cancer risk was found in males (HR 4.24, 95% CI 2.03-8.84) and females (HR 3.32, 95% CI 1.44-7.63) with polydactyly+. Isolated polydactyly was associated with cancer in childhood (HR 1.87, 95% CI 1.05-3.33) and young adulthood (HR 2.30, 95% CI 1.17-4.50) in males but not in females. The increased cancer risk remained after exclusion of two known CPS: Down syndrome and neurofibromatosis. The highest site-specific cancer risk was observed for kidney cancer and leukaemia. CONCLUSIONS: An increased cancer risk was found in individuals with polydactyly, especially in males and in individuals with polydactyly+. We encourage future research about polydactyly and cancer associations and emphasise the importance of clinical phenotyping.

Identification of truncated variants in GLI family zinc finger 3 (GLI3) associated with polydactyly.

BACKGROUND: Polydactyly is a prevalent congenital anomaly with an incidence of 2.14 per 1000 live births in China. GLI family zinc finger 3 (GLI3) is a classical causative gene of polydactyly, and serves as a pivotal transcription factor in the hedgehog signaling pathway, regulating the development of the anterior-posterior axis in limbs. METHODS: Three pedigrees of polydactyly patients were enrolled from Hunan Province, China. Pathogenic variants were identified by whole-exome sequencing (WES) and Sanger sequencing. RESULTS: Three variants in GLI3 were identified in three unrelated families, including a novel deletion variant (c.1372del, p.Thr458GlnfsTer44), a novel insertion-deletion (indel) variant (c.1967_1968delinsAA, p.Ser656Ter), and a nonsense variant (c.2374 C > T, p.Arg792Ter). These variants were present exclusively in patients but not in healthy individuals. CONCLUSIONS: We identified three pathogenic GLI3 variants in polydactyly patients, broadening the genetic spectrum of GLI3 and contributing significantly to genetic counseling and diagnosis for polydactyly.

A novel homozygous FAM92A gene (CIBAR1) variant further confirms its association with non-syndromic postaxial polydactyly type A9 (PAPA9).

Polydactyly is a very common digit anomaly, having extra digits in hands and/or toes. Non-syndromic polydactyly in both autosomal dominant and autosomal recessive forms are caused by disease-causing variants in several genes, including GLI1, GLI3, ZNF141, FAM92A, IQCE, KIAA0825, MIPOL1, STKLD1, PITX1, and DACH1. Whole exome sequencing (WES) followed by bi-directional Sanger sequencing was performed for the single affected individual (II-1) of the family to reveal the disease causative variant/gene. 3D protein modeling and structural molecular docking was performed to determine the effect of the identified mutation on the overall protein structure. WES revealed a novel biallelic missense variant (c.472G>C; p.Ala158Pro) in exon 6 of the FAM92A gene. The identified variant segregated perfectly with the disease phenotype using Sanger sequencing. Furthermore, Insilco analysis revealed that the variant significantly changes the protein secondary structure, and substantially impact the stability of FAM92A. We report the second FAM92A disease-causing mutation associated with recessive non-syndromic postaxial polydactyly. The data further confirms the contribution of FAM92A in limb development and patterning.

Treatment for Lateral Ray Polydactyly with Brachydactyly of the Foot.

Surgical methods for lateral ray polydactyly with brachydactyly of the foot include simple toe ablation and toe lengthening. However, there are few reports on comparative studies, and there is no standard treatment. We retrospectively investigated cases of lateral ray polydactyly with brachydactyly treated at our department and related facilities. In our study, the prevalence of Hirai-Togashi classification type IV was 8.8% (13/147 toes). Five patients did not request toe lengthening and underwent simple ablation, resulting in a shortened remaining toe in these 5 patients. The surgical methods for toe lengthening were pedicle bone grafting in 2 cases and on-top formation in 6 cases. Good results can be obtained in the most common phalangeal type cases, but care must be taken in cases with block-shaped metatarsal heads to avoid poor toe alignment.

Phenotypic and genotypic analysis of 11 fetal cases with Bardet-Biedl syndrome.

OBJECTIVE: To present the prenatal sonographic features and genomic spectrum of pregnancies with fetal Bardet-Biedl syndrome (BBS). METHODS: This was a retrospective study of 11 cases with BBS diagnosed by prenatal ultrasound and confirmed by genetic testing. Clinical and laboratory data were collected and reviewed for these cases, including maternal demographics, prenatal sonographic findings, molecular testing sequencing results, and pregnancy outcomes. RESULTS: All cases had unremarkable first-trimester ultrasound scans without reporting limb malformations. All had second-trimester abnormal ultrasounds: postaxial polydactyly in nine cases (9/11), renal abnormalities in seven (7/11), reduced amniotic fluid volume in two (2/11), central nervous system anomalies in two (2/11), and ascites in three (3/11). Ten fetuses presented with at least two-system anomalies, and one (Case 11) presented with only postaxial polydactyly. Variants were detected in five genes, including BBS2, ARL6/BBS3, BBS7, CEP290/BBS14 and IFT74/BBS22. Ten pregnancies were terminated in the second trimester, while one continued to term. CONCLUSION: Enlarged hyperechogenic kidneys and postaxial polydactyly are the two most common sonographic features of fetal BBS. Prenatal diagnosis of BBS can be done with ultrasound and genetic testing although the diagnosis may be made in the second trimester.

A novel missense variant located within the zinc finger domain of the GLI3 gene was identified in a Vietnamese pedigree with index finger polydactyly.

BACKGROUND: Polydactyly, particularly of the index finger, remains an intriguing anomaly for which no specific gene or locus has been definitively linked to this phenotype. In this study, we conducted an investigation of a three-generation family displaying index finger polydactyly. METHODS: Exome sequencing was conducted on the patient, with a filtration to identify potential causal variation. Validation of the obtained variant was conducted by Sanger sequencing, encompassing all family members. RESULTS: Exome analysis uncovered a novel heterozygous missense variant (c.1482A>T; p.Gln494His) at the zinc finger DNA-binding domain of the GLI3 protein within the proband and all affected family members. Remarkably, the variant was absent in unaffected individuals within the pedigree, underscoring its association with the polydactyly phenotype. Computational analyses revealed that GLI3 p.Gln494His impacts a residue that is highly conserved across species. CONCLUSION: The GLI3 zinc finger DNA-binding region is an essential part of the Sonic hedgehog signaling pathway, orchestrating crucial aspects of embryonic development through the regulation of target gene expression. This novel finding not only contributes valuable insights into the molecular pathways governing polydactyly during embryonic development but also has the potential to enhance diagnostic and screening capabilities for this condition in clinical settings.

Operative Management of Y-Shaped Metatarsal with Biphalangeal Sixth Toe.

BACKGROUND: In the literature, there is no consensus regarding the surgical management of postaxial polydactyly, and few cases of polymetatarsia with polydactyly have been reported. Treatment of the complete deformity will prevent further foot and gait disorders. OBJECTIVE: To identify literature relevant to the operative management of Y-shaped metatarsal with biphalangeal sixth toe and related skin and wound care to improve surgical treatment protocols from a clinical experience perspective. DATA SOURCES: The authors searched several electronic databases in December 2022 for articles related to postaxial polysyndactyly in the feet and polymetatarsia. Databases searched included PubMed, SciELO, ScienceDirect, Cochrane Database of Systematic Reviews, and Google Scholar gray literature. STUDY SELECTION: Two independent researchers conducted the searches and read the article titles and abstracts. Studies were included if they were narrative reviews, case studies, or observational studies; written in English or Spanish; and published between 2012 and 2022. Nonhuman studies were excluded. Studies that met the inclusion criteria were fully evaluated. Disagreements between reviewers were resolved by consensus, and when there was no consensus, a senior researcher was consulted. DATA EXTRACTION: The following data were extracted from the included studies using a standardized form: author and year of publication, study type, number of participants, sex, polydactyly location, polymetatarsia, type of polydactyly, participants' history of hereditary associated diseases or malformations, treatment, removal criteria, and timing of surgery. DATA SYNTHESIS: Authors evaluated 11 studies of postaxial polydactyly that included a total of 153 participants (64 men, 89 women). They also document their clinical experience with a surgical technique used in cases of bilateral postaxial polydactyly of the foot with a Y-shaped metatarsal with biphalangeal sixth toe. CONCLUSIONS: Surgical correction with lateral removal of the sixth toe is a resolutive treatment to improve the functionality of the foot, its aesthetic appearance, and the patient's quality of life. Case-specific treatment should be applied and tailored to meet the individual needs. The biomechanics of gait and shoe problems in these patients improve with surgical treatment, without presenting secondary aesthetic problems in skin care.

Single-cell transcriptome analysis reveals characteristic transcription factors in polydactyly.

OBJECTIVE: The study aims to provide guidance on the identification of multiple-digit malformations as potential biomarkers and therapeutic targets. MATERIALS AND METHODS: Single-cell RNA sequencing (scRNA-seq) data of four multiple-finger malformation samples were downloaded from the GEO public database. Fibroblasts and keratinocytes were divided into cellular subpopulations and the transcription factors of different subpopulations were analyzed. The regulatory network of transcription factors and their target genes were constructed to analyze the functionality of regulons. RESULTS: Examination of the transcriptional profile data from 11,806 single cells uncovered significant associations between regulons and cell function in polydactyly. Specifically, the analysis highlighted the involvement of HOX family members and GLI2 transcription factors, including HOXD13, MSX2, LHX2, EMX2, LEF1, CREB3L2, and LHX2, in the polydactyly process within fibroblast cells. Furthermore, it sheds light on the roles of HES2 and GLIS1 in the formation and development of keratinocytes. CONCLUSIONS: Significant presence of transcription factors, especially HOXD13, MSX2, and LHX2, may be strongly related to the development of polydactyly.

A de novo Mutation (p.Gln277X) of Cyclin D2 is Responsible for a Child with Megalencephaly-Polymicrogyria-Polydactyly-Hydrocephalus Syndrome.

Megalencephaly-polymicrogyria-polydactyly-hydrocephalus syndrome (MPPH), a type of overgrowth syndrome, is characterized by progressive megalencephaly, cortical brain malformations, and distal limb anomalies. Previous studies have revealed that the overactivity of the phosphatidylinositol 3-kinase-Protein kinase B pathway and the increased cyclin D2 (CCND2) expression were the main factors contributing to this disease. Here, we present the case of a patient who exhibited megalencephaly, polymicrogyria, abnormal neuronal migration, and developmental delay. Serum tandem mass spectrometry and chromosome examination did not detect any metabolic abnormalities or copy number variants. However, whole-exome sequencing and Sanger sequencing revealed a de novo nonsense mutation (NM_001759.3: c.829C>T; p.Gln277X) in the CCND2 gene of the patient. Bioinformatics analysis predicted that this mutation may disrupt the structure and surface charge of the CCND2 protein. This disruption could potentially prevent polyubiquitination of CCND2, leading to its resistance against degradation. Consequently, this could drive cell division and growth by altering the activity of key cell cycle regulatory nodes, ultimately contributing to the development of MPPH. This study not only presents a new case of MPPH and expands the mutation spectrum of CCND2 but also enhances our understanding of the mechanisms connecting CCND2 with overgrowth syndromes.

Season, household registry and isolated birth defects: a population-based case-control study in Danyang, China.

BACKGROUND: A birth population-based study was conducted in Danyang, Jiangsu Province, to evaluate major birth defects in emerging regions in China with similar maternal and neonatal care conditions. METHODS: We conducted a population-based study in a cohort of infants born in Danyang from 2014 to 2021, including 55 709 perinatal infants. Four categories of isolated birth defects were defined as cases: congenital heart defects (CHDs; n=2138), polydactyly (n=145), cleft lip with or without palate (CL/P; n=76) and accessory auricles (n=93). Infants with congenital malformations were identified by the Chinese Birth Defects Monitoring Network. RESULTS: Compared with autumn, conception in spring (OR=1.31 [1.16-1.48]) and winter (OR=1.39 [1.23-1.58]) was associated with an increased risk of CHD. Increased risk of CHD, CL/P and accessory auricles was significantly associated with non-local registered residence (OR=1.17 [1.07-1.28], OR=2.73 [1.52-4.88] and OR=2.11 [1.20-3.71], respectively). Individuals of Han nationality were less likely to have polydactyly (OR=0.23 [0.05-0.98]). CONCLUSIONS: The season of pregnancy was significantly associated with CHDs. Offspring of mothers with non-local registered hometown had greater risks of CHDs, CL/P and accessory auricles.

Streamlined, single-step non-viral CRISPR-Cas9 knockout strategy enhances gene editing efficiency in primary human chondrocyte populations.

BACKGROUND: CRISPR-Cas9-based genome engineering represents a powerful therapeutic tool for cartilage tissue engineering and for understanding molecular pathways driving cartilage diseases. However, primary chondrocytes are difficult to transfect and rapidly dedifferentiate during monolayer (2D) cell culture, making the lengthy expansion of a single-cell-derived edited clonal population not feasible. For this reason, functional genetics studies focused on cartilage and rheumatic diseases have long been carried out in cellular models that poorly recapitulate the native molecular properties of human cartilaginous tissue (e.g., cell lines, induced pluripotent stem cells). Here, we set out to develop a non-viral CRISPR-Cas9, bulk-gene editing method suitable for chondrocyte populations from different cartilaginous sources. METHODS: We screened electroporation and lipid nanoparticles for ribonucleoprotein (RNP) delivery in primary polydactyly chondrocytes, and optimized RNP reagents assembly. We knocked out RELA (also known as p65), a subunit of the nuclear factor kappa B (NF-κB), in polydactyly chondrocytes and further characterized knockout (KO) cells with RT-qPCR and Western Blot. We tested RELA KO in chondrocytes from diverse cartilaginous sources and characterized their phenotype with RT-qPCR. We examined the chondrogenic potential of wild-type (WT) and KO cell pellets in presence and absence of interleukin-1ß (IL-1ß). RESULTS: We established electroporation as the optimal transfection technique for chondrocytes enhancing transfection and editing efficiency, while preserving high cell viability. We knocked out RELA with an unprecedented efficiency of ~90%, confirming lower inflammatory pathways activation upon IL-1ß stimulation compared to unedited cells. Our protocol could be easily transferred to primary human chondrocytes harvested from osteoarthritis (OA) patients, human FE002 chondroprogenitor cells, bovine chondrocytes, and a human chondrocyte cell line, achieving comparable mean RELA KO editing levels using the same protocol. All KO pellets from primary human chondrocytes retained chondrogenic ability equivalent to WT cells, and additionally displayed enhanced matrix retention under inflamed conditions. CONCLUSIONS: We showcased the applicability of our bulk gene editing method to develop effective autologous and allogeneic off-the-shelf gene therapies strategies and to enable functional genetics studies in human chondrocytes to unravel molecular mechanisms of cartilage diseases.

Epidemiology of congenital polydactyly and syndactyly in Hunan Province, China.

OBJECTIVE: To describe the prevalence and epidemiology of congenital polydactyly and syndactyly in Hunan Province, China, 2016-2020. METHODS: Data were obtained from the Birth Defects Surveillance System in Hunan Province, China, 2016-2020. Prevalence of birth defects (polydactyly or syndactyly) is the number of cases per 1000 births (unit: ‰). Prevalence and 95% confidence intervals (CI) were calculated by the log-binomial method. Chi-square trend tests (χ2trend) were used to determine trends in prevalence by year. Crude odds ratios (ORs) were calculated to examine the association of each demographic characteristic with polydactyly and syndactyly. RESULTS: Our study included 847,755 births, and 14,459 birth defects were identified, including 1,888 polydactyly and 626 syndactyly cases, accounting for 13.06% and 4.33% of birth defects, respectively. The prevalences of total birth defects, polydactyly, and syndactyly were 17.06‰ (95%CI: 16.78-17.33), 2.23‰ (95%CI: 2.13-2.33), and 0.74‰ (95%CI: 0.68-0.80), respectively. Most polydactyly (96.77%) and syndactyly (95.69%) were diagnosed postnatally (within 7 days). From 2016 to 2020, the prevalences of polydactyly were 1.94‰, 2.07‰, 2.20‰, 2.54‰, and 2.48‰, respectively, showing an upward trend (χ2trend = 19.48, P < 0.01); The prevalences of syndactyly were 0.62‰, 0.66‰, 0.77‰, 0.81‰, and 0.89‰, respectively, showing an upward trend (χ2trend = 10.81, P = 0.03). Hand polydactyly (2.26‰ vs. 1.33‰, OR = 1.69, 95%CI: 1.52-1.87) and hand syndactyly (0.43‰ vs. 0.28‰, OR = 1.42, 95%CI: 1.14-1.76) were more common in males than females. Polydactyly (2.67‰ vs. 1.93‰, OR = 1.38, 95%CI: 1.26-1.51) and syndactyly (0.91‰ vs. 0.62‰, OR = 1.47, 95%CI: 1.26-1.72) were more common in urban areas than in rural areas. Compared to maternal age 25-29, hand polydactyly was more common in maternal age < 20 (2.48‰ vs. 1.74‰, OR = 1.43, 95%CI: 1.01-2.02) or ≥ 35 (2.25‰ vs. 1.74‰, OR = 1.30, 95%CI: 1.12-1.50). CONCLUSION: In summary, we have described the prevalence and epidemiology of polydactyly and syndactyly from hospital-based surveillance in Hunan Province, China, 2016-2020. Our findings make some original contributions to the field, which may be valuable for future research.