Skeletal Defects Research Articles

An analysis of factors affecting survival in prenatally diagnosed omphalocele.

To estimate factors affecting survival in prenatally diagnosed omphalocele, factors predicting genetic abnormalities, and association of omphalocele and specific groups of anomalies. A retrospective observational study was performed, analyzing data of all omphalocele cases diagnosed prenatally in the perinatology clinic of a referral center. Demographic data, characteristics of the omphalocele (size,content, associated anomalies), results of genetic testing, pregnancy outcomes and postnatal outcomes were analyzed. Sixty-nine fetuses with omphalocele were included. The prevalence of omphalocele in livebirth was 0.007 %. Overall survival during the study period was 73.9 %.Twenty-eight (71.7 %) out of 39 cases with associated anomalies who were born live, survived, whereas survival was 85.7 % in the isolated cases. The most common anomaly associated with omphalocele were cardiac defects with 42 %; followed by placental or umbilical cord anomalies (28.9 %), skeletal defects (27.5), genitourinary anomalies (20.2 %), central nervous system (18.8 %) and facial anomalies (7.2 %), respectively. Eighty-five percent of the fetuses had at least one additional anomaly or ultrasound finding. Skeletal abnormalities and staged surgical repair of omphalocele were associated with survival. Associated skeletal anomalies and staged repair significantly increase the risk of postnatal death (OR: 4.6 95 % CI (1.1-19.5) and (OR: 10.3 95 % CI (1.6-63.9), respectively). Associated skeletal abnormalities and staged surgical repair are negatively associated with postnatal survival.

A Ready-To-Use Purified Exosome Product for Volumetric Muscle Loss and Functional Recovery.

Large skeletal muscle defects due to trauma or following tumor extirpation can result in substantial functional impairment. Purified exosomes are now available clinically and have been used for wound healing. The objective of this study was to evaluate the regenerative capacity of commercially available exosomes on an animal model of volumetric muscle loss (VML) and potential translation to human muscle injury. An established VML rat model was used. In the in-vitro experiment, rat myoblasts were isolated and co-cultured with 5% purified exosome product (PEP) to validate uptake. Myoblast proliferation and migration was evaluated with increasing concentrations of PEP (2.5%, 5%, and 10%) in comparison to control media(F10) and myoblast growth media (MGM). In the in-vivo experiment, a lateral gastrocnemius-VML defect was made in the rat hindlimb. Animals were randomized into four experimental groups; defects were treated with either surgery alone, fibrin sealant, fibrin sealant and PEP, or platelet rich plasma (PRP). The groups were further randomized into four recovery timepoints (14, 28, 45, or 90 days). The isometric tetanic force (ITF), which was measured as a percentage of force compared to normal limb, was used for functional evaluation. Florescence microscopy confirmed that 5% PEP demonstrated cellular uptake around 8-12 hrs. Compared to control, myoblasts showed faster proliferation with PEP irrespective of concentration. PEP concentrations of 2.5 and 5% promoted myoblast migration faster compared to control (<0.05). At 90 days postop, both the PEP and fibrin sealant and PRP groups showed greater ITF compared to control and fibrin sealant alone (<0.05). At 45 days postop, PEP with fibrin sealant had greater cellularity compared to control (<0.05). At 90 days postop, both PEP with fibrin sealant and the PRP treated groups had greater cellularity compared to fibrin sealant and control (<0.05). PEP promoted myoblast proliferation and migration. When delivered to a wound with a fibrin sealant, PEP allowed for muscle regeneration producing greater functional recovery and more cellularity in vivo compared to untreated animals. PEP may promote muscle regeneration in cases of VML, further research is warranted to evaluate PEP for the treatment of clinical muscle defects.

Physiochemical Dissolution Governs Early Modifications in Acid-Exposed Murine Bone with Long-term Recovery

Metabolic acidosis (MA), a disease affecting millions annually, is clinically characterized by a decrease in systemic pH and bicarbonate. MA has been shown to have skeletal consequences, such as increased bone loss and fractures. However, the mechanisms impacting the mechanics of bone remain unknown. To determine this, we characterized the skeletal phenotypes resulting from two independent regimes of ammonium chloride (NH4Cl) acid-dosing in skeletally mature CD-1 mice for 1, 3, 7, and 14 days to establish the temporal progression of materials-level skeletal defects. The classical, flat-dose model resulted in a temporary decrease in blood pH but did not reduce blood bicarbonate levels. Minimal alterations were observed in bone composition, structure, mechanics, and cellular behavior. However, the graded administration of NH4Cl maintained MA for up to 14 days. This resulted in decreased bone mineral content, collagen organization, and bone volume while increasing mineral crystallinity at early time points followed by a return to baseline. Changes to the matrix lead to a potential decrease in bone toughness at early time points, which may explain the increased fracture risk seen clinically. However, there were no changes in bone formation rate nor osteoclast number or activity during any time point. After 14 days, the bones were fully recovered even though the mice were still under acidotic conditions. Ultimately, we conclude that physicochemical dissolution is the main mechanism for early changes in murine bone, which is influenced by the dosing of exogenous acid loading. This knowledge will aid the discovery of treatments that prevent acidosis-induced bone dissolution.

Icariin Promotes Osteogenic Differentiation in a Cell Model with NF1 Gene Knockout by Activating the cAMP/PKA/CREB Pathway

Neurofibromatosis type 1 is a rare autosomal dominant genetic disorder, with up to 50% of patients clinically displaying skeletal defects. Currently, the pathogenesis of bone disorders in NF1 patients is unclear, and there are no effective preventive and treatment measures. In this study, we found that knockout of the NF1 gene reduced cAMP levels and osteogenic differentiation in an osteoblast model, and icariin activated the cAMP/PKA/CREB pathway to promote osteoblast differentiation of the NF1 gene knockout cell model by increasing intracellular cAMP levels. The PKA selective inhibitor H89 significantly impaired the stimulatory effect of icariin on osteogenesis in the NF1 cell model. In this study, an osteoblast model of NF1 was successfully constructed, and icariin was applied to the cell model for the first time. The results will help to elucidate the molecular mechanism of NF1 bone disease and provide new ideas for the clinical prevention and treatment of NF1 bone disease and drug development in the future.

Dolutegravir-induced neural tube defects in mice are folate responsive.

In 2018, the Botswana Tsepamo Study reported a nine-fold increased risk of neural tube defects in infants whose mothers were treated with dolutegravir (DTG) from the time of conception. As maternal folate supplementation and status is a well known modifier of neural tube defect (NTD) risk, we sought to evaluate birth outcomes in mice fed normal and low folic acid diets treated with DTG during pregnancy. DTG was evaluated for developmental toxicity using pregnant mice fed normal or low folic acid diet. CD-1 mice were provided diet with normal (3 mg/kg) or low (0.3 mg/kg) folic acid. They were treated with water, a human therapeutic-equivalent dose, or supratherapeutic dose of DTG from mouse embryonic day E6.5 to E12.5. Pregnant dams were sacrificed at term (E18.5) and fetuses were inspected for gross, internal, and skeletal defects. Fetuses with exencephaly, an NTD, were present in both therapeutic human equivalent and supratherapeutic exposures in dams fed low folic acid diet. Cleft palates were also found under both folate conditions. Recommended dietary folic acid levels during mouse pregnancy ameliorate developmental defects that arise from DTG exposure. Since low folate status in mice exposed to DTG increases the risk for NTDs, it is possible that DTG exposures in people living with HIV with low folate status during pregnancy may explain, at least in part, the elevated NTD risk signal observed in Botswana. Based on these results, future studies should consider folate status as a modifier for DTG-associated NTD risk.

Pathogenic RAB34 variants impair primary cilium assembly and cause a novel oral-facial-digital syndrome.

Oral-facial-digital syndromes (OFDS) are a group of clinically and genetically heterogeneous disorders characterized by defects in the development of the face and oral cavity along with digit anomalies. Pathogenic variants in over twenty genes encoding ciliary proteins have been found to cause OFDS through deleterious structural or functional impacts on primary cilia. We identified by exome sequencing bi-allelic missense variants in a novel disease-causing ciliary gene RAB34 in four individuals from three unrelated families. Affected individuals presented a novel form of OFDS (OFDS-RAB34) accompanied by cardiac, cerebral, skeletal, and anorectal defects. RAB34 encodes a member of the Rab GTPase superfamily and was recently identified as a key mediator of ciliary membrane formation. Unlike many genes required for cilium assembly, RAB34 acts selectively in cell types that use the intracellular ciliogenesis pathway, in which nascent cilia begin to form in the cytoplasm. We find that the protein products of these pathogenic variants, which are clustered near the RAB34 C-terminus, exhibit a strong loss of function. Although some variants retain the ability to be recruited to the mother centriole, cells expressing mutant RAB34 exhibit a significant defect in cilium assembly. While many Rab proteins have been previously linked to ciliogenesis, our studies establish RAB34 as the first small GTPase involved in OFDS and reveal the distinct clinical manifestations caused by impairment of intracellular ciliogenesis.

Three-Dimensional Scaffolds for Bone Tissue Engineering.

Immobilization using external or internal splints is a standard and effective procedure to treat minor skeletal fractures. In the case of major skeletal defects caused by extreme trauma, infectious diseases or tumors, the surgical implantation of a bone graft from external sources is required for a complete cure. Practical disadvantages, such as the risk of immune rejection and infection at the implant site, are high in xenografts and allografts. Currently, an autograft from the iliac crest of a patient is considered the "gold standard" method for treating large-scale skeletal defects. However, this method is not an ideal solution due to its limited availability and significant reports of morbidity in the harvest site (30%) as well as the implanted site (5-35%). Tissue-engineered bone grafts aim to create a mechanically strong, biologically viable and degradable bone graft by combining a three-dimensional porous scaffold with osteoblast or progenitor cells. The materials used for such tissue-engineered bone grafts can be broadly divided into ceramic materials (calcium phosphates) and biocompatible/bioactive synthetic polymers. This review summarizes the types of materials used to make scaffolds for cryo-preservable tissue-engineered bone grafts as well as the distinct methods adopted to create the scaffolds, including traditional scaffold fabrication methods (solvent-casting, gas-foaming, electrospinning, thermally induced phase separation) and more recent fabrication methods (fused deposition molding, stereolithography, selective laser sintering, Inkjet 3D printing, laser-assisted bioprinting and 3D bioprinting). This is followed by a short summation of the current osteochondrogenic models along with the required scaffold mechanical properties for in vivo applications. We then present a few results of the effects of freezing and thawing on the structural and mechanical integrity of PLLA scaffolds prepared by the thermally induced phase separation method and conclude this review article by summarizing the current regulatory requirements for tissue-engineered products.

Harnessing Biofabrication Strategies to Re-Surface Osteochondral Defects: Repair, Enhance, and Regenerate.

Osteochondral tissue (OC) is a complex and multiphasic system comprising cartilage and subchondral bone. The discrete OC architecture is layered with specific zones characterized by different compositions, morphology, collagen orientation, and chondrocyte phenotypes. To date, the treatment of osteochondral defects (OCD) remains a major clinical challenge due to the low self-regenerative capacity of damaged skeletal tissue, as well as the critical lack of functional tissue substitutes. Current clinical approaches fail to fully regenerate damaged OC recapitulating the zonal structure while granting long-term stability. Thus, the development of new biomimetic treatment strategies for the functional repair of OCDs is urgently needed. Here, we review recent developments in the preclinical investigation of novel functional approaches for the resurfacing of skeletal defects. The most recent studies on preclinical augmentation of OCDs and highlights on novel studies for the in vivo replacement of diseased cartilage are presented.

Sall4 restricts glycolytic metabolism in limb buds through transcriptional regulation of glycolytic enzyme genes

Recent studies illustrate the importance of regulation of cellular metabolism, especially glycolysis and pathways branching from glycolysis, during vertebrate embryo development. For example, glycolysis generates cellular energy ATP. Glucose carbons are also directed to the pentose phosphate pathway, which is needed to sustain anabolic processes in the rapidly growing embryos. However, our understanding of the exact status of glycolytic metabolism as well as genes that regulate glycolytic metabolism are still incomplete. Sall4 is a zinc finger transcription factor that is highly expressed in undifferentiated cells in developing mouse embryos, such as blastocysts and the post-implantation epiblast. TCre; Sall4 conditional knockout mouse embryos exhibit various defects in the posterior part of the body, including hindlimbs. Using transcriptomics approaches, we found that many genes encoding glycolytic enzymes are upregulated in the posterior trunk, including the hindlimb-forming region, of Sall4 conditional knockout mouse embryos. In situ hybridization and qRT-PCR also confirmed upregulation of expression of several glycolytic genes in hindlimb buds. A fraction of those genes are bound by SALL4 at the promoters, gene bodies or distantly-located regions, suggesting that Sall4 directly regulates expression of several glycolytic enzyme genes in hindlimb buds. To further gain insight into the metabolic status associated with the observed changes at the transcriptional level, we performed a comprehensive analysis of metabolite levels in limb buds in wild type and Sall4 conditional knockout embryos by high-resolution mass spectrometry. We found that the levels of metabolic intermediates of glycolysis are lower, but glycolytic end-products pyruvate and lactate did not exhibit differences in Sall4 conditional knockout hindlimb buds. The increased expression of glycolytic genes would have caused accelerated glycolytic flow, resulting in low levels of intermediates. This condition may have prevented intermediates from being re-directed to other pathways, such as the pentose phosphate pathway. Indeed, the change in glycolytic metabolite levels is associated with reduced levels of ATP and metabolites of the pentose phosphate pathway. To further test whether glycolysis regulates limb patterning downstream of Sall4, we conditionally inactivated Hk2, which encodes a rate-limiting enzyme gene in glycolysis and is regulated by Sall4. The TCre; Hk2 conditional knockout hindlimb exhibited a short femur, and a lack of tibia and anterior digits in hindlimbs, which are defects similarly found in the TCre; Sall4 conditional knockout. The similarity of skeletal defects in Sall4 mutants and Hk2 mutants suggests that regulation of glycolysis plays a role in hindlimb patterning. These data suggest that Sall4 restricts glycolysis in limb buds and contributes to patterning and regulation of glucose carbon flow during development of limb buds.

Haploinsufficiency of the essential gene Rps12 causes defects in erythropoiesis and hematopoietic stem cell maintenance.

Ribosomal protein (Rp) gene haploinsufficiency can result in Diamond-Blackfan Anemia (DBA), characterized by defective erythropoiesis and skeletal defects. Some mouse Rp mutations recapitulate DBA phenotypes, although others lack erythropoietic or skeletal defects. We generated a conditional knockout mouse to partially delete Rps12. Homozygous Rps12 deletion resulted in embryonic lethality. Mice inheriting the Rps12KO/+ genotype had growth and morphological defects, pancytopenia, and impaired erythropoiesis. A striking reduction in hematopoietic stem cells (HSCs) and progenitors in the bone marrow (BM) was associated with decreased ability to repopulate the blood system after competitive and non-competitive BM transplantation. Rps12KO/+ lost HSC quiescence, experienced ERK and MTOR activation, and increased global translation in HSC and progenitors. Post-natal heterozygous deletion of Rps12 in hematopoietic cells using Tal1-Cre-ERT also resulted in pancytopenia with decreased HSC numbers. However, post-natal Cre-ERT induction led to reduced translation in HSCs and progenitors, suggesting that this is the most direct consequence of Rps12 haploinsufficiency in hematopoietic cells. Thus, RpS12 has a strong requirement in HSC function, in addition to erythropoiesis.

Analysis of GPR126 polymorphisms and their relationship with scoliosis in Marfan syndrome and Marfan-like syndrome in Mexican patients.

Marfan syndrome (MFS) is an inherited connective tissue disorder. As the spinal growth depends on delicate balance of forces, conditions that affect musculoskeletal matrix often lead to spinal deformities. A large cross-sectional study revealed a 63% prevalence of scoliosis among patients with MFS. Multi-ethnic genome-wide association studies and analyses of human genetic mutations showed that variations and mutations of G protein-coupled receptor 126 (GPR126)locus are associated with multiple skeletal defects, including shorter stature and adolescent idiopathic scoliosis. The study included 54 patients with MFS and 196 control patients. The DNA was extracted from peripheral blood using the saline expulsion method and single nucleotide polymorphism (SNP) determination was carried out using TaqMan probes. Allelic discrimination was performed by RT-qPCR. Significant differences in genotype frequencies were found for SNP rs6570507 in relation to MFS and sex (recessive model, OR 2.46, 95% CI 1.03 -5.87; P = 0.03) and rs7755109 (overdominant model, OR 0.39, 95% CI 0.16-0.91; P = 0.03). The most significant association was found in SNP rs7755109, where the frequency of genotype AG was significantly different between MFS patients with scoliosis and those without (OR 5.68, 95% CI 1.09-29.48; P=0.04). This study, for the first time, examined the genetic association of SNP GPR126 with the risk of scoliosis in patients with connective tissue diseases. The study revealed that SNP rs7755109 is associated with the presence of scoliosis in Mexican patients with MFS.

FREE VASCULARIZED FIBULA GRAFTING AS MANAGEMENT FOR LARGE SKELETAL DEFECTS IN HUMERAL OSTEOMYELITIS IN NORTHERN IRELAND

IntroductionThe purpose of this study is to evaluate the radiological and clinical outcomes in Northern Ireland of free vascularised fibular bone grafting for the treatment of humeral bone loss secondary to osteomyelitis. Upper limb skeletal bone loss due to osteomyelitis is a devastating and challenging complication to manage for both surgeon and patient. Patients can be left with life altering disability and functional impairment. This limb threatening complication raises the question of salvage versus amputation and the associated risk and benefits of each. Free vascularised fibula grafting is a recognised treatment option for large skeletal defects in long bones but is not without significant risk. The benefit of vascularised over non-vascularised fibula grafts include preservation of blood supply lending itself to improved remodeling and osteointegration.Materials &amp; MethodsSixteen patients in Northern Ireland had free vascularised fibula grafting. Inclusion criteria included grafting to humeral defects secondary to osteomyelitis. Six patients were included in this study. Patients were contacted to complete DASH (Disabilities of the Arm, Shoulder and Hand) questionnaires as our primary outcome measure. Secondary outcome measures included radiological evaluation of osteointegration and associated operative complications. Complications were assessed via review of Electronic Care Record outpatient and in-patient documents.ResultsSix patients in Northern Ireland have been treated with free fibula grafting for humeral defects secondary to osteomyelitis. Regarding aetiology of osteomyelitis, five patients had sustained humeral fractures which were initially managed with open reduction internal fixation and complicated by infected non-union. These cases required removal of metalwork, segmental resection and placement of cement spacer with prolonged antibiotic therapy before return to theatre for vascularised fibula grafting. One patient developed native humeral osteomyelitis, the exact cause of which was unclear. Four patients were contactable for outcome score completion. The average DASH score was 45.5. (range 11.1 – 70.4). The median score was 50.2. With regard to post-operative complications, four of the six patients required return to theatre. Reasons were flap congestion, failure of metalwork at three months, donor site skin grafting for delayed healing and external fixation of graft fracture following trauma. Three patients experienced donor site morbidity in the form of delayed healing or infection requiring admission for intravenous antibiotics. One patient died due to cerebrovascular accident (CVA) within one month of return to theatre for a graft related complication - radiographic osteointegration was not assessed for this patient. At the time of this study, three of the remaining five patients have achieved graft osteointegration. Two patients are within 6 months of grafting procedure therefore too early to comment on radiographic osteoingreration. Radiographs to date are encouraging.ConclusionsFree vascularised fibula grafting for humeral shaft bone defects is a viable option for limb salvage but carries significant risk of complications. Our study highlights the need for proper informed consent regarding risk for this uncommon complex procedure which should only be undertaken by a multidisciplinary team in a specialist orthoplastic unit. This procedure is performed infrequently and our study has given us more information on risk and clinical outcomes to provide full and informed consent for patients in the future.

The genetic landscape and clinical spectrum of nephronophthisis and related ciliopathies

Nephronophthisis (NPH) is an autosomal-recessive ciliopathy representing one of the most frequent causes of kidney failure in childhood characterized by a broad clinical and genetic heterogeneity. Applied to one of the worldwide largest cohorts of patients with NPH, genetic analysis encompassing targeted and whole exome sequencing identified disease-causing variants in 600 patients from 496 families with a detection rate of 71%. Of 788 pathogenic variants, 40 known ciliopathy genes were identified. However, the majority of patients (53%) bore biallelic pathogenic variants in NPHP1. NPH-causing gene alterations affected all ciliary modules defined by structural and/or functional subdomains. Seventy six percent of these patients had progressed to kidney failure, of which 18% had an infantile form (under five years) and harbored variants affecting the Inversin compartment or intraflagellar transport complex A. Forty eight percent of patients showed a juvenile (5-15 years) and 34% a late-onset disease (over 15 years), the latter mostly carrying variants belonging to the Transition Zone module. Furthermore, while more than 85% of patients with an infantile form presented with extra-kidney manifestations, it only concerned half of juvenile and late onset cases. Eye involvement represented a predominant feature, followed by cerebellar hypoplasia and other brain abnormalities, liver and skeletal defects. The phenotypic variability was in a large part associated with mutation types, genes and corresponding ciliary modules with hypomorphic variants in ciliary genes playing a role in early steps of ciliogenesis associated with juvenile-to-late onset NPH forms. Thus, our data confirm a considerable proportion of late-onset NPH suggesting an underdiagnosis in adult chronic kidney disease.

Multiomics of Bohring-Opitz syndrome truncating ASXL1 mutations identify canonical and noncanonical Wnt signaling dysregulation.

ASXL1 (additional sex combs-like 1) plays key roles in epigenetic regulation of early developmental gene expression. De novo protein-truncating mutations in ASXL1 cause Bohring-Opitz syndrome (BOS; OMIM #605039), a rare neurodevelopmental condition characterized by severe intellectual disabilities, distinctive facial features, hypertrichosis, increased risk of Wilms tumor, and variable congenital anomalies, including heart defects and severe skeletal defects giving rise to a typical BOS posture. These BOS-causing ASXL1 variants are also high-prevalence somatic driver mutations in acute myeloid leukemia. We used primary cells from individuals with BOS (n = 18) and controls (n = 49) to dissect gene regulatory changes caused by ASXL1 mutations using comprehensive multiomics assays for chromatin accessibility (ATAC-seq), DNA methylation, histone methylation binding, and transcriptome in peripheral blood and skin fibroblasts. Our data show that regardless of cell type, ASXL1 mutations drive strong cross-tissue effects that disrupt multiple layers of the epigenome. The data showed a broad activation of canonical Wnt signaling at the transcriptional and protein levels and upregulation of VANGL2, which encodes a planar cell polarity pathway protein that acts through noncanonical Wnt signaling to direct tissue patterning and cell migration. This multiomics approach identifies the core impact of ASXL1 mutations and therapeutic targets for BOS and myeloid leukemias.

Two‐dimensional materials for bone‐tissue engineering

AbstractThere are diverse diseases such as some infections, trauma, and tumor resections during cancer surgery that can cause bone damage or skeletal defects in persons. Most of the time, these defects cannot heal spontaneously due to several medical conditions that patients encounter, like diabetes, hormone‐related problems, and autoimmune disorders. This issue is even worse for older people and some special treatments should be provided for them. Bone‐tissue engineering has emerged to tackle these challenges. By investigating bone repair strategies, studying bone structures and biomechanics, and employing appropriate growth factors, suitable scaffolds, and biomaterial‐centered regenerative approaches can be employed to treat bone defects more effectively. This study reviews some recent bone‐tissue‐engineering strategies relying on two‐dimensional (2D) materials, including graphene and its derivatives, black phosphorus, and MXenes that are exhibiting a great potential in regenerative medicine.

Stress Trajectory Variations During Occlusal Loading in Human Skull with a Maxillofacial Defect: A Finite Element Analysis.

Biting forces from the teeth are distributed to the facial bones and to the skull through the stress trajectories. The presence of a bony defect in either the maxilla or mandible might lead to variations in the stress distribution. The aim of this study was to evaluate the stress distribution and variations in stress trajectories from biting forces in a human skull with maxillofacial defect using a finite element (FE) model. In this study, a cone beam computed tomography (CBCT) of an adult male patient with a maxillectomy defect consequent to surgical treatment of mucormycosis is evaluated for the stress distribution from the biting forces of the remaining posterior teeth. Finite element model without the mandible was constructed from the patient's CBCT data. Occlusal loading forces of 600N were applied on each side of the maxillary teeth. Stress trajectories were visualised through the stress distribution pattern. The results showed deviations in the normal stress distribution during occlusal loading and variations in zygomatic and pterygoid stress trajectories in the maxillofacial and skull regions in our FE model due to the bony defect. We conclude that a skeletal maxillofacial defect should be reconstructed to resume proper stress distribution during functional forces to maintain a healthy craniofacial skeleton.

The non-canonical Wnt receptor Ror2 is required for cartilage cell polarity and morphogenesis of the craniofacial skeleton in zebrafish.

Non-canonical/β-catenin-independent Wnt signaling plays crucial roles in tissue/cell polarity in epithelia, but its functions have been less well studied in mesenchymal tissues, such as the skeleton. Mutations in non-canonical Wnt signaling pathway genes cause human skeletal diseases such as Robinow syndrome and Brachydactyly Type B1, which disrupt bone growth throughout the endochondral skeleton. Ror2 is one of several non-canonical Wnt receptor/co-receptors. Here, we show that ror2-/- mutant zebrafish have craniofacial skeletal defects, including disruptions of chondrocyte polarity. ror1-/- mutants appear to be phenotypically wild type, but loss of both ror1 and ror2 leads to more severe cartilage defects, indicating partial redundancy. Skeletal defects in ror1/2 double mutants resemble those of wnt5b-/- mutants, suggesting that Wnt5b is the primary Ror ligand in zebrafish. Surprisingly, the proline-rich domain of Ror2, but not its kinase domain, is required to rescue its function in mosaic transgenic experiments in ror2-/- mutants. These results suggest that endochondral bone defects in ROR-related human syndromes reflect defects in cartilage polarity and morphogenesis.

Toxic effects of 70% ethanol extract of Moringa stenopetala leaf (Baker f.) Cufod. (Moringaceae) on fetus and placenta of pregnant Wistar rats

BackgroundMoringa stenopetala leaves (Baker f.) Cufod. (Moringaceae) are used as a staple food and traditional medicine for treating various diseases like malaria, hypertension, stomach pain, diabetes, elevated cholesterol, and removing the retained placenta. Its prenatal toxicity study is minimal. Thus, this study aimed to assess the toxic effects of a 70% ethanol extract of Moringa stenopetala leaf on the fetuses and placentas of pregnant Wistar rats.MethodFresh leaves of Moringa stenopetala were collected, dried at room temperature, ground to powder, and extracted using 70% ethanol. For this study, five groups of animals, each containing ten pregnant rats, were used. Groups I–III were experimental groups and treated with 250, 500, and 1000 mg/kg body weight of Moringa stenopetala leaf extract, respectively. Groups IV and V were pair-fed and ad libitum control groups. The extract was given during gestation days 6 to 12. The fetuses were recovered at day 20 of gestation and examined for the presence of developmental delays, gross external malformations, skeletal and visceral defects. Gross and histopathological changes in the placenta were also evaluated.ResultsCompared to the pair-fed control group, maternal daily food intake and weight gain were reduced in the 1000 mg/kg-treated group during the treatment and post-treatment periods. A significantly higher number of fetal resorptions was also seen in the 1000 mg/kg treatment group. The crown-rump length and fetal and placental weights were all significantly reduced in pregnant rats given 1000 mg/kg. However, there were no visible malformations in the visceral organs as well as external genitalia in all the treatment and control groups. About 40.7% of the fetuses in the 1000 mg/kg treated rats had no proximal hindlimb phalanges. In addition, light microscopic investigations of the placenta in the high-dose treated rats revealed structural changes in the decidual basalis, trophoblastic zone, and labyrinthine zones.ConclusionIn conclusion, consumption of M. stenopetala leaves at a higher dose may have toxic effects on the development of rat fetuses. At a higher dose, the plant extract increased the number of fetal resorptions, reduced the number of fetuses, decreased the fetal and placental weights, and alter the placental histopathology. Thus, it is recommended to limit the excess feeding of M. stenopetala leaves during gestation.

A Rare Case of Cutis Aplasia Congenita in a Newborn

A Rare Case of Cutis Aplasia Congenita in a Newborn

Carbonic anhydrase II deficiency.

Carbonic anhydrase II deficiency (OMIM # 259730), initially called "osteopetrosis with renal tubular acidosis and cerebral calcification syndrome", reveals an important role for the enzyme carbonic anhydrase II (CA II) in osteoclast and renal tubule function. Discovered in 1972 and subsequently given various names, CA II deficiency now describes >100 affected individuals encountered predominantly from the Middle East and Mediterranean region. In 1983, CA II deficiency emerged as the first osteopetrosis (OPT) understood metabolically, and in 1991 the first understood molecularly. CA II deficiency is the paradigm OPT featuring failure of osteoclasts to resorb bone due to inability to acidify their pericellular milieu. The disorder presents late in infancy or early in childhood with fracturing, developmental delay, weakness, short stature, and/or cranial nerve compression and palsy. Mental retardation is common. The skeletal findings may improve by adult life, and CA II deficiency can be associated with a normal life-span. Therefore, it has been considered an "intermediate" type of OPT. In CA II deficiency, OPT is uniquely accompanied by renal tubular acidosis (RTA) of proximal, distal, or combined type featuring hyperchloremic metabolic acidosis, rarely with hypokalemia and paralysis. Cerebral calcification uniquely appears in early childhood. The etiology is bi-allelic loss-of-function mutations of CA2 that encodes CA II. Prenatal diagnosis requires mutational analysis of CA2. Although this enzymopathy reveals how CA II is important for the skeleton and kidney tubule, the pathogenesis of the mental subnormality and cerebral calcification is less well understood. Several mouse models of CA II deficiency have shown growth hormone deficiency, yet currently there is no standard pharmacologic therapy for patients. Treatment of the systemic acidosis is often begun when growth is complete. Although CA II deficiency is an "osteoclast-rich" OPT, and therefore transplantation of healthy osteoclasts can improve the skeletal disease, the RTA and central nervous system difficulties persist.